Involvement of Sphingosine-1-Phosphate/RhoA/Rho-Kinase Signaling Pathway in Corporal Fibrosis Following Cavernous Nerve Injury in Male Rats

INTRODUCTION: Postprostatectomy erectile dysfunction (ED) is thought to be due primarily to injury to cavernous nerve (CN) during surgery. The molecular mechanisms leading to ED after CN injury are poorly understood. AIM: We determined whether transforming growth factor-β(1) (TGF-β(1)), sphingosine-1-phosphate (S1P) and RhoA/Rho-kinase (ROCK) signaling pathways were involved in corporal fibrosis after bilateral CN injury in rats. METHODS: Forty-eight 10-week-old male Sprague-Dawley rats were equally divided into the following four groups: normal control group (C); sham surgery group (S); bilateral CN crush injury group (I); and bilateral CN transection group (T). Within each of the four groups, two subgroups were analyzed as a function of time (1 and 8 weeks postoperatively). MAIN OUTCOME MEASURES: Penile tissue was processed for immunoblot (RhoA, ROCK1, phospho-myosin phosphatase target subunit [MYPT1]), reverse transcription-polymerase chain reaction (RT-PCR) (TGF-β(1), sphingosine kinase type 1 [SphK1], and S1P(2)), immunohistochemistry (alpha smooth muscle actin [α-SMA]), and Masson's trichrome staining. RESULTS: At 1 and 8 weeks postoperatively, the I and T groups had a significantly decreased smooth muscle cell/collagen ratio, the expression of α-SMA and phospho-MYPT1 compared to the C group. Densitometry revealed a significantly higher expression of RhoA and ROCK1 in the T group compared to the C group at 1 and 8 weeks postoperatively. For the I group, the expression of RhoA significantly increased starting from 1 week postoperatively, but the expression of ROCK1 significantly increased as late as 8 weeks following injury. The expression of TGF-β(1) and S1P(2) mRNA in the I or T group remained significantly increased up to 8 weeks compared to the C group, despite significant reduction at 8 weeks compared to 1 week postoperatively. The expression of SphK1 mRNA in the I and T groups was significantly increased at 1 week but not 8 weeks postoperatively. CONCLUSIONS: Our data suggest that S1P and RhoA/ROCK1 signaling may be involved in corporal fibrosis associated with loss of smooth muscle through coordination with TGF-β(1) after CN injury.