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L-Arginine metabolism Nitric oxide Synthases Dimethylarginines Effects Arginine Supplementation Infusion/ Oral Endothelial Function Epidemiology 2 Source of Arginine
L-Arginine is considered a semiessential amino acid: it becomes essential in growing children, during pregnancy or after injury. A Western diet provides about 4-6 g/day of which 40-50% is absorbed.
4 The liver produces considerable amounts of arginine during the urea cycle, but little is available for synthesis. The intestines produce citrulline which is converted by other tissues (kidney, 80%) into L-arginine which is then made available to other tissues.
5 NH2 C=NH2+ NH CH2 CH2 CH2 H C NH3+ COOL-Arginine 2O2 2O2 NO + H2O Nitric Oxide NADPH+ H+ NADP NADP+
+ NH2 C=O NH CH2 CH2 CH2 H C NH3+ COOL-Citrulline The synthesis of nitric oxide (NO) is catalyzed by nitric oxide synthase (NOS). The reaction is more complicated than indicated in the figure.
6 Nitric Oxide Synthase
Three Isoforms Neuronal (constitutive, calcium dependent) Endothelial (constitutive, calcium dependent) Macrophages (inducible, calcium independent). Can lead to high levels of NO being formed.
7 Nitric Oxide Effects (via formation of cGMP)
Relaxes smooth muscle Inhibits platelet aggregation and activation Neurotransmitter Tumoricidal and bactericidal agent from macrophages (excess can damage healthy tissue)
8 Many studies involve infusion or dietary supplements of L-arginine in both animals and humans. The physiological effects elicited were unexpected as the km of NOS for L-arginine is about 2 M whereas the circulating levels of L-arginine are about 100 M.
9 The explanation may involve the presence of naturally occurring inhibitors of NOS (ADMA and NMA). These two analogues of L-arginine plus SDMA are also competitors for the y+ transport system that delivers L-arginine to NOS. 10 Structure of L-arginine and Circulatory Analogues
CH3 NH C NH CH2 CH2 CH2 CH NH2 COOH NH2 L-NMA NH2 NH C NH CH2 CH2 CH2 CH COOH NH2 ADMA NH NH C NH CH2 CH2 CH2 CH COOH NH2 CH3 CH3 N CH3 N C NH CH2 CH2 CH2 CH COOH CH3 NH L-ARGININE SDMA 11 As shown in the metabolic pathway (slide 14):
L-arginine is methylated while a component of proteins by: PRMT (type I): occurs in nucleus, many substrates forms ADMA and NMA PRMT (type II) : specific for myelin basic protein, forms SDMA and NMA The methylated analogues are released by hydrolysis during normal protein turnover 12 The methylated analogues are removed by renal excretion or catabolism DDAH type I associated with neural NOS DDAH type II associated with endothelial NOS Neither DDAH is active on SDMA DPT (a minor pathway) acts on all three analogues The enzymes are particularly active in kidney
13 Protein PRMT (types I and II) Modified Protein Containing ADMA+ SDMA+ NMA Hydrolysis ADMA +SDMA +NMA DDAH (types I and II) Citrulline + Methylamines Acetylated Products DPT -keto acid products Renal Excretion PRMT: Protein arginine methyltransferase ADMA: Asymmetrical dimethylarginine SDMA: Symmetrical dimethylarginine NMA: N-monomethylarginine DDAH: Dimethylaminohydrolase DPT: Dimethylarginine pyruvate transferase 14 In slides 18 and 19, results from Cooke et al (1992) are shown. The investigators fed male rabbits either (a) normal chow (control) or (b) 1% cholesterol diet; or (c) 1% cholesterol diet supplemented with drinking water containing 2.25% L-arginine HCl. After 10 weeks of dietary intervention, analyses indicated:
16 Endothelium dependent relaxation of the thoracic aortae elicited by acetylcholine was reduced in cholesterol-fed animals and the response was significantly ameliorated by L-arginine. L-arginine also significantly reduced the lesion surface area in the descending thoracic aorta elicited by cholesterol diets (intima thickness also reduced)
17 120 % Nonrepineprine Precontraction 100 80 60 40 20 0 9 8 7 6 5 4 ACh (-Log M) Control Cholesterol Arginine Cooke, JP, et al. Journal of Clinical Investigation (1992) 90:1168-1172. 18 40 Plaque (% of Total Surface 20 Area) 10 0 Cholesterol Arginine 30 Cooke, JP, et al. Journal of Clinical Investigation (1992) 90:1168-1172. 19 Candipan et al (1996) fed rabbits either normal chow (controls) or 0.5% cholesterol chow for 10 weeks and then the cholesterol group received either vehicle or L-arginine (2.25% in water) (arginine group) for an additional 13 weeks.
20 Histomorphic measurements indicated a gradual deterioration in the cholesterol group (intima-media thickness) and this was ameliorated by L-arginine at 18 weeks but not at 23 weeks (slide 22). This may indicate that the effects of L-arginine are not sustained.
21 Intima, mm2
Week CHOL ARG 10 14 18 23 0.920.36 0.980.16 1.800.37 2.331.36 3.580.71 1.510.64* 4.210.74 4.181.71 Candipan, RC, et al. Arteriosclerosis, Thrombosis, and Vascular *p< 0.05 Biology (1996) 16(1): 44-50. 22 In human studies, Drexler et al (1994) infused 18 cardiac transplant recipients with acetylcholine (10-6, 105 , 10-4 mol/L) before and after intravenous with L-arginine (10 mg/ kg. min. for 20 minutes). (slide 24) Acetylcholine elicited a dosedependent constriction of the coronary artery that was attenuated by L-arginine (p <0.01 at 10-4 acetylcholine)
23 2 1 0 Response to ACh -1 (% change from -2 baseline) -3 -4 -5 -6 -7 Before L-arginine
Before L-Arginine After L-Arginine ach 10-6 ach 10-5 ach10-4 After L-arginine Drexler, H, et al. Circulation (1994) 89(4):1615 24 Bger et al (1998) reported that infusion of L-arginine ameliorated the clinical symptoms of intermittent claudication in patients with peripheral arterial occlusive disease. 13 patients received two intravenous infusions of L-arginine (8 g each) for 3 weeks.
25 13 patients received no infusions (control group) Both groups maintained normal walking exercises. Results indicated that L-arginine improved pain-free walking distance (slide 27) by 230 63% (p < 0.05). Absolute walking distance also improved by 155 48% (p < 0.05).
26 Pain-free Walking Distance (m) Bger, RH, et al. J Am Coll Cardiol (1998) 32(5): 1336-44. 27 Physiological effects have also been elicited by infusion of NOS inhibitors. Vallance et al (1992) infused ADMA(8 mol/min for 5 min into 5 volunteers) and observed a decrease in forearm blood flow (slide 30) 28 McVeigh et al (2001) infused L-NAME (NG- nitro-L-arginine methyl ester) into 15 healthy men and observed an increase in systemic vascular resistance (slide 31) and a decrease in small artery compliance (slide 32) The effects were ameliorated by infusion of L-arginine but not by Darginine.
29 Vallance, P, et al. Lancet (1992) 339:572-575. 30 p < 0.01 versus control p < 0.01 D-arginine versus L-arginine McVeigh, GE, et al. Clinical Science. (2001)100: 387-393. 31 * p < 0.05 versus controls p < 0.01 versus controls p < 0.01 D-arginine versus L-arginine
32 McVeigh, GE et al. Clinical Science. (2001) 100: 387-393. Epidemiological studies have observed associations between ADMA concentrations and subclinical and clinical measures of atherosclerosis. Miyazaki et al (1999) studied 116 subjects with no symptoms of coronary or peripheral artery disease and not taking medications. Results indicated:
34 Plasma ADMA levels were significantly correlated with intima-media thickness (slide 36) Stepwise multiple regression analysis indicated plasma ADMA was a significant determinant of the intimamedia thickness (slide 37) 35 Miyazaki, H, et al. Circulation (1999) 99(9): 1141-1146. 36 Variable Coefficient p Age Plasma ADMA Mean arterial pressure glucose Smoking Total cholesterol Positive family history 0.005 0.29 0.16 0.14 -0.05 0.02 -0.02 0.0001 0.03 0.1 0.16 0.56 0.81 0.94 r= 0.41. Miyazaki, H, et al. Circulation (1999) 99(9):1141-1146. 37 Zoccalli et al (2001) studied 225 haemodialysis patients with endstage renal disease. Plasma ADMA was significantly and independently correlated with allcause mortality and fatal and nonfatal cardiovascular events. No significant associations were observed for plasma SDMA or Larginine (slides 40 and 41).
38 Valkonen et al (2001), in a prospective case-control study analyzed the association between ADMA and the risk of acute coronary events. Among non-smoking men, ADMA was a significant risk factor for acute coronary events. The conclusions were dependent on presence or absence of a history of coronary heart disease (CHD): not significant in the absence of a history of CHD; significant in the presence of a history of CHD (slide 42).
39 All-cause mortality Unit of increase Hazard ratio* (95%Cl) p Fully adjusted hazard ratio* (95% Cl) p ADMA 1.28(1.16-1.41) 1 mol/L <0.0001 1.26(1.11-1.41) 0.0001 SDMA 1 mol/L 1.02(0.93-1.11) 1.01(0.89-1.14) 0.73 0.92 1.06(0.94-1.18) 0.92(0.80-1.05) 0.34 0.22 L-arginine 10 mmol/L Zoccalli, C, et al. Lancet (2001) 358: 2113-2117. 40 Fatal and non-fatal cardiovascular events
Hazard ratio* p Fully adjusted hazard ratio (95% Cl) 0.0001 1.17 (1.04-1.33) 0.008 p Unit of increase ADMA 1 mol/L (95% Cl) 1.21 (1.10-1.32) SDMA Larginine 1 mol/L 10mmol/ L 0.97 (0.88-1.07) 1.06 (0.94-1.19) 0.61 1.00 (0.88-1.14) 0.37 1.00 (0.87-1.15) 0.98 0.97 Zoccalli, C, et al. Lancet (2001) 358: 2113-2117. 41 All (n=50) No history of coronary heart disease* (n=80) p Odds ratio (95% Cl) p History of coronary heart disease* (n=70) Odds ratio (95% Cl) p Odds ratio (95% Cl) Baseline characteristic ADMA, highest quartile (>0.62 mol/L) Family history of coronary heart disease 3.92 (1.25-12.3) 0.02 2.39 (0.54-10.5) 0.25 21.8 (1.4-348.5) 0.03 3.02 (1.14-7.96) 0.03 2.53 (0.75-8.54) 0.14 3.3 (0.6-19.1) 0.19 Valkonen, V-P, et al. Lancet (2001) 358: 2127-8. 42 Are the effects of L-arginine supplementation sustained? Will L-arginine supplementation be of clinical benefit? How general will any benefit be?
43 I would like to thank Ms. Meghan Dabkowski for her assistance in the preparation of this presentation. 44 ...
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This note was uploaded on 02/23/2012 for the course PHARM 290 taught by Professor Staff during the Fall '10 term at Rutgers.
- Fall '10