RhoA/Rho激酶信号通路在高脂血症导致阴茎海绵体纤维化重构过程中的作用机制的研究

The relationship between hyperlipidemia and erectile dysfunction(ED) is complex but interconnected. Besides the endothelial dysfunction as a common mechanism, we proposed another hypothesis of “remodeling” based on our previous research to explain the process of ED, that is， hyperlipidemia can induce hypoxia in the penis corpus cavernosum, leading to smooth muscle cell(SMC) apoptosis and finally fibrosis. However, the mechanism of hypoxia induced by hyperlipidemia needs to be further investigated. For this reason, we hypothesis that the RhoA/Rho-kinase signaling pathway plays an important role in resulting chronic hypoxia in erectile tissues under hyperlipidemia situations, as this pathway can enhance the contraction of SMCs by increasing Ca2+ sensitivity. To demonstrate it, we use hyperlipidemia rabbits as models to detect the expression of Rho-kinase, hypoxia induced factor1-α and casepase3 in the penis corpus cavernosum SMCs by RT-PCR, IHC and Western Blot. Combining with other techniques such as laser confocal microscopy, myotonometer and patch clamp, we try to illustrate the relationships among the protein expression, SMC apoptosis and SMC tones. Moreover, we use Rho-kinase inhibitors to support our hypothesis and in vitro we try to study where that the RhoA/Rho-kinase signaling pathway plays roles on in the “remolding” process. Our research may uncover a new mechanism of ED induced by hyperlipidemia, clarify the important role of RhoA/Rho-kinase signaling pathway in the “remolding” process. Thus, it is expected to provide not only new ideas and new therapeutic targets for ED diagnosis and treatments, but also the basis for future explorations of pathogenesis in other ED types.

高脂血症可导致阴茎勃起功能障碍（ED）发生，具体机制未明。我们过去的研究中提出了“海绵体重构”假说：即高脂引起的海绵体内慢性低氧触发纤维化重构，最终发展为ED。然而高脂引起低氧的机制需进一步研究。本课题提出高脂激活RhoA/Rho激酶通路是造成海绵体平滑肌收缩亢进、海绵窦供血减少、慢性缺氧的重要原因。为此以高脂家兔为ED模型，运用蛋白组学、激光共聚焦、肌张力测定仪和膜片钳等技术观察高脂对阴茎海绵体内Rho激酶活性与平滑肌张力的影响，以及同低氧、海绵体重构程度之间的关系。同时通过抑制Rho激酶活性，从反向证实该信号通路在高脂导致ED发生进展中的重要作用。另外，体外实验进一步探索该信号通路在阴茎“海绵体重构”中的作用环节，证实高脂引起海绵体平滑肌收缩亢进继而引起慢性低氧的假设。本课题有望发现高脂导致ED的新机制，为ED的治疗提供新靶点和新思路，也为将来探索为其他类型ED的发病机制提供理论依据。

高脂血症可导致阴茎勃起功能障碍（ED）的发生，是 ED 的重要危险因子之一。前期研究运用高脂饮食结合髂内动脉内膜球囊剥脱术制作高脂血症家兔 ED 模型，发现可引起海绵体内低氧，平滑肌细胞凋亡增加，胶原沉积等纤维化病变，平滑肌舒张功能减退。然而进来临床和研究发现高脂血症也可引起非动脉性 ED，机制则尚不清楚。本项目应用高脂饮食喂养制成家兔高脂血症模型，运用海绵体平滑肌肌张力测定证实了平滑肌舒张功能降低，海绵体内压则测定确认了阴茎勃起能力下降。通过免疫组化和免疫印迹研究发现阴茎海绵体RhoA/Rho 激酶信号通路激活，ROCK-2蛋白表达上调，伴随海绵体平滑肌掉凋亡水平增加、细胞外基质沉积及平滑肌表型发生转换，暨由顺应性较好的收缩型转变为顺应性较差的合成型。抑制 RhoA 的治疗可改善上述海绵体病变及阴茎勃起功能。说明上述海绵体内的病变以及勃起功能的下降是由RhoA/Rho 激酶信号通路激活而引起的。此外，对高脂血症家兔的阴茎海绵体运用弹力超声剪切波技术（SWE）检测，发现测定值 SWQ 显著高于正常对照组。这一改变与阴茎海绵体平滑肌的病变相关，能有效反应海绵体组织的质地变化，有望在临床取代海绵体活检，成为一种全新的无创、简便、可重复的 ED 诊断工具。体外细胞实验证实在低氧条件下，体外细胞培养的阴茎海绵体平滑肌细胞发生自噬程度增强的变化以及细胞膜表面氯离子通道电流密度降低，细胞兴奋性增强，收缩能力增加。RhoA 过表达能增强上述变化，抑制 RhoA 则显著缓解上述病变。证实低氧通过RhoA/Rho 激酶信号通路引起上述细胞变化。以上发现为 ED 的治疗找到新靶点，具有一定的临床应用价值和社会意义。