Clinical Significances of Lipoprotein Metabolism
Fortunately, few individuals carry the inherited defects in lipoprotein metabolism that lead to hyper- or hypolipoproteinemias (see Tables below for brief descriptions). Persons suffering from diabetes mellitus, hypothyroidism and kidney disease often exhibit abnormal lipoprotein metabolism as a result of secondary effects of their disorders. For example, because lipoprotein lipase (LPL) synthesis is regulated by insulin, LPL deficiencies leading to Type I hyperlipoproteinemia may occur as a secondary outcome of diabetes mellitus. Additionally, insulin and thyroid hormones positively affect hepatic LDL-receptor interactions; therefore, the hypercholesterolemia and increased risk of athersclerosis associated with uncontrolled diabetes or hypothyroidism is likely due to decreased hepatic LDL uptake and metabolism.
Of the many disorders of lipoprotein metabolism, familial hypercholesterolemia (FH) may be the most prevalent in the general population. Heterozygosity at the FH locus occurs in 1:500 individuals, whereas, homozygosity is observed in 1:1,000,000 individuals. FH is an inherited disorder comprising four different classes of mutation in the LDL receptor gene. The class 1 defect (the most common) results in a complete loss of receptor synthesis. The class 2 defect results in the synthesis of a receptor protein that is not properly processed in the Golgi apparatus and therefore is not transported to the plasma membrane. The class 3 defect results in an LDL receptor that is incapable of binding LDLs. The class 4 defect results in receptors that bind LDLs but do not cluster in coated pits and are, therefore, not internalized.
FH sufferers may be either heterozygous or homologous for a particular mutation in the receptor gene. Homozygotes exhibit grossly elevated serum cholesterol (primarily in LDLs). The elevated levels of LDLs result in their phagocytosis by macrophages. These lipid-laden phagocytic cells tend to deposit within the skin and tendons, leading to xanthomas. A greater complication results from cholesterol deposition within the arteries, leading to atherosclerosis, the major contributing factor of nearly all cardiovascular diseases.
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Hyperlipoproteinemias
Disorder
Defect
Comments
Type I
(familial LPL deficiency, familial hyperchylomicronemia) (a) deficiency of LPL;
(b) production of abnormal LPL;
(c) apoC-II deficiency slow chylomicron clearance, reduced LDL and HDL levels; treated by low fat/complex carbohydrate diet; no increased risk of coronary artery disease
Type II
(familial hypercholesterolemia, FH) 4 classes of LDL receptor defect reduced LDL clearance leads to hypercholesterolemia, resulting in athersclerosis and coronary artery disease
Type III
(familial dysbetalipoproteinemia, remnant removal disease, broad beta disease, apolipoprotein E deficiency) hepatic remnant clearance impaired due to apoE abnormality; patients only express the apoE2 isoform that interacts poorly with the apoE receptor causes xanthomas, hypercholesterolemia and athersclerosis in peripheral and coronary arteries due to elevated levels of chylomicrons and VLDLs
Type IV
(familial hypertriacylglycerolemia) elevated production of VLDL associated with glucose intolerance and hyperinsulinemia frequently associated with type-II non-insulin dependent diabetes mellitus, obesity, alcoholism or administration of progestational hormones; elevated cholesterol as a result of increased VLDLs
Type V familial elevated chylomicrons and VLDLs due to unknown cause hypertriacylglycerolemia and hypercholesterolemia with decreased LDLs and HDLs
Familial hyperalphalipoproteinemia increased level of HDLs a rare condition that is beneficial for health and longevity
Type II
Familial hyperbetalipoproteinemia increased LDL production and delayed clearance of triacylglycerols and fatty acids strongly associated with increased risk of coronary artery disease
Familial ligand-defective apoB 2 different mutations: Gln for Arg (amino acid 3500) or Cys for Arg (amino acid 3531); both lead to reduced affinity of LDL for LDL receptor dramatic increase in LDL levels; no affect on HDL, VLDL or plasma triglyceride levels; significant cause of hypercholesterolemia and premature coronary artery disease
Familial LCAT deficiency absence of LCAT leads to inability of HDLs to take up cholesterol
(reverse cholesterol transport) decreased levels of plasma cholesteryl esters and lysolecithin; abnormal LDLs (Lp-X) and VLDLs; symptoms also found associated with cholestasis
Wolman disease
(cholesteryl ester storage disease) defect in lysosomal cholesteryl ester hydrolase; affects metabolism of LDLs reduced LDL clearance leads to hypercholesterolemia, resulting in athersclerosis and coronary artery disease
heparin-releasable hepatic triglyceride lipase deficiency deficiency of the lipase leads to accumulation of triacylglycerol-rich HDLs and VLDL remnants (IDLs) causes xanthomas and coronary artery disease
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Hypolipoproteinemias
Disorder Defect Comments
Abetalipoproteinemia
(acanthocytosis, Bassen-Kornzweig syndrome) no chylomicrons, VLDLs or LDLs due to defect in apoB expression rare defect; intestine and liver accumulate, malabsorption of fat, retinitis pigmentosa, ataxic neuropathic disease, erythrocytes have thorny appearance
Familial hypobetalipoproteinemia at least 20 different apoB gene mutations identified, LDL concentrations 10-20% of normal, VLDL slightly lower, HDL normal mild or no pathological changes
Familial alpha-lipoprotein deficiency
(Tangier disease, Fish-eye disease, apoA-I and -C-III deficiencies) all of these related syndromes have reduced HDL concentrations, no effect on chylomicron or VLDL production tendency to hypertriacylglycerolemia; some elevation in VLDLs; Fish-eye disease characterized by severe corneal opacity
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Pharmacologic Intervention
Drug treatment to lower plasma lipoproteins and/or cholesterol is primarily aimed at reducing the risk of athersclerosis and subsequent coronary artery disease that exists in patients with elevated circulating lipids. Drug therapy usually is considered as an option only if non-pharmacologic interventions (altered diet and exercise) have failed to lower plasma lipids.
Atorvastatin (Lipotor®), Simvastatin (Zocor®), Lovastatin (Mevacor®): These drugs are fungal HMG-CoA reductase (HMGR) inhibitors and are members of the family of drugs referred to as the statins. The net result of treatment is an increased cellular uptake of LDLs, since the intracellular synthesis of cholesterol is inhibited and cells are therefore dependent on extracellular sources of cholesterol. However, since mevalonate (the product of the HMG-CoA reductase reaction) is required for the synthesis of other important isoprenoid compounds besides cholesterol, long-term treatments carry some risk of toxicity.
The statins have become recognized as a class of drugs capable of more pharmacologic benefits than just lowering blood cholesterol levels via their actions on HMGR. Part of the cardiac benefit of the statins relates to their ability to regulate the production of S-nitrosylated COX-2. COX-2 is an inducible enzyme involved in the synthesis of the prostaglandins and thromboxanes as well as the lipoxins and resolvins. The latter two classes of compounds are anti-inflammatory lipids discussed in the Aspirin page. Evidence has shown that statins activate inducible nitric oxide synthase (iNOS) leading to nitrosylation of COX-2. The S-nitrosylated COX-2 enzyme produces the lipid compound 15R-hydroxyeicosatetraenoic acid (15R-HETE) which is then converted via the action of 5-lipoxygenase (5-LOX) to the epimeric lipoxin, 15-epi-LXA4. This latter compound is the same as the aspirin-triggered lipoxin (ATL) that results from the aspirin-induced acetylation of COX-2. Therefore, part of the beneficial effects of the statins are exerted via the actions of the lipoxin family of anti-inflammatory lipids.
Nicotinic acid: Nicotinic acid reduces the plasma levels of both VLDLs and LDLs by inhibiting hepatic VLDL secretion, as well as suppressing the flux of FFA release from adipose tissue by inhibiting lipolysis. Because of its ability to cause large reductions in circulating levels of cholesterol, nicotinic acid is used to treat Type II, III, IV and V hyperlipoproteinemias.
Gemfibrozil (Lopid®), Fenofibrate (TriCor®): These compounds (called fibrates) are derivatives of fibric acid and although used clinically since the 1930's were only recently discovered to exert some of their lipid-lowering effects via the activation of peroxisome proliferation. Specifically, the fibrates were found to be activators of the peroxisome proliferator-activated receptor-α (PPAR-α) class of proteins that are classified as co-activators. The naturally occurring ligands for PPAR-α are leukotriene B4 (LTB4, see the Lipid Synthesis page), unsaturated fatty acids and oxidized components of VLDLs and LDLs. The PPARs interact with another receptor family called the retinoid X receptors (RXRs) that bind 9-cis-retinoic acid. Activation of PPARs results in modulation of the expression of genes involved in lipid metabolism. In addition the PPARs modulate carbohydrate metabolism and adipose tissue differentiation. Fibrates result in the activation of PPAR-α in liver and muscle. In the liver this leads to increased β-oxidation of fatty acids, thereby decreasing the liver's secretion of triacylglycerol- and cholesterol-rich VLDLs, as well as increased clearance of chylomicron remnants, increased levels of HDLs and increased lipoprotein lipase activity which in turn promotes rapid VLDL turnover.
Cholestyramine or colestipol (resins): These compounds are nonabsorbable resins that bind bile acids which are then not reabsorbed by the liver but excreted. The drop in hepatic reabsorption of bile acids releases a feedback inhibitory mechanism that had been inhibiting bile acid synthesis. As a result, a greater amount of cholesterol is converted to bile acids to maintain a steady level in circulation. Additionally, the synthesis of LDL receptors increases to allow increased cholesterol uptake for bile acid synthesis, and the overall effect is a reduction in plasma cholesterol. This treatment is ineffective in homozygous FH patients, since they are completely deficient in LDL receptors.
Tuesday, February 17, 2009
serum cholestrol values
Lipid Profile Values
Standard fasting blood tests for cholesterol and lipid profiles will include values for total cholesterol, HDL cholesterol (so-called "good" cholesterol), LDL cholesterol (so-called "bad" cholesterol) and triglycerides. Family history and life style, including factors such as blood pressure and whether or not one smokes, affect what would be considered ideal versus non-ideal values for fasting blood lipid profiles. Included here are the values for various lipids that indicate low to high risk for coronary artery disease.
Total Serum Cholesterol
<200mg/dL = desired values
200 - 239mg/dL = borderline to high risk
240mg/dL and above = high risk
HDL Cholesterol
With HDL cholesterol the higher the better.
<40mg/dL for men and <50mg/dL for women = higher risk
40-50mg/dL for men and 50-60mg/dL for woman = normal values
>60mg/dL is associated with some level of protection against heart disease
LDL Cholesterol
With LDL cholesterol the lower the better.
<100mg/dL = optimal values
100mg/dL - 129mg/dL = optimal to near optimal
130mg/dL - 159mg/dL = borderline high risk
160mg/dL - 189mg/dL = high risk
190mg/dL and higher = very high risk
Triglycerides
With triglycerides the lower the better.
<150mg/dL = normal
150mg/dL - 199mg/dL = borderline to high risk
200mg/dL - 499mg/dL = high risk
>500mg/dL = very high risk
Standard fasting blood tests for cholesterol and lipid profiles will include values for total cholesterol, HDL cholesterol (so-called "good" cholesterol), LDL cholesterol (so-called "bad" cholesterol) and triglycerides. Family history and life style, including factors such as blood pressure and whether or not one smokes, affect what would be considered ideal versus non-ideal values for fasting blood lipid profiles. Included here are the values for various lipids that indicate low to high risk for coronary artery disease.
Total Serum Cholesterol
<200mg/dL = desired values
200 - 239mg/dL = borderline to high risk
240mg/dL and above = high risk
HDL Cholesterol
With HDL cholesterol the higher the better.
<40mg/dL for men and <50mg/dL for women = higher risk
40-50mg/dL for men and 50-60mg/dL for woman = normal values
>60mg/dL is associated with some level of protection against heart disease
LDL Cholesterol
With LDL cholesterol the lower the better.
<100mg/dL = optimal values
100mg/dL - 129mg/dL = optimal to near optimal
130mg/dL - 159mg/dL = borderline high risk
160mg/dL - 189mg/dL = high risk
190mg/dL and higher = very high risk
Triglycerides
With triglycerides the lower the better.
<150mg/dL = normal
150mg/dL - 199mg/dL = borderline to high risk
200mg/dL - 499mg/dL = high risk
>500mg/dL = very high risk
clinical significance of bile salts
Clinical Significance of Bile Acid Synthesis
Bile acids perform four primary physiologically significant functions:
1. their synthesis and subsequent excretion in the feces represent the only significant mechanism for the elimination of excess cholesterol.
2. bile acids and phospholipids solubilize cholesterol in the bile, thereby preventing the precipitation of cholesterol in the gallbladder.
3. they facilitate the digestion of dietary triacylglycerols by acting as emulsifying agents that render fats accessible to pancreatic lipases.
4. they facilitate the intestinal absorption of fat-soluble vitamins.
Over the past several years new insights into the biological activities of the bile acids have been elucidated. Following the isolation and characterization of the farnesoid X receptors (FXRs, see above), for which the bile acids are physiological ligands, the functions of bile acids in the regulation of lipid and glucose homeostasis has begun to emerge. As indicated above, the binding of bile acids to FXRs results in the attenuated expression of several genes involved in overall bile acid homeostasis. However, genes involved in bile acid metabolism are not the only ones that are regulated by FXR action as a consequence of binding bile acid. In the liver, FXR action is known to regulate the expression of genes involved in lipoprotein metabolism (e.g. apoC-II), glucose metabolism (e.g. PEPCK), and hepatoprotection (e.g. CYP3A4, which was originally identified as nifedipine oxidase; nifedipine being a member of the calcium channel blocker drugs). Thus, it is becoming clear that bile acids serve functions not only as intestinal lipid emulsifiers but as significant participants in numerous biochemical and physiological processes.
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Bile acids perform four primary physiologically significant functions:
1. their synthesis and subsequent excretion in the feces represent the only significant mechanism for the elimination of excess cholesterol.
2. bile acids and phospholipids solubilize cholesterol in the bile, thereby preventing the precipitation of cholesterol in the gallbladder.
3. they facilitate the digestion of dietary triacylglycerols by acting as emulsifying agents that render fats accessible to pancreatic lipases.
4. they facilitate the intestinal absorption of fat-soluble vitamins.
Over the past several years new insights into the biological activities of the bile acids have been elucidated. Following the isolation and characterization of the farnesoid X receptors (FXRs, see above), for which the bile acids are physiological ligands, the functions of bile acids in the regulation of lipid and glucose homeostasis has begun to emerge. As indicated above, the binding of bile acids to FXRs results in the attenuated expression of several genes involved in overall bile acid homeostasis. However, genes involved in bile acid metabolism are not the only ones that are regulated by FXR action as a consequence of binding bile acid. In the liver, FXR action is known to regulate the expression of genes involved in lipoprotein metabolism (e.g. apoC-II), glucose metabolism (e.g. PEPCK), and hepatoprotection (e.g. CYP3A4, which was originally identified as nifedipine oxidase; nifedipine being a member of the calcium channel blocker drugs). Thus, it is becoming clear that bile acids serve functions not only as intestinal lipid emulsifiers but as significant participants in numerous biochemical and physiological processes.
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vertibrate hormones
Common Vertebrate Hormones
Hormone
Structure
Functions
Pituitary Hormones
Oxytocin polypeptide of 9 amino acids CYIQNCPLG (C's are disulfide bonded) uterine contraction, causes milk ejection in lactating females, responds to suckling reflex and estradiol, lowers steroid synthesis in testes
Vasopressin
(antidiuretic hormone, ADH) polypeptide of 9 amino acids CYFQNCPRG (C's are disulfide bonded) responds to osmoreceptor which senses extracellular [Na+], blood pressure regulation, increases H2O readsorption from distal tubules in kidney
Melanocyte-stimulating hormones (MSH) α polypeptide = 13 amino acids
β polypeptide = 18 amino acids
γ polypeptide = 12 amino acids pigmentation
Corticotropin (adrenocorticotropin, ACTH) polypeptide = 39 amino acids stimulates cells of adrenal gland to increase steroid synthesis and secretion
Lipotropin (LPH) β polypeptide = 93 amino acids
γ polypeptide = 60 amino acids increases fatty acid release from adipocytes
Thyrotropin (thyroid-stimulating hormone, TSH) 2 proteins: α is 96 amino acids; β is 112 acts on thyroid follicle cells to stimulate throid hormone synthesis
Growth hormone (GH, or somatotropin) protein of 191 amino acids general anabolic stimulant, increases release of insulin-like growth factor-I (IGF-I), cell growth and bone sulfation
Prolactin (PRL) protein of 197 amino acids stimulates differentiation of secretory cells of mammary gland and stimulates milk synthesis
Luteinizing hormone (LH); human chorionic gonadotropin (hCG) is similar and produced in placenta 2 proteins: α is 96 amino acids; β is 121 increases ovarian progesterone synthesis, luteinization; acts on Leydig cells of testes to increase testosterone synthesis and release and increases interstitial cell development
Follicle-stimulating hormone (FSH) 2 proteins: α is 96 amino acids; β is 120 ovarian follicle development and ovulation, increases estrogen production; acts on Sertoli cells of semiferous tubule to increase spermatogenesis
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Hypothalamic Hormones
Corticotropin-releasing factor (CRF or CRH) protein of 41 amino acids acts on corticotrope to release ACTH and β-endorphin (lipotropin)
Gonadotropin-releasing factor (GnRF or GnRH) polypeptide of 10 amino acids acts on gonadotrope to release LH and FSH
Prolactin-releasing factor (PRF) this may be TRH acts on lactotrope to release prolactin
Prolactin-release inhibiting factor (PIF) may be derived from GnRH precursor, 56 amino acids acts on lactotrope to inhibit prolactin release
Growth hormone-releasing factor (GRF or GRH) protein of 40 and 44 amino acids stimulates GH secretion
Somatostatin (SIF, also called growth hormone-release inhibiting factor, GIF) polypeptide of 14 and 28 amino acids inhibits GH and TSH secretion
Thyrotropin-releasing factor (TRH or TRF) peptide of 3 amino acids: EHP stimulates TSH and prolactin secretion
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Thyroid Hormones
Thyroxine and triiodothyronine iodinated dityrosin derivatives responds to TSH and stimulates oxidations in many cells
Calcitonin protein of 32 amino acids produced in parafollicular C cells of the thyroid, regulation of Ca2+ and Pi metabolism
Calcitonin gene-related peptide (CGRP) protein of 37 amino acids, product of the calcitonin gene derived by alternative splicing of the precursor mRNA in the brain acts as a vasodilator
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Parathyroid Hormone
Parathyroid hormone (PTH) protein of 84 amino acids regulation of Ca2+ and Pi metabolism, stimulates bone resorption thus increasing serum [Ca2+], stimulates Pi secretion by kidneys
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Adipose Tissue Hormones
additional discussion of adipocyte hormones and cytokines
Leptin 167 amino acid precursor processed to 146 amino acids regulation of overall body weight by limiting food intake and increasing energy expenditure, regulation of the neuroendocrine axis, inflammatory responses, blood pressure, and bone mass
Adiponectin 244 amino acid protein with 4 distinct functional domains major biological actions are increases in insulin sensitivity and fatty acid oxidation
Resistin 108 amino acid pre-protein in humans induces insulin resistance
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Hormones and Peptides of the Gut
additional discussion of gastrointestinal hormones
Glucagon-like peptide 1 (GLP-1) formerly called enteroglucagon Two forms: 31 amino acids, GLP-1(7-37) and 30 amino acids, GLP-1(7-36)amide potentiates glucose-dependent insulin secretion, inhibits glucagon secretion, inhibits gastric emptying
Glucose-dependent insulinotropic polypeptide (GIP) originally called gastric inhibitory polypeptide polypeptide of 42 amino acids inhibits secretion of gastric acid, enhances insulin secretion
Ghrelin 28 amino acids derived from preproghrelin protein; acylated on Ser3 with n-octanoic acid, non-acylated forms found in circulation also but not bioactive appetite stimulation, stimulates NPY release, regulation of energy homeostasis, glucose metabolism, gastric secretion and emptying, insulin secretion
Obestatin 23 amino acids derived from preproghrelin protein acts in opposition to ghrelin action on appetite
Gastrin 17 amino acids produced by stomach antrum, stimulates acid and pepsin secretion, also stimulates pancreatic secretions
Secretin 27 amino acids secreted from duodenum at pH values below 4.5, stimulates pancreatic acinar cells to release bicarbonate and H2O
Cholecystokinin, CCK 33 amino acids stimulates gallbladder contraction and bile flow, increases secretion of digestive enzymes from pancreas
Motilin 22 amino acids controls gastrointestinal muscles
Vasoactive intestinal peptide (VIP) 28 amino acids produced by hypothalamus and GI tract, relaxes the GI, inhibits acid and pepsin secretion, acts as a neurotransmitter in peripheral autonomic nervous system, increases secretion of H2O and electrolytes from pancreas and gut
Somatostatin 14 amino acid version inhibits release and action of numerous gut peptides, e.g. CKK, gastrin, secretin, motilin, GIP; also inhibits insulin and glucagon secretion from pancreas
Substance P, a member of the tachykinin family that includes neurokinin A (NKA) and neurokinin B (NKB) 11 amino acids CNS function in pain (nociception), involved in vomit reflex, stimulates salivary secretions, induces vasodilation
antagonists have anti-depressant properties
PP, PYY and NPY constitute the Pancreatic Polypeptide family of 36 amino acid peptides.
PP and PYY exhibit endocrine functions.
NPY functions as a neuropeptide
Pancreatic Polypeptide, PP 36 amino acids suppresses glucose-induced insulin secretion, inhibits bicarbonate and protein secretion from pancreas
Peptide Tyrosine Tyrosine, PYY 36 amino acids inhibits gastric motility by inhibiting cholinergic neurotransmission, inhibits gastric acid secretion
Neuropeptide Tyrosine, NPY 36 amino acids, 6 receptors effects on hypothalamic function in appetite, controls feeding behavior and energy homeostasis, levels increase during starvation to induce food intake
Amphiregulin 2 peptides: 78 amino acid truncated form and 84 amino acid form with 6 additional N-terminal amino acids homology to EGF and binds to the EGF receptor (EGFR)
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Pancreatic Hormones
Insulin disulfide bonded dipeptide of 21 and 30 amino acids produced by β-cells of the pancreas, increases glucose uptake and utilization, increases lipogenesis, general anabolic effects
Glucagon polypeptide of 29 amino acids produced by α-cells of the pancreas, increases lipid mobilization and glycogenolysis in order to increase blood glucose levels
Pancreatic polypeptide polypeptide of 36 amino acids increases glycogenolysis, regulation of gastrointestinal activity
Somatostatin 14 amino acid version inhibition of glucagon and somatotropin release
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Placental Hormones
Estrogens steroids maintenance of pregnancy
Progestins steroids mimic action of progesterone
Chorionic gonadotropin 2 proteins: α is 96 amino acids; β is 147 activity similar to LH
Placental lactogen protein of 191 amino acids acts like prolactin and GH
Relaxin 2 proteins of 22 and 32 amino acids produced in ovarian corpus luteum, inhibits myometrial contractions, secretion increases durin gestation
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Gonadal Hormones
Estrogens (ovarian) steroids: estradiol and estrone maturation and function of female secondary sex organs
Progestins (ovarian) steroid: progesterone implantation of ovum and maintenance of pregnancy
Androgens (testicular) steroid: testosterone maturation and function of male secondary sex organs
Inhibins A and B 1 protein (α is 134 amino acids; β is 115 and 116 amino acids) inhibition of FSH secretion
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Adrenal Cortical Hormones
Glucocorticoids steroids: cortisol and corticosterone diverse effects on inflammation and protein synthesis
Mineralocorticoids steroids: aldosterone maintenance of salt balance
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Adrenal Medullary Hormones
Epinephrine (adrenalin) derived from tyrosine classic "fight-or-flight" response, increases glycogenolysis, lipid mobilization, smooth muscle contraction, cardiac function, binds to all classes of catecholamine receptors (α- and β-adrenergic)
Norepinephrine (noradrenalin) derived from tyrosine classic "fight-or-flight" response, lipid mobilization, arteriole contraction, also acts as neurotransmitter in the CNS, released from noradrenergic neurons, binds all catecholamine receptors except β2-adrenergic
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Liver Hormones
Angiotensin II polypeptide of 8 amino acids derived from angiotensinogen (present in the α2-globulin fraction of plasma) which is cleaved by the kidney enzyme renin to give the decapeptide, angiotensin I, the C-terminal 2 amino acids are then released (by action of angiotensin-converting enzyme, ACE) to yield angiotensin II responsible for essential hypertension through stimulated synthesis and release of aldosterone from adrenal cells
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Kidney Hormones
Calcitriol [1,25-(OH)2-vitamin D3] derived from 7-dehydrocholesterol responsible for maintenance of calcium and phosphorous homeostasis, increases intestinal Ca2+ uptake, regulates bone mineralization
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Cardiac Hormones
Atrial natriuretic peptide (ANP) several active peptides cleaved from a 126 amino acid precursor released from heart atria in response to hypovolemia, acts on outer adrenal cells to decrease aldosterone production; smooth muscle relaxation
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Pineal Hormones
Melatonin N-acetyl-5-methoxytryptamine regulation of circadian rhythms
Hormone
Structure
Functions
Pituitary Hormones
Oxytocin polypeptide of 9 amino acids CYIQNCPLG (C's are disulfide bonded) uterine contraction, causes milk ejection in lactating females, responds to suckling reflex and estradiol, lowers steroid synthesis in testes
Vasopressin
(antidiuretic hormone, ADH) polypeptide of 9 amino acids CYFQNCPRG (C's are disulfide bonded) responds to osmoreceptor which senses extracellular [Na+], blood pressure regulation, increases H2O readsorption from distal tubules in kidney
Melanocyte-stimulating hormones (MSH) α polypeptide = 13 amino acids
β polypeptide = 18 amino acids
γ polypeptide = 12 amino acids pigmentation
Corticotropin (adrenocorticotropin, ACTH) polypeptide = 39 amino acids stimulates cells of adrenal gland to increase steroid synthesis and secretion
Lipotropin (LPH) β polypeptide = 93 amino acids
γ polypeptide = 60 amino acids increases fatty acid release from adipocytes
Thyrotropin (thyroid-stimulating hormone, TSH) 2 proteins: α is 96 amino acids; β is 112 acts on thyroid follicle cells to stimulate throid hormone synthesis
Growth hormone (GH, or somatotropin) protein of 191 amino acids general anabolic stimulant, increases release of insulin-like growth factor-I (IGF-I), cell growth and bone sulfation
Prolactin (PRL) protein of 197 amino acids stimulates differentiation of secretory cells of mammary gland and stimulates milk synthesis
Luteinizing hormone (LH); human chorionic gonadotropin (hCG) is similar and produced in placenta 2 proteins: α is 96 amino acids; β is 121 increases ovarian progesterone synthesis, luteinization; acts on Leydig cells of testes to increase testosterone synthesis and release and increases interstitial cell development
Follicle-stimulating hormone (FSH) 2 proteins: α is 96 amino acids; β is 120 ovarian follicle development and ovulation, increases estrogen production; acts on Sertoli cells of semiferous tubule to increase spermatogenesis
back to Table of Contents
Hypothalamic Hormones
Corticotropin-releasing factor (CRF or CRH) protein of 41 amino acids acts on corticotrope to release ACTH and β-endorphin (lipotropin)
Gonadotropin-releasing factor (GnRF or GnRH) polypeptide of 10 amino acids acts on gonadotrope to release LH and FSH
Prolactin-releasing factor (PRF) this may be TRH acts on lactotrope to release prolactin
Prolactin-release inhibiting factor (PIF) may be derived from GnRH precursor, 56 amino acids acts on lactotrope to inhibit prolactin release
Growth hormone-releasing factor (GRF or GRH) protein of 40 and 44 amino acids stimulates GH secretion
Somatostatin (SIF, also called growth hormone-release inhibiting factor, GIF) polypeptide of 14 and 28 amino acids inhibits GH and TSH secretion
Thyrotropin-releasing factor (TRH or TRF) peptide of 3 amino acids: EHP stimulates TSH and prolactin secretion
back to Table of Contents
Thyroid Hormones
Thyroxine and triiodothyronine iodinated dityrosin derivatives responds to TSH and stimulates oxidations in many cells
Calcitonin protein of 32 amino acids produced in parafollicular C cells of the thyroid, regulation of Ca2+ and Pi metabolism
Calcitonin gene-related peptide (CGRP) protein of 37 amino acids, product of the calcitonin gene derived by alternative splicing of the precursor mRNA in the brain acts as a vasodilator
back to Table of Contents
Parathyroid Hormone
Parathyroid hormone (PTH) protein of 84 amino acids regulation of Ca2+ and Pi metabolism, stimulates bone resorption thus increasing serum [Ca2+], stimulates Pi secretion by kidneys
back to Table of Contents
Adipose Tissue Hormones
additional discussion of adipocyte hormones and cytokines
Leptin 167 amino acid precursor processed to 146 amino acids regulation of overall body weight by limiting food intake and increasing energy expenditure, regulation of the neuroendocrine axis, inflammatory responses, blood pressure, and bone mass
Adiponectin 244 amino acid protein with 4 distinct functional domains major biological actions are increases in insulin sensitivity and fatty acid oxidation
Resistin 108 amino acid pre-protein in humans induces insulin resistance
back to Table of Contents
Hormones and Peptides of the Gut
additional discussion of gastrointestinal hormones
Glucagon-like peptide 1 (GLP-1) formerly called enteroglucagon Two forms: 31 amino acids, GLP-1(7-37) and 30 amino acids, GLP-1(7-36)amide potentiates glucose-dependent insulin secretion, inhibits glucagon secretion, inhibits gastric emptying
Glucose-dependent insulinotropic polypeptide (GIP) originally called gastric inhibitory polypeptide polypeptide of 42 amino acids inhibits secretion of gastric acid, enhances insulin secretion
Ghrelin 28 amino acids derived from preproghrelin protein; acylated on Ser3 with n-octanoic acid, non-acylated forms found in circulation also but not bioactive appetite stimulation, stimulates NPY release, regulation of energy homeostasis, glucose metabolism, gastric secretion and emptying, insulin secretion
Obestatin 23 amino acids derived from preproghrelin protein acts in opposition to ghrelin action on appetite
Gastrin 17 amino acids produced by stomach antrum, stimulates acid and pepsin secretion, also stimulates pancreatic secretions
Secretin 27 amino acids secreted from duodenum at pH values below 4.5, stimulates pancreatic acinar cells to release bicarbonate and H2O
Cholecystokinin, CCK 33 amino acids stimulates gallbladder contraction and bile flow, increases secretion of digestive enzymes from pancreas
Motilin 22 amino acids controls gastrointestinal muscles
Vasoactive intestinal peptide (VIP) 28 amino acids produced by hypothalamus and GI tract, relaxes the GI, inhibits acid and pepsin secretion, acts as a neurotransmitter in peripheral autonomic nervous system, increases secretion of H2O and electrolytes from pancreas and gut
Somatostatin 14 amino acid version inhibits release and action of numerous gut peptides, e.g. CKK, gastrin, secretin, motilin, GIP; also inhibits insulin and glucagon secretion from pancreas
Substance P, a member of the tachykinin family that includes neurokinin A (NKA) and neurokinin B (NKB) 11 amino acids CNS function in pain (nociception), involved in vomit reflex, stimulates salivary secretions, induces vasodilation
antagonists have anti-depressant properties
PP, PYY and NPY constitute the Pancreatic Polypeptide family of 36 amino acid peptides.
PP and PYY exhibit endocrine functions.
NPY functions as a neuropeptide
Pancreatic Polypeptide, PP 36 amino acids suppresses glucose-induced insulin secretion, inhibits bicarbonate and protein secretion from pancreas
Peptide Tyrosine Tyrosine, PYY 36 amino acids inhibits gastric motility by inhibiting cholinergic neurotransmission, inhibits gastric acid secretion
Neuropeptide Tyrosine, NPY 36 amino acids, 6 receptors effects on hypothalamic function in appetite, controls feeding behavior and energy homeostasis, levels increase during starvation to induce food intake
Amphiregulin 2 peptides: 78 amino acid truncated form and 84 amino acid form with 6 additional N-terminal amino acids homology to EGF and binds to the EGF receptor (EGFR)
back to Table of Contents
Pancreatic Hormones
Insulin disulfide bonded dipeptide of 21 and 30 amino acids produced by β-cells of the pancreas, increases glucose uptake and utilization, increases lipogenesis, general anabolic effects
Glucagon polypeptide of 29 amino acids produced by α-cells of the pancreas, increases lipid mobilization and glycogenolysis in order to increase blood glucose levels
Pancreatic polypeptide polypeptide of 36 amino acids increases glycogenolysis, regulation of gastrointestinal activity
Somatostatin 14 amino acid version inhibition of glucagon and somatotropin release
back to Table of Contents
Placental Hormones
Estrogens steroids maintenance of pregnancy
Progestins steroids mimic action of progesterone
Chorionic gonadotropin 2 proteins: α is 96 amino acids; β is 147 activity similar to LH
Placental lactogen protein of 191 amino acids acts like prolactin and GH
Relaxin 2 proteins of 22 and 32 amino acids produced in ovarian corpus luteum, inhibits myometrial contractions, secretion increases durin gestation
back to Table of Contents
Gonadal Hormones
Estrogens (ovarian) steroids: estradiol and estrone maturation and function of female secondary sex organs
Progestins (ovarian) steroid: progesterone implantation of ovum and maintenance of pregnancy
Androgens (testicular) steroid: testosterone maturation and function of male secondary sex organs
Inhibins A and B 1 protein (α is 134 amino acids; β is 115 and 116 amino acids) inhibition of FSH secretion
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Adrenal Cortical Hormones
Glucocorticoids steroids: cortisol and corticosterone diverse effects on inflammation and protein synthesis
Mineralocorticoids steroids: aldosterone maintenance of salt balance
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Adrenal Medullary Hormones
Epinephrine (adrenalin) derived from tyrosine classic "fight-or-flight" response, increases glycogenolysis, lipid mobilization, smooth muscle contraction, cardiac function, binds to all classes of catecholamine receptors (α- and β-adrenergic)
Norepinephrine (noradrenalin) derived from tyrosine classic "fight-or-flight" response, lipid mobilization, arteriole contraction, also acts as neurotransmitter in the CNS, released from noradrenergic neurons, binds all catecholamine receptors except β2-adrenergic
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Liver Hormones
Angiotensin II polypeptide of 8 amino acids derived from angiotensinogen (present in the α2-globulin fraction of plasma) which is cleaved by the kidney enzyme renin to give the decapeptide, angiotensin I, the C-terminal 2 amino acids are then released (by action of angiotensin-converting enzyme, ACE) to yield angiotensin II responsible for essential hypertension through stimulated synthesis and release of aldosterone from adrenal cells
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Kidney Hormones
Calcitriol [1,25-(OH)2-vitamin D3] derived from 7-dehydrocholesterol responsible for maintenance of calcium and phosphorous homeostasis, increases intestinal Ca2+ uptake, regulates bone mineralization
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Cardiac Hormones
Atrial natriuretic peptide (ANP) several active peptides cleaved from a 126 amino acid precursor released from heart atria in response to hypovolemia, acts on outer adrenal cells to decrease aldosterone production; smooth muscle relaxation
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Pineal Hormones
Melatonin N-acetyl-5-methoxytryptamine regulation of circadian rhythms
Diabetic ketoacidosis
Definition of Diabetic Ketoacidosis
The most severe and life threatening complication of poorly controlled type 1 diabetes is diabetic ketoacidosis (DKA). DKA is characterized by metabolic acidosis, hyperglycemia and hyperketonemia. Diagnosis of DKA is accomplished by detection of hyperketonemia and metabolic acidosis (as measured by anion gap) in the presence of hyperglycemia. The primary ions that make up the anion gap are sodium (Na+), chloride (Cl–) and bicarbonate (HCO3–) and it is defined as [Na+ – (Cl– + HCO3–)] where the sodium and chloride concentrations are measured as mEq/L and the bicarbonate concentration is mmol/L. Normal anion gap is between 8 and 12 and a higher number is diagnostic of metabolic acidosis. Rapid and aggressive treatment is necessary as the metabolic acidosis will result in cerebral edema and coma eventually leading to death.
The hyperketonemia in DKA is the result of insulin deficiency and unregulated glucagon secretion from α-cells of the pancreas. Circulating glucagon stimulates the adipose tissue to release fatty acids stored in triglycerides. The free fatty acids enter the circulation and are taken up primarily by the liver where they undergo fatty acid oxidation to acetylCoA. Normally, acetyl CoA is completely oxidized to CO2 and water in the TCA cycle. However, the level of fatty acid oxidation is in excess of the livers' ability to fully oxidize the excess acetyl CoA and, thus, the compound is diverted into the ketogenesis pathway. The ketones (ketone bodies) are β-hydroxybutyrate and acetoacetate with β-hydroxybutyrate being the most abundant. Acetoacetate will spontaneously (non-enzymatic) decarboxylate to acetone. Acetone is volatile and is released from the lungs giving the characteristic sweet smell to the breath of someone with hyperketonemia. The ketones are released into the circulation and because they are acidic lower the pH of the blood resulting in metabolic acidosis.
Insulin deficiency also causes increased triglyceride and protein metabolism in skeletal muscle. This leads to increased release of glycerol (from triglyceride metabolism) and alanine (from protein metabolism) to the circulation. These substances then enter the liver where they are used as substrates for gluconeogenesis which is enhanced in the absence of insulin and the elevated glucagon. The increased rate of glucose production in the liver, coupled with the glucagon-mediated inhibition of glucose storage into glycogen results in the increased glucose release from the liver and consequent hyperglycemia. The resultant hyperglycemia produces an osmotic diuresis that leads to loss of water and electrolytes in the urine. The ketones are also excreted in the urine and this results in an obligatory loss of Na+ and K+. The loss in K+ is large, sometimes exceeding 300 mEq/L/24 h. Initial serum K+ is typically normal or elevated because of the extracellular migration of K+ in response to the metabolic acidosis. The level of K+ will fall further during treatment as insulin therapy drives K+ into cells. If serum K+ is not monitored and replaced as needed (see below), life-threatening hypokalemia may develop.
Treatment of Diabetic Ketoacidosis
The following is not intended to be considered as routine orders for the diagnosis and treatment of all cases of DKA but is presented only as one possible treatment regimen. Each case of DKA must be treated on an individual basis.
Initial Assessment of DKA
blood glucose > 250mg/dL
arterial pH <7.3
serum bicarbonate <15mEq/L
urinary ketones ≥ 3+ and/or serum ketones are positive
Monitoring
vital signs every hour
serum glucose every hour and as needed
blood gas pH every 2 hrs (use arterial for 1st measurement then can use venous)
electrolytes every 1–2 hrs
urine ketones on each void
fluid input and output continuously
magnesium and phopshorous immediately and then every 1–2 hrs
Fluid Management
start normal saline at 1L/hr or 15–20ml/kg/hr initially
determine hydration status, goal being to replace 50% of estimated volume loss in the 1st 4hrs then remainder over next 8–12 hrs
infuse normal saline 125–500 ml/hr, rate dependent on hydration status
once serum Na+ is corrected infuse ½ normal saline at 4–14ml/kg/hr
when serum glucose reaches 250mg/dL change fluid to D5W ½ normal saline at same rate
Insulin Management
discontinue all oral diabetic medications and previous insulin orders
give regular insulin iv bolus of 10 units
start insulin infusion usually at a rate of 0.15units/kg
insulin administration goal is to reduce serum glucose 50–70mg/dL/hr
when serum glucose is ≤ 150mg/dL then can switch to adult sq insulin with basal insulin
Potassium Management
if serum K+ is <3.3 give 40mEq/hr until it is >3.3
if serum K+ is >3.3 but <5.0 give 20–30mEq/L of iv fluids to keep serum K+ between 4–5mEq/L
if serum K+ is ≥5.0 do not give K+ but check serum levels every 2hrs
when replacing K+ both potassium chloride and potassium phosphate can be used
hold K+ replacement if patient urine output is <30ml/hr
Bicarbonate Management
assess need for bicarbonate by arterial pH measurement
if pH <6.9 give 100mEq sodium bicarbonate in 1L D5W and infuse at 200ml/hr
if pH is 6.9 – 7.0 give 50mEq sodium bicarbonate in 1L D5W and infuse at 200ml/hr
if pH >7.0 do not give bicarbonate
continue sodium bicarbonate administration until pH is >7.0
monitor serum K+
The most severe and life threatening complication of poorly controlled type 1 diabetes is diabetic ketoacidosis (DKA). DKA is characterized by metabolic acidosis, hyperglycemia and hyperketonemia. Diagnosis of DKA is accomplished by detection of hyperketonemia and metabolic acidosis (as measured by anion gap) in the presence of hyperglycemia. The primary ions that make up the anion gap are sodium (Na+), chloride (Cl–) and bicarbonate (HCO3–) and it is defined as [Na+ – (Cl– + HCO3–)] where the sodium and chloride concentrations are measured as mEq/L and the bicarbonate concentration is mmol/L. Normal anion gap is between 8 and 12 and a higher number is diagnostic of metabolic acidosis. Rapid and aggressive treatment is necessary as the metabolic acidosis will result in cerebral edema and coma eventually leading to death.
The hyperketonemia in DKA is the result of insulin deficiency and unregulated glucagon secretion from α-cells of the pancreas. Circulating glucagon stimulates the adipose tissue to release fatty acids stored in triglycerides. The free fatty acids enter the circulation and are taken up primarily by the liver where they undergo fatty acid oxidation to acetylCoA. Normally, acetyl CoA is completely oxidized to CO2 and water in the TCA cycle. However, the level of fatty acid oxidation is in excess of the livers' ability to fully oxidize the excess acetyl CoA and, thus, the compound is diverted into the ketogenesis pathway. The ketones (ketone bodies) are β-hydroxybutyrate and acetoacetate with β-hydroxybutyrate being the most abundant. Acetoacetate will spontaneously (non-enzymatic) decarboxylate to acetone. Acetone is volatile and is released from the lungs giving the characteristic sweet smell to the breath of someone with hyperketonemia. The ketones are released into the circulation and because they are acidic lower the pH of the blood resulting in metabolic acidosis.
Insulin deficiency also causes increased triglyceride and protein metabolism in skeletal muscle. This leads to increased release of glycerol (from triglyceride metabolism) and alanine (from protein metabolism) to the circulation. These substances then enter the liver where they are used as substrates for gluconeogenesis which is enhanced in the absence of insulin and the elevated glucagon. The increased rate of glucose production in the liver, coupled with the glucagon-mediated inhibition of glucose storage into glycogen results in the increased glucose release from the liver and consequent hyperglycemia. The resultant hyperglycemia produces an osmotic diuresis that leads to loss of water and electrolytes in the urine. The ketones are also excreted in the urine and this results in an obligatory loss of Na+ and K+. The loss in K+ is large, sometimes exceeding 300 mEq/L/24 h. Initial serum K+ is typically normal or elevated because of the extracellular migration of K+ in response to the metabolic acidosis. The level of K+ will fall further during treatment as insulin therapy drives K+ into cells. If serum K+ is not monitored and replaced as needed (see below), life-threatening hypokalemia may develop.
Treatment of Diabetic Ketoacidosis
The following is not intended to be considered as routine orders for the diagnosis and treatment of all cases of DKA but is presented only as one possible treatment regimen. Each case of DKA must be treated on an individual basis.
Initial Assessment of DKA
blood glucose > 250mg/dL
arterial pH <7.3
serum bicarbonate <15mEq/L
urinary ketones ≥ 3+ and/or serum ketones are positive
Monitoring
vital signs every hour
serum glucose every hour and as needed
blood gas pH every 2 hrs (use arterial for 1st measurement then can use venous)
electrolytes every 1–2 hrs
urine ketones on each void
fluid input and output continuously
magnesium and phopshorous immediately and then every 1–2 hrs
Fluid Management
start normal saline at 1L/hr or 15–20ml/kg/hr initially
determine hydration status, goal being to replace 50% of estimated volume loss in the 1st 4hrs then remainder over next 8–12 hrs
infuse normal saline 125–500 ml/hr, rate dependent on hydration status
once serum Na+ is corrected infuse ½ normal saline at 4–14ml/kg/hr
when serum glucose reaches 250mg/dL change fluid to D5W ½ normal saline at same rate
Insulin Management
discontinue all oral diabetic medications and previous insulin orders
give regular insulin iv bolus of 10 units
start insulin infusion usually at a rate of 0.15units/kg
insulin administration goal is to reduce serum glucose 50–70mg/dL/hr
when serum glucose is ≤ 150mg/dL then can switch to adult sq insulin with basal insulin
Potassium Management
if serum K+ is <3.3 give 40mEq/hr until it is >3.3
if serum K+ is >3.3 but <5.0 give 20–30mEq/L of iv fluids to keep serum K+ between 4–5mEq/L
if serum K+ is ≥5.0 do not give K+ but check serum levels every 2hrs
when replacing K+ both potassium chloride and potassium phosphate can be used
hold K+ replacement if patient urine output is <30ml/hr
Bicarbonate Management
assess need for bicarbonate by arterial pH measurement
if pH <6.9 give 100mEq sodium bicarbonate in 1L D5W and infuse at 200ml/hr
if pH is 6.9 – 7.0 give 50mEq sodium bicarbonate in 1L D5W and infuse at 200ml/hr
if pH >7.0 do not give bicarbonate
continue sodium bicarbonate administration until pH is >7.0
monitor serum K+
Biochemistry of nerve transmission
Table of Neurotransmitters
Synaptic Transmission
Neuromuscular Transmission
Neurotransmitter Receptors
Acetylcholine
Cholinergic Agonists and Antagonists
Catecholamines
Serotonin
GABA
Web themedicalbiochemistrypage.org
Table of Neurotransmitters
Transmitter Molecule
Derived From
Site of Synthesis
Acetylcholine Choline CNS, parasympathetic nerves
Serotonin
5-Hydroxytryptamine (5-HT) Tryptophan CNS, chromaffin cells of the gut, enteric cells
GABA Glutamate CNS
Glutamate CNS
Aspartate CNS
Glycine spinal cord
Histamine Histidine hypothalamus
Epinephrine
synthesis pathway Tyrosine adrenal medulla, some CNS cells
Norpinephrine
synthesis pathway Tyrosine CNS, sympathetic nerves
Dopamine
synthesis pathway Tyrosine CNS
Adenosine ATP CNS, peripheral nerves
ATP sympathetic, sensory and enteric nerves
Nitric oxide, NO Arginine CNS, gastrointestinal tract
Many other neurotransmitters are derived from precursor proteins, the so-called peptide neurotransmitters. As many as 50 different peptides have been shown to exert their effects on neural cell function. Several of these peptide transmitters are derived from the larger protein pre-opiomelanocortin (POMC). Neuropeptides are responsible for mediating sensory and emotional responses including hunger, thirst, sex drive, pleasure and pain.
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Synaptic Transmission
Synaptic transmission refers to the propagation of nerve impulses from one nerve cell to another. This occurs at a specialized cellular structure known as the synapse, a junction at which the axon of the presynaptic neuron terminates at some location upon the postsynaptic neuron. The end of a presynaptic axon, where it is juxtaposed to the postsynaptic neuron, is enlarged and forms a structure known as the terminal button. An axon can make contact anywhere along the second neuron: on the dendrites (an axodendritic synapse), the cell body (an axosomatic synapse) or the axons (an axo-axonal synapse).
Nerve impulses are transmitted at synapses by the release of chemicals called neurotransmitters. As a nerve impulse, or action potential, reaches the end of a presynaptic axon, molecules of neurotransmitter are released into the synaptic space. The neurotransmitters are a diverse group of chemical compounds ranging from simple amines such as dopamine and amino acids such as γ-aminobutyrate (GABA), to polypeptides such as the enkephalins. The mechanisms by which they elicit responses in both presynaptic and postsynaptic neurons are as diverse as the mechanisms employed by growth factor and cytokine receptors.
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Neuromuscular Transmission
A different type of nerve transmission occurs when an axon terminates on a skeletal muscle fiber, at a specialized structure called the neuromuscular junction. An action potential occurring at this site is known as neuromuscular transmission. At a neuromuscular junction, the axon subdivides into numerous terminal buttons that reside within depressions formed in the motor end-plate. The particular transmitter in use at the neuromuscular junction is acetylcholine.
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Neurotransmitter Receptors
Once the molecules of neurotransmitter are released from a cell as the result of the firing of an action potential, they bind to specific receptors on the surface of the postsynaptic cell. In all cases in which these receptors have been cloned and characterized in detail, it has been shown that there are numerous subtypes of receptor for any given neurotransmitter. As well as being present on the surfaces of postsynaptic neurons, neurotransmitter receptors are found on presynaptic neurons. In general, presynaptic neuron receptors act to inhibit further release of neurotransmitter.
The vast majority of neurotransmitter receptors belong to a class of proteins known as the serpentine receptors. This class exhibits a characteristic transmembrane structure: that is, it spans the cell membrane, not once but seven times. The link between neurotransmitters and intracellular signaling is carried out by association either with G-proteins (small GTP-binding and hydrolyzing proteins) or with protein kinases, or by the receptor itself in the form of a ligand-gated ion channel (for example, the acetylcholine receptor). One additional characteristic of neurotransmitter receptors is that they are subject to ligand-induced desensitization: That is, they can become unresponsive upon prolonged exposure to their neurotransmitter.
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Acetylcholine
Acetylcholine (ACh) is a simple molecule synthesized from choline and acetyl-CoA through the action of choline acetyltransferase. Neurons that synthesize and release ACh are termed cholinergic neurons. When an action potential reaches the terminal button of a presynaptic neuron a voltage-gated calcium channel is opened. The influx of calcium ions, Ca2+, stimulates the exocytosis of presynaptic vesicles containing ACh, which is thereby released into the synaptic cleft. Once released, ACh must be removed rapidly in order to allow repolarization to take place; this step, hydrolysis, is carried out by the enzyme, acetylcholinesterase. The acetylcholinesterase found at nerve endings is anchored to the plasma membrane through a glycolipid.
Synthesis of Acetylcholine
ACh receptors are ligand-gated cation channels composed of four different polypeptide subunits arranged in the form [(α2)(β)(γ)(δ)]. Two main classes of ACh receptors have been identified on the basis of their responsiveness to the toadstool alkaloid, muscarine, and to nicotine, respectively: the muscarinic receptors and the nicotinic receptors. Both receptor classes are abundant in the human brain. Nicotinic receptors are further divided into those found at neuromuscular junctions and those found at neuronal synapses. The activation of ACh receptors by the binding of ACh leads to an influx of Na+ into the cell and an efflux of K+, resulting in a depolarization of the postsynaptic neuron and the initiation of a new action potential.
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Cholinergic Agonists and Antagonists
Numerous compounds have been identified that act as either agonists or antagonists of cholinergic neurons. The principal action of cholinergic agonists is the excitation or inhibition of autonomic effector cells that are innervated by postganglionic parasympathetic neurons and as such are referred to as parasympathomimetic agents. The cholinergic agonists include choline esters (such as ACh itself) as well as protein- or alkaloid-based compounds. Several naturally occurring compounds have been shown to affect cholinergic neurons, either positively or negatively.
The responses of cholinergic neurons can also be enhanced by administration of cholinesterase (ChE) inhibitors. ChE inhibitors have been used as components of nerve gases but also have significant medical application in the treatment of disorders such as glaucoma and myasthenia gravis as well as in terminating the effects of neuromuscular blocking agents such as atropine.
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Natural Cholinergic Agonist and Antagonists
Source of Compound
Mode of Action
Agonists
Nicotine alkaloid prevalent in the tobacco plant activates nicotinic class of ACh receptors, locks the channel open
Muscarine alkaloid produced by Amanita muscaria mushrooms activates muscarinic class of ACh receptors
α-Latrotoxin protein produced by the black widow spider induces massive ACh release, possibly by acting as a Ca2+ ionophore
Antagonists
atropine (and related compound Scopolamine) alkaloid produced by the deadly nightshade, Atropa belladonna blocks ACh actions only at muscarinic receptors
Botulinus toxin eight proteins produced by Clostridium botulinum inhibits the release of ACh
α-Bungarotoxin protein produced by Bungarus genus of snakes prevents ACh receptor channel opening
d-Tubocurarine active ingredient of curare prevents ACh receptor channel opening at motor end-plate
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Catecholamines
The principal catecholamines are norepinephrine, epinephrine and dopamine. These compounds are formed from phenylalanine and tyrosine. Tyrosine is produced in the liver from phenylalanine through the action of phenylalanine hydroxylase. The tyrosine is then transported to catecholamine-secreting neurons where a series of reactions convert it to dopamine, to norepinephrine and finally to epinephrine (see also Specialized Products of Amino Acids).
Synthesis of the Catecholamines from Tyrosine.
Catecholamines exhibit peripheral nervous system excitatory and inhibitory effects as well as actions in the CNS such as respiratory stimulation and an increase in psychomotor activity. The excitatory effects are exerted upon smooth muscle cells of the vessels that supply blood to the skin and mucous membranes. Cardiac function is also subject to excitatory effects, which lead to an increase in heart rate and in the force of contraction. Inhibitory effects, by contrast, are exerted upon smooth muscle cells in the wall of the gut, the bronchial tree of the lungs, and the vessels that supply blood to skeletal muscle.
In addition to their effects as neurotransmitters, norepinephrine and epinephrine can influence the rate of metabolism. This influence works both by modulating endocrine function such as insulin secretion and by increasing the rate of glycogenolysis and fatty acid mobilization.
The catecholamines bind to two different classes of receptors termed the α- and β-adrenergic receptors. The catecholamines therefore are also known as adrenergic neurotransmitters; neurons that secrete them are adrenergic neurons. Norepinephrine-secreting neurons are noradrenergic. The adrenergic receptors are classical serpentine receptors that couple to intracellular G-proteins. Some of the norepinephrine released from presynaptic noradrenergic neurons recycled in the presynaptic neuron by a reuptake mechanism.
Catecholamine Catabolism
Epinephrine and norepinephrine are catabolized to inactive compounds through the sequential actions of catecholamine-O-methyltransferase (COMT) and monoamine oxidase (MAO). Compounds that inhibit the action of MAO have been shown to have beneficial effects in the treatment of clinical depression, even when tricyclic antidepressants are ineffective. The utility of MAO inhibitors was discovered serendipitously when patients treated for tuberculosis with isoniazid showed signs of an improvement in mood; isoniazid was subsequently found to work by inhibiting MAO.
Metabolism of the catecholamine neurotransmitters. Only clinically important enzymes are included in this diagram. The catabolic byproducts of the catecholamines, whose levels in the cerebrospinal fluid are indicative of defects in catabolism, are in blue underlined text. Abbreviations: TH = tyrosine hydroxylase, DHPR = dihydropteridine reductase, H2B = dihydrobiopterin, H4B = tetrahydrobiopterin, MAO = monoamine oxidase, COMT = catecholamine-O-methyltransferase, MHPG = 3-methoxy-4-hydroxyphenylglycol, DOPAC = dihydroxyphenylacetic acid.
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Serotonin
Serotonin (5-hydroxytryptamine, 5HT) is formed by the hydroxylation and decarboxylation of tryptophan (see also Specialized Products of Amino Acids).
Pathway for serotonin synthesis from tryptophan. Abbreviations: THP = tryptophan hydroxylase, DHPR = dihydropteridine reductase, H2B = dihydrobiopterin, H4B = tetrahyrobiopterin, 5-HT = 5-hydroxytryptophan, AADC = aromatic L-amino acid decarboxylase.
The greatest concentration of 5HT (90%) is found in the enterochromaffin cells of the gastrointestinal tract. Most of the remainder of the body's 5HT is found in platelets and the CNS. The effects of 5HT are felt most prominently in the cardiovascular system, with additional effects in the respiratory system and the intestines. Vasoconstriction is a classic response to the administration of 5HT.
Neurons that secrete 5HT are termed serotonergic. Following the release of 5HT, a portion is taken back up by the presynaptic serotonergic neuron in a manner similar to that of the reuptake of norepinephrine.
The function of serotonin is exerted upon its interaction with specific receptors. Several serotonin receptors have been cloned and are identified as 5HT1, 5HT2, 5HT3, 5HT4, 5HT5, 5HT6, and 5HT7. Within the 5HT1 group there are subtypes 5HT1A, 5HT1B, 5HT1D, 5HT1E, and 5HT1F. There are three 5HT2 subtypes, 5HT2A, 5HT2B, and 5HT2C as well as two 5HT5 subtypes, 5HT5a and 5HT5B. Most of these receptors are coupled to G-proteins that affect the activities of either adenylate cyclase or phospholipase Cγ. The 5HT3 class of receptors are ion channels.
Some serotonin receptors are presynaptic and others postsynaptic. The 5HT2A receptors mediate platelet aggregation and smooth muscle contraction. The 5HT2C receptors are suspected in control of food intake as mice lacking this gene become obese from increased food intake and are also subject to fatal seizures. The 5HT3 receptors are present in the gastrointestinal tract and are related to vomiting. Also present in the gastrointestinal tract are 5HT4 receptors where they function in secretion and peristalsis. The 5HT6 and 5HT7 receptors are distributed throughout the limbic system of the brain and the 5HT6 receptors have high affinity for antidepressant drugs.
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GABA
Several amino acids have distinct excitatory or inhibitory effects upon the nervous system. The amino acid derivative, γ-aminobutyrate, also called 4-aminobutyrate, (GABA) is a well-known inhibitor of presynaptic transmission in the CNS, and also in the retina. Neurons that secrete GABA are termed GABAergic.
The formation of GABA occurs by the decarboxylation of glutamate catalyzed by glutamate decarboxylase (GAD). GAD is present in many nerve endings of the brain as well as in the β-cells of the pancreas. The activity of GAD requires pyridoxal phosphate (PLP) as a cofactor. PLP is generated from the B6 vitamins (pyridoxine, pyridoxal, and pyridoxamine) through the action of pyridoxal kinase. Pyridoxal kinase itself requires zinc for activation. A deficiency in zinc or defects in pyridoxal kinase can lead to seizure disorders, particularly in seizure-prone preeclamptic patients (hypertensive condition in late pregnancy).
GABA Synthesis
GABA exerts its effects by binding to two distinct receptors, GABA-A and GABA-B. The GABA-A receptors form a Cl– channel. The binding of GABA to GABA-A receptors increases the Cl- conductance of presynaptic neurons. The anxiolytic drugs of the benzodiazepine family exert their soothing effects by potentiating the responses of GABA-A receptors to GABA binding. The GABA-B receptors are coupled to an intracellular G-protein and act by increasing conductance of an associated K+ channel.
Synaptic Transmission
Neuromuscular Transmission
Neurotransmitter Receptors
Acetylcholine
Cholinergic Agonists and Antagonists
Catecholamines
Serotonin
GABA
Web themedicalbiochemistrypage.org
Table of Neurotransmitters
Transmitter Molecule
Derived From
Site of Synthesis
Acetylcholine Choline CNS, parasympathetic nerves
Serotonin
5-Hydroxytryptamine (5-HT) Tryptophan CNS, chromaffin cells of the gut, enteric cells
GABA Glutamate CNS
Glutamate CNS
Aspartate CNS
Glycine spinal cord
Histamine Histidine hypothalamus
Epinephrine
synthesis pathway Tyrosine adrenal medulla, some CNS cells
Norpinephrine
synthesis pathway Tyrosine CNS, sympathetic nerves
Dopamine
synthesis pathway Tyrosine CNS
Adenosine ATP CNS, peripheral nerves
ATP sympathetic, sensory and enteric nerves
Nitric oxide, NO Arginine CNS, gastrointestinal tract
Many other neurotransmitters are derived from precursor proteins, the so-called peptide neurotransmitters. As many as 50 different peptides have been shown to exert their effects on neural cell function. Several of these peptide transmitters are derived from the larger protein pre-opiomelanocortin (POMC). Neuropeptides are responsible for mediating sensory and emotional responses including hunger, thirst, sex drive, pleasure and pain.
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Synaptic Transmission
Synaptic transmission refers to the propagation of nerve impulses from one nerve cell to another. This occurs at a specialized cellular structure known as the synapse, a junction at which the axon of the presynaptic neuron terminates at some location upon the postsynaptic neuron. The end of a presynaptic axon, where it is juxtaposed to the postsynaptic neuron, is enlarged and forms a structure known as the terminal button. An axon can make contact anywhere along the second neuron: on the dendrites (an axodendritic synapse), the cell body (an axosomatic synapse) or the axons (an axo-axonal synapse).
Nerve impulses are transmitted at synapses by the release of chemicals called neurotransmitters. As a nerve impulse, or action potential, reaches the end of a presynaptic axon, molecules of neurotransmitter are released into the synaptic space. The neurotransmitters are a diverse group of chemical compounds ranging from simple amines such as dopamine and amino acids such as γ-aminobutyrate (GABA), to polypeptides such as the enkephalins. The mechanisms by which they elicit responses in both presynaptic and postsynaptic neurons are as diverse as the mechanisms employed by growth factor and cytokine receptors.
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Neuromuscular Transmission
A different type of nerve transmission occurs when an axon terminates on a skeletal muscle fiber, at a specialized structure called the neuromuscular junction. An action potential occurring at this site is known as neuromuscular transmission. At a neuromuscular junction, the axon subdivides into numerous terminal buttons that reside within depressions formed in the motor end-plate. The particular transmitter in use at the neuromuscular junction is acetylcholine.
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Neurotransmitter Receptors
Once the molecules of neurotransmitter are released from a cell as the result of the firing of an action potential, they bind to specific receptors on the surface of the postsynaptic cell. In all cases in which these receptors have been cloned and characterized in detail, it has been shown that there are numerous subtypes of receptor for any given neurotransmitter. As well as being present on the surfaces of postsynaptic neurons, neurotransmitter receptors are found on presynaptic neurons. In general, presynaptic neuron receptors act to inhibit further release of neurotransmitter.
The vast majority of neurotransmitter receptors belong to a class of proteins known as the serpentine receptors. This class exhibits a characteristic transmembrane structure: that is, it spans the cell membrane, not once but seven times. The link between neurotransmitters and intracellular signaling is carried out by association either with G-proteins (small GTP-binding and hydrolyzing proteins) or with protein kinases, or by the receptor itself in the form of a ligand-gated ion channel (for example, the acetylcholine receptor). One additional characteristic of neurotransmitter receptors is that they are subject to ligand-induced desensitization: That is, they can become unresponsive upon prolonged exposure to their neurotransmitter.
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Acetylcholine
Acetylcholine (ACh) is a simple molecule synthesized from choline and acetyl-CoA through the action of choline acetyltransferase. Neurons that synthesize and release ACh are termed cholinergic neurons. When an action potential reaches the terminal button of a presynaptic neuron a voltage-gated calcium channel is opened. The influx of calcium ions, Ca2+, stimulates the exocytosis of presynaptic vesicles containing ACh, which is thereby released into the synaptic cleft. Once released, ACh must be removed rapidly in order to allow repolarization to take place; this step, hydrolysis, is carried out by the enzyme, acetylcholinesterase. The acetylcholinesterase found at nerve endings is anchored to the plasma membrane through a glycolipid.
Synthesis of Acetylcholine
ACh receptors are ligand-gated cation channels composed of four different polypeptide subunits arranged in the form [(α2)(β)(γ)(δ)]. Two main classes of ACh receptors have been identified on the basis of their responsiveness to the toadstool alkaloid, muscarine, and to nicotine, respectively: the muscarinic receptors and the nicotinic receptors. Both receptor classes are abundant in the human brain. Nicotinic receptors are further divided into those found at neuromuscular junctions and those found at neuronal synapses. The activation of ACh receptors by the binding of ACh leads to an influx of Na+ into the cell and an efflux of K+, resulting in a depolarization of the postsynaptic neuron and the initiation of a new action potential.
back to the top
Cholinergic Agonists and Antagonists
Numerous compounds have been identified that act as either agonists or antagonists of cholinergic neurons. The principal action of cholinergic agonists is the excitation or inhibition of autonomic effector cells that are innervated by postganglionic parasympathetic neurons and as such are referred to as parasympathomimetic agents. The cholinergic agonists include choline esters (such as ACh itself) as well as protein- or alkaloid-based compounds. Several naturally occurring compounds have been shown to affect cholinergic neurons, either positively or negatively.
The responses of cholinergic neurons can also be enhanced by administration of cholinesterase (ChE) inhibitors. ChE inhibitors have been used as components of nerve gases but also have significant medical application in the treatment of disorders such as glaucoma and myasthenia gravis as well as in terminating the effects of neuromuscular blocking agents such as atropine.
back to the top
Natural Cholinergic Agonist and Antagonists
Source of Compound
Mode of Action
Agonists
Nicotine alkaloid prevalent in the tobacco plant activates nicotinic class of ACh receptors, locks the channel open
Muscarine alkaloid produced by Amanita muscaria mushrooms activates muscarinic class of ACh receptors
α-Latrotoxin protein produced by the black widow spider induces massive ACh release, possibly by acting as a Ca2+ ionophore
Antagonists
atropine (and related compound Scopolamine) alkaloid produced by the deadly nightshade, Atropa belladonna blocks ACh actions only at muscarinic receptors
Botulinus toxin eight proteins produced by Clostridium botulinum inhibits the release of ACh
α-Bungarotoxin protein produced by Bungarus genus of snakes prevents ACh receptor channel opening
d-Tubocurarine active ingredient of curare prevents ACh receptor channel opening at motor end-plate
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Catecholamines
The principal catecholamines are norepinephrine, epinephrine and dopamine. These compounds are formed from phenylalanine and tyrosine. Tyrosine is produced in the liver from phenylalanine through the action of phenylalanine hydroxylase. The tyrosine is then transported to catecholamine-secreting neurons where a series of reactions convert it to dopamine, to norepinephrine and finally to epinephrine (see also Specialized Products of Amino Acids).
Synthesis of the Catecholamines from Tyrosine.
Catecholamines exhibit peripheral nervous system excitatory and inhibitory effects as well as actions in the CNS such as respiratory stimulation and an increase in psychomotor activity. The excitatory effects are exerted upon smooth muscle cells of the vessels that supply blood to the skin and mucous membranes. Cardiac function is also subject to excitatory effects, which lead to an increase in heart rate and in the force of contraction. Inhibitory effects, by contrast, are exerted upon smooth muscle cells in the wall of the gut, the bronchial tree of the lungs, and the vessels that supply blood to skeletal muscle.
In addition to their effects as neurotransmitters, norepinephrine and epinephrine can influence the rate of metabolism. This influence works both by modulating endocrine function such as insulin secretion and by increasing the rate of glycogenolysis and fatty acid mobilization.
The catecholamines bind to two different classes of receptors termed the α- and β-adrenergic receptors. The catecholamines therefore are also known as adrenergic neurotransmitters; neurons that secrete them are adrenergic neurons. Norepinephrine-secreting neurons are noradrenergic. The adrenergic receptors are classical serpentine receptors that couple to intracellular G-proteins. Some of the norepinephrine released from presynaptic noradrenergic neurons recycled in the presynaptic neuron by a reuptake mechanism.
Catecholamine Catabolism
Epinephrine and norepinephrine are catabolized to inactive compounds through the sequential actions of catecholamine-O-methyltransferase (COMT) and monoamine oxidase (MAO). Compounds that inhibit the action of MAO have been shown to have beneficial effects in the treatment of clinical depression, even when tricyclic antidepressants are ineffective. The utility of MAO inhibitors was discovered serendipitously when patients treated for tuberculosis with isoniazid showed signs of an improvement in mood; isoniazid was subsequently found to work by inhibiting MAO.
Metabolism of the catecholamine neurotransmitters. Only clinically important enzymes are included in this diagram. The catabolic byproducts of the catecholamines, whose levels in the cerebrospinal fluid are indicative of defects in catabolism, are in blue underlined text. Abbreviations: TH = tyrosine hydroxylase, DHPR = dihydropteridine reductase, H2B = dihydrobiopterin, H4B = tetrahydrobiopterin, MAO = monoamine oxidase, COMT = catecholamine-O-methyltransferase, MHPG = 3-methoxy-4-hydroxyphenylglycol, DOPAC = dihydroxyphenylacetic acid.
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Serotonin
Serotonin (5-hydroxytryptamine, 5HT) is formed by the hydroxylation and decarboxylation of tryptophan (see also Specialized Products of Amino Acids).
Pathway for serotonin synthesis from tryptophan. Abbreviations: THP = tryptophan hydroxylase, DHPR = dihydropteridine reductase, H2B = dihydrobiopterin, H4B = tetrahyrobiopterin, 5-HT = 5-hydroxytryptophan, AADC = aromatic L-amino acid decarboxylase.
The greatest concentration of 5HT (90%) is found in the enterochromaffin cells of the gastrointestinal tract. Most of the remainder of the body's 5HT is found in platelets and the CNS. The effects of 5HT are felt most prominently in the cardiovascular system, with additional effects in the respiratory system and the intestines. Vasoconstriction is a classic response to the administration of 5HT.
Neurons that secrete 5HT are termed serotonergic. Following the release of 5HT, a portion is taken back up by the presynaptic serotonergic neuron in a manner similar to that of the reuptake of norepinephrine.
The function of serotonin is exerted upon its interaction with specific receptors. Several serotonin receptors have been cloned and are identified as 5HT1, 5HT2, 5HT3, 5HT4, 5HT5, 5HT6, and 5HT7. Within the 5HT1 group there are subtypes 5HT1A, 5HT1B, 5HT1D, 5HT1E, and 5HT1F. There are three 5HT2 subtypes, 5HT2A, 5HT2B, and 5HT2C as well as two 5HT5 subtypes, 5HT5a and 5HT5B. Most of these receptors are coupled to G-proteins that affect the activities of either adenylate cyclase or phospholipase Cγ. The 5HT3 class of receptors are ion channels.
Some serotonin receptors are presynaptic and others postsynaptic. The 5HT2A receptors mediate platelet aggregation and smooth muscle contraction. The 5HT2C receptors are suspected in control of food intake as mice lacking this gene become obese from increased food intake and are also subject to fatal seizures. The 5HT3 receptors are present in the gastrointestinal tract and are related to vomiting. Also present in the gastrointestinal tract are 5HT4 receptors where they function in secretion and peristalsis. The 5HT6 and 5HT7 receptors are distributed throughout the limbic system of the brain and the 5HT6 receptors have high affinity for antidepressant drugs.
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GABA
Several amino acids have distinct excitatory or inhibitory effects upon the nervous system. The amino acid derivative, γ-aminobutyrate, also called 4-aminobutyrate, (GABA) is a well-known inhibitor of presynaptic transmission in the CNS, and also in the retina. Neurons that secrete GABA are termed GABAergic.
The formation of GABA occurs by the decarboxylation of glutamate catalyzed by glutamate decarboxylase (GAD). GAD is present in many nerve endings of the brain as well as in the β-cells of the pancreas. The activity of GAD requires pyridoxal phosphate (PLP) as a cofactor. PLP is generated from the B6 vitamins (pyridoxine, pyridoxal, and pyridoxamine) through the action of pyridoxal kinase. Pyridoxal kinase itself requires zinc for activation. A deficiency in zinc or defects in pyridoxal kinase can lead to seizure disorders, particularly in seizure-prone preeclamptic patients (hypertensive condition in late pregnancy).
GABA Synthesis
GABA exerts its effects by binding to two distinct receptors, GABA-A and GABA-B. The GABA-A receptors form a Cl– channel. The binding of GABA to GABA-A receptors increases the Cl- conductance of presynaptic neurons. The anxiolytic drugs of the benzodiazepine family exert their soothing effects by potentiating the responses of GABA-A receptors to GABA binding. The GABA-B receptors are coupled to an intracellular G-protein and act by increasing conductance of an associated K+ channel.
blood coagulation
Introduction
Platelet Activation and von Willebrand Factor (vWF)
Description of Clotting Factors
Image of the Clotting Cascade
Kallikrein-Kinin System in Coagulation
Intrinsic (contact activation) Clotting Cascade
Extrinsic (tissue factor) Clotting Cascade
Activation of Thrombin
Regulation of Thrombin Levels
Activation of Fibrin
Dissolution of Fibrin Clots
Clinical Significances of Hemostasis: The Bleeding Disorders
Pharmacological Intervention in Bleeding
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Introduction
The ability of the body to control the flow of blood following vascular injury is paramount to continued survival. The process of blood clotting and then the subsequent dissolution of the clot, following repair of the injured tissue, is termed hemostasis. Hemostasis, composed of 4 major events that occur in a set order following the loss of vascular integrity:
1. The initial phase of the process is vascular constriction. This limits the flow of blood to the area of injury.
2. Next, platelets become activated by thrombin and aggregate at the site of injury, forming a temporary, loose platelet plug. The protein fibrinogen is primarily responsible for stimulating platelet clumping. Platelets clump by binding to collagen that becomes exposed following rupture of the endothelial lining of vessels. Upon activation, platelets release the nucleotide, ADP and the eicosanoid, TXA2 (both of which activate additional platelets), serotonin, phospholipids, lipoproteins, and other proteins important for the coagulation cascade. In addition to induced secretion, activated platelets change their shape to accommodate the formation of the plug.
3. To insure stability of the initially loose platelet plug, a fibrin mesh (also called the clot) forms and entraps the plug. If the plug contains only platelets it is termed a white thrombus; if red blood cells are present it is called a red thrombus
4. Finally, the clot must be dissolved in order for normal blood flow to resume following tissue repair. The dissolution of the clot occurs through the action of plasmin
Two pathways lead to the formation of a fibrin clot: the intrinsic and extrinsic pathway. Although they are initiated by distinct mechanisms, the two converge on a common pathway that leads to clot formation. Both pathways are complex and involve numerous different proteins termed clotting factors. Fibrin clot formation in response to tissue injury is the most clinically relevant event of hemostasis under normal physiological conditions. This process is the result of the activation of the extrinsic pathway. The formation of a red thrombus or a clot in response to an abnormal vessel wall in the absence of tissue injury is the result of the intrinsic pathway. The intrinsic pathway has low significance under normal physiological conditions. Most significant clinically is the activation of the intrinsic pathway by contact of the vessel wall with lipoprotein particles, VLDLs and chylomicrons. This process clearly demonstrates the role of hyperlipidemia in the generation of atherosclerosis. The intrinsic pathway can also be activated by vessel wall contact with bacteria.
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Platelet Activation and von Willebrand Factor (vWF)
In order for hemostasis to occur, platelets must adhere to exposed collagen, release the contents of their granules, and aggregate. The adhesion of platelets to the collagen exposed on endothelial cell surfaces is mediated by von Willebrand factor (vWF). Inherited deficiencies of vWF are the causes of von Willebrand disease, (vWD) (also see below for more details). The function of vWF is to act as a bridge between a specific glycoprotein complex on the surface of platelets (GPIb-GPIX-GPV) and collagen fibrils. The importance of this interaction between vWF and the GPIb-GPIX-GPV complex of platelets is demonstrated by the inheritance of bleeding disorders caused by defects in any one of the four the proteins of the complex, the most common of which is Bernard-Soulier syndrome (also called giant platelet syndrome). The GPIb part of the complex is composed of two proteins, GPIbα and GPIbβ encoded by separate genes.
In addition to its role as a bridge between platelets and exposed collagen on endothelial surfaces, vWF binds to and stabilizes coagulation factor VIII. Binding of factor VIII by vWF is required for normal survival of factor VIII in the circulation.
von Willebrand factor is a complex multimeric glycoprotein that is produced by and stored in the α-granules of platelets. It is also synthesized by megakaryocytes and found associated with subendothelial connective tissue.
The initial activation of platelets is induced by thrombin binding to specific receptors on the surface of platelets, thereby initiating a signal transduction cascade. The thrombin receptor is coupled to a G-protein that, in turn, activates phospholipase C-γ (PLC-γ). PLC-γ hydrolyzes phosphatidylinositol-4,5-bisphosphate (PIP2) leading to the formation of inositol trisphosphate (IP3) and diacylglycerol (DAG). IP3 induces the release of intracellular Ca2+ stores, and DAG activates protein kinase C (PKC).
The collagen to which platelets adhere as well as the release of intracellular Ca2+ leads to the activation of phospholipase A2 (PLA2), which then hydrolyzes membrane phospholipids, leading to liberation of arachidonic acid. The arachidonic acid release leads to an increase in the production and subsequent release of thromboxane A2 (TXA2). TXA2 is a potent vasoconstrictor and inducer of platelet aggregation that functions by binding to receptors that function through the PLC-γ pathway.
Another enzyme activated by the released intracellular Ca2+ stores is myosin light chain kinase (MLCK). Activated MLCK phosphorylates the light chain of myosin which then interacts with actin, resulting in altered platelet morphology and motility.
One of the many effects of PKC is the phosphorylation and activation of a specific 47,000-Dalton platelet protein. This activated protein induces the release of platelet granule contents; one of which is ADP. ADP further stimulates platelets increasing the overall activation cascade. The important role of ADP in platelet activation can be appreciated from the use of the ADP receptor antagonist, Plavix® (clopidogrel), in the control of thrombosis (see below). ADP also modifies the platelet membranes leading to exposure platelet glycoprotein receptor complex: GPIIb-GPIIIa. GPIIb-GPIIIa constitutes a receptor for vWF and fibrinogen, resulting in fibrinogen-induced platelet aggregation. The GPIIb-GPIIIa complex is a member of the integrin family of cell-surface receptors that interact with the extracellular matrix. The GPIIb-GPIIIa complex is also called integrin αIIb-β3. The importance of the GPIIb-GPIIIa in platelet activation and coagulation is demonstrated by the fact that bleeding disorders result from inherited defects in this glycoprotein complex. The most commonly inherited platelet dysfunction is Glanzmann thrombasthenia which results from defects in the GPIIb protein of this complex. In addition, the importance of this complex in overall hemostasis is demonstrated by the use of antibodies that block this receptor as anti-coagulants (e.g. ReoPro®, abciximab: see below).
Activation of platelets is required for their consequent aggregation to a platelet plug. However, equally significant is the role of activated platelet surface phospholipids in the activation of the coagulation cascade.
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Primary Factors
Factor
Trivial Name(s)
Pathway
Characteristic
Prekallikrein (PK) Fletcher factor Intrinsic Functions with HMWK and factor XII
High molecular weight kininogen (HMWK) contact activation cofactor; Fitzgerald, Flaujeac Williams factor Intrinsic Co-factor in kallikrein and factor XII activation, necessary in factor XIIa activation of XI, precursor for bradykinin (a potent vasodilator and inducer of smooth muscle contraction
I Fibrinogen Both
II Prothrombin Both Contains N-term. gla segment
III Tissue Factor Extrinsic
IV Calcium Both
V Proaccelerin, labile factor, accelerator (Ac-) globulin Both Protein cofactor
VI (same as Va) Accelerin Both This is Va, redundant to Factor V
VII Proconvertin, serum prothrombin conversion accelerator (SPCA), cothromboplastin Extrinsic Endopeptidase with gla residues
VIII Antihemophiliac factor A, antihemophilic globulin (AHG) Intrinsic Protein cofactor
IX Christmas Factor, antihemophilic factor B,plasma thromboplastin component (PTC) Intrinsic Endopeptidase with gla residues
X Stuart-Prower Factor Both Endopeptidase with gla residues
XI Plasma thromboplastin antecedent (PTA) Intrinsic Endopeptidase
XII Hageman Factor Intrinsic Endopeptidase
XIII Protransglutaminase, fibrin stabilizing factor (FSF), fibrinoligase Both Transpeptidase
Functional Classification of Clotting Factors
Zymogens of Serine Proteases
Activities
Factor XII binds to exposed collagen at site of vessel wall injury, activated by high-MW kininogen and kallikrein
Factor XI activated by factor XIIa
Factor IX activated by factor XIa in presence of Ca2+
Factor VII activated by thrombin in presence of Ca2+
Factor X activated on surface of activated platelets by tenase complex and by factor VIIa in presence of tissue factor and Ca2+
Factor II activated on surface of activated platelets by prothrombinase complex
Cofactors
Activities
Factor VIII activated by thrombin; factor VIIIa is a cofactor in the activation of factor X by factor IXa
Factor V activated by thrombin; factor Va is a cofactor in the activation of prothrombin by factor Xa
Factor III (tissue factor) a subendothelial cell-surface glycoprotein that acts as a cofactor for factor VII
Fibrinogen
Activity
Factor I cleaved by thrombin to form fibrin clot
Transglutaminase
Activity
Factor XIII activated by thrombin in presence of Ca2+; stabilizes fibrin clot by covalent cross-linking
Regulatory/Other Proteins
Activities
von Willebrand factor associated with subendothelial connective tissue; serves as a bridge between platelet glycoprotein GPIb/IX and collagen
Protein C activated to protein Ca by thrombin bound to thrombomodulin; then degrades factors VIIIa and Va
Protein S acts as a cofactor of protein C; both proteins contain gla residues
Thrombomodulin protein on the surface of endothelial cells; binds thrombin, which then activates protein C
Antithrombin III most important coagulation inhibitor, controls activities of thrombin, and factors IXa, Xa, XIa and XIIa
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The Clotting Cascades
The clotting cascades: The intrinsic cascade (which has less in vivo significance in normal physiological circumstances than the extrinsic cascade) is initiated when contact is made between blood and exposed negatively charged surfaces. The extrinsic pathway is initiated upon vascular injury which leads to exposure of tissue factor, TF (also identified as factor III), a subendothelial cell-surface glycoprotein that binds phospholipid. The green dotted arrow represents a point of cross-over between the extrinsic and intrinsic pathways. The two pathways converge at the activation of factor X to Xa. Factor Xa has a role in the further activation of factor VII to VIIa as depicted by the green arrow. Active factor Xa hydrolyzes and activates prothrombin to thrombin. Thrombin can then activate factors XI, VIII and V furthering the cascade. Ultimately the role of thrombin is to convert fribrinogen to fibrin and to activate factor XIII to XIIIa. Factor XIIIa (also termed transglutaminase) cross-links fibrin polymers solidifying the clot. HMWK = high molecular weight kininogen. PK = prekallikrein. PL = phospholipid.
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The Kallikrein-Kinin System in Coagulation
The kallikrein-kinin system comprises a complex of proteins that when activated leads to the release of vasoactive kinins. The kinins are released from both high molecular weight kininogen (HMWK) and low molecular weight kininogen (LMWK) as a result of activation of either tissue kallikrein or plasma kallikrein. The kallekreins themselves exist in inactive pre-forms. The kinins are involved in many physiological and pathological processes including regulation of blood pressure and flow (via modulation of the renin-angiotensin pathway), blood coagulation, cellular proliferation and growth, angiogenesis, apoptosis, and inflammation. Kinin action on endothelial cells leads to vasodilation, increased vascular permeability, release of tissue plasminogen activator (tPA), production of nitric oxide (NO), and the mobilization of arachidonic acid, primarily resulting in prostacyclin (PGI2) production by endothelial cells. Although the activities of the kallikrein-kinin system are involved in numerous processes, this section will deal only with their function in blood coagulation. With respect to hemostasis the most important kinin is bradykinin which is released from HMWK.
The two forms of prekallikrein, plasma and tissue, are derived from distinct genes on different chromosomes. The plasma kallikrein gene (symbol KLKB1) is on chromosome 4q34-q35 and the tissue kallikrein gene (symbol KLK1) located on chromosome 19q13.2-q13.4. These two kallikreins are serine proteases whose major substrates are HMWK (plasma kallikrein) and LMWK (tissue kallikrein). When plasma prekallikrein is activated to kallikrein it cleaves HMWK in a two-step process that ultimately releases bradykinin. Bradykinin is a 9-amino acid vasoactive peptide that induces vasodilation and increases vascular permeability. Activated tissue kallikrein cleaves lysyl-bradykinin (also called kallidin) from LMWK. Lysyl-bradykinin is bradykinin with a lysine residue at the N-terminus making it a 10-amino acid vasoactive peptide. Its activities are essentially identical to those of bradykinin.
Both HMWK and LMWK are derived from the same gene on chromosome 3q26-qter which is composed of 11 exons. Exons 1 to 9 encode the heavy chain of both kininogens. Exon 10 encodes bradykinin as well as the light chain of HMWK. Exon 11 encodes the light chain of LMWK. The heavy and light chain nomenclature refers to the disulfide-bonded structure of each kininogen after their activation by through kallikrein cleavage.
HMWK is considered an α-globulin and is composed of six functional domains. The protein circulates in the plasma as single-chain polypeptide with a molecular weight of 88 - 120 kDa dependent upon the level of glycosylation. The heavy chain is 64 kDa and contains domains 1, 2, and 3 whereas the light chain is 45 - 56 kDa and comprises domains 5 and 6. The heavy and light chains are linked together through domain 4 which also contains the bradykinin sequence. Domain 1 contains a low affinity calcium-binding site. Domains 2 and 3 contain amino acid sequences (QVVAG) that inhibit cysteine proteases. Domain 3 also has platelet and endothelial cell-binding activity. Domain 5 has sequences for heparin binding, cell-binding sites, and antiangiogenic properties. The binding of HMWK to negatively charged surfaces occurs through a histidine region of the light chain which is in domain 5. Domain 6 contains the prekallikrein and factor XI-binding sites. By being able to bind to charged surfaces via domain 5 and simultaneously bind factor XI and prekallikrein via domain 6, HMWK can serve as the cofactor in contact activation of plasma. LMWK is considered a β-globulin and has a molecular weight of 50 - 68 kDa. The structure of LMWK is similar to that of HMWK, however, the light chain is only 4 - 5 kDa and has no contact activation nor prekallikrein-binding sites.
The plasma kinin forming system is called the contact system of plasma and is composed of factor XII, factor XI, prekallikrein and HMWK. Factor XII, prekallikrein, and HMWK saturably and reversibly bind to endothelial cells, platelets, and granulocytes in a zinc-dependent reaction. When plasma makes contact with a negatively charged surface factor XII binds and is autoactivated to factor XIIa (the "a" signifies the activated factor). Factor XIIa then activates prekallikrein to kallikrein and kallikrein cleaves HMWK releasing bradykinin. There is also reciprocal activation of factor XII by kallikrein resulting in amplification of the system. The actual surface that leads to factor XII autoactivation is unknown however, several physiologic substances support the process. These substances include hematin, skin, fatty acids, sodium urate crystals, protoporphyrin, sulfatides, heparins, chondroitin sulfates, articular cartilage, endotoxin, L-homocysteine, and amyloid β-protein. Once the contact system is activated the intrinsic pathway (described below) is initiated.
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The Intrinsic Clotting Cascade
The intrinsic pathway (also called the contact activation pathway) is much less significant to hemostasis under normal physiological conditions than is the extrinsic pathway. However, abnormal physiology such as hyperlipidemic states or bacterial infiltration can lead to activation of thrombosis via the intrinsic clotting cascade.
The intrinsic pathway requires the clotting factors VIII, IX, X, XI, and XII. Also required are the proteins prekallikrein (PK) and high-molecular-weight kininogen (HK or HMWK), as well as calcium ions and phospholipids secreted from platelets. The role of PK and HMWK is described in the above section. Each of these pathway constituents leads to the conversion of factor X (inactive) to factor Xa. Initiation of the intrinsic pathway occurs when prekallikrein, high-molecular-weight kininogen, factor XI and factor XII are exposed to a negatively charged surface. This is termed the contact phase and can occur as a result of interaction with the phospholipids (primarily phosphatidylethanolamine, PE) of circulating lipoprotein particles such as chylomicrons and VLDLs. This is the basis of the role of hyperlipidemia in the promotion of a pro-thrombotic state and the development of atherosclerosis. Contact activation of the intrinsic pathway can also occur on the surface of bacteria.
The assemblage of contact phase components results in conversion of prekallikrein to kallikrein, which in turn activates factor XII to factor XIIa. Factor XIIa then activates factor XI to factor XIa. Factor XIIa will also hydrolyze more prekallikrein to kallikrein, establishing a reciprocal activation cascade. Kallikrein acts upon HMWK leading to the release of bradykinin, a potent vasodilator.
In the presence of Ca2+, factor XIa activates factor IX to factor IXa. Factor IX is a proenzyme that contains vitamin K-dependent γ-carboxyglutamate (gla) residues, whose serine protease activity is activated following Ca2+ binding to these gla residues. Several of the serine proteases of the cascade (II, VII, IX, and X) are gla-containing proenzymes. Active factor IXa cleaves factor X at an internal arg-ile (R-I) bond leading to its activation to factor Xa.
The activation of factor Xa requires assemblage of the tenase complex (Ca2+ and factors VIIIa, IXa and X) on the surface of activated platelets. One of the responses of platelets to activation is the presentation of phosphatidylserine (PS) and phosphatidylinositol (PI) on their surfaces. The exposure of these phospholipids allows the tenase complex to form. The role of factor VIII in this process is to act as a receptor, in the form of factor VIIIa, for factors IXa and X. Factor VIIIa is termed a cofactor in the clotting cascade. The activation of factor VIII to factor VIIIa (the actual receptor) occurs in the presence of minute quantities of thrombin. As the concentration of thrombin increases, factor VIIIa is ultimately cleaved by thrombin and inactivated. This dual action of thrombin, upon factor VIII, acts to limit the extent of tenase complex formation and thus the extent of the coagulation cascade.
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Extrinsic Clotting Cascade
Activated factor Xa is the site at which the intrinsic and extrinsic coagulation cascades converge. The extrinsic pathway is initiated at the site of injury in response to the release of tissue factor (factor III) and thus, is also known as the tissue factor pathway. Tissue factor is a cofactor in the factor VIIa-catalyzed activation of factor X. Factor VIIa, a gla residue containing serine protease, cleaves factor X to factor Xa in a manner identical to that of factor IXa of the intrinsic pathway. The activation of factor VII occurs through the action of thrombin or factor Xa. The ability of factor Xa to activate factor VII creates a link between the intrinsic and extrinsic pathways. An additional link between the two pathways exists through the ability of tissue factor and factor VIIa to activate factor IX. The formation of complex between factor VIIa and tissue factor is believed to be a principal step in the overall clotting cascade. Evidence for this stems from the fact that persons with hereditary deficiencies in the components of the contact phase of the intrinsic pathway do not exhibit clotting problems. A major mechanism for the inhibition of the extrinsic pathway occurs at the tissue factor-factor VIIa-Ca2+-Xa complex. The protein, lipoprotein-associated coagulation inhibitor, LACI specifically binds to this complex. LACI is also referred to as extrinsic pathway inhibitor, EPI or tissue factor pathway inhibitor, TFPI and was formerly named anticonvertin. LACI is composed of 3 tandem protease inhibitor domains. Domain 1 binds to factor Xa and domain 2 binds to factor VIIa only in the presence of factor Xa.
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Activation of Prothrombin to Thrombin
The common point in both pathways is the activation of factor X to factor Xa. Factor Xa activates prothrombin (factor II) to thrombin (factor IIa). Thrombin, in turn, converts fibrinogen to fibrin. The activation of thrombin occurs on the surface of activated platelets and requires formation of a prothrombinase complex. This complex is composed of the platelet phospholipids, phosphatidylinositol and phosphatidylserine, Ca2+, factors Va and Xa, and prothrombin. Factor V is a cofactor in the formation of the prothrombinase complex, similar to the role of factor VIII in tenase complex formation. Like factor VIII activation, factor V is activated to factor Va by means of minute amounts and is inactivated by increased levels of thrombin. Factor Va binds to specific receptors on the surfaces of activated platelets and forms a complex with prothrombin and factor Xa.
Prothrombin is a 72,000-Dalton, single-chain protein containing ten gla residues in its N-terminal region. Within the prothrombinase complex, prothrombin is cleaved at 2 sites by factor Xa. This cleavage generates a 2-chain active thrombin molecule containing an A and a B chain which are held together by a single disulfide bond.
In addition to its role in activation of fibrin clot formation, thrombin plays an important regulatory role in coagulation. Thrombin combines with thrombomodulin present on endothelial cell surfaces forming a complex that converts protein C to protein Ca. The cofactor protein S and protein Ca degrade factors Va and VIIIa, thereby limiting the activity of these 2 factors in the coagulation cascade.
Thrombin also binds to a class of G-protein-coupled receptors called protease activated receptors (PARs), specifically PAR-1, -3 and -4. PARs utilize a unique mechanism to convert the result of extracellular proteolytic cleavage into an intracellular signaling event. PARs carry their own ligand which remains inactive until protease cleavage, such as by thrombin, "unmasks" the ligand. Following thrombin cleavage the unmasked ligand is still a part of the intact PAR but is now capable of interacting with the ligand-binding domain of the PAR resulting in the activation of numerous signaling cascades. Because the activation of PARs requires proteolytic cleavage the activation process is irreversible. The signaling cascades activated by thrombin-activated PARs include release of the interleukins, (ILs), IL-1 and IL-6, increased secretion of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). The thrombin-induced signaling also leads to increased platelet activation and leukocyte adhesion.
Thrombin also activates thrombin-activatable fibrinolysis inhibitor (TAFI) thus modulating fibrinolysis (degradation of fibrin clots). TAFI is also known as carboxypeptidase U (CPU) whose activity leads to removal of C-terminal lysines from partially degraded fibrin. This leads to an impairment of plasminogen activation, thereby reducing the rate of fibrin clot dissolution (i.e. fibrinolysis).
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Control of Thrombin Levels
The inability of the body to control the circulating level of active thrombin would lead to dire consequences. There are 2 principal mechanisms by which thrombin activity is regulated. The predominant form of thrombin in the circulation is the inactive prothrombin, whose activation requires the pathways of proenzyme activation described above for the coagulation cascade. At each step in the cascade, feedback mechanisms regulate the balance between active and inactive enzymes.
The activation of thrombin is also regulated by 4 specific thrombin inhibitors Antithrombin III is the most important since it can also inhibit the activities of factors IXa, Xa, XIa and XIIa. The activity of antithrombin III is potentiated in the presence of heparin by the following means: heparin binds to a specific site on antithrombin III, producing an altered conformation of the protein, and the new conformation has a higher affinity for thrombin as well as its other substrates. This effect of heparin is the basis for its clinical use as an anticoagulant. The naturally occurring heparin activator of antithrombin III is present as heparan and heparan sulfate on the surface of vessel endothelial cells. It is this feature that controls the activation of the intrinsic coagulation cascade.
However, thrombin activity is also inhibited by α2-macroglobulin, heparin cofactor II and α1-antitrypsin. Although a minor player in thrombin regulation α1-antitrypsin is the primary serine protease inhibitor of human plasma. Its physiological significance is demonstrated by the fact that lack of this protein plays a causative role in the development of emphysema.
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Activation of Fibrinogen to Fibrin
Fibrinogen (factor I) consists of 3 pairs of polypeptides ([Aα][Bβ][γ])2. The 6 chains are covalently linked near their N-terminals through disulfide bonds. The A and B portions of the Aα and Bβ chains comprise the fibrinopeptides, A and B, respectively. The fibrinopeptide regions of fibrinogen contain several glutamate and aspartate residues imparting a high negative charge to this region and aid in the solubility of fibrinogen in plasma. Active thrombin is a serine protease that hydrolyses fibrinogen at four arg-gly (R-G) bonds between the fibrinopeptide and the a and b portions of the protein.
Thrombin-mediated release of the fibrinopeptides generates fibrin monomers with a subunit structure (αβγ)2. These monomers spontaneously aggregate in a regular array, forming a somewhat weak fibrin clot. In addition to fibrin activation, thrombin converts factor XIII to factor XIIIa, a highly specific transglutaminase that introduces cross-links composed of covalent bonds between the amide nitrogen of glutamines and e-amino group of lysines in the fibrin monomers.
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Dissolution of Fibrin Clots
Degradation of fibrin clots is the function of plasmin, a serine protease that circulates as the inactive proenzyme, plasminogen. Any free circulating plasmin is rapidly inhibited by α2-antiplasmin. Plasminogen binds to both fibrinogen and fibrin, thereby being incorporated into a clot as it is formed. Tissue plasminogen activator (tPA) and, to a lesser degree, urokinase are serine proteases which convert plasminogen to plasmin. Inactive tPA is released from vascular endothelial cells following injury; it binds to fibrin and is consequently activated. Urokinase is produced as the precursor, prourokinase by epithelial cells lining excretory ducts. The role of urokinase is to activate the dissolution of fibrin clots that may be deposited in these ducts.
Active tPA cleaves plasminogen to plasmin which then digests the fibrin; the result is soluble degradation product to which neither plasmin nor plasminogen can bind. Following the release of plasminogen and plasmin they are rapidly inactivated by their respective inhibitors. The inhibition of tPA activity results from binding to specific inhibitory proteins. At least 4 distinct inhibitors have been identified, of which 2: plasminogen activator-inhibitors type 1 (PAI-1) and type 2 (PAI-2) are of greatest physiological significance.
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Clinical Significances of Hemostasis
The Bleeding Disorders
Defects in the process of hemostasis, leading to bleeding disorders, have been identified at the level of the proteins of the clotting cascades, platelet activation and function, contact activation and antithrombin function This list is not all inclusive and for more details please visit the Inborn Errors: Clotting Factors page.
Hemophilia A
Hemophilia A is classic hemophilia (a disease referring to the inability to clot blood). It is an X-linked disorder resulting from a deficiency in factor VIII, a key component of the coagulation cascade. There are severe, moderate and mild forms of hemophilia A that reflect the level of active factor VIII in the plasma.
Hemophilia A arises from a variety of mutations. Some 150 different point mutations have been characterized in the factor VIII gene in hemophilia A. Inheritence of the disorder occurs with a frequency of 1:5,000 to 1:10,000 males in all populations. Factor VIII is a cofactor in the activation of factor X to factor Xa in a reaction catalyzed by factor IXa. Activation of factor VIII occurs via proteolytic cleavage by thrombin and factor Xa. Inactivaqtion of factor VIIIa occurs by limited proteolysis by factor Xa or activated protein C.
Individuals with deficiencies in factor VIII suffer joint and muscle hemorrhage, easy bruising and prolonged bleeding from wounds. Treatment of hemophilia A is accomplished by infusion of factor VIII concentrates prepared from either human plasma or by recombinant DNA technology.
Hemophilia B
Hemophilia B results from deficiencies in factor IX. The prevalence of hemophilia B is approximately one-tenth that of hemophilia A. All patients with hemophilia B have prolonged coagulation time and decreased factor IX clotting activity. Like hemophilia A, there are severe, moderate and mild forms of hemophilia B and reflect the factor IX activity in plasma.
At least 300 unique factor IX mutations have been identified, 85% are point mutations, 3% are short nucleotide deletions or insertions and 12% are gross gene alterations.
Disorders of Fibrinogen and Factor XIII
Several cardivascular risk factors are associated with abnormalities in fibrinogen. As a result of the acute-phase response or through other poorly understood mechanisms, elevated plasma fibrinogen levels have been observed in patients with coronary artery disease, diabetes, hypertension, peripheral artery disease, hyperlipoproteinemia and hypertriglyceridemia. In addition, pregnancy, menopause, hypercholesterolemia, use of oral contraceptives and smoking lead to increased plasma fibrinogen levels.
Although rare, there are inherited disorders in fibrinogen. These disorders include afibrinogenemia (a complete lack of fibrinogen), hypofibrinogenemia (reduced levels of fibrinogen) and dysfibrinogenemia (presence of dysfunctional fibrinogen). Afibrinogenemia is characterized by neonatal umbilical cord hemorrhage, ecchymoses, mucosal hemorrhage, internal hemorrhage, and recurrent abortion. The disorder is inherited in an autosomal recessive manner. Hypofibrinogenemia is characterized by fibrinogen levels below 100mg/dL (normal is 250-350mg/dL) and can be either aquired or inherited. Symptoms of hypofibrinogememia are similar to, but less severe than, afibrinogenemia. Dysfibrinogenemias are extremely heterogeneous affecting any of the functional properties of fibrinogen. Clinical consequences of dysfibrinogenemias include hemorrhage, spontaneous abortion and thromboembolism.
Factor XIII is the proenzyme form of plasma transglutaminase and is activated by thrombin in the presence of calcium ions. Active factor XIII catalyzes the cross-linking of fibrin monomers. Factor XIII is a tetramer of two different peptides, A and B (forming A2B2). Hereditary deficiencies (autosomal recessive) occur resulting in the absence of either subunit. Clinical manifestation of factor XIII deficiency is delayed bleeding although primary hemostasis is normal. Deficiency leads to neonatal umbilical cord bleeding, intracranial hemorrhage and soft tissue hematomas.
von Willebrand Disease
von Willebrand disease (vWD) is due to inherited deficiency in von Willebrand factor (vWF). vWD is the most common inherited bleeding disorder of humans. Using sensitive laboratory testing, abnormalities in vWF can be detected in approximately 8000 people per million. Clinically significant vWD occurs in approximatley 125 people per million. This is a frequency at least twice that of hemophilia A.
Deficiency of vWF results in defective platelet adhesion and causes a secondary deficiency in factor VIII. The result is that vWF deficiency can cause bleeding that appears similar to that caused by platelet dysfunction or hemophilia. vWD is an extremely heterogeneous disorder that has been classified into several major subtypes. Type I vWD is the most common and is inherited as an autosomal dominant trait. This variant is due to simple quantitative deficiency of all vWF multimers. Type 2 vWD is also subdivided further dependent upon whether the dysfunctional protein has decreased or paradoxically increased function in certain laboratory tests of binding to platelets. Type 3 vWD is clinically severe and is characterized by recessive inheritance and virtual absence of vWF.
Factor XI and Contact Activation
When blood makes contact with negatively charged surfaces it triggers a series of interactions that involve factor XI, prekallikrein and high molecular weight kininogen leading to blood coagulation. This process is referred to as contact activation. Deficiency in factor XI confers an injury-related bleeding tendency. This deficiency was identified in 1953 and originally termed hemophilia C. Factor XI deficiency is very common in Ashkenazic Jews and is inherited as an autosomal disorder with either homozygosity or compound heterozygosity. Three independent point mutations in factor XI have been identified.
Antithrombin Deficiency
Antithrombin functions to inhibit several activated coagulation factors including thrombin, factor IXa and factor Xa, by forming a stable complex with the various factors. Heparin and heparan sulfates increase the activity of antithrombin at least 1000 fold.
Deficiency in antithrombin is seen in approximately 2% of patients with venous thromboembolic disease. Inheritance occurs as an autosomal dominant trait. The prevalence of symptomatic antithrombin deficiency ranges from 1 per 2000 to 1 per 5000 in the general population. Deficiencies results from mutations that affect synthesis or stability of antithrombin or from mutations that affect the protease and/or heparin binding sites of antithrombin.
Clinical manifestations of antithrombin deficiency include deep vein thrombosis and pulmonary embolism. Arterial thrombosis is rare in anththrombin deficiency. Thrombosis may occur spontaneously or in association with surgery, trauma or pregnancy. Treatment of acute episodes of thrombosis is by infusion of heparin (for 5-7 days) followed by oral anticoagulant therapy.
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Pharmacological Intervention in Bleeding
Coumarin drugs (based on the chemical benzopyrone), such as warfarin (trade name Coumadin®) as well as the glycosaminoglycans, heparin and heparan sulfate, are useful as anticoagulants. Heparin is useful as an anticoagulant because it binds to, and activates, antithrombin III which then inhibits the serine proteases of the coagulation cascade. Heparin is abundant in granules of the mast cells that line the vasculature. In response to injury, the heparin is released and inhibits coagulation. The coumarin drugs inhibit coagulation by inhibiting the vitamin K-dependent γ-carboxylation reactions necessary to the function of thrombin, and factors VII, IX, and X as well as proteins C and S. These drugs act by inhibiting the reduction of the quinone derivatives of vitamin K to their active hydroquinone forms. Because of the mode of action of coumarin drugs, it takes several days for their maximum effect to be realized. For this reason, heparin is normally administered first followed by warfarin or warfarin-related drugs.
The plasminogen activators also are useful for controlling coagulation. Because tPA is highly selective for the degradation of fibrin in clots, it is extremely useful in restoring the patency of the coronary arteries following thrombosis, in particular during the short period following myocardial infarct. Streptokinase (an enzyme from the Streptococci bacterium) is another plasminogen activator useful from a therapeutic standpoint. However, it is less selective than tPA, being able to activate circulating plasminogen as well as that bound to a fibrin clot.
Aspirin is an important inhibitor of platelet activation. By virtue of inhibiting the activity of cyclooxygenase, aspirin reduces the production of TXA2 by platelets. Aspirin also reduces endothelial cell production of prostacyclin (PGI2), an inhibitor of platelet aggregation and a vasodilator. Localized to the site of coagulation is a balance between the levels of platelet derived TXA2 and endothelial cell derived PGI2. This allows for platelet aggregation and clot formation but preventing excessive accumulation of the clot, thus maintaining blood flow around the site of the clot. Endothelial cells regenerate active cyclooxygenase faster than platelets because mature platelets cannot synthesize the enzyme, requiring new platelets to enter the circulation (platelet half-life is approximately 4 days). Therefore, PGI2 synthesis is greater than that of TXA2. The net effect of aspirin is more in favor of endothelial cell-mediated inhibition of the coagulation cascade. This reflects one of the the cardiovascular benefits to low dose administration of aspirin. Aspirin also has important effects on inflammatory processes that impact cardiovascular systems (see the Aspirin page for more details).
Newer classes of anticoagulation drugs are being developed that function by inhibiting the activation of platelets and their subsequent aggregation. The drug clopidogrel: Plavix® (Bristol-Myers Squibb) is an irreversible inhibitor of the ADP receptor on platelet membranes. When ADP binds to platelets they are activated and aggregate leading to amplification of the coagulation response, thus Plavix interferes with this process. Plavix is prescribed for the treatment of peripheral vascular and cerebrovascular disease as well as coronary artery disease to prevent the formation of thrombotic plaques.
Another target of pharmacologic intervention in coagulation involving platelets is the role of GPIIb-GPIIIa in fibrinogen-induced platelet aggregation. Glanzmann thrombasthenia is an inherited platelet function defect characterized by a lack of platelet aggregation in response to all physiological agonists. This disorder is due to a lack of the GPIIb-GPIIIa receptor complex on platelets. Patients with this disorder present with significantly increased bleeding times. Although a rare disorder, study of the pathophysiology of the disease led to the development of anticoagulant drugs that inhibit platelet aggregation regardless of the agonist. Therefore, the GPIIb-GPIIIa antagonists more completely inhibit platelet aggregation than do aspirin or Plavix. The current family of these drugs includes ReoPro® (abciximab: a human monoclonal antibody), Integrilin® (eptifibatide: a cyclic hexapeptide derived from a protein found in the venom of the southeastern pygmy rattlesnake) and Aggrastat® (tirofiban: a synthetic organic non-peptide molecule).
Platelet Activation and von Willebrand Factor (vWF)
Description of Clotting Factors
Image of the Clotting Cascade
Kallikrein-Kinin System in Coagulation
Intrinsic (contact activation) Clotting Cascade
Extrinsic (tissue factor) Clotting Cascade
Activation of Thrombin
Regulation of Thrombin Levels
Activation of Fibrin
Dissolution of Fibrin Clots
Clinical Significances of Hemostasis: The Bleeding Disorders
Pharmacological Intervention in Bleeding
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Introduction
The ability of the body to control the flow of blood following vascular injury is paramount to continued survival. The process of blood clotting and then the subsequent dissolution of the clot, following repair of the injured tissue, is termed hemostasis. Hemostasis, composed of 4 major events that occur in a set order following the loss of vascular integrity:
1. The initial phase of the process is vascular constriction. This limits the flow of blood to the area of injury.
2. Next, platelets become activated by thrombin and aggregate at the site of injury, forming a temporary, loose platelet plug. The protein fibrinogen is primarily responsible for stimulating platelet clumping. Platelets clump by binding to collagen that becomes exposed following rupture of the endothelial lining of vessels. Upon activation, platelets release the nucleotide, ADP and the eicosanoid, TXA2 (both of which activate additional platelets), serotonin, phospholipids, lipoproteins, and other proteins important for the coagulation cascade. In addition to induced secretion, activated platelets change their shape to accommodate the formation of the plug.
3. To insure stability of the initially loose platelet plug, a fibrin mesh (also called the clot) forms and entraps the plug. If the plug contains only platelets it is termed a white thrombus; if red blood cells are present it is called a red thrombus
4. Finally, the clot must be dissolved in order for normal blood flow to resume following tissue repair. The dissolution of the clot occurs through the action of plasmin
Two pathways lead to the formation of a fibrin clot: the intrinsic and extrinsic pathway. Although they are initiated by distinct mechanisms, the two converge on a common pathway that leads to clot formation. Both pathways are complex and involve numerous different proteins termed clotting factors. Fibrin clot formation in response to tissue injury is the most clinically relevant event of hemostasis under normal physiological conditions. This process is the result of the activation of the extrinsic pathway. The formation of a red thrombus or a clot in response to an abnormal vessel wall in the absence of tissue injury is the result of the intrinsic pathway. The intrinsic pathway has low significance under normal physiological conditions. Most significant clinically is the activation of the intrinsic pathway by contact of the vessel wall with lipoprotein particles, VLDLs and chylomicrons. This process clearly demonstrates the role of hyperlipidemia in the generation of atherosclerosis. The intrinsic pathway can also be activated by vessel wall contact with bacteria.
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Platelet Activation and von Willebrand Factor (vWF)
In order for hemostasis to occur, platelets must adhere to exposed collagen, release the contents of their granules, and aggregate. The adhesion of platelets to the collagen exposed on endothelial cell surfaces is mediated by von Willebrand factor (vWF). Inherited deficiencies of vWF are the causes of von Willebrand disease, (vWD) (also see below for more details). The function of vWF is to act as a bridge between a specific glycoprotein complex on the surface of platelets (GPIb-GPIX-GPV) and collagen fibrils. The importance of this interaction between vWF and the GPIb-GPIX-GPV complex of platelets is demonstrated by the inheritance of bleeding disorders caused by defects in any one of the four the proteins of the complex, the most common of which is Bernard-Soulier syndrome (also called giant platelet syndrome). The GPIb part of the complex is composed of two proteins, GPIbα and GPIbβ encoded by separate genes.
In addition to its role as a bridge between platelets and exposed collagen on endothelial surfaces, vWF binds to and stabilizes coagulation factor VIII. Binding of factor VIII by vWF is required for normal survival of factor VIII in the circulation.
von Willebrand factor is a complex multimeric glycoprotein that is produced by and stored in the α-granules of platelets. It is also synthesized by megakaryocytes and found associated with subendothelial connective tissue.
The initial activation of platelets is induced by thrombin binding to specific receptors on the surface of platelets, thereby initiating a signal transduction cascade. The thrombin receptor is coupled to a G-protein that, in turn, activates phospholipase C-γ (PLC-γ). PLC-γ hydrolyzes phosphatidylinositol-4,5-bisphosphate (PIP2) leading to the formation of inositol trisphosphate (IP3) and diacylglycerol (DAG). IP3 induces the release of intracellular Ca2+ stores, and DAG activates protein kinase C (PKC).
The collagen to which platelets adhere as well as the release of intracellular Ca2+ leads to the activation of phospholipase A2 (PLA2), which then hydrolyzes membrane phospholipids, leading to liberation of arachidonic acid. The arachidonic acid release leads to an increase in the production and subsequent release of thromboxane A2 (TXA2). TXA2 is a potent vasoconstrictor and inducer of platelet aggregation that functions by binding to receptors that function through the PLC-γ pathway.
Another enzyme activated by the released intracellular Ca2+ stores is myosin light chain kinase (MLCK). Activated MLCK phosphorylates the light chain of myosin which then interacts with actin, resulting in altered platelet morphology and motility.
One of the many effects of PKC is the phosphorylation and activation of a specific 47,000-Dalton platelet protein. This activated protein induces the release of platelet granule contents; one of which is ADP. ADP further stimulates platelets increasing the overall activation cascade. The important role of ADP in platelet activation can be appreciated from the use of the ADP receptor antagonist, Plavix® (clopidogrel), in the control of thrombosis (see below). ADP also modifies the platelet membranes leading to exposure platelet glycoprotein receptor complex: GPIIb-GPIIIa. GPIIb-GPIIIa constitutes a receptor for vWF and fibrinogen, resulting in fibrinogen-induced platelet aggregation. The GPIIb-GPIIIa complex is a member of the integrin family of cell-surface receptors that interact with the extracellular matrix. The GPIIb-GPIIIa complex is also called integrin αIIb-β3. The importance of the GPIIb-GPIIIa in platelet activation and coagulation is demonstrated by the fact that bleeding disorders result from inherited defects in this glycoprotein complex. The most commonly inherited platelet dysfunction is Glanzmann thrombasthenia which results from defects in the GPIIb protein of this complex. In addition, the importance of this complex in overall hemostasis is demonstrated by the use of antibodies that block this receptor as anti-coagulants (e.g. ReoPro®, abciximab: see below).
Activation of platelets is required for their consequent aggregation to a platelet plug. However, equally significant is the role of activated platelet surface phospholipids in the activation of the coagulation cascade.
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Primary Factors
Factor
Trivial Name(s)
Pathway
Characteristic
Prekallikrein (PK) Fletcher factor Intrinsic Functions with HMWK and factor XII
High molecular weight kininogen (HMWK) contact activation cofactor; Fitzgerald, Flaujeac Williams factor Intrinsic Co-factor in kallikrein and factor XII activation, necessary in factor XIIa activation of XI, precursor for bradykinin (a potent vasodilator and inducer of smooth muscle contraction
I Fibrinogen Both
II Prothrombin Both Contains N-term. gla segment
III Tissue Factor Extrinsic
IV Calcium Both
V Proaccelerin, labile factor, accelerator (Ac-) globulin Both Protein cofactor
VI (same as Va) Accelerin Both This is Va, redundant to Factor V
VII Proconvertin, serum prothrombin conversion accelerator (SPCA), cothromboplastin Extrinsic Endopeptidase with gla residues
VIII Antihemophiliac factor A, antihemophilic globulin (AHG) Intrinsic Protein cofactor
IX Christmas Factor, antihemophilic factor B,plasma thromboplastin component (PTC) Intrinsic Endopeptidase with gla residues
X Stuart-Prower Factor Both Endopeptidase with gla residues
XI Plasma thromboplastin antecedent (PTA) Intrinsic Endopeptidase
XII Hageman Factor Intrinsic Endopeptidase
XIII Protransglutaminase, fibrin stabilizing factor (FSF), fibrinoligase Both Transpeptidase
Functional Classification of Clotting Factors
Zymogens of Serine Proteases
Activities
Factor XII binds to exposed collagen at site of vessel wall injury, activated by high-MW kininogen and kallikrein
Factor XI activated by factor XIIa
Factor IX activated by factor XIa in presence of Ca2+
Factor VII activated by thrombin in presence of Ca2+
Factor X activated on surface of activated platelets by tenase complex and by factor VIIa in presence of tissue factor and Ca2+
Factor II activated on surface of activated platelets by prothrombinase complex
Cofactors
Activities
Factor VIII activated by thrombin; factor VIIIa is a cofactor in the activation of factor X by factor IXa
Factor V activated by thrombin; factor Va is a cofactor in the activation of prothrombin by factor Xa
Factor III (tissue factor) a subendothelial cell-surface glycoprotein that acts as a cofactor for factor VII
Fibrinogen
Activity
Factor I cleaved by thrombin to form fibrin clot
Transglutaminase
Activity
Factor XIII activated by thrombin in presence of Ca2+; stabilizes fibrin clot by covalent cross-linking
Regulatory/Other Proteins
Activities
von Willebrand factor associated with subendothelial connective tissue; serves as a bridge between platelet glycoprotein GPIb/IX and collagen
Protein C activated to protein Ca by thrombin bound to thrombomodulin; then degrades factors VIIIa and Va
Protein S acts as a cofactor of protein C; both proteins contain gla residues
Thrombomodulin protein on the surface of endothelial cells; binds thrombin, which then activates protein C
Antithrombin III most important coagulation inhibitor, controls activities of thrombin, and factors IXa, Xa, XIa and XIIa
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The Clotting Cascades
The clotting cascades: The intrinsic cascade (which has less in vivo significance in normal physiological circumstances than the extrinsic cascade) is initiated when contact is made between blood and exposed negatively charged surfaces. The extrinsic pathway is initiated upon vascular injury which leads to exposure of tissue factor, TF (also identified as factor III), a subendothelial cell-surface glycoprotein that binds phospholipid. The green dotted arrow represents a point of cross-over between the extrinsic and intrinsic pathways. The two pathways converge at the activation of factor X to Xa. Factor Xa has a role in the further activation of factor VII to VIIa as depicted by the green arrow. Active factor Xa hydrolyzes and activates prothrombin to thrombin. Thrombin can then activate factors XI, VIII and V furthering the cascade. Ultimately the role of thrombin is to convert fribrinogen to fibrin and to activate factor XIII to XIIIa. Factor XIIIa (also termed transglutaminase) cross-links fibrin polymers solidifying the clot. HMWK = high molecular weight kininogen. PK = prekallikrein. PL = phospholipid.
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The Kallikrein-Kinin System in Coagulation
The kallikrein-kinin system comprises a complex of proteins that when activated leads to the release of vasoactive kinins. The kinins are released from both high molecular weight kininogen (HMWK) and low molecular weight kininogen (LMWK) as a result of activation of either tissue kallikrein or plasma kallikrein. The kallekreins themselves exist in inactive pre-forms. The kinins are involved in many physiological and pathological processes including regulation of blood pressure and flow (via modulation of the renin-angiotensin pathway), blood coagulation, cellular proliferation and growth, angiogenesis, apoptosis, and inflammation. Kinin action on endothelial cells leads to vasodilation, increased vascular permeability, release of tissue plasminogen activator (tPA), production of nitric oxide (NO), and the mobilization of arachidonic acid, primarily resulting in prostacyclin (PGI2) production by endothelial cells. Although the activities of the kallikrein-kinin system are involved in numerous processes, this section will deal only with their function in blood coagulation. With respect to hemostasis the most important kinin is bradykinin which is released from HMWK.
The two forms of prekallikrein, plasma and tissue, are derived from distinct genes on different chromosomes. The plasma kallikrein gene (symbol KLKB1) is on chromosome 4q34-q35 and the tissue kallikrein gene (symbol KLK1) located on chromosome 19q13.2-q13.4. These two kallikreins are serine proteases whose major substrates are HMWK (plasma kallikrein) and LMWK (tissue kallikrein). When plasma prekallikrein is activated to kallikrein it cleaves HMWK in a two-step process that ultimately releases bradykinin. Bradykinin is a 9-amino acid vasoactive peptide that induces vasodilation and increases vascular permeability. Activated tissue kallikrein cleaves lysyl-bradykinin (also called kallidin) from LMWK. Lysyl-bradykinin is bradykinin with a lysine residue at the N-terminus making it a 10-amino acid vasoactive peptide. Its activities are essentially identical to those of bradykinin.
Both HMWK and LMWK are derived from the same gene on chromosome 3q26-qter which is composed of 11 exons. Exons 1 to 9 encode the heavy chain of both kininogens. Exon 10 encodes bradykinin as well as the light chain of HMWK. Exon 11 encodes the light chain of LMWK. The heavy and light chain nomenclature refers to the disulfide-bonded structure of each kininogen after their activation by through kallikrein cleavage.
HMWK is considered an α-globulin and is composed of six functional domains. The protein circulates in the plasma as single-chain polypeptide with a molecular weight of 88 - 120 kDa dependent upon the level of glycosylation. The heavy chain is 64 kDa and contains domains 1, 2, and 3 whereas the light chain is 45 - 56 kDa and comprises domains 5 and 6. The heavy and light chains are linked together through domain 4 which also contains the bradykinin sequence. Domain 1 contains a low affinity calcium-binding site. Domains 2 and 3 contain amino acid sequences (QVVAG) that inhibit cysteine proteases. Domain 3 also has platelet and endothelial cell-binding activity. Domain 5 has sequences for heparin binding, cell-binding sites, and antiangiogenic properties. The binding of HMWK to negatively charged surfaces occurs through a histidine region of the light chain which is in domain 5. Domain 6 contains the prekallikrein and factor XI-binding sites. By being able to bind to charged surfaces via domain 5 and simultaneously bind factor XI and prekallikrein via domain 6, HMWK can serve as the cofactor in contact activation of plasma. LMWK is considered a β-globulin and has a molecular weight of 50 - 68 kDa. The structure of LMWK is similar to that of HMWK, however, the light chain is only 4 - 5 kDa and has no contact activation nor prekallikrein-binding sites.
The plasma kinin forming system is called the contact system of plasma and is composed of factor XII, factor XI, prekallikrein and HMWK. Factor XII, prekallikrein, and HMWK saturably and reversibly bind to endothelial cells, platelets, and granulocytes in a zinc-dependent reaction. When plasma makes contact with a negatively charged surface factor XII binds and is autoactivated to factor XIIa (the "a" signifies the activated factor). Factor XIIa then activates prekallikrein to kallikrein and kallikrein cleaves HMWK releasing bradykinin. There is also reciprocal activation of factor XII by kallikrein resulting in amplification of the system. The actual surface that leads to factor XII autoactivation is unknown however, several physiologic substances support the process. These substances include hematin, skin, fatty acids, sodium urate crystals, protoporphyrin, sulfatides, heparins, chondroitin sulfates, articular cartilage, endotoxin, L-homocysteine, and amyloid β-protein. Once the contact system is activated the intrinsic pathway (described below) is initiated.
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The Intrinsic Clotting Cascade
The intrinsic pathway (also called the contact activation pathway) is much less significant to hemostasis under normal physiological conditions than is the extrinsic pathway. However, abnormal physiology such as hyperlipidemic states or bacterial infiltration can lead to activation of thrombosis via the intrinsic clotting cascade.
The intrinsic pathway requires the clotting factors VIII, IX, X, XI, and XII. Also required are the proteins prekallikrein (PK) and high-molecular-weight kininogen (HK or HMWK), as well as calcium ions and phospholipids secreted from platelets. The role of PK and HMWK is described in the above section. Each of these pathway constituents leads to the conversion of factor X (inactive) to factor Xa. Initiation of the intrinsic pathway occurs when prekallikrein, high-molecular-weight kininogen, factor XI and factor XII are exposed to a negatively charged surface. This is termed the contact phase and can occur as a result of interaction with the phospholipids (primarily phosphatidylethanolamine, PE) of circulating lipoprotein particles such as chylomicrons and VLDLs. This is the basis of the role of hyperlipidemia in the promotion of a pro-thrombotic state and the development of atherosclerosis. Contact activation of the intrinsic pathway can also occur on the surface of bacteria.
The assemblage of contact phase components results in conversion of prekallikrein to kallikrein, which in turn activates factor XII to factor XIIa. Factor XIIa then activates factor XI to factor XIa. Factor XIIa will also hydrolyze more prekallikrein to kallikrein, establishing a reciprocal activation cascade. Kallikrein acts upon HMWK leading to the release of bradykinin, a potent vasodilator.
In the presence of Ca2+, factor XIa activates factor IX to factor IXa. Factor IX is a proenzyme that contains vitamin K-dependent γ-carboxyglutamate (gla) residues, whose serine protease activity is activated following Ca2+ binding to these gla residues. Several of the serine proteases of the cascade (II, VII, IX, and X) are gla-containing proenzymes. Active factor IXa cleaves factor X at an internal arg-ile (R-I) bond leading to its activation to factor Xa.
The activation of factor Xa requires assemblage of the tenase complex (Ca2+ and factors VIIIa, IXa and X) on the surface of activated platelets. One of the responses of platelets to activation is the presentation of phosphatidylserine (PS) and phosphatidylinositol (PI) on their surfaces. The exposure of these phospholipids allows the tenase complex to form. The role of factor VIII in this process is to act as a receptor, in the form of factor VIIIa, for factors IXa and X. Factor VIIIa is termed a cofactor in the clotting cascade. The activation of factor VIII to factor VIIIa (the actual receptor) occurs in the presence of minute quantities of thrombin. As the concentration of thrombin increases, factor VIIIa is ultimately cleaved by thrombin and inactivated. This dual action of thrombin, upon factor VIII, acts to limit the extent of tenase complex formation and thus the extent of the coagulation cascade.
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Extrinsic Clotting Cascade
Activated factor Xa is the site at which the intrinsic and extrinsic coagulation cascades converge. The extrinsic pathway is initiated at the site of injury in response to the release of tissue factor (factor III) and thus, is also known as the tissue factor pathway. Tissue factor is a cofactor in the factor VIIa-catalyzed activation of factor X. Factor VIIa, a gla residue containing serine protease, cleaves factor X to factor Xa in a manner identical to that of factor IXa of the intrinsic pathway. The activation of factor VII occurs through the action of thrombin or factor Xa. The ability of factor Xa to activate factor VII creates a link between the intrinsic and extrinsic pathways. An additional link between the two pathways exists through the ability of tissue factor and factor VIIa to activate factor IX. The formation of complex between factor VIIa and tissue factor is believed to be a principal step in the overall clotting cascade. Evidence for this stems from the fact that persons with hereditary deficiencies in the components of the contact phase of the intrinsic pathway do not exhibit clotting problems. A major mechanism for the inhibition of the extrinsic pathway occurs at the tissue factor-factor VIIa-Ca2+-Xa complex. The protein, lipoprotein-associated coagulation inhibitor, LACI specifically binds to this complex. LACI is also referred to as extrinsic pathway inhibitor, EPI or tissue factor pathway inhibitor, TFPI and was formerly named anticonvertin. LACI is composed of 3 tandem protease inhibitor domains. Domain 1 binds to factor Xa and domain 2 binds to factor VIIa only in the presence of factor Xa.
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Activation of Prothrombin to Thrombin
The common point in both pathways is the activation of factor X to factor Xa. Factor Xa activates prothrombin (factor II) to thrombin (factor IIa). Thrombin, in turn, converts fibrinogen to fibrin. The activation of thrombin occurs on the surface of activated platelets and requires formation of a prothrombinase complex. This complex is composed of the platelet phospholipids, phosphatidylinositol and phosphatidylserine, Ca2+, factors Va and Xa, and prothrombin. Factor V is a cofactor in the formation of the prothrombinase complex, similar to the role of factor VIII in tenase complex formation. Like factor VIII activation, factor V is activated to factor Va by means of minute amounts and is inactivated by increased levels of thrombin. Factor Va binds to specific receptors on the surfaces of activated platelets and forms a complex with prothrombin and factor Xa.
Prothrombin is a 72,000-Dalton, single-chain protein containing ten gla residues in its N-terminal region. Within the prothrombinase complex, prothrombin is cleaved at 2 sites by factor Xa. This cleavage generates a 2-chain active thrombin molecule containing an A and a B chain which are held together by a single disulfide bond.
In addition to its role in activation of fibrin clot formation, thrombin plays an important regulatory role in coagulation. Thrombin combines with thrombomodulin present on endothelial cell surfaces forming a complex that converts protein C to protein Ca. The cofactor protein S and protein Ca degrade factors Va and VIIIa, thereby limiting the activity of these 2 factors in the coagulation cascade.
Thrombin also binds to a class of G-protein-coupled receptors called protease activated receptors (PARs), specifically PAR-1, -3 and -4. PARs utilize a unique mechanism to convert the result of extracellular proteolytic cleavage into an intracellular signaling event. PARs carry their own ligand which remains inactive until protease cleavage, such as by thrombin, "unmasks" the ligand. Following thrombin cleavage the unmasked ligand is still a part of the intact PAR but is now capable of interacting with the ligand-binding domain of the PAR resulting in the activation of numerous signaling cascades. Because the activation of PARs requires proteolytic cleavage the activation process is irreversible. The signaling cascades activated by thrombin-activated PARs include release of the interleukins, (ILs), IL-1 and IL-6, increased secretion of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). The thrombin-induced signaling also leads to increased platelet activation and leukocyte adhesion.
Thrombin also activates thrombin-activatable fibrinolysis inhibitor (TAFI) thus modulating fibrinolysis (degradation of fibrin clots). TAFI is also known as carboxypeptidase U (CPU) whose activity leads to removal of C-terminal lysines from partially degraded fibrin. This leads to an impairment of plasminogen activation, thereby reducing the rate of fibrin clot dissolution (i.e. fibrinolysis).
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Control of Thrombin Levels
The inability of the body to control the circulating level of active thrombin would lead to dire consequences. There are 2 principal mechanisms by which thrombin activity is regulated. The predominant form of thrombin in the circulation is the inactive prothrombin, whose activation requires the pathways of proenzyme activation described above for the coagulation cascade. At each step in the cascade, feedback mechanisms regulate the balance between active and inactive enzymes.
The activation of thrombin is also regulated by 4 specific thrombin inhibitors Antithrombin III is the most important since it can also inhibit the activities of factors IXa, Xa, XIa and XIIa. The activity of antithrombin III is potentiated in the presence of heparin by the following means: heparin binds to a specific site on antithrombin III, producing an altered conformation of the protein, and the new conformation has a higher affinity for thrombin as well as its other substrates. This effect of heparin is the basis for its clinical use as an anticoagulant. The naturally occurring heparin activator of antithrombin III is present as heparan and heparan sulfate on the surface of vessel endothelial cells. It is this feature that controls the activation of the intrinsic coagulation cascade.
However, thrombin activity is also inhibited by α2-macroglobulin, heparin cofactor II and α1-antitrypsin. Although a minor player in thrombin regulation α1-antitrypsin is the primary serine protease inhibitor of human plasma. Its physiological significance is demonstrated by the fact that lack of this protein plays a causative role in the development of emphysema.
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Activation of Fibrinogen to Fibrin
Fibrinogen (factor I) consists of 3 pairs of polypeptides ([Aα][Bβ][γ])2. The 6 chains are covalently linked near their N-terminals through disulfide bonds. The A and B portions of the Aα and Bβ chains comprise the fibrinopeptides, A and B, respectively. The fibrinopeptide regions of fibrinogen contain several glutamate and aspartate residues imparting a high negative charge to this region and aid in the solubility of fibrinogen in plasma. Active thrombin is a serine protease that hydrolyses fibrinogen at four arg-gly (R-G) bonds between the fibrinopeptide and the a and b portions of the protein.
Thrombin-mediated release of the fibrinopeptides generates fibrin monomers with a subunit structure (αβγ)2. These monomers spontaneously aggregate in a regular array, forming a somewhat weak fibrin clot. In addition to fibrin activation, thrombin converts factor XIII to factor XIIIa, a highly specific transglutaminase that introduces cross-links composed of covalent bonds between the amide nitrogen of glutamines and e-amino group of lysines in the fibrin monomers.
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Dissolution of Fibrin Clots
Degradation of fibrin clots is the function of plasmin, a serine protease that circulates as the inactive proenzyme, plasminogen. Any free circulating plasmin is rapidly inhibited by α2-antiplasmin. Plasminogen binds to both fibrinogen and fibrin, thereby being incorporated into a clot as it is formed. Tissue plasminogen activator (tPA) and, to a lesser degree, urokinase are serine proteases which convert plasminogen to plasmin. Inactive tPA is released from vascular endothelial cells following injury; it binds to fibrin and is consequently activated. Urokinase is produced as the precursor, prourokinase by epithelial cells lining excretory ducts. The role of urokinase is to activate the dissolution of fibrin clots that may be deposited in these ducts.
Active tPA cleaves plasminogen to plasmin which then digests the fibrin; the result is soluble degradation product to which neither plasmin nor plasminogen can bind. Following the release of plasminogen and plasmin they are rapidly inactivated by their respective inhibitors. The inhibition of tPA activity results from binding to specific inhibitory proteins. At least 4 distinct inhibitors have been identified, of which 2: plasminogen activator-inhibitors type 1 (PAI-1) and type 2 (PAI-2) are of greatest physiological significance.
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Clinical Significances of Hemostasis
The Bleeding Disorders
Defects in the process of hemostasis, leading to bleeding disorders, have been identified at the level of the proteins of the clotting cascades, platelet activation and function, contact activation and antithrombin function This list is not all inclusive and for more details please visit the Inborn Errors: Clotting Factors page.
Hemophilia A
Hemophilia A is classic hemophilia (a disease referring to the inability to clot blood). It is an X-linked disorder resulting from a deficiency in factor VIII, a key component of the coagulation cascade. There are severe, moderate and mild forms of hemophilia A that reflect the level of active factor VIII in the plasma.
Hemophilia A arises from a variety of mutations. Some 150 different point mutations have been characterized in the factor VIII gene in hemophilia A. Inheritence of the disorder occurs with a frequency of 1:5,000 to 1:10,000 males in all populations. Factor VIII is a cofactor in the activation of factor X to factor Xa in a reaction catalyzed by factor IXa. Activation of factor VIII occurs via proteolytic cleavage by thrombin and factor Xa. Inactivaqtion of factor VIIIa occurs by limited proteolysis by factor Xa or activated protein C.
Individuals with deficiencies in factor VIII suffer joint and muscle hemorrhage, easy bruising and prolonged bleeding from wounds. Treatment of hemophilia A is accomplished by infusion of factor VIII concentrates prepared from either human plasma or by recombinant DNA technology.
Hemophilia B
Hemophilia B results from deficiencies in factor IX. The prevalence of hemophilia B is approximately one-tenth that of hemophilia A. All patients with hemophilia B have prolonged coagulation time and decreased factor IX clotting activity. Like hemophilia A, there are severe, moderate and mild forms of hemophilia B and reflect the factor IX activity in plasma.
At least 300 unique factor IX mutations have been identified, 85% are point mutations, 3% are short nucleotide deletions or insertions and 12% are gross gene alterations.
Disorders of Fibrinogen and Factor XIII
Several cardivascular risk factors are associated with abnormalities in fibrinogen. As a result of the acute-phase response or through other poorly understood mechanisms, elevated plasma fibrinogen levels have been observed in patients with coronary artery disease, diabetes, hypertension, peripheral artery disease, hyperlipoproteinemia and hypertriglyceridemia. In addition, pregnancy, menopause, hypercholesterolemia, use of oral contraceptives and smoking lead to increased plasma fibrinogen levels.
Although rare, there are inherited disorders in fibrinogen. These disorders include afibrinogenemia (a complete lack of fibrinogen), hypofibrinogenemia (reduced levels of fibrinogen) and dysfibrinogenemia (presence of dysfunctional fibrinogen). Afibrinogenemia is characterized by neonatal umbilical cord hemorrhage, ecchymoses, mucosal hemorrhage, internal hemorrhage, and recurrent abortion. The disorder is inherited in an autosomal recessive manner. Hypofibrinogenemia is characterized by fibrinogen levels below 100mg/dL (normal is 250-350mg/dL) and can be either aquired or inherited. Symptoms of hypofibrinogememia are similar to, but less severe than, afibrinogenemia. Dysfibrinogenemias are extremely heterogeneous affecting any of the functional properties of fibrinogen. Clinical consequences of dysfibrinogenemias include hemorrhage, spontaneous abortion and thromboembolism.
Factor XIII is the proenzyme form of plasma transglutaminase and is activated by thrombin in the presence of calcium ions. Active factor XIII catalyzes the cross-linking of fibrin monomers. Factor XIII is a tetramer of two different peptides, A and B (forming A2B2). Hereditary deficiencies (autosomal recessive) occur resulting in the absence of either subunit. Clinical manifestation of factor XIII deficiency is delayed bleeding although primary hemostasis is normal. Deficiency leads to neonatal umbilical cord bleeding, intracranial hemorrhage and soft tissue hematomas.
von Willebrand Disease
von Willebrand disease (vWD) is due to inherited deficiency in von Willebrand factor (vWF). vWD is the most common inherited bleeding disorder of humans. Using sensitive laboratory testing, abnormalities in vWF can be detected in approximately 8000 people per million. Clinically significant vWD occurs in approximatley 125 people per million. This is a frequency at least twice that of hemophilia A.
Deficiency of vWF results in defective platelet adhesion and causes a secondary deficiency in factor VIII. The result is that vWF deficiency can cause bleeding that appears similar to that caused by platelet dysfunction or hemophilia. vWD is an extremely heterogeneous disorder that has been classified into several major subtypes. Type I vWD is the most common and is inherited as an autosomal dominant trait. This variant is due to simple quantitative deficiency of all vWF multimers. Type 2 vWD is also subdivided further dependent upon whether the dysfunctional protein has decreased or paradoxically increased function in certain laboratory tests of binding to platelets. Type 3 vWD is clinically severe and is characterized by recessive inheritance and virtual absence of vWF.
Factor XI and Contact Activation
When blood makes contact with negatively charged surfaces it triggers a series of interactions that involve factor XI, prekallikrein and high molecular weight kininogen leading to blood coagulation. This process is referred to as contact activation. Deficiency in factor XI confers an injury-related bleeding tendency. This deficiency was identified in 1953 and originally termed hemophilia C. Factor XI deficiency is very common in Ashkenazic Jews and is inherited as an autosomal disorder with either homozygosity or compound heterozygosity. Three independent point mutations in factor XI have been identified.
Antithrombin Deficiency
Antithrombin functions to inhibit several activated coagulation factors including thrombin, factor IXa and factor Xa, by forming a stable complex with the various factors. Heparin and heparan sulfates increase the activity of antithrombin at least 1000 fold.
Deficiency in antithrombin is seen in approximately 2% of patients with venous thromboembolic disease. Inheritance occurs as an autosomal dominant trait. The prevalence of symptomatic antithrombin deficiency ranges from 1 per 2000 to 1 per 5000 in the general population. Deficiencies results from mutations that affect synthesis or stability of antithrombin or from mutations that affect the protease and/or heparin binding sites of antithrombin.
Clinical manifestations of antithrombin deficiency include deep vein thrombosis and pulmonary embolism. Arterial thrombosis is rare in anththrombin deficiency. Thrombosis may occur spontaneously or in association with surgery, trauma or pregnancy. Treatment of acute episodes of thrombosis is by infusion of heparin (for 5-7 days) followed by oral anticoagulant therapy.
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Pharmacological Intervention in Bleeding
Coumarin drugs (based on the chemical benzopyrone), such as warfarin (trade name Coumadin®) as well as the glycosaminoglycans, heparin and heparan sulfate, are useful as anticoagulants. Heparin is useful as an anticoagulant because it binds to, and activates, antithrombin III which then inhibits the serine proteases of the coagulation cascade. Heparin is abundant in granules of the mast cells that line the vasculature. In response to injury, the heparin is released and inhibits coagulation. The coumarin drugs inhibit coagulation by inhibiting the vitamin K-dependent γ-carboxylation reactions necessary to the function of thrombin, and factors VII, IX, and X as well as proteins C and S. These drugs act by inhibiting the reduction of the quinone derivatives of vitamin K to their active hydroquinone forms. Because of the mode of action of coumarin drugs, it takes several days for their maximum effect to be realized. For this reason, heparin is normally administered first followed by warfarin or warfarin-related drugs.
The plasminogen activators also are useful for controlling coagulation. Because tPA is highly selective for the degradation of fibrin in clots, it is extremely useful in restoring the patency of the coronary arteries following thrombosis, in particular during the short period following myocardial infarct. Streptokinase (an enzyme from the Streptococci bacterium) is another plasminogen activator useful from a therapeutic standpoint. However, it is less selective than tPA, being able to activate circulating plasminogen as well as that bound to a fibrin clot.
Aspirin is an important inhibitor of platelet activation. By virtue of inhibiting the activity of cyclooxygenase, aspirin reduces the production of TXA2 by platelets. Aspirin also reduces endothelial cell production of prostacyclin (PGI2), an inhibitor of platelet aggregation and a vasodilator. Localized to the site of coagulation is a balance between the levels of platelet derived TXA2 and endothelial cell derived PGI2. This allows for platelet aggregation and clot formation but preventing excessive accumulation of the clot, thus maintaining blood flow around the site of the clot. Endothelial cells regenerate active cyclooxygenase faster than platelets because mature platelets cannot synthesize the enzyme, requiring new platelets to enter the circulation (platelet half-life is approximately 4 days). Therefore, PGI2 synthesis is greater than that of TXA2. The net effect of aspirin is more in favor of endothelial cell-mediated inhibition of the coagulation cascade. This reflects one of the the cardiovascular benefits to low dose administration of aspirin. Aspirin also has important effects on inflammatory processes that impact cardiovascular systems (see the Aspirin page for more details).
Newer classes of anticoagulation drugs are being developed that function by inhibiting the activation of platelets and their subsequent aggregation. The drug clopidogrel: Plavix® (Bristol-Myers Squibb) is an irreversible inhibitor of the ADP receptor on platelet membranes. When ADP binds to platelets they are activated and aggregate leading to amplification of the coagulation response, thus Plavix interferes with this process. Plavix is prescribed for the treatment of peripheral vascular and cerebrovascular disease as well as coronary artery disease to prevent the formation of thrombotic plaques.
Another target of pharmacologic intervention in coagulation involving platelets is the role of GPIIb-GPIIIa in fibrinogen-induced platelet aggregation. Glanzmann thrombasthenia is an inherited platelet function defect characterized by a lack of platelet aggregation in response to all physiological agonists. This disorder is due to a lack of the GPIIb-GPIIIa receptor complex on platelets. Patients with this disorder present with significantly increased bleeding times. Although a rare disorder, study of the pathophysiology of the disease led to the development of anticoagulant drugs that inhibit platelet aggregation regardless of the agonist. Therefore, the GPIIb-GPIIIa antagonists more completely inhibit platelet aggregation than do aspirin or Plavix. The current family of these drugs includes ReoPro® (abciximab: a human monoclonal antibody), Integrilin® (eptifibatide: a cyclic hexapeptide derived from a protein found in the venom of the southeastern pygmy rattlesnake) and Aggrastat® (tirofiban: a synthetic organic non-peptide molecule).
Growth factors & cytokines
Introduction
Growth Factor Table
Interleukin and Cytokine Table
Adipocytokine Table
Discussion of Individual Factors
Epidermal Growth Factor (EGF)
Platelet-Derived Growth Factor (PDGF)
Fibroblast Growth Factors (FGFs)
Transforming Growth Factors-β TGFs-β)
Transforming Growth Factor-α (TGF-α)
Erythropoietin (Epo)
Insulin-Like Growth Factor-I (IGF-I)
Insulin-Like Growth Factor-II (IGF-II)
Interleukin-1 (IL-1)
Interleukin-2 (IL-2)
Interleukin-6 (IL-6)
Interleukin-8 (IL-8)
Tumor Necrosis Factor-α (TNF-α)
Tumor Necrosis Factor-β (TNF-β)
Interferon-γ (INF-γ)
Colony Stimulating Factors (CSFs)
For a more comprehensive listing see: The C.O.P.E. Site
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Introduction
Growth factors are proteins that bind to receptors on the cell surface, with the primary result of activating cellular proliferation and/or differentiation. Many growth factors are quite versatile, stimulating cellular division in numerous different cell types; while others are specific to a particular cell-type.
Cytokines are a class of signaling proteins that are used extensively in cellular communication, immune function and embryogenesis. Cytokines are produced by a variety of hematopoietic and non-hematopoietic cell types and can exert autocrine, paracrine and endocrine effects as do the hormones. They are, therefore, more correctly related to hormones than to growth factors in their overall functions. However, many cytokines also exhibit growth factor activity so they are discussed here as well as in the Peptide Hormones page.
The lists in the following Tables as well as the descriptions of several factors are not intended to be comprehensive nor complete but a look at some of the more commonly known factors and their principal activities.
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Growth Factors
Factor
Principal Source
Primary Activity
Comments
PDGF platelets, endothelial cells, placenta promotes proliferation of connective tissue, glial and smooth muscle cells two different protein chains form 3 distinct dimer forms; AA, AB and BB
EGF submaxillary gland, Brunners gland promotes proliferation of mesenchymal, glial and epithelial cells
TGF-α common in transformed cells may be important for normal wound healing related to EGF
FGF wide range of cells; protein is associated with the ECM promotes proliferation of many cells; inhibits some stem cells; induces mesoderm to form in early embryos at least 19 family members, 4 distinct receptors
NGF promotes neurite outgrowth and neural cell survival several related proteins first identified as proto-oncogenes; trkA ("trackA"), trkB, trkC
Erythropoietin kidney promotes proliferation and differentiation of erythrocytes
TGF-β activated Th1 cells (T-helper) and natural killer (NK) cells anti-inflammatory (suppresses cytokine production and class II MHC expression), promotes wound healing, inhibits macrophage and lymphocyte proliferation at least 100 different family members
IGF-I primarily liver promotes proliferation of many cell types related to IGF-II and proinsulin, also called Somatomedin C
IGF-II variety of cells promotes proliferation of many cell types primarily of fetal origin related to IGF-I and proinsulin
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Interleukins and Cytokines
Cytokines are a unique family of growth factors. Secreted primarily from leukocytes, cytokines stimulate both the humoral and cellular immune responses, as well as the activation of phagocytic cells. Cytokines that are secreted from lymphocytes are termed lymphokines, whereas those secreted by monocytes or macrophages are termed monokines. A large family of cytokines are produced by various cells of the body. Many of the lymphokines are also known as interleukins (ILs), since they are not only secreted by leukocytes but also able to affect the cellular responses of leukocytes. Specifically, interleukins are growth factors targeted to cells of hematopoietic origin. The list of identified interleukins grows continuously with the total number of individual activities now at 22 (18 are listed in the Table below).
Interleukins
Principal Source
Primary Activity
IL1-α and -β macrophages and other antigen presenting cells (APCs) costimulation of APCs and T cells, inflammation and fever, acute phase response, hematopoiesis
IL-2 activated Th1 cells, NK cells proliferation of B cells and activated T cells, NK functions
IL-3 activated T cells growth of hematopoietic progenitor cells
IL-4 Th2 and mast cells B cell proliferation, eosinophil and mast cell growth and function, IgE and class II MHC expression on B cells, inhibition of monokine production
IL-5 Th2 and mast cells eosinophil growth and function
IL-6 activated Th2 cells, APCs, other somatic cells such as hepatocytes and adipocytes acute phase response, B cell proliferation, thrombopoiesis, synergistic with IL-1 and TNF on T cells
IL-7 thymic and marrow stromal cells T and B lymphopoiesis
IL-8 macrophages, other somatic cells chemoattractant for neutrophils and T cells
IL-9 T cells hematopoietic and thymopoietic effects
IL-10 activated Th2 cells, CD8+ T and B cells, macrophages inhibits cytokine production, promotes B cell proliferation and antibody production, suppresses cellular immunity, mast cell growth
IL-11 stromal cells synergisitc hematopoietic and thrombopoietic effects
IL-12 B cells, macrophages proliferation of NK cells, INF-γ production, promotes cell-mediated immune functions
IL-13 Th2 cells, B cells, macrophages stimulates growth and proliferation of B cells, inhibits production of macrophage inflammatory cytokines
IL-14 T cells and malignant B cells regulates the growth and proliferation of B cells
IL-15 virus infected macrophages, mononuclear phagocytes induces production of NK cells
IL-16 eosinophils, CD8+ T cells, lymphocytes, epithelial cells chemoattractant for CD4+ cells
IL-17 subsets of T cells increaases production of inflammatory cytokines, angiogenesis
IL-18 macrophages increases NK cell activity, induces production of INF-γ
Interferons
Principal Source
Primary Activity
INF-α and -β macrophages, neutrophils and some somatic cells antiviral effects, induction of class I MHC on all somatic cells, activation of NK cells and macrophages
INF-γ activated Th1 and NK cells induces of class I MHC on all somatic cells, induces class II MHC on APCs and somatic cells, activates macrophages, neutrophils, NK cells, promotes cell-mediated immunity, antiviral effects
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Adipocytokines
Adipose tissue is not merely an organ designed to passively store excess carbon in the form of fatty acids esterified to glycerol (triacylglycerols). Mature adipocytes synthesize and secrete numerous enzymes, growth factors, cytokines and hormones that are involved in overall energy homeostasis. Many of the factors that influence adipogenesis are also involved in diverse processes in the body including lipid homeostasis and modulation of inflammatory responses. In addition, a number of proteins secreted by adipocytes play important roles in these same processes. In fact recent evidence has demonstrated that many factors secreted from adipocytes are proinflammatory mediators and these proteins have been termed adipocytokines or adipokines. Members of this class of protein secreted from adipocytes include TNF-α, IL-6 and leptin. Listed in the Table below is only a subset of proteins known to be secreted by adipose tissue and the focus is on those that effect overall metabolic homeostasis and modulate inflammatory processes. As is clear from the Table, not all the proteins are unique to adipose tissue.
Factor Principal Source Major Action
Leptin predominantly adipocytes, mammary gland, intestine, muscle, placenta see Peptide Hormones page
Adiponectin; also called adipocyte complement factor 1q-related protein (ACRP30), and adipoQ adipocytes see Peptide Hormones page
IL-6 adipocytes, hepatocytes, activated Th2 cells, and antigen-presenting cells (APCs) acute phase response, B cell proliferation, thrombopoiesis, synergistic with IL-1 and TNF on T cells
TNFα primarily activated macrophages, adipocytes induces expression of other autocrine growth factors, increases cellular responsiveness to growth factors and induces signaling pathways that lead to proliferation
Resistin adipocytes, spleen, monocytes, macrophages, lung, kidney, bone marrow, placenta see Peptide Hormones page
Visfatin; also called pre-B cell-enhancing factor (PBEF) visceral white adipocyte tissue binds to and activates the insulin receptor thus acting as an insulin mimetic; inhibits neutrophil apoptosis
Adipsin (also called complement factor D) adipocytes, liver, monocytes, macrophages rate limiting enzyme in complement activation
monocyte chemotactic protein-1 (MCP-1) leukocytes, adipocytes is a chemokine defined as CCL2 (C-C motif, ligand 2); recruits monocytes, T cells, and dendritic cells to sites of infection and tissue injury
plasminogen-activator inhibitor-1 (PAI-1) adipocytes, monocytes, placenta, platelets, endometrium see the Blood Coagulation page for more details
C-reactive protein (CRP) hepatocytes, adipocytes is a member of the pentraxin family of calcium-dependent ligand binding proteins; assists complement interaction with foreign and damaged cells; enhances phagocytosis by macrophages; levels of expression regulated by circulating IL-6; modulates endothelial cell functions by inducing expression of various cell adhesion molecules, e.g. ICAM-1, VCAM-1, and selectins; induces MCP-1 expression in endothelium; attenuates NO production by downregulating NOS expression; increase expression and activity of PAI-1
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Epidermal Growth Factor (EGF)
EGF, like all growth factors, binds to specific high-affinity, low-capacity receptors on the surface of responsive cells. Intrinsic to the EGF receptor is tyrosine kinase activity, which is activated in response to EGF binding. The kinase domain of the EGF receptor phosphorylates the EGF receptor itself (autophosphorylation) as well as other proteins, in signal transduction cascades, that associate with the receptor following activation. Experimental evidence has shown that the NEU proto-oncogene is a homologue of the EGF receptor.
EGF has proliferative effects on cells of both mesodermal and ectodermal origin, particularly keratinocytes and fibroblasts. EGF exhibits negative growth effects on certain carcinomas as well as hair follicle cells. Growth-related responses to EGF include the induction of nuclear proto-oncogene expression, such as FOS, JUN and MYC. EGF also has the effect of decreasing gastric acid secretion.
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Platelet-Derived Growth Factor (PDGF)
PDGF is composed of two distinct polypeptide chains, A and B, that form homodimers (AA or BB) or heterodimers (AB). The SIS proto-oncogene has been shown to be homologous to the PDGF A chain. Only the dimeric forms of PDGF interact with the PDGF receptor. Two distinct classes of PDGF receptor have been cloned, one specific for AA homodimers and another that binds BB and AB type dimers. Like the EGF receptor, the PDGF receptors have intrinsic tyrosine kinase activity. Following autophosphorylation of the PDGF receptor, numerous signal-transducing proteins associate with the receptor and are subsequently tyrosine phosphorylated.
Proliferative responses to PDGF action are exerted on many mesenchymal cell types. Other growth-related responses to PDGF include cytoskeletal rearrangement and increased polyphosphoinositol turnover. Again, like EGF, PDGF induces the expression of a number of nuclear localized proto-oncogenes, such as FOS, MYC and JUN. The primary effects of TGF-β are due to the induction, by TGF-β, of PDGF expression.
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Fibroblast Growth Factors (FGFs)
There are at least 19 distinct members of the FGF family of growth factors. The two originally characterized FGFs were identified by biological assay and are termed FGF1 (acidic-FGF, aFGF) and FGF2 (basic-FGF, bFGF). Kaposi's sarcoma cells (prevalent in patients with AIDS) secrete a homologue of FGF called the K-FGF proto-oncogene. In mice the mammary tumor virus integrates at two predominant sites in the mouse genome identified as Int-1 and Int-2. The protein encoded by the Int-2 locus is a homologue of the FGF family of growth factors.
Studies of human disorders as well as gene knock-out studies in mice show the prominent role for FGFs is in the development of the skeletal system and nervous system in mammals. FGFs also are neurotrophic for cells of both the peripheral and central nervous system. Additionally, several members of the FGF family are potent inducers of mesodermal differentiation in early embryos. Non-proliferative effects include regulation of pituitary and ovarian cell function.
The FGFs interact with specific cell-surface receptors. There have been identified 4 distinct receptor types identified as FGFR1 - FGFR4. Each of these receptors has intrinsic tyrosine kinase activity like both the EGF and PDGF receptors. As with all transmembrane receptors that have tyrosine kinase activity, autophosphorylation of the receptor is the immediate response to FGF binding. Following activation of FGF receptors, numerous signal-transducing proteins associate with the receptor and become tyrosine-phosphorylated. The FLG proto-oncogene is a homologue of the FGF receptor family. The FGFR1 receptor also has been shown to be the portal of entry into cells for herpes viruses. FGFs also bind to cell-surface heparan-sulfated proteoglycans with low affinity relative to that of the specific receptors. The purpose in binding of FGFs to theses proteoglycans is not completely understood but may allow the growth factor to remain associated with the extracellular surface of cells that they are intended to stimulate under various conditions.
The FGF receptors are widley expressed in developing bone and several common autosomal dominant disorders of bone growth have been shown to result from mutations in the FGFR genes. The most prevalent is achondroplasia, ACH. ACH is characterized by disproportionate short stature, where the limbs are shorter than the trunk, and macrocephaly (excessive head size). Almost all persons with ACH exhibit a glycine to arginine substitution in the transmembrane domain of FGFR3. This mutation results in ligand-independent activation of the receptor. FGFR3 is predominantly expressed in quiescent chondrocytes where it is responsible for restricting chondrocyte proliferation and differentiation. In mice with inactivating mutations in FGFR3 there is an expansion of long bone growth and zones of proliferating cartilage further demonstrating that FGFR3 is necessary to control the rate and amount of chondrocyte growth.
Several other disorders of bone growth collectively identified as craniosynostosis syndromes have been shown to result from mutations in FGFR1, FGFR2 and FGFR3. Sometimes the same mutation can cause two or more different craniosynostosis syndromes. A cysteine to tyrosine substitution in FGFR2 can cause either Pfeiffer or Crouzon syndrome. This phenomenon indicates that additional factors are likely responsible for the different phenotypes.
Affected Receptor
Syndrome
Phenotypes
FGFR1 Pfeiffer broad first digits, hypertelorism
FGFR2 Apert mid-face hypoplasia, fusion of digits
FGFR2 Beare-Stevenson mid-face hypoplasia, corrugated skin
FGFR2 Crouzon mid-face hypoplasia, ocular proptosis
FGFR2 Jackson-Weiss mid-face hypoplasia, foot anamolies
FGFR2 Pfeiffer same as for FGFR1 mutations
FGFR3 Crouzon mid-face hypoplasia, acanthosis nigricans, ocular proptosis
FGFR3 Non-syndromatic craniosynostosis digit defects, hearing loss
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Transforming Growth Factors-β (TGFs-β)
A more detailed description of the TGF-β family of growth factors and associated signaling pathways can be found on the Signaling by Wnts and TGFs-β/BMP page.
TGF-β was originally characterized as a protein (secreted from a tumor cell line) that was capable of inducing a transformed phenotype in non-neoplastic cells in culture. This effect was reversible, as demonstrated by the reversion of the cells to a normal phenotype following removal of the TGF-β. Subsequently, many proteins homologous to TGF-β have been identified. The four closest relatives are TGF-β-1 (the original TGF-β) through TGF-β-5 (TGF-β-1 = TGF-β-4). All four of these proteins share extensive regions of similarity in their amino acids. Many other proteins, possessing distinct biological functions, have stretches of amino-acid homology to the TGF-β family of proteins, particularly the C-terminal region of these proteins.
The TGF-β-related family of proteins includes the activin and inhibin proteins. There are activin A, B and AB proteins, as well as an inhibin A and inhibin B protein. The Mullerian inhibiting substance (MIS) is also a TGF-β-related protein, as are members of the bone morphogenetic protein (BMP) family of bone growth-regulatory factors. Indeed, the TGF-β family may comprise as many as 100 distinct proteins, all with at least one region of amino-acid sequence homology.
There are several classes of cell-surface receptors that bind different TGFs-β with differing affinities. There also are cell-type specific differences in receptor sub-types. Unlike the EGF, PDGF and FGF receptors, the TGF-β family of receptors all have intrinsic serine/threonine kinase activity and, therefore, induce distinct cascades of signal transduction.
TGFs-β have proliferative effects on many mesenchymal and epithelial cell types. Under certain conditions TGFs-β will demonstrate anti-proliferative effects on endothelial cells, macrophages, and T- and B-lymphocytes. Such effects include decreasing the secretion of immunoglobulin and suppressing hematopoiesis, myogenesis, adipogenesis and adrenal steroidogenesis. Several members of the TGF-β family are potent inducers of mesodermal differentiation in early embryos, in particular TGF-β and activin A.
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Transforming Growth Factor-α (TGF-α)
TGF-α, like the β form, was first identified as a substance secreted from certain tumor cells that, in conjunction with TGF-β-1, could reversibly transform certain types of normal cells in culture. TGF-α binds to the EGF receptor, as well as its own distinct receptor, and it is this interaction that is thought to be responsible for the growth factor's effect. The predominant sources of TGF-α are carcinomas, but activated macrophages and keratinocytes (and possibly other epithelial cells) also secrete TGF-α. In normal cell populations, TGF-α is a potent keratinocyte growth factor; forming an autocrine growth loop by virtue of the protein activating the very cells that produce it.
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Erythropoietin (EPO)
EPO is synthesized by the kidney and is the primary regulator of erythropoiesis. EPO stimulates the proliferation and differentiation of immature erythrocytes; it also stimulates the growth of erythoid progenitor cells (e.g. erythrocyte burst-forming and colony-forming units) and induces the differentiation of erythrocyte colony-forming units into proerythroblasts. When patients suffering from anemia due to kidney failure are given EPO, the result is a rapid and significant increase in red blood cell count.
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Insulin-Like Growth Factor-I (IGF-I)
IGF-I (originally called somatomedin C) is a growth factor structurally related to insulin. IGF-I is the primary protein involved in responses of cells to growth hormone (GH): that is, IGF-I is produced in response to GH and then induces subsequent cellular activities, particularly on bone growth. It is the activity of IGF-I in response to GH that gave rise to the term somatomedin. Subsequent studies have demonstrated, however, that IGF-I has autocrine and paracrine activities in addition to the initially observed endocrine activities on bone. The IGF-I receptor, like the insulin receptor, has intrinsic tyrosine kinase activity. Owing to their structural similarities IGF-I can bind to the insulin receptor but does so at a much lower affinity than does insulin itself.
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Insulin-Like Growth Factor-II (IGF-II)
IGF-II is almost exclusively expressed in embryonic and neonatal tissues. Following birth, the level of detectable IGF-II protein falls significantly. For this reason IGF-II is thought to be a fetal growth factor. The IGF-II receptor is identical to the mannose-6-phosphate receptor that is responsible for the integration of lysosomal enzymes (which contain mannose-6-phosphate residues) to the lysosomes.
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Interleukin-1 (IL-1)
IL-1 is one of the most important immune response-- modifying interleukins. The predominant function of IL-1 is to enhance the activation of T-cells in response to antigen. The activation of T-cells, by IL-1, leads to increased T-cell production of IL-2 and of the IL-2 receptor, which in turn augments the activation of the T-cells in an autocrine loop. IL-1 also induces expression of interferon-γ (IFN-γ) by T-cells. This effect of T-cell activation by IL-1 is mimicked by TNF-α which is another cytokine secreted by activated macrophages. There are 2 distinct IL-1 proteins, termed IL-1α and -1β, that are 26% homologous at the amino acid level. The IL-1s are secreted primarily by macrophages but also from neutrophils, endothelial cells, smooth muscle cells, glial cells, astrocytes, B- and T-cells, fibroblasts and keratinocytes. Production of IL-1 by these different cell types occurs only in response to cellular stimulation. In addition to its effects on T-cells, IL-1 can induce proliferation in non-lymphoid cells.
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Interleukin-2 (IL-2)
IL-2, produced and secreted by activated T-cells, is the major interleukin responsible for clonal T-cell proliferation. IL-2 also exerts effects on B-cells, macrophages, and natural killer (NK) cells. The production of IL-2 occurs primarily by CD4+ T-helper cells. As indicated above, the expression of both IL-2 and the IL-2 receptor by T-cells is induced by IL-1. Indeed, the IL-2 receptor is not expressed on the surface of resting T-cells and is present only transiently on the surface of T-cells, disappearing within 6-10 days of antigen presentation. In contrast to T-helper cells, NK cells constitutively express IL-2 receptors and will secrete TNF-α, IFN-γ and GM-CSF in response to IL-2, which in turn activate macrophages.
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Interleukin-6 (IL-6)
IL-6 is produced by macrophages, fibroblasts, endothelial cells and activated T-helper cells. IL-6 acts in synergy with IL-1 and TNF-( in many immune responses, including T-cell activation. In particular, IL-6 is the primary inducer of the acute-phase response in liver. IL-6 also enhances the differentiation of B-cells and their consequent production of immunoglobulin. Glucocorticoid synthesis is also enhanced by IL-6. Unlike IL-1, IL-2 and TNF-α, IL-6 does not induce cytokine expression; its main effects, therefore, are to augment the responses of immune cells to other cytokines.
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Interleukin-8 (IL-8)
IL-8 is an interleukin that belongs to an ever-expanding family of proteins that exert chemoattractant activity to leukocytes and fibroblasts. This family of proteins is termed the chemokines. IL-8 is produced by monocytes, neutrophils, and NK cells and is chemoattractant for neutrophils, basophils and T-cells. In addition, IL-8 activates neutrophils to degranulate.
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Tumor Necrosis Factor-α (TNF-α)
TNF-α (also called cachectin), like IL-1 is a major immune response-modifying cytokine produced primarily by activated macrophages. Like IL-1, TNF-α induces the expression of other autocrine growth factors, increases cellular responsiveness to growth factors and induces signaling pathways that lead to proliferation. TNF-α acts synergistically with EGF and PDGF on some cell types. Like other growth factors, TNF-α induces expression of a number of nuclear proto-oncogenes as well as of several interleukins.
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Tumor Necrosis Factor-β (TNF-β)
TNF-β (also called lymphotoxin) is characterized by its ability to kill a number of different cell types, as well as the ability to induce terminal differentiation in others. One significant non-proliferative response to TNF-β is an inhibition of lipoprotein lipase present on the surface of vascular endothelial cells. The predominant site of TNF-β synthesis is T-lymphocytes, in particular the special class of T-cells called cytotoxic T-lymphocytes (CTL cells). The induction of TNF-β expression results from elevations in IL-2 as well as the interaction of antigen with T-cell receptors.
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Interferon-γ (INF-γ)
IFN-α, IFN-β and IFN-ω are known as type I interferons: they are predominantly responsible for the antiviral activities of the interferons. In contrast, IFN-γ is a type II or immune interferon. Although IFN-γ has antiviral activity, it is significantly less active at this function than the type I IFNs. Unlike the type I IFNs, IFN-γ is not induced by infection nor by double-stranded RNAs. IFN-γ is secreted primarily by CD8+ T-cells. Nearly all cells express receptors for IFN-γ and respond to IFN-γ binding by increasing the surface expression of class I MHC proteins, thereby promoting the presentation of antigen to T-helper (CD4+) cells. IFN-γ also increases the presentation of class II MHC proteins on class II cells further enhancing the ability of cells to present antigen to T-cells.
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Colony Stimulating Factors (CSFs)
CSFs are cytokines that stimulate the proliferation of specific pluripotent stem cells of the bone marrow in adults. Granulocyte-CSF (G-CSF) is specific for proliferative effects on cells of the granulocyte lineage. Macrophage-CSF (M-CSF) is specific for cells of the macrophage lineage. Granulocyte-macrophage-CSF (GM-CSF) has proliferative effects on both classes of lymphoid cells. Epo is also considered a CSF as well as a growth factor, since it stimulates the proliferation of erythrocyte colony-forming units. IL-3 (secreted primarily from T-cells) is also known as multi-CSF, since it stimulates stem cells to produce all forms of hematopoietic cells.
Growth Factor Table
Interleukin and Cytokine Table
Adipocytokine Table
Discussion of Individual Factors
Epidermal Growth Factor (EGF)
Platelet-Derived Growth Factor (PDGF)
Fibroblast Growth Factors (FGFs)
Transforming Growth Factors-β TGFs-β)
Transforming Growth Factor-α (TGF-α)
Erythropoietin (Epo)
Insulin-Like Growth Factor-I (IGF-I)
Insulin-Like Growth Factor-II (IGF-II)
Interleukin-1 (IL-1)
Interleukin-2 (IL-2)
Interleukin-6 (IL-6)
Interleukin-8 (IL-8)
Tumor Necrosis Factor-α (TNF-α)
Tumor Necrosis Factor-β (TNF-β)
Interferon-γ (INF-γ)
Colony Stimulating Factors (CSFs)
For a more comprehensive listing see: The C.O.P.E. Site
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Introduction
Growth factors are proteins that bind to receptors on the cell surface, with the primary result of activating cellular proliferation and/or differentiation. Many growth factors are quite versatile, stimulating cellular division in numerous different cell types; while others are specific to a particular cell-type.
Cytokines are a class of signaling proteins that are used extensively in cellular communication, immune function and embryogenesis. Cytokines are produced by a variety of hematopoietic and non-hematopoietic cell types and can exert autocrine, paracrine and endocrine effects as do the hormones. They are, therefore, more correctly related to hormones than to growth factors in their overall functions. However, many cytokines also exhibit growth factor activity so they are discussed here as well as in the Peptide Hormones page.
The lists in the following Tables as well as the descriptions of several factors are not intended to be comprehensive nor complete but a look at some of the more commonly known factors and their principal activities.
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Growth Factors
Factor
Principal Source
Primary Activity
Comments
PDGF platelets, endothelial cells, placenta promotes proliferation of connective tissue, glial and smooth muscle cells two different protein chains form 3 distinct dimer forms; AA, AB and BB
EGF submaxillary gland, Brunners gland promotes proliferation of mesenchymal, glial and epithelial cells
TGF-α common in transformed cells may be important for normal wound healing related to EGF
FGF wide range of cells; protein is associated with the ECM promotes proliferation of many cells; inhibits some stem cells; induces mesoderm to form in early embryos at least 19 family members, 4 distinct receptors
NGF promotes neurite outgrowth and neural cell survival several related proteins first identified as proto-oncogenes; trkA ("trackA"), trkB, trkC
Erythropoietin kidney promotes proliferation and differentiation of erythrocytes
TGF-β activated Th1 cells (T-helper) and natural killer (NK) cells anti-inflammatory (suppresses cytokine production and class II MHC expression), promotes wound healing, inhibits macrophage and lymphocyte proliferation at least 100 different family members
IGF-I primarily liver promotes proliferation of many cell types related to IGF-II and proinsulin, also called Somatomedin C
IGF-II variety of cells promotes proliferation of many cell types primarily of fetal origin related to IGF-I and proinsulin
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Interleukins and Cytokines
Cytokines are a unique family of growth factors. Secreted primarily from leukocytes, cytokines stimulate both the humoral and cellular immune responses, as well as the activation of phagocytic cells. Cytokines that are secreted from lymphocytes are termed lymphokines, whereas those secreted by monocytes or macrophages are termed monokines. A large family of cytokines are produced by various cells of the body. Many of the lymphokines are also known as interleukins (ILs), since they are not only secreted by leukocytes but also able to affect the cellular responses of leukocytes. Specifically, interleukins are growth factors targeted to cells of hematopoietic origin. The list of identified interleukins grows continuously with the total number of individual activities now at 22 (18 are listed in the Table below).
Interleukins
Principal Source
Primary Activity
IL1-α and -β macrophages and other antigen presenting cells (APCs) costimulation of APCs and T cells, inflammation and fever, acute phase response, hematopoiesis
IL-2 activated Th1 cells, NK cells proliferation of B cells and activated T cells, NK functions
IL-3 activated T cells growth of hematopoietic progenitor cells
IL-4 Th2 and mast cells B cell proliferation, eosinophil and mast cell growth and function, IgE and class II MHC expression on B cells, inhibition of monokine production
IL-5 Th2 and mast cells eosinophil growth and function
IL-6 activated Th2 cells, APCs, other somatic cells such as hepatocytes and adipocytes acute phase response, B cell proliferation, thrombopoiesis, synergistic with IL-1 and TNF on T cells
IL-7 thymic and marrow stromal cells T and B lymphopoiesis
IL-8 macrophages, other somatic cells chemoattractant for neutrophils and T cells
IL-9 T cells hematopoietic and thymopoietic effects
IL-10 activated Th2 cells, CD8+ T and B cells, macrophages inhibits cytokine production, promotes B cell proliferation and antibody production, suppresses cellular immunity, mast cell growth
IL-11 stromal cells synergisitc hematopoietic and thrombopoietic effects
IL-12 B cells, macrophages proliferation of NK cells, INF-γ production, promotes cell-mediated immune functions
IL-13 Th2 cells, B cells, macrophages stimulates growth and proliferation of B cells, inhibits production of macrophage inflammatory cytokines
IL-14 T cells and malignant B cells regulates the growth and proliferation of B cells
IL-15 virus infected macrophages, mononuclear phagocytes induces production of NK cells
IL-16 eosinophils, CD8+ T cells, lymphocytes, epithelial cells chemoattractant for CD4+ cells
IL-17 subsets of T cells increaases production of inflammatory cytokines, angiogenesis
IL-18 macrophages increases NK cell activity, induces production of INF-γ
Interferons
Principal Source
Primary Activity
INF-α and -β macrophages, neutrophils and some somatic cells antiviral effects, induction of class I MHC on all somatic cells, activation of NK cells and macrophages
INF-γ activated Th1 and NK cells induces of class I MHC on all somatic cells, induces class II MHC on APCs and somatic cells, activates macrophages, neutrophils, NK cells, promotes cell-mediated immunity, antiviral effects
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Adipocytokines
Adipose tissue is not merely an organ designed to passively store excess carbon in the form of fatty acids esterified to glycerol (triacylglycerols). Mature adipocytes synthesize and secrete numerous enzymes, growth factors, cytokines and hormones that are involved in overall energy homeostasis. Many of the factors that influence adipogenesis are also involved in diverse processes in the body including lipid homeostasis and modulation of inflammatory responses. In addition, a number of proteins secreted by adipocytes play important roles in these same processes. In fact recent evidence has demonstrated that many factors secreted from adipocytes are proinflammatory mediators and these proteins have been termed adipocytokines or adipokines. Members of this class of protein secreted from adipocytes include TNF-α, IL-6 and leptin. Listed in the Table below is only a subset of proteins known to be secreted by adipose tissue and the focus is on those that effect overall metabolic homeostasis and modulate inflammatory processes. As is clear from the Table, not all the proteins are unique to adipose tissue.
Factor Principal Source Major Action
Leptin predominantly adipocytes, mammary gland, intestine, muscle, placenta see Peptide Hormones page
Adiponectin; also called adipocyte complement factor 1q-related protein (ACRP30), and adipoQ adipocytes see Peptide Hormones page
IL-6 adipocytes, hepatocytes, activated Th2 cells, and antigen-presenting cells (APCs) acute phase response, B cell proliferation, thrombopoiesis, synergistic with IL-1 and TNF on T cells
TNFα primarily activated macrophages, adipocytes induces expression of other autocrine growth factors, increases cellular responsiveness to growth factors and induces signaling pathways that lead to proliferation
Resistin adipocytes, spleen, monocytes, macrophages, lung, kidney, bone marrow, placenta see Peptide Hormones page
Visfatin; also called pre-B cell-enhancing factor (PBEF) visceral white adipocyte tissue binds to and activates the insulin receptor thus acting as an insulin mimetic; inhibits neutrophil apoptosis
Adipsin (also called complement factor D) adipocytes, liver, monocytes, macrophages rate limiting enzyme in complement activation
monocyte chemotactic protein-1 (MCP-1) leukocytes, adipocytes is a chemokine defined as CCL2 (C-C motif, ligand 2); recruits monocytes, T cells, and dendritic cells to sites of infection and tissue injury
plasminogen-activator inhibitor-1 (PAI-1) adipocytes, monocytes, placenta, platelets, endometrium see the Blood Coagulation page for more details
C-reactive protein (CRP) hepatocytes, adipocytes is a member of the pentraxin family of calcium-dependent ligand binding proteins; assists complement interaction with foreign and damaged cells; enhances phagocytosis by macrophages; levels of expression regulated by circulating IL-6; modulates endothelial cell functions by inducing expression of various cell adhesion molecules, e.g. ICAM-1, VCAM-1, and selectins; induces MCP-1 expression in endothelium; attenuates NO production by downregulating NOS expression; increase expression and activity of PAI-1
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Epidermal Growth Factor (EGF)
EGF, like all growth factors, binds to specific high-affinity, low-capacity receptors on the surface of responsive cells. Intrinsic to the EGF receptor is tyrosine kinase activity, which is activated in response to EGF binding. The kinase domain of the EGF receptor phosphorylates the EGF receptor itself (autophosphorylation) as well as other proteins, in signal transduction cascades, that associate with the receptor following activation. Experimental evidence has shown that the NEU proto-oncogene is a homologue of the EGF receptor.
EGF has proliferative effects on cells of both mesodermal and ectodermal origin, particularly keratinocytes and fibroblasts. EGF exhibits negative growth effects on certain carcinomas as well as hair follicle cells. Growth-related responses to EGF include the induction of nuclear proto-oncogene expression, such as FOS, JUN and MYC. EGF also has the effect of decreasing gastric acid secretion.
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Platelet-Derived Growth Factor (PDGF)
PDGF is composed of two distinct polypeptide chains, A and B, that form homodimers (AA or BB) or heterodimers (AB). The SIS proto-oncogene has been shown to be homologous to the PDGF A chain. Only the dimeric forms of PDGF interact with the PDGF receptor. Two distinct classes of PDGF receptor have been cloned, one specific for AA homodimers and another that binds BB and AB type dimers. Like the EGF receptor, the PDGF receptors have intrinsic tyrosine kinase activity. Following autophosphorylation of the PDGF receptor, numerous signal-transducing proteins associate with the receptor and are subsequently tyrosine phosphorylated.
Proliferative responses to PDGF action are exerted on many mesenchymal cell types. Other growth-related responses to PDGF include cytoskeletal rearrangement and increased polyphosphoinositol turnover. Again, like EGF, PDGF induces the expression of a number of nuclear localized proto-oncogenes, such as FOS, MYC and JUN. The primary effects of TGF-β are due to the induction, by TGF-β, of PDGF expression.
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Fibroblast Growth Factors (FGFs)
There are at least 19 distinct members of the FGF family of growth factors. The two originally characterized FGFs were identified by biological assay and are termed FGF1 (acidic-FGF, aFGF) and FGF2 (basic-FGF, bFGF). Kaposi's sarcoma cells (prevalent in patients with AIDS) secrete a homologue of FGF called the K-FGF proto-oncogene. In mice the mammary tumor virus integrates at two predominant sites in the mouse genome identified as Int-1 and Int-2. The protein encoded by the Int-2 locus is a homologue of the FGF family of growth factors.
Studies of human disorders as well as gene knock-out studies in mice show the prominent role for FGFs is in the development of the skeletal system and nervous system in mammals. FGFs also are neurotrophic for cells of both the peripheral and central nervous system. Additionally, several members of the FGF family are potent inducers of mesodermal differentiation in early embryos. Non-proliferative effects include regulation of pituitary and ovarian cell function.
The FGFs interact with specific cell-surface receptors. There have been identified 4 distinct receptor types identified as FGFR1 - FGFR4. Each of these receptors has intrinsic tyrosine kinase activity like both the EGF and PDGF receptors. As with all transmembrane receptors that have tyrosine kinase activity, autophosphorylation of the receptor is the immediate response to FGF binding. Following activation of FGF receptors, numerous signal-transducing proteins associate with the receptor and become tyrosine-phosphorylated. The FLG proto-oncogene is a homologue of the FGF receptor family. The FGFR1 receptor also has been shown to be the portal of entry into cells for herpes viruses. FGFs also bind to cell-surface heparan-sulfated proteoglycans with low affinity relative to that of the specific receptors. The purpose in binding of FGFs to theses proteoglycans is not completely understood but may allow the growth factor to remain associated with the extracellular surface of cells that they are intended to stimulate under various conditions.
The FGF receptors are widley expressed in developing bone and several common autosomal dominant disorders of bone growth have been shown to result from mutations in the FGFR genes. The most prevalent is achondroplasia, ACH. ACH is characterized by disproportionate short stature, where the limbs are shorter than the trunk, and macrocephaly (excessive head size). Almost all persons with ACH exhibit a glycine to arginine substitution in the transmembrane domain of FGFR3. This mutation results in ligand-independent activation of the receptor. FGFR3 is predominantly expressed in quiescent chondrocytes where it is responsible for restricting chondrocyte proliferation and differentiation. In mice with inactivating mutations in FGFR3 there is an expansion of long bone growth and zones of proliferating cartilage further demonstrating that FGFR3 is necessary to control the rate and amount of chondrocyte growth.
Several other disorders of bone growth collectively identified as craniosynostosis syndromes have been shown to result from mutations in FGFR1, FGFR2 and FGFR3. Sometimes the same mutation can cause two or more different craniosynostosis syndromes. A cysteine to tyrosine substitution in FGFR2 can cause either Pfeiffer or Crouzon syndrome. This phenomenon indicates that additional factors are likely responsible for the different phenotypes.
Affected Receptor
Syndrome
Phenotypes
FGFR1 Pfeiffer broad first digits, hypertelorism
FGFR2 Apert mid-face hypoplasia, fusion of digits
FGFR2 Beare-Stevenson mid-face hypoplasia, corrugated skin
FGFR2 Crouzon mid-face hypoplasia, ocular proptosis
FGFR2 Jackson-Weiss mid-face hypoplasia, foot anamolies
FGFR2 Pfeiffer same as for FGFR1 mutations
FGFR3 Crouzon mid-face hypoplasia, acanthosis nigricans, ocular proptosis
FGFR3 Non-syndromatic craniosynostosis digit defects, hearing loss
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Transforming Growth Factors-β (TGFs-β)
A more detailed description of the TGF-β family of growth factors and associated signaling pathways can be found on the Signaling by Wnts and TGFs-β/BMP page.
TGF-β was originally characterized as a protein (secreted from a tumor cell line) that was capable of inducing a transformed phenotype in non-neoplastic cells in culture. This effect was reversible, as demonstrated by the reversion of the cells to a normal phenotype following removal of the TGF-β. Subsequently, many proteins homologous to TGF-β have been identified. The four closest relatives are TGF-β-1 (the original TGF-β) through TGF-β-5 (TGF-β-1 = TGF-β-4). All four of these proteins share extensive regions of similarity in their amino acids. Many other proteins, possessing distinct biological functions, have stretches of amino-acid homology to the TGF-β family of proteins, particularly the C-terminal region of these proteins.
The TGF-β-related family of proteins includes the activin and inhibin proteins. There are activin A, B and AB proteins, as well as an inhibin A and inhibin B protein. The Mullerian inhibiting substance (MIS) is also a TGF-β-related protein, as are members of the bone morphogenetic protein (BMP) family of bone growth-regulatory factors. Indeed, the TGF-β family may comprise as many as 100 distinct proteins, all with at least one region of amino-acid sequence homology.
There are several classes of cell-surface receptors that bind different TGFs-β with differing affinities. There also are cell-type specific differences in receptor sub-types. Unlike the EGF, PDGF and FGF receptors, the TGF-β family of receptors all have intrinsic serine/threonine kinase activity and, therefore, induce distinct cascades of signal transduction.
TGFs-β have proliferative effects on many mesenchymal and epithelial cell types. Under certain conditions TGFs-β will demonstrate anti-proliferative effects on endothelial cells, macrophages, and T- and B-lymphocytes. Such effects include decreasing the secretion of immunoglobulin and suppressing hematopoiesis, myogenesis, adipogenesis and adrenal steroidogenesis. Several members of the TGF-β family are potent inducers of mesodermal differentiation in early embryos, in particular TGF-β and activin A.
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Transforming Growth Factor-α (TGF-α)
TGF-α, like the β form, was first identified as a substance secreted from certain tumor cells that, in conjunction with TGF-β-1, could reversibly transform certain types of normal cells in culture. TGF-α binds to the EGF receptor, as well as its own distinct receptor, and it is this interaction that is thought to be responsible for the growth factor's effect. The predominant sources of TGF-α are carcinomas, but activated macrophages and keratinocytes (and possibly other epithelial cells) also secrete TGF-α. In normal cell populations, TGF-α is a potent keratinocyte growth factor; forming an autocrine growth loop by virtue of the protein activating the very cells that produce it.
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Erythropoietin (EPO)
EPO is synthesized by the kidney and is the primary regulator of erythropoiesis. EPO stimulates the proliferation and differentiation of immature erythrocytes; it also stimulates the growth of erythoid progenitor cells (e.g. erythrocyte burst-forming and colony-forming units) and induces the differentiation of erythrocyte colony-forming units into proerythroblasts. When patients suffering from anemia due to kidney failure are given EPO, the result is a rapid and significant increase in red blood cell count.
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Insulin-Like Growth Factor-I (IGF-I)
IGF-I (originally called somatomedin C) is a growth factor structurally related to insulin. IGF-I is the primary protein involved in responses of cells to growth hormone (GH): that is, IGF-I is produced in response to GH and then induces subsequent cellular activities, particularly on bone growth. It is the activity of IGF-I in response to GH that gave rise to the term somatomedin. Subsequent studies have demonstrated, however, that IGF-I has autocrine and paracrine activities in addition to the initially observed endocrine activities on bone. The IGF-I receptor, like the insulin receptor, has intrinsic tyrosine kinase activity. Owing to their structural similarities IGF-I can bind to the insulin receptor but does so at a much lower affinity than does insulin itself.
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Insulin-Like Growth Factor-II (IGF-II)
IGF-II is almost exclusively expressed in embryonic and neonatal tissues. Following birth, the level of detectable IGF-II protein falls significantly. For this reason IGF-II is thought to be a fetal growth factor. The IGF-II receptor is identical to the mannose-6-phosphate receptor that is responsible for the integration of lysosomal enzymes (which contain mannose-6-phosphate residues) to the lysosomes.
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Interleukin-1 (IL-1)
IL-1 is one of the most important immune response-- modifying interleukins. The predominant function of IL-1 is to enhance the activation of T-cells in response to antigen. The activation of T-cells, by IL-1, leads to increased T-cell production of IL-2 and of the IL-2 receptor, which in turn augments the activation of the T-cells in an autocrine loop. IL-1 also induces expression of interferon-γ (IFN-γ) by T-cells. This effect of T-cell activation by IL-1 is mimicked by TNF-α which is another cytokine secreted by activated macrophages. There are 2 distinct IL-1 proteins, termed IL-1α and -1β, that are 26% homologous at the amino acid level. The IL-1s are secreted primarily by macrophages but also from neutrophils, endothelial cells, smooth muscle cells, glial cells, astrocytes, B- and T-cells, fibroblasts and keratinocytes. Production of IL-1 by these different cell types occurs only in response to cellular stimulation. In addition to its effects on T-cells, IL-1 can induce proliferation in non-lymphoid cells.
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Interleukin-2 (IL-2)
IL-2, produced and secreted by activated T-cells, is the major interleukin responsible for clonal T-cell proliferation. IL-2 also exerts effects on B-cells, macrophages, and natural killer (NK) cells. The production of IL-2 occurs primarily by CD4+ T-helper cells. As indicated above, the expression of both IL-2 and the IL-2 receptor by T-cells is induced by IL-1. Indeed, the IL-2 receptor is not expressed on the surface of resting T-cells and is present only transiently on the surface of T-cells, disappearing within 6-10 days of antigen presentation. In contrast to T-helper cells, NK cells constitutively express IL-2 receptors and will secrete TNF-α, IFN-γ and GM-CSF in response to IL-2, which in turn activate macrophages.
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Interleukin-6 (IL-6)
IL-6 is produced by macrophages, fibroblasts, endothelial cells and activated T-helper cells. IL-6 acts in synergy with IL-1 and TNF-( in many immune responses, including T-cell activation. In particular, IL-6 is the primary inducer of the acute-phase response in liver. IL-6 also enhances the differentiation of B-cells and their consequent production of immunoglobulin. Glucocorticoid synthesis is also enhanced by IL-6. Unlike IL-1, IL-2 and TNF-α, IL-6 does not induce cytokine expression; its main effects, therefore, are to augment the responses of immune cells to other cytokines.
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Interleukin-8 (IL-8)
IL-8 is an interleukin that belongs to an ever-expanding family of proteins that exert chemoattractant activity to leukocytes and fibroblasts. This family of proteins is termed the chemokines. IL-8 is produced by monocytes, neutrophils, and NK cells and is chemoattractant for neutrophils, basophils and T-cells. In addition, IL-8 activates neutrophils to degranulate.
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Tumor Necrosis Factor-α (TNF-α)
TNF-α (also called cachectin), like IL-1 is a major immune response-modifying cytokine produced primarily by activated macrophages. Like IL-1, TNF-α induces the expression of other autocrine growth factors, increases cellular responsiveness to growth factors and induces signaling pathways that lead to proliferation. TNF-α acts synergistically with EGF and PDGF on some cell types. Like other growth factors, TNF-α induces expression of a number of nuclear proto-oncogenes as well as of several interleukins.
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Tumor Necrosis Factor-β (TNF-β)
TNF-β (also called lymphotoxin) is characterized by its ability to kill a number of different cell types, as well as the ability to induce terminal differentiation in others. One significant non-proliferative response to TNF-β is an inhibition of lipoprotein lipase present on the surface of vascular endothelial cells. The predominant site of TNF-β synthesis is T-lymphocytes, in particular the special class of T-cells called cytotoxic T-lymphocytes (CTL cells). The induction of TNF-β expression results from elevations in IL-2 as well as the interaction of antigen with T-cell receptors.
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Interferon-γ (INF-γ)
IFN-α, IFN-β and IFN-ω are known as type I interferons: they are predominantly responsible for the antiviral activities of the interferons. In contrast, IFN-γ is a type II or immune interferon. Although IFN-γ has antiviral activity, it is significantly less active at this function than the type I IFNs. Unlike the type I IFNs, IFN-γ is not induced by infection nor by double-stranded RNAs. IFN-γ is secreted primarily by CD8+ T-cells. Nearly all cells express receptors for IFN-γ and respond to IFN-γ binding by increasing the surface expression of class I MHC proteins, thereby promoting the presentation of antigen to T-helper (CD4+) cells. IFN-γ also increases the presentation of class II MHC proteins on class II cells further enhancing the ability of cells to present antigen to T-cells.
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Colony Stimulating Factors (CSFs)
CSFs are cytokines that stimulate the proliferation of specific pluripotent stem cells of the bone marrow in adults. Granulocyte-CSF (G-CSF) is specific for proliferative effects on cells of the granulocyte lineage. Macrophage-CSF (M-CSF) is specific for cells of the macrophage lineage. Granulocyte-macrophage-CSF (GM-CSF) has proliferative effects on both classes of lymphoid cells. Epo is also considered a CSF as well as a growth factor, since it stimulates the proliferation of erythrocyte colony-forming units. IL-3 (secreted primarily from T-cells) is also known as multi-CSF, since it stimulates stem cells to produce all forms of hematopoietic cells.
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