Diabetes‐induced activation of the RhoA/Rho kinase pathway in cardiomyocytes is sustained by a positive feedback loop involving PKC beta2 and iNOS

We previously reported that acute inhibition of Rho kinase (ROCK) normalized contractile function of diabetic rat hearts. However, the mechanism responsible for the sustained ROCK activation in diabetes is unclear. We recently showed that PKCβ 2 induces iNOS expression, which in turn upregulates RhoA and increases ROCK activity. However, we found that ROCK also controls RhoA expression. This led us to hypothesize that ROCK upregulates RhoA by activating PKCβ 2 and subsequently promoting iNOS expression, forming a positive feedback loop. This was investigated in ventricular cardiomyocytes isolated from streptozotocin‐diabetic and control rat hearts. Incubation of diabetic cardiomyocytes with the Rho inhibitor C3 toxin or ROCK inhibitor Y‐27632 not only attenuated the diabetes‐induced increase in ROCK activity and RhoA expression but also markedly decreased PKCβ 2 Thr 641 phosphorylation (index of activity). The same treatments had no effect in control cells. Inhibition of PKCβ with LY333531 or iNOS with 1400W also blunted the increase in RhoA expression and ROCK activity in diabetic cardiomyocytes. Blockade of ROCK or PKCβ significantly decreased iNOS expression, while inhibition of iNOS attenuated PKCβ 2 phosphorylation. The data suggest that RhoA/ROCK pathway activation in diabetic cardiomyopathy is sustained by a positive feedback loop involving PKCβ 2 activation and subsequent iNOS upregulation.