He Caigu, Liu Guang, Zhuang Shuting, Zhang Jialin, Chen Yangtao, Li Hetian, Huang Zhengping, Zheng Yanfang
Department of Histology and Embryology, Fujian University of Traditional Chinese Medicine, Fuzhou, Fujian 350122, People's Republic of China.
Department of Biochemistry, Fujian University of Traditional Chinese Medicine, Fuzhou, Fujian 350122, People's Republic of China.
Diabetes Metab Syndr Obes. 2020 Jun 18;13:2081-2092. doi: 10.2147/DMSO.S253494. eCollection 2020.
Yu Nu compound (YNJ) is a traditional Chinese medicine widely utilized to treat type 2 diabetes possibly through mediating autophagy. Abnormal podocyte autophagy and apoptosis could result in podocyte loss in diabetics nephropathy (DN). The mechanism of Yu Nu compound in DN is still unclear. Therefore, the study aims to investigate the effects of Yu Nu compound and analyze the potential mechanism.
Goto-Kakizaki (GK) rats were administered using YNJ with different doses once a day by gavage for 4 weeks. The renal cortex injury was observed by HE staining and electron microscope. Cell apoptosis of renal cortex was analyzed by TUNNEL staining. The mTOR, autophagy-related proteins and apoptosis-related proteins were detected by Western blot or real-time PCR in vivo and vitro. MPC5 cells were exposed to high glucose (HG, 30mM) for 12h to simulate podocyte injury in DN. MPC5 cells were treated by serum containing YNJ with different dosages. Cell activities and apoptosis were, respectively, detected through Cell Counting Kit-8 (CCK8) assay and flow cytometry.
The results showed that the medium dose of YNJ had better effects on decreasing blood glucose and improving renal injury in GK rats, followed by decreasing mTOR levels. The autophagy levels were enhanced in renal cortex, accompanied with the increase of cell apoptosis in vivo. Besides, the proteins regulating autophagy and apoptosis were significantly modulated by YNJ in GK rats. Then, we found that the decreasing endogenous mTOR could reverse the effects of YNJ on podocyte apoptosis and autophagy in vivo.
The study suggested that YNJ recovered normal autophagy and suppressed apoptosis through regulating mTOR. The maintenance of normal basal autophagic activity possibly based on the effect of YNJ on multiple target was essential for maintaining podocyte function.
玉女煎（YNJ）是一种广泛用于治疗2型糖尿病的中药，可能通过介导自噬发挥作用。足细胞自噬和凋亡异常可导致糖尿病肾病（DN）中足细胞丢失。玉女煎治疗DN的机制尚不清楚。因此，本研究旨在探讨玉女煎的作用并分析其潜在机制。
将不同剂量的玉女煎每日一次经口灌胃给予Goto-Kakizaki（GK）大鼠，持续4周。通过HE染色和电子显微镜观察肾皮质损伤。采用TUNNEL染色分析肾皮质细胞凋亡情况。通过蛋白质免疫印迹法或实时PCR在体内和体外检测mTOR、自噬相关蛋白和凋亡相关蛋白。将MPC5细胞暴露于高糖（HG，30mM）环境12小时以模拟DN中的足细胞损伤。用含不同剂量玉女煎的血清处理MPC5细胞。分别通过细胞计数试剂盒-8（CCK8）检测法和流式细胞术检测细胞活性和凋亡情况。
结果显示，中剂量的玉女煎对降低GK大鼠血糖和改善肾损伤效果更佳，其次是降低mTOR水平。肾皮质自噬水平增强，同时体内细胞凋亡增加。此外，玉女煎在GK大鼠中显著调节了自噬和凋亡相关蛋白。然后，我们发现降低内源性mTOR可逆转玉女煎对体内足细胞凋亡和自噬的影响。
本研究表明，玉女煎通过调节mTOR恢复正常自噬并抑制凋亡。基于玉女煎对多个靶点的作用维持正常基础自噬活性对于维持足细胞功能至关重要。
