Isoform-specific regulation by N(G),N(G)-dimethylarginine dimethylaminohydrolase of rat serum asymmetric dimethylarginine and vascular endothelium-derived relaxing factor/NO.
Asymmetric dimethylarginine (ADMA), which inhibits NO synthase, is inactivated by N(G),N(G)-dimethylarginine dimethylaminohydrolase (DDAH). We tested whether DDAH-1 or -2 regulates serum ADMA (S(ADMA)) and/or endothelium-derived relaxing factor (EDRF)/NO. Small inhibitory (si)RNAs targeting DDAH-1 or -2, or an siRNA control were given intravenously to rats. After 72 hours, EDRF/NO was assessed from acetylcholine-induced, NO synthase-dependent relaxation and 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate for NO activity in isolated mesenteric resistance vessels (MRVs). Expression of mRNA for DDAH-1 versus -2 was 2- and 7-fold higher in the kidney cortex and liver, respectively, whereas expression of DDAH-2 versus -1 was 5-fold higher in MRVs. The proteins and mRNAs for DDAH-1 or -2 were reduced selectively by 35% to 85% in the kidney cortex, liver, and MRVs 72 hours following the corresponding siRNA. S(ADMA) was increased only after siDDAH-1 (266+/-25 versus 342+/-39 [mean+/-SD] nmol x L(-1); P<0.005), whereas EDRF/NO responses and NO activity were not changed consistently by siDDAH-1 but were greatly reduced after siDDAH-2. Mean arterial pressure was not changed significantly by any siRNA. In conclusion, S(ADMA) is regulated by DDAH-1, which is expressed at sites of ADMA metabolism in the kidney cortex and liver, whereas EDRF/NO is regulated primarily by DDAH-2, which is expressed strongly in blood vessels. This implies specific functions of DDAH isoforms.
Division of Nephrology and Hypertension, Georgetown University, Washington, DC 20007, USA.
Dan
D
Wang
Pritmohinder S
PS
Gill
Tinatin
T
Chabrashvili
Maristela L
ML
Onozato
Julie
J
Raggio
Margarida
M
Mendonca
Kathryn
K
Dennehy
Min
M
Li
Paul
P
Modlinger
James
J
Leiper
Patrick
P
Vallance
Oscar
O
Adler
Anna
A
Leone
Akihiro
A
Tojo
William J
WJ
Welch
Christopher S
CS
Wilcox
D000109:Acetylcholine / Q000494:pharmacology; D000581:Amidohydrolases / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001120:Arginine / Q000031:analogs & derivatives* / Q000097:blood / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D045462:Endothelium-Dependent Relaxing Factors / Q000097:blood / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D007527:Isoenzymes / Q000378:metabolism; D007672:Kidney Cortex / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D008297:Male; D008638:Mesenteric Arteries / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D009569:Nitric Oxide / Q000097:blood / Q000378:metabolism*; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013997:Time Factors; D014664:Vasodilation / Q000187:drug effects; D014665:Vasodilator Agents / Q000494:pharmacology
D045462:Endothelium-Dependent Relaxing Factors; D007527:Isoenzymes; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D014665:Vasodilator Agents; D009569:Nitric Oxide; C018524:N,N-dimethylarginine; D001120:Arginine; D000581:Amidohydrolases; C059999:dimethylargininase; D000109:Acetylcholine