CD131在CEPO抑制高糖诱导的肾小管上皮细胞凋亡中的作用及机制

Erythropoietin and its derivatives have been confirmed to play non hematological tissue protective effects through anti apoptosis. Carbamylated erythropoietin, which has a long half-life, and has no hematopoietic activity, is considered to be one of the erythropoietin derivatives with better prospects for clinical application. Unlike its erythropoiesis effect, which need a homodimer of two erythropoietin receptors, erythropoietin and its derivatives need heterodimers consisted of erythropoietin receptor and CD131 to play non hematological tissue protective effects in the central nervous system and cardiovascular system. However, there is still controversy about the existence of the heterodimers in above tissues. In previous work, we found the expression of erythropoietin receptor in rat renal tubular cells, and our preliminary experiment showed carbamylated erythropoietin can inhibit rat renal tubular cells apoptosis induced by high glucose. So, we hypothesize the presence of the heterodimers consisted of erythropoietin receptor and CD131 in renal tubular cells, and carbamylated erythropoietin can inhibit renal tubular cells apoptosis induced by high glucose through the heterodimers in vitro and in vivo, thereby play a role in renal protection in diabetic rats. To demonstrate this hypothesis, we will observe the effect of carbamoyl erythropoietin on renal tubular cell apoptosis induced by high glucose in vitro and in vivo, and the possible role of CD131 in this process. Finally, this work will provide experimental basis for the exploration of new treatments for diabetic kidney disease.

EPO及其衍生物被证实可通过抗细胞凋亡等效应发挥非血液学的组织保护作用，CEPO半衰期较长，同时无造血活性，被认为是具有更好临床应用前景的EPO衍生物之一。与EPO需要通过EPOR-EPOR同二聚体介导发挥促红细胞生成作用不同，近期有研究发现EPO及其衍生物发挥组织保护作用可能需要通过EPOR-CD131异二聚体介导，但目前对这一受体系统的存在与否仍有争论。在前期的工作中，我们发现大鼠肾小管细胞表达EPOR，且预实验显示CEPO可抑制高糖诱导的大鼠肾小管细胞凋亡。我们推测肾小管细胞内存在EPOR-CD131异二聚体，CEPO可以通过EPOR-CD131介导，抑制糖尿病时高血糖状态导致的肾小管细胞凋亡，发挥肾脏保护作用。为验证这一假说，本项目拟通过细胞培养、动物实验研究CEPO对高糖诱导的肾小管细胞凋亡的影响及CD131在此过程中的可能作用，为探索新的糖尿病肾脏病治疗方案提供实验依据。

目前，随着我国居民生活水平的提高，日常饮食结构的改变，糖尿病的发病率日益增高。糖尿病肾脏病（Diabetic Kidney Disease，DKD）作为糖尿病的常见微血管并发症，其发病率亦日益增高，已成为导致终末期肾病的主要原因之一，给家庭和社会带来巨大的医疗负担。但DKD 的发病机制较复杂，目前尚无特效疗法，患者一旦进入临床DKD期，病情将不可逆发展，所以对DKD的早期防治具有重要意义。近年的研究发现肾小管间质病变在DKD的发生发展过程中发挥着关键作用，且肾小管间质病变早于肾小球病变出现，部分研究者甚至认为占据肾脏体积90%的肾小管间质病变的不断加重诱发了DKD时的肾小球病变。糖尿病时的高血糖状态、晚期糖基化终末产物、过量活性氧等因素均可导致肾小管上皮细胞凋亡，导致肾小管萎缩、变性，进而引起肾小管及间质纤维化。作为一种主要由肾脏成纤维样细胞合成分泌的糖蛋白激素，促红细胞生成素（Erythropoietin，EPO）的非血液学效应越来越受到关注。目前的研究认为，EPO及其衍生物发挥非血液学组织保护作用主要是通过抗细胞凋亡和抑制炎症反应实现。本课题主要研究氨甲酰EPO（Carbamylated erythropoietin，CEPO）是否可以抑制糖尿病时高血糖导致的肾小管上皮细胞凋亡，发挥肾脏保护作用。同时对CEPO发挥抗肾小管上皮细胞凋亡的机制进行探讨。我们首先通过RT-PCR、Westernblot等实验方法证实了大鼠肾小管上皮细胞表达EPOR、CD131，并通过免疫荧光共定位、免疫共沉淀、阻断肽封闭及siRNA干扰等实验方法证实大鼠肾小管上皮细胞存在EPOR-CD131异二聚体受体系统，而CEPO可通过EPOR-CD131异二聚体受体系统介导，调节bcl-2家族相关凋亡基因的表达，抑制高糖诱导的肾小管上皮细胞凋亡，发挥肾脏保护作用。本课题的研究结果提高了对EPO这一传统激素的认识，证实了其衍生物CEPO对DKD肾脏病变的保护作用，为DKD的防治提供了新的策略。