摘要
目的探索3-甲基腺嘌呤(3-MA)对大鼠骨髓源性内皮祖细胞(EPCs)超微结构和自噬体膜型(LC3-Ⅱ)蛋白表达的影响。方法采用密度梯度法取大鼠骨髓EPCs,体外诱导分化并鉴定;分成对照组和4个3-MA浓度组(分别加入1.25、2.5、5、10mmol/L 3-MA)。采用单丹(磺)酰戊二胺(MDC)荧光染色和透射电镜观察3-MA给药后EPCs自噬的发生。Western blot检测3-MA给药后EPCs内LC3-Ⅱ蛋白表达的变化。结果 MDC荧光染色和透射电镜均观察到5、10 mmol/L组EPCs超微结构明显区别于正常及死亡细胞,是典型的凋亡形态图。Western blot检测3-MA给药24h后EPCs LC3-Ⅱ蛋白表达降低;与对照组比较,5、10 mmol/L组LC3-Ⅱ蛋白表达明显降低(P<0.05)。结论 5、10mmol/L的3-MA对EPCs超微结构和LC3-Ⅱ蛋白表达均有影响,推测浓度≥5mmol/L的3-MA能抑制EPCs的自噬。
Objective To investigate the effect of 3-methlyadenine（3-MA ） on ultrastructure and LC3-Ⅱ protein expression in rat bone marrow-derived endothelial progenitor cells （EPCs）. Methods Mononuclear cells （MNCs） were isolated from rat bone marrow; the EPCs were induced from cultured MNCs and identified.The induced rat EPCs were treated with different concentrations of 3-MA （0, 1.25, 2.5, 5, 10mmol/L）. Monodansylcadaverine staining and transmission electron microscopy （TEM） were used to examine cell autophagy induced by 3-MA in rat EPCs.Western blot analysis was used to detect the LC3-Ⅱ protein expression. Results Autophagy inhibition was observed in EPCs by 3-MA at concentration of 5mmol/L and 10mmol/L by both MDC staining and TEM. LC3-Ⅱ protein expression in EPCs was decreased aftertreatment of 5mmol/L and 10mmol/L 3-MA as detected by western blot （P〈0.05）. Conclusion High dose of 3-MA caninduce the ultrastructural changes and reduce LC3-Ⅱ expression in rat bone marrow-derived epithelial progenitor cells.
出处
《浙江医学》
CAS
2016年第8期531-533,537,共4页
Zhejiang Medical Journal
基金
国家自然科学基金资助项目(30972941)
