目的 研究从地黄Rehmannia glutinosa中分离得到的地黄苷D对皮质酮诱导的肾上腺嗜铬细胞瘤细胞（PC-12细胞）的保护作用及机制。方法 以500 μmol/L皮质酮处理PC-12细胞24 h建立损伤模型，同时给予氟西汀和地黄苷D进行干预，采用MTT法检测细胞存活率；采用流式细胞术检测细胞凋亡、活性氧（reactive oxygen species，ROS）以及线粒体膜电位水平；采用In-Cell Western法检测细胞内凋亡相关蛋白半胱氨酸天冬氨酸蛋白酶-3（cystein-asparate protease-3，Caspase-3）、剪切型Caspase-3（cleaved Caspase-3）、B淋巴细胞瘤2（B-cell lymphoma 2，Bcl-2）和Bcl-2相关X蛋白（Bcl-2 associated X protein，Bax）表达；采用高内涵细胞成像系统检测细胞内脑源性神经营养因子（brain-derived neurotrophic factor，BDNF）蛋白表达。结果 地黄苷D明显提高皮质酮诱导的PC-12细胞存活率（P＜0.05、0.01）；抑制细胞凋亡和细胞内ROS水平（P＜0.01）；升高线粒体膜电位（P＜0.01）；下调细胞Bax/Bcl-2蛋白表达（P＜0.05）；上调BDNF蛋白表达（P＜0.01）。酪氨酸激酶受体B（tyrosine kinase receptor B，Trk B）拮抗剂K252a能够拮抗地黄苷D对PC-12细胞凋亡的抑制作用。结论 地黄苷D可能是通过激活BDNF-Trk B通路，抑制细胞凋亡通路，从而保护皮质酮诱导的PC-12细胞损伤。

Objective To study the protective effect and mechanism of rehmangoside D isolated from Rehmannia glutinosa on PC-12 cells induced by corticosterone. Methods PC-12 cells were treated with 500 μmol/L corticosterone for 24 h to establish an injury model, fluoxetine and rehmangoside D were given for intervention at the same time. Cell viability was detected by MTT method; Intracellular apoptosis, reactive oxygen species (ROS) and mitochondrial membrane potential levels were determined by flow cytometry; In-Cell Western method was used to detect intracellular apoptosis-related protein cysteine-aspartate protease-3 (Caspase-3), cleaved Caspase-3, B-cell lymphoma 2 (Bcl-2) and Bcl-2 associated X protein (Bax) expressions; High-content cell imaging system was used to detect intracellular brain-derived neurotrophic factor (BDNF) protein expression. Results Rehmangoside D significantly increased the survival rate of PC-12 cells induced by corticosterone (P < 0.05, 0.01), inhibited cell apoptosis and intracellular ROS level (P < 0.01), increased mitochondrial membrane potential (P < 0.01); Expression of Bax/Bcl-2 protein was down-regulated (P < 0.05); BDNF protein expression was up-regulated (P < 0.01). Tyrosine kinase receptor B (Trk B) antagonist K252a could antagonize the inhibitory effect of rehmangoside D on apoptosis of PC-12 cells. Conclusion Rehmangoside D may protect PC-12 cells from corticosterone-induced injury by activating BDNF-Trk B pathway and inhibiting apoptosis pathway.

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国家重点研发计划项目(2017YFC1702800);河南省重大科技专项(171100310500);河南省高层次人才特殊支持"中原千人计划"项目(ZYQR201810080);河南省教育厅河南省科技攻关项目(212102311106);河南省中医药科学研究专项课题(20-21ZY2151);国家留学基金委访问学者项目(201908410093);河南中医药大学2018年度博士科研基金资助项目(BSJJ2018-04)