It has been shown that blockade of
Rho kinase with pharmacologic inhibitors inhibits renal
fibrosis. This study examined the role of
Rho kinase in renal
fibrosis in the unilateral
ureteral obstruction (UUO) model in mice that do not express the ROCK1 gene, a critical downstream mediator of
Rho GTPase. Unexpected, real-time PCR, Western blot, and immunohistochemistry demonstrated that, compared with the wild-type mice, mice with ROCK1 knockout (KO) were not protected against renal
fibrosis at both the early (day 5) and late (day 10) UUO, as determined by histology and expression of both
mRNA and
protein levels of alpha-smooth muscle actin,
collagen types I and III, and
fibronectin within the diseased kidney. Then the mechanisms of loss of protective effect on renal
fibrosis in ROCK1 KO mice were investigated. It is interesting that mice that lacked ROCK1 did not have altered expression of ROCK2 but significantly increased
TGF-beta expression and Smad2/3 activation (phosphorylation and nuclear translocation) in the diseased kidney at day 5, which remained high at day 10 of UUO. Similarly, primary cultures of kidney fibroblasts that were obtained from both ROCK1 wild-type and KO mice showed that deletion of ROCK1 did not prevent
TGF-beta-induced activation of Smad2/3 and
collagen I expression. This also was observed in the presence of
Rho kinase inhibitor
Y-27632. Taken together, results from this study suggest that Rho/
Rho kinase may not be a necessary or a central pathway for renal
fibrosis in the UUO model. The interplay between the Rho/
Rho kinase pathway and the Smad signaling pathway may be a key mechanism by which loss of ROCK1 does not prevent renal
fibrosis in the UUO model.