Knockdown of arginase I restores NO signaling in the vasculature of old rats.
Arginase, expressed in endothelial cells and upregulated in aging blood vessels, competes with NO synthase (NOS) for l-arginine, thus modulating vasoreactivity and attenuating NO signaling. Moreover, arginase inhibition restores endothelial NOS signaling and l-arginine responsiveness in old rat aorta. The arginase isoform responsible for modulating NOS, however, remains unknown. Because isoform-specific arginase inhibitors are unavailable, we used an antisense (AS) oligonucleotide approach to knockdown arginase I (Arg I). Western blot and quantitative PCR confirmed that Arg I is the predominant isoform expressed in endothelialized aortic rings and is upregulated in old rats compared with young. Aortic rings from 22-month-old rats were incubated for 24 hours with sense (S), AS oligonucleotides, or medium alone (C). Immunohistochemistry, immunoblotting, and enzyme assay confirmed a significant knockdown of Arg I protein and arginase activity in AS but not S or C rings. Conversely, calcium-dependent NOS activity and vascular metabolites of NO was increased in AS versus S or C rings. Acetylcholine (endothelial-dependent) vasorelaxant responses were enhanced in AS versus S or C treated rings. In addition, 1H-oxadiazolo quinoxalin-1-one (10 micromol/L), a soluble guanylyl cyclase inhibitor, increased the phenylephrine response in AS compared with S and C rings suggesting increased NO bioavailability. Finally, l-arginine (0.1 mmol/L)-induced relaxation was increased in AS versus C rings. These data support our hypothesis that Arg I plays a critical role in the pathobiology of age-related endothelial dysfunction. AS oligonucleotides may, therefore, represent a novel therapeutic strategy against age-related vascular endothelial dysfunction.
White
Anthony R
AR
Department of Anesthesiology and Critical Care Medicine, The Johns Hopkins Medical Institutions, Baltimore, MD, USA.
Ryoo
Sungwoo
S
Li
Dechun
D
Champion
Hunter C
HC
Steppan
Jochen
J
Wang
Danming
D
Nyhan
Daniel
D
Shoukas
Artin A
AA
Hare
Joshua M
JM
Berkowitz
Dan E
DE
D000109:Acetylcholine / Q000494:pharmacology; D000375:Aging / Q000378:metabolism*; D000818:Animals; D001011:Aorta / Q000187:drug effects / Q000378:metabolism*; D001119:Arginase / Q000235:genetics*; D001120:Arginine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D066298:In Vitro Techniques; D008297:Male; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D010069:Oxadiazoles / Q000494:pharmacology; D010656:Phenylephrine / Q000494:pharmacology; D011810:Quinoxalines / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction / Q000187:drug effects*; D015854:Up-Regulation; D014662:Vasoconstrictor Agents / Q000494:pharmacology; D014664:Vasodilation; D014665:Vasodilator Agents / Q000494:pharmacology
C095284:1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one; D004791:Enzyme Inhibitors; D016376:Oligonucleotides, Antisense; D010069:Oxadiazoles; D011810:Quinoxalines; D014662:Vasoconstrictor Agents; D014665:Vasodilator Agents; D010656:Phenylephrine; D009569:Nitric Oxide; D001120:Arginine; D019001:Nitric Oxide Synthase; D001119:Arginase; D000109:Acetylcholine; D002118:Calcium