Acute Rho-kinase inhibition improves coronary dysfunction in vivo, in the early diabetic microcirculation.
OBJECTIVE
Activation of RhoA/Rho-kinase (ROCK) is increasingly implicated in acute vasospasm and chronic vasoconstriction in major organ systems. Therefore we aimed to ascertain whether an increase in ROCK activity plays a role in the deterioration of coronary vascular function in early stage diabetes.


METHODS
Synchrotron radiation microangiography was used to determine in vivo coronary responses in diabetic (3 weeks post streptozotocin 65 mg/kg ip) and vehicle treated male Sprague-Dawley rats (n = 8 and 6). Changes in vessel number and calibre during vasodilator stimulation before and after blockade of nitric oxide synthase and cyclooxygenase were compared between rats. Acute responses to ROCK inhibitor, fasudil (10 mg/kg iv) was evaluated. Further, perivascular and myocardial fibrosis, arterial intimal thickening were assessed by histology, and capillary density, nitrotyrosine and ROCK1/2 expressions were evaluated by immunohistochemical staining.


RESULTS
Diabetic rats had significantly elevated plasma glucose (P < 0.001 vs control), but did not differ in fibrotic scores, media to lumen ratio, capillary density or baseline visible vessel number or calibre. Responses to acetylcholine and sodium nitroprusside stimulation were similar between groups. However, in comparison to control rats the diabetic rats showed more segmental constrictions during blockade, which were not completely alleviated by acetylcholine, but were alleviated by fasudil. Further, second order vessel branches in diabetic rats were significantly more dilated relative to baseline (37% vs 12% increase, P < 0.05) after fasudil treatment compared to control rats, while visible vessel number increased in both groups. ROCK2 expression was borderline greater in diabetic rat hearts (P < 0.053).


CONCLUSIONS
We found that ahead of the reported decline in coronary endothelial vasodilator function in diabetic rats there was moderate elevation in ROCK expression, more widespread segmental constriction when nitric oxide and prostacyclin production were inhibited and notably, increased calibre in second and third order small arteries-arterioles following ROCK inhibition. Based on nitrotyrosine staining oxidative stress was not significantly elevated in early diabetic rats. We conclude that tonic ROCK mediated vasoconstriction contributes to coronary vasomotor tone in early diabetes.
Department of Physiology, Monash University, Melbourne, Australia. james.pearson@monash.edu.
James T
JT
Pearson
Mathew J
MJ
Jenkins
Amanda J
AJ
Edgley
Takashi
T
Sonobe
Mandar
M
Joshi
Mark T
MT
Waddingham
Yutaka
Y
Fujii
Daryl O
DO
Schwenke
Hirotsugu
H
Tsuchimochi
Misa
M
Yoshimoto
Keiji
K
Umetani
Darren J
DJ
Kelly
Mikiyasu
M
Shirai
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D017023:Coronary Angiography; D003326:Coronary Circulation / Q000187:drug effects*; D003331:Coronary Vessels / Q000000981:diagnostic imaging / Q000187:drug effects* / Q000201:enzymology / Q000503:physiopathology; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D003921:Diabetes Mellitus, Experimental / Q000150:complications*; D003925:Diabetic Angiopathies / Q000000981:diagnostic imaging / Q000188:drug therapy* / Q000201:enzymology / Q000209:etiology / Q000503:physiopathology; D011464:Epoprostenol / Q000378:metabolism; D005355:Fibrosis; D008297:Male; D008833:Microcirculation / Q000187:drug effects*; D009206:Myocardium / Q000201:enzymology / Q000473:pathology; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism; D014661:Vasoconstriction / Q000187:drug effects; D014664:Vasodilation / Q000187:drug effects; D014665:Vasodilator Agents / Q000494:pharmacology*; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism
D016861:Cyclooxygenase Inhibitors; D047428:Protein Kinase Inhibitors; D014665:Vasodilator Agents; D009569:Nitric Oxide; C002744:3-nitrotyrosine; D014443:Tyrosine; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D011464:Epoprostenol; D019001:Nitric Oxide Synthase; D054460:rho-Associated Kinases; C049347:fasudil