Investigation of cellular mechanisms of the possible interaction of L-cysteine/ hydrogen sulfide pathway with RHOA/RHO-kinase in mouse bladder smooth muscle

In our study, the cellular mechanisms of the possible interaction of the L-cysteine/hydrogen sulfide (H2S) pathway with RhoA/Rho-kinase in isolated mouse bladder tissue were investigated. For this purpose, cumulative carbachol (10-8-10-4 M) contractile responses were obtained in bladder tissues isolated from Swiss-albino male mice. Contractile responses were inhibited in the presence of L-cysteine (endogenous H2S; 10-2 M), sodium hydrogen sulfide (NaHS) (exogenous H2S; 10-3 M), Y-27632 (Rho-kinase inhibitor; 10-6 M), GF-109203x (protein kinase C inhibitor 10-6 M). Although Y-27632 had no effect on the inhibition of NaHS or GF-109203x, the inhibitory effect of L-cysteine was reversed in the presence of propargylglycine (PAG; cystathionine-gamma-lyase (CSE) inhibitor; 10–2 M), aminooxyacetic acid (AOAA; cystathionine-β-synthase (CBS) inhibitor; 10–2 M), Y-27632 and GF-109203x. The inhibitory effects of Y-27632 and GF-109203x were reversed in the presence of PAG, but AOAA had no effect on the Y-27632 response. Basal H2S release was detected and while L-cysteine, Y-27632 and GF-109203x increased H2S levels, PAG showed a significant decrease in the measurements of endogenous H2S levels. Increased H2S levels of Y-27632 and GF-109203x groups were inhibited in the presence of L-cysteine. In addition, protein expressions of CSE, CBS and 3-mercaptopurivate sülfürtansferase (3-MST) enzymes responsible for endogenous H2S synthesis were demonstrated by western blot method. In our current study, it has been suggested that the interaction between L-cysteine/H2S and RhoA/Rho-kinase and/or PKC pathway may occur via the CSE/H2S pathway.