BKca下调RhoA/ROCK信号通路改善糖尿病大鼠阴茎勃起功能

Improve of erectile function in diabetic rats through regulation of RhoA/ROCK signaling pathway by Bkca

摘要:

目的：观察大电导钙激活钾通道（big conductance Ca2+-activated K+ channel，BKca）对糖尿病勃起功能障碍（diabetes mellitus-induced erectile dysfunction，DMED）大鼠阴茎海绵体平滑肌RhoA/ROCK信号通路的影响。方法：实验大鼠分为空白对照组8只，DMED组8只，NS1619组（DMED治疗组）7只，NS1619组大鼠采用特异性BKca激活剂（NS1619）阴茎海绵体注入2周。观察各组勃起行为，并电刺激检测阴茎海绵体内压（intracavernous pressure，ICP）/平均动脉压（mean arterial pressure，MAP），取各组阴茎海绵体平滑肌检测肌张力，采用Western blot和定量PCR（real-time PCR）方法检测RhoA、ROCK1、ROCK2表达，反应RhoA/ROCK信号通路差异，同时检测MYPT-1表达反应肌球蛋白轻链磷酸酶（myosin light chain phosphatase，MLCP）磷酸化程度。结果：成功构建DMED大鼠模型，NS1619组大鼠与DMED组大鼠比较，勃起次数和ICP/MAP明显改善（P=0.025、0.024），阴茎海绵体平滑肌舒缩顺应性提高（P=0.031、0.024）并且RhoA、ROCK2以及肌球蛋白磷酸酶靶向结合亚基（myosin phosphatase target subunit，MYPT）-1蛋白和mRNA表达水平下调（P=0.029、0.003、0.002），但仍未达到正常对照组大鼠水平（P=0.018、0.040、0.003），ROCK1表达无明显差异（P=0.533）。结论：DMED大鼠因RhoA/ROCK信号通路上调和MLCP磷酸化增强导致阴茎海绵体平滑肌舒缩顺应性降低以致勃起功能障碍发生，激活BKca可以通过下调RhoA/ROCK信号通路和抑制MLCP磷酸化，改善勃起功能。

Abstract:

Objective：To investigate the effect of big conductance Ca2+-activated K+ channel（BKca）on RhoA/ROCK signaling pathway of diabetes mellitus-induced erectile dysfunction（DMED） rats. Methods：Rats were divided into control group（n=8），DMED group（n=8） and NS1619 group（treatment group，n=7）. The NS1619 group was treated with BKca specific opener（NS1619） for two weeks. Rats underwent cavernous nerve stimulation to determine the value of intracavernous pressure（ICP）/mean arterial pressure（MAP） and the number of erection was recorded. Corpus cavernosum smooth muscle was isolated for detection of contractile force. Western blot and real-time PCR were used to detect the expression of RhoA，ROCK1，ROCK2，which determined the differences of RhoA/ROCK signaling pathway and phosphorylation of myosin light chain phosphatase（MLCP）. Results：DMED rat models were successfully estab－lished. While erectile function and compliance of smooth muscle was decreased in rats with DMED，administration of NS1619 im－proved erectile responses（P=0.025，0.024） and compliance（P=0.031，0.024）. Treatment significantly decreased the expression of RhoA，ROCK2 and MYPT-1（P=0.029，0.003，0.002），but not ROCK1（P=0.533）. Compared with those in control group the NS1619 group have a higher expression levels of those protein（P=0.018，0.040，0.003）. Conclusion：Up-regulation of the RhoA/ROCK sig－naling pathway is harmful to erectile function. Activation of BKca can improve erectile function by down-regulation the level of RhoA/ROCK and phosphorylation of MLCP.