Atg5 通过激活cathepsins-caspase 信号通路参与缺氧缺糖诱导的小鼠胚胎成纤维细胞的凋亡
荣加国，倪勇，陈洁茹，张慧灵
基金项目：国家自然科学基金项目（30973510）及教育部留学回国人员基金项（K513400110）
作者简介：荣加国（1987- ），男，硕士研究生，研究方向：脑血管病药理学
通信联系人：张慧灵（1965- ），女，教授，博导，研究方向：脑血管病药理学.
（苏州大学药学院药理系，苏州，215123）
摘要：目的：在Atg5-/-小鼠胚胎成纤维细胞上，观察Atg5 是否通过激活cathepsins-caspase信号通路参与缺糖缺氧诱导的小鼠胚胎成纤维细胞的凋亡。方法：采用氧糖剥夺(oxygen-glucose deprivation, OGD)方法建立缺氧缺糖细胞模型。光镜法观察OGD 诱导的
MEFs 细胞形态学变化； LDH 法检测OGD 诱导的MEFs 乳酸脱氢酶(LDH) 漏出率；Westernblot 法检测cathepsins-caspase 信号通路相关蛋白的表达。结果：在OGD 诱导的MEFs 损伤模型中，与野生型小鼠胚胎成纤维细胞(WT MEFs) 相比，Atg5-/- MEFs 的细胞形态明显改善，细胞数目显著增加，LDH 漏出率下降。Active-cathepsin B、active-cathepsin L、tBid、active-caspase 3 的蛋白水平下调， 线粒体Cyt-c 上调和细胞浆Cyt-c 下调。结论：Atg5 参与OGD 诱导的MEFs 细胞凋亡, 其机制与抑制cathepsins-caspase 信号通路的激活有关。
关键词：Atg5; MEFs; OGD; 细胞凋亡; cathepsins; caspase 3
中图分类号：R966
Atg5 is involved in the mouse embryo fibroblasts apoptosis via activation of the cathepsins-caspase signaling pathway
RONG Jiaguo, NI Yong, CHEN Jieru, ZHANG Huiling
(Department of Pharmacology and Laboratory of Cerebrovascular Pharmacology, College of Pharmaceutical Science, Soochow University, Suzhou 215123)
Abstract: Objective: To investigate the role of Atg5 in OGD-induced apoptosis and its activation of cathepsins-caspase signaling pathway in Atg5-deficient mouse embryo fibroblasts (MEFs).Methods: MEFs were exposed to oxygen-glucose deprivation (OGD). The morphological changes of the MEFs induced by OGD were observed by light microscope, cell death was determined b y a LDH assay. The cathepsins-caspase signaling pathway related proteins were detected by western
blot analysis. Results: In the OGD-induced MEFs injury model, compared with wild type (WT) MEFs, Atg5-deficient significantly improved the morphology of MEFs, incresed the number of MEFs and decreased the LDH leakage. Western blot analysis showed that OGD treatment induced a decrease in active-cathepsin B and L, tBid, active-caspase 3 and cytoplastic Cyt-c, and an increase in mitochondrial Cyt-c. Conclusion: Atg5-deficient protects MEFs against OGD-induced apoptosis, and its mechanisms is associated with inhibition of cathepsins-caspase signaling pathway.
Key words: Atg5; MEFs; OGD; apoptosis; cathepsins; caspase 3