Post on 24-Dec-2015
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ContentContent::••L-Arginine metabolismL-Arginine metabolism••Nitric oxide Nitric oxide
••SynthasesSynthases ••DimethylargininesDimethylarginines
••EffectsEffects••Arginine SupplementationArginine Supplementation
••Infusion/ OralInfusion/ Oral••Endothelial FunctionEndothelial Function
••EpidemiologyEpidemiology
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Source of ArginineSource of Arginine
L-Arginine is considered a semi-L-Arginine is considered a semi-essential amino acid: it becomes essential amino acid: it becomes essential in growing children, during essential in growing children, during pregnancy or after injury. pregnancy or after injury.
A Western diet provides about 4-6 A Western diet provides about 4-6 g/day of which 40-50% is absorbed. g/day of which 40-50% is absorbed.
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••The liver produces considerable The liver produces considerable amounts of arginine during the urea amounts of arginine during the urea cycle, but little is available for cycle, but little is available for synthesis.synthesis.
• • The intestines produce citrulline The intestines produce citrulline
which is converted by other tissues which is converted by other tissues (kidney, 80%) into L-arginine which (kidney, 80%) into L-arginine which is then made available to other is then made available to other tissues. tissues.
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NADP+
2O2
The synthesis of nitric oxide (NO) is catalyzed by nitric oxide synthase (NOS). The reaction is more complicated than indicated in the figure.
NH2
C=NH2+
NH
CH2
CH2
CH2
H C NH3+
COO-
L-Arginine
NH2
C=O
NH
CH2
CH2
CH2
H C NH3+
COO-
L-Citrulline
NADPH+ H+ NADP+
2O2 NO + H2O Nitric Oxide
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Nitric Oxide SynthaseNitric Oxide Synthase
Three IsoformsThree Isoforms
Neuronal (constitutive, calcium Neuronal (constitutive, calcium dependent)dependent)
Endothelial (constitutive, calcium Endothelial (constitutive, calcium dependent)dependent)
Macrophages (inducible, calcium Macrophages (inducible, calcium independent). Can lead to high independent). Can lead to high levels of NO being formed. levels of NO being formed.
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Nitric Oxide Effects Nitric Oxide Effects (via formation of cGMP)(via formation of cGMP)Relaxes smooth muscleRelaxes smooth muscle
Inhibits platelet aggregation and Inhibits platelet aggregation and activationactivation
NeurotransmitterNeurotransmitter
Tumoricidal and bactericidal agent Tumoricidal and bactericidal agent from macrophages (excess can from macrophages (excess can damage healthy tissue)damage healthy tissue)
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• • Many studies involve infusion or Many studies involve infusion or dietary supplements of L-arginine in dietary supplements of L-arginine in both animals and humans. both animals and humans.
• • The physiological effects elicited The physiological effects elicited were unexpected as the kwere unexpected as the kmm of NOS for of NOS for
L-arginine is about 2 L-arginine is about 2 μμM whereas the M whereas the circulating levels of L-arginine are circulating levels of L-arginine are about 1about 100 00 μμM. M.
1010
• • The explanation may involve the The explanation may involve the presence of naturally occurring presence of naturally occurring inhibitors of NOS (ADMA and NMA). inhibitors of NOS (ADMA and NMA). These two analogues of L-arginine These two analogues of L-arginine plus SDMA are also competitors for plus SDMA are also competitors for the ythe y+ + transport system that delivers transport system that delivers L-arginine to NOS. L-arginine to NOS.
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Structure of L-arginine and Circulatory AnaloguesStructure of L-arginine and Circulatory Analogues
NH NH2
C
NH
CH2
CH2
CH2
CH
NH2 COOH
L-ARGININE L-NMA ADMA SDMA
NH NH
CH3
C
NH
CH2
CH2
CH2
CH
NH2 COOH
NH N
CH3 CH3
C
NH
CH2
CH2
CH2
CH
NH2 COOH
C
N NH
CH3 CH3
NH
CH2
CH2
CH2
CH NH2 COOH
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As shown in the metabolic pathway As shown in the metabolic pathway (slide 14):(slide 14):
••L-arginine is methylated while a L-arginine is methylated while a component of proteins by:component of proteins by:••PRMT (type I): occurs in nucleus, PRMT (type I): occurs in nucleus, many substrates forms ADMA and many substrates forms ADMA and NMANMA••PRMT (type II) : specific for myelin PRMT (type II) : specific for myelin basic protein, forms SDMA and NMAbasic protein, forms SDMA and NMA
••The methylated analogues are The methylated analogues are released by hydrolysis during normal released by hydrolysis during normal protein turnoverprotein turnover
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• • The methylated analogues are removed The methylated analogues are removed by renal excretion or catabolismby renal excretion or catabolism••DDAH type I associated with neural DDAH type I associated with neural NOSNOS••DDAH type II associated with DDAH type II associated with endothelial NOSendothelial NOS••Neither DDAH is active on SDMANeither DDAH is active on SDMA••DPT (a minor pathway) acts on all DPT (a minor pathway) acts on all three analoguesthree analogues••The enzymes are particularly active in The enzymes are particularly active in kidneykidney
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Protein
PRMT (types I and II)
Modified Protein Containing ADMA+ SDMA+ NMA
Hydrolysis
ADMA +SDMA +NMA
Acetylated Products α-keto
acid products
Citrulline + Methylamines
DDAH (types I and II) Renal
Excretion
PRMT: Protein arginine methyltransferaseADMA: Asymmetrical dimethylarginineSDMA: Symmetrical dimethylarginine
NMA: N-monomethylarginineDDAH: Dimethylaminohydrolase
DPT: Dimethylarginine pyruvate transferase
DPT
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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:
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Endothelium dependent relaxation of Endothelium dependent relaxation of the thoracic aortae elicited by the thoracic aortae elicited by acetylcholine was reduced in acetylcholine was reduced in cholesterol-fed animals and the cholesterol-fed animals and the response was significantly response was significantly ameliorated by L-arginine.ameliorated by L-arginine.L-arginine also significantly reduced L-arginine also significantly reduced the lesion surface area in the the lesion surface area in the descending thoracic aorta elicited by descending thoracic aorta elicited by cholesterol diets (intima thickness cholesterol diets (intima thickness also reduced)also reduced)
1818
0
20
40
60
80
100
120
9 8 7 6 5 4
ACh (-Log M)
% N
on
rep
inep
rin
e P
reco
ntr
acti
on
Control
Cholesterol
Arginine
Cooke, JP, et al. Journal of Clinical Investigation (1992) 90:1168-1172.
1919
0
10
20
30
40
Plaque (% of Total Surface
Area)
Cholesterol Arginine
Cooke, JP, et al. Journal of Clinical Investigation (1992) 90:1168-1172.
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••Candipan et al (1996) fed rabbits Candipan et al (1996) fed rabbits either normal chow (controls) or 0.5% either normal chow (controls) or 0.5% cholesterol chow for 10 weeks and cholesterol chow for 10 weeks and then the cholesterol group received then the cholesterol group received either vehicle or L-arginine (2.25% in either vehicle or L-arginine (2.25% in water) (arginine group) for an water) (arginine group) for an additional 13 weeks. additional 13 weeks.
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••Histomorphic measurements indicated Histomorphic measurements indicated a gradual deterioration in the a gradual deterioration in the cholesterol group (intima-media cholesterol group (intima-media thickness) and this was ameliorated thickness) and this was ameliorated by L-arginine at 18 weeks but not at by L-arginine at 18 weeks but not at 23 weeks (slide 22). 23 weeks (slide 22).
••This may indicate that the This may indicate that the effects of L-arginine are not effects of L-arginine are not sustained. sustained.
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Candipan, RC, et al. Arteriosclerosis, Thrombosis, and Vascular Biology (1996) 16(1): 44-50.
Intima, mm2
WeekWeek CHOLCHOL
ARG ARG
1010 0.92±0.360.92±0.36 0.98±0.160.98±0.16
1414 1.80±0.371.80±0.37 2.33±1.362.33±1.36
1818 3.58±0.713.58±0.71 1.51±0.64*1.51±0.64*
2323 4.21±0.744.21±0.74 4.18±1.714.18±1.71
*p< 0.05*p< 0.05
2323
In human studies, Drexler et al (1994) In human studies, Drexler et al (1994) infused 18 cardiac transplant infused 18 cardiac transplant recipients with acetylcholine (10recipients with acetylcholine (10-6,-6, 10 10--
55, 10, 10-4-4 mol/L) before and after mol/L) before and after intravenous with L-arginine (10 mg/ intravenous with L-arginine (10 mg/ kg. min. for 20 minutes). (slide 24)kg. min. for 20 minutes). (slide 24)
Acetylcholine elicited a dose-Acetylcholine elicited a dose-dependent constriction of the dependent constriction of the coronary artery that was attenuated coronary artery that was attenuated by L-arginine (p <0.01 at 10by L-arginine (p <0.01 at 10-4 -4
acetylcholine)acetylcholine)
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-7
-6
-5
-4
-3
-2
-1
0
1
2
Response to ACh (% change from
baseline)
ach 10-6
ach 10-5
ach10-4
Before L-arginine After L-arginine
Before L-Arginine After L-Arginine
Drexler, H, et al. Circulation (1994) 89(4):1615
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• • BBööger et al (1998) reported that infusion ger et al (1998) reported that infusion of L-arginine ameliorated the clinical of L-arginine ameliorated the clinical symptoms of intermittent claudication in symptoms of intermittent claudication in patients with peripheral arterial occlusive patients with peripheral arterial occlusive disease. disease.
• • 13 patients received two 13 patients received two intravenous infusions of L-arginine (8 g intravenous infusions of L-arginine (8 g each) for 3 weeks. each) for 3 weeks.
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••13 patients received no infusions 13 patients received no infusions (control group)(control group)
••Both groups maintained normal Both groups maintained normal walking exercises. walking exercises.
••Results indicated that L-arginine Results indicated that L-arginine improved pain-free walking distance improved pain-free walking distance (slide 27) by 230± 63% (p < 0.05). (slide 27) by 230± 63% (p < 0.05). Absolute walking distance also Absolute walking distance also improved by 155 ±48% (p < 0.05). improved by 155 ±48% (p < 0.05).
2727
Pai
n-f
ree
Wal
kin
g D
ista
nce
(m
)
Böger, RH, et al. J Am Coll Cardiol (1998) 32(5): 1336-44.
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••Physiological effects have also been Physiological effects have also been elicited by infusion of NOS inhibitors.elicited by infusion of NOS inhibitors.
••Vallance et al (1992) infused ADMA(8 Vallance et al (1992) infused ADMA(8 μμmol/min for 5 min into 5 volunteers) mol/min for 5 min into 5 volunteers) and observed a decrease in forearm and observed a decrease in forearm blood flow (slide 30)blood flow (slide 30)
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••McVeigh et al (2001) infused L-NAME McVeigh et al (2001) infused L-NAME (N(NGG- nitro-L-arginine methyl ester) into - nitro-L-arginine methyl ester) into 15 healthy men and observed an 15 healthy men and observed an increase in systemic vascular increase in systemic vascular resistance (slide 31) and a decrease resistance (slide 31) and a decrease in small artery compliance (slide 32)in small artery compliance (slide 32)
••The effects were ameliorated by The effects were ameliorated by infusion of L-arginine but not by D-infusion of L-arginine but not by D-arginine. arginine.
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Vallance, P, et al. Lancet (1992) 339:572-575.
3131 McVeigh, GE, et al. Clinical Science. (2001)100: 387-393.
† p < 0.01 versus control
‡ p < 0.01 D-arginine versus L-arginine
3232 McVeigh, GE et al. Clinical Science. (2001) 100: 387-393.
* p < 0.05 versus controls
† p < 0.01 versus controls
‡ p < 0.01 D-arginine versus L-arginine
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•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:
3535
••Plasma ADMA levels were significantly Plasma ADMA levels were significantly correlated with intima-media correlated with intima-media thickness (slide 36)thickness (slide 36)
••Stepwise multiple regression analysis Stepwise multiple regression analysis indicated plasma ADMA was a indicated plasma ADMA was a significant determinant of the intima-significant determinant of the intima-media thickness (slide 37)media thickness (slide 37)
3636 Miyazaki, H, et al. Circulation (1999) 99(9): 1141-1146.
3737 Miyazaki, H, et al. Circulation (1999) 99(9):1141-1146.
Variable Variable CoefficientCoefficient
p p
AgeAge 0.0050.005 0.00010.0001
Plasma ADMAPlasma ADMA 0.290.29 0.030.03
Mean arterialMean arterialpressurepressure 0.160.16 0.10.1
Σ glucoseΣ glucose 0.140.14 0.160.16
SmokingSmoking -0.05-0.05 0.560.56
Total cholesterolTotal cholesterol 0.020.02 0.810.81
Positive family Positive family -0.02-0.02 0.940.94
historyhistory
r²= 0.41.r²= 0.41.
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• •Zoccalli et al (2001) studied 225 Zoccalli et al (2001) studied 225 haemodialysis patients with end-haemodialysis patients with end-stage renal disease. stage renal disease.
••Plasma ADMA was significantly and Plasma ADMA was significantly and independently correlated with all-independently correlated with all-cause mortality and fatal and non-cause mortality and fatal and non-fatal cardiovascular events. No fatal cardiovascular events. No significant associations were significant associations were observed for plasma SDMA or L-observed for plasma SDMA or L-arginine (slides 40 and 41).arginine (slides 40 and 41).
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• • Valkonen et al (2001), in a prospective Valkonen et al (2001), in a prospective case-control study analyzed the case-control study analyzed the association between ADMA and the risk of association between ADMA and the risk of acute coronary events. acute coronary events.
• •Among non-smoking men, ADMA was a Among non-smoking men, ADMA was a significant risk factor for acute coronary significant risk factor for acute coronary events. The conclusions were dependent events. The conclusions were dependent on presence or absence of a history of on presence or absence of a history of coronary heart disease (CHD): not coronary heart disease (CHD): not significant in the absence of a history of significant in the absence of a history of CHD; significant in the presence of a CHD; significant in the presence of a history of CHD (slide 42). history of CHD (slide 42).
4040 Zoccalli, C, et al. Lancet (2001) 358: 2113-2117.
0.220.22 0.92(0.80-1.05)0.92(0.80-1.05)0.920.921.01(0.89-1.14)1.01(0.89-1.14) 10 10 mmol/L mmol/L L-arginineL-arginine
0.340.34 1.06(0.94-1.18)1.06(0.94-1.18)0.730.731.02(0.93-1.11)1.02(0.93-1.11)
11 μmol/L μmol/LSDMASDMA
0.00010.0001 1.26(1.11-1.41)1.26(1.11-1.41)
<<0.00010.00011.28(1.16-1.41)1.28(1.16-1.41)
11 μmol/L μmol/L ADMAADMA
(95% Cl)(95% Cl)
hazard ratio*hazard ratio*(95%Cl)(95%Cl)
pp Fully adjusted Fully adjusted ppHazard ratio*Hazard ratio* Unit of Unit of increaseincrease
All-cause mortalityAll-cause mortality
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Hazard ratio*Hazard ratio* ppFully adjusted Fully adjusted hazardhazard pp
Unit ofUnit ofincreaseincrease (95% Cl)(95% Cl) ratio (95% Cl)ratio (95% Cl)
ADMAADMA 1 μmol/L 1 μmol/L 1.21 (1.10-1.32)1.21 (1.10-1.32) 0.00010.0001 1.17 (1.04-1.33)1.17 (1.04-1.33) 0.0080.008
SDMASDMA 1 μmol/L 1 μmol/L 0.97 (0.88-1.07)0.97 (0.88-1.07) 0.610.61 1.00 (0.88-1.14)1.00 (0.88-1.14) 0.980.98
L-L-argininearginine
10mmol/10mmol/LL 1.06 (0.94-1.19)1.06 (0.94-1.19) 0.370.37 1.00 (0.87-1.15)1.00 (0.87-1.15) 0.970.97
Fatal and non-fatal cardiovascular eventsFatal and non-fatal cardiovascular events
Zoccalli, C, et al. Lancet (2001) 358: 2113-2117.
4242 Valkonen, V-P, et al. Lancet (2001) 358: 2127-8.
All (n=50)All (n=50) No history of No history of coronary heart coronary heart disease* (n=80)disease* (n=80)
History of History of coronary heart coronary heart disease* (n=70)disease* (n=70)
Odds ratio (95% Odds ratio (95% Cl)Cl)
pp Odds ratio (95% Odds ratio (95% Cl)Cl)
pp Odds ratio (95% Odds ratio (95% Cl)Cl)
pp
BaselineBaseline
characteristiccharacteristic
ADMA, highest ADMA, highest quartile quartile
((>0.62 >0.62 μμmol/L)mol/L)
3.92 (1.25-12.3)3.92 (1.25-12.3) 0.020.02 2.39 (0.54-10.5)2.39 (0.54-10.5) 0.250.25 21.8 (1.4-348.5)21.8 (1.4-348.5) 0.030.03
Family history of Family history of
coronary heart coronary heart diseasedisease
3.02 (1.14-7.96)3.02 (1.14-7.96) 0.030.03 2.53 (0.75-8.54)2.53 (0.75-8.54) 0.140.14 3.3 (0.6-19.1)3.3 (0.6-19.1) 0.190.19
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Are the effects of L-arginine Are the effects of L-arginine supplementation sustained?supplementation sustained?
Will L-arginine supplementation be of Will L-arginine supplementation be of clinical benefit?clinical benefit?
How general will any benefit be?How general will any benefit be?
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I would like to thank Ms. Meghan I would like to thank Ms. Meghan Dabkowski for her assistance in the Dabkowski for her assistance in the
preparation of this presentation. preparation of this presentation.