Restoration of Cavernous Veno-Occlusive Function through Chronic Administration of a Jun-Amino Terminal Kinase Inhibitor and a LIM-Kinase 2 Inhibitor by Suppressing Cavernous Apoptosis and Fibrosis in a Rat Model of Cavernous Nerve Injury: A Comparison with a Phosphodiesterase Type 5 Inhibitor

Purpose: To determine if chronic administration of Jun-amino terminal kinase (JNK)-inhibitors and LIM-kinase 2 (LIMK2)-inhibitors from the immediate post-injury period in a rat model of cavernous-nerve-crush-injury could normalize cavernousveno-occlusive-function, and to compare it with phosphodiesterase type 5 (PDE5)-inhibitors. Materials and Methods: A total of 75 12-week-old male Sprague–Dawley-rats were randomized into five groups: sham-surgery(S), cavernous-nerve-crush-injury (I), cavernous-nerve-crush-injury treated with 10.0 mg/kg LIMK2-inhibitor (L) or 10.0 mg/kg JNK-inhibitor and 10.0 mg/kg LIMK2-inhibitor (J+L) or 20.0 mg/kg udenafil (P) for five-weeks. Five-weeks after surgery,dynamic-infusion-cavernosometry, histological-studies, caspase-3-activity-assay, and Western-blot were investigated. Results: Group-I had lower papaverine-response, higher maintenance-rate and higher drop-rate, compared to Group-S.Group-L, Group-J+L and Group-P showed improvement in the three dynamic-infusion-cavernosometry parameters. The papaverine-response and drop-rate in Group-J+L and Group-P recovered to sham-control level, but those in Group-L did not.Regarding apoptosis, Group-I had decreased content of α-smooth-muscle-actin, increased caspase-3 activity and increased cJun-phosphorylation. The cJun-phosphorylation improved only in Group-J+L. The α-smooth-muscle-actin content andcaspase-3-activity in Group-J+L and Group-P improved, but those in Group-L were not. Regarding fibrosis, Group-I had decreased smooth muscle (SM)/collagen-ratio, increased protein-expression of fibronectin, and increased Cofilin-phosphorylation.Cofilin-phosphorylation was normalized in Group-L and Group-J+L, but not in Group-P. SM/collagen-ratio and proteinexpression of fibronectin in Group-L, Group-J+L and Group-P improved. Conclusions: Our data indicate that chronic inhibition of JNK and LIMK2 can restore cavernous-veno-occlusive-function by suppressing cavernous-apoptosis and cavernous-fibrosis, comparable to the results by PDE5-inhibitors. Chronic inhibition of JNK and LIMK2 might be a potential mechanism-specific targeted therapy for cavernous-veno-occlusive-dysfunction induced by cavernous nerve-injury.

研究目的：明确在海绵体神经挤压伤大鼠模型中，于损伤即刻开始长期给予c-Jun氨基末端激酶（Jun-amino terminal kinase, JNK）抑制剂及LIM激酶2（LIM-kinase 2, LIMK2）抑制剂，是否可恢复海绵体静脉闭塞功能，并与5型磷酸二酯酶（phosphodiesterase type 5, PDE5）抑制剂进行对比。
材料与方法：将75只12周龄雄性Sprague-Dawley大鼠随机分为5组：假手术组（S组）、海绵体神经挤压伤组（I组）、10.0 mg/kg LIMK2抑制剂处理组（L组）、10.0 mg/kg JNK抑制剂联合10.0 mg/kg LIMK2抑制剂处理组（J+L组）以及20.0 mg/kg乌地那非（udenafil）处理组（P组），连续给药5周。术后5周，开展动态灌注海绵体测压、组织学研究、半胱天冬氨酸蛋白酶-3（caspase-3）活性测定及蛋白质印迹（Western blot）检测。
结果：与S组相比，I组的罂粟碱反应率更低、维持压率及压降率更高。L组、J+L组及P组的三项动态灌注海绵体测压参数均得到改善。J+L组与P组的罂粟碱反应率和压降率恢复至假手术对照组水平，但L组未达到该效果。在细胞凋亡层面，I组的α-平滑肌肌动蛋白（α-smooth-muscle-actin）含量降低、caspase-3活性升高及c-Jun磷酸化水平升高。仅J+L组的c-Jun磷酸化水平得到改善。J+L组与P组的α-平滑肌肌动蛋白含量及caspase-3活性得到恢复，但L组未出现明显改善。在纤维化层面，I组的平滑肌（SM）/胶原蛋白比值降低、纤连蛋白（fibronectin）的蛋白表达升高及丝切蛋白（Cofilin）磷酸化水平升高。L组与J+L组的丝切蛋白磷酸化水平恢复至正常水平，但P组未恢复。L组、J+L组及P组的平滑肌/胶原蛋白比值及纤连蛋白蛋白表达均得到改善。
结论：本研究数据表明，长期抑制JNK与LIMK2可通过抑制海绵体凋亡与海绵体纤维化恢复海绵体静脉闭塞功能，其效果与PDE5抑制剂相当。长期抑制JNK与LIMK2或可成为海绵体神经损伤诱导的海绵体静脉闭塞功能障碍的潜在机制特异性靶向治疗方案。