淫羊藿与女贞子配伍对自噬激活状态ASMCs的影响  

Effects of the Combination of Herba Epimedii Combined with Fructus Ligustri Lucidi on Autophagy Activation State of ASMCs

作　　者：王瀚 马紫童 龙玉婷 李玉曼 马再娜 于萍[1] 刘仁慧[1] WANG Han;MA Zitong;LONG Yuting(School of Traditional Chinese Medicine,Capital Medical University,Beijing 100069,China)

机构地区：[1]首都医科大学中医药学院、中医络病研究北京市重点实验室,1000699

出　　处：《医学研究杂志》2023年第12期36-40,70,共6页Journal of Medical Research

基　　金：国家自然科学基金资助项目(81873115);北京市自然科学基金资助项目(7202014)。

摘　　要：目的研究淫羊藿、女贞子配伍对自噬激活剂雷帕霉素(rapamycin,RAP)诱导的气道平滑肌细胞(airway smooth muscle cells,ASMCs)自噬、凋亡及增殖活性的调节作用。方法培养大鼠原代ASMCs,随机分为正常组、RAP组、淫羊藿组、女贞子组、配伍组。根据分组分别加入正常大鼠血清或含药大鼠血清;除正常组外,其余各组同时加入RAP诱导的ASMCs,48h后进行检测。电镜观察细胞超微结构;MTT法检测细胞活力;免疫细胞化学法检测增殖蛋白ki-67蛋白表达,评价细胞增殖活性;Annexin V-FITC/PI法检测细胞凋亡活性;Western blot法和细胞免疫荧光法检测LC3、Beclin-1、mTOR、p-mTOR、Bax、Bcl-2、caspase-3及p53蛋白表达。结果淫羊藿、女贞子配伍能显著抑制RAP诱导的ASMCs自噬,下调LC3-Ⅱ/Ⅰ比值(P<0.05)及Beclin-1蛋白表达(P<0.01);改善RAP诱导的细胞活力、增殖活性降低(P<0.01);抑制凋亡水平升高(P<0.01),且优于淫羊藿、女贞子单用组(P<0.05或P<0.01)。此外,配伍组能上调caspase-3蛋白表达(P<0.05),下调p53蛋白表达(P<0.01);但对mTOR及p-mTOR、Bax及Bcl-2蛋白表达无显著影响。结论淫羊藿、女贞子配伍能抑制RAP诱导的ASMCs的过度自噬/凋亡水平,恢复ASMCs的细胞活力及增殖活性,避免ASMCs过度损伤。Objective To study the regulatory effect of herba epimedii(HE)combined with fructus ligustri lucidi(FLL)on autophagy,apoptosis and proliferation activity of airway smooth muscle cells(ASMCs)induced by rapamycin(RAP).Methods Primary ASMCs of rats were cultured and randomly divided into five groups:normal group,RAP group,HE group,FLL group and compatible group.Normal rat serum or medicated rat serum was added separately according to the group.Except for normal group,all other groups were simultaneously added with RAP-induced ASMCs,which were examined after 48h.Transmission electron microscope was used to observe cell microstructure;MTT assay was used to detect cell viability;immunocytochemistry was used to detect protein expression of proliferation protein ki-67 to evaluate the proliferation activity of cells;Annexin V-FITC/PI assay was used to detect the apoptosis activity of cells;Western blot and cell immunofluorescence were used to detect the protein expressions of LC3,Beclin-1,mTOR,p-mTOR,Bax,Bcl-2,caspase-3 and p53.Results The compatibility of HE and FLL significantly inhibited RAP-induced autophagy in ASMCs,and down-regulated LC3-Ⅱ/Ⅰprotein ratio(P<0.05)and Beclin-1 protein expression(P<0.01),alleviated the reduction of cell viability and proliferation activity(all P<0.01),and depressed apoptosis activity(P<0.01).The above effects of compatibility of HE and FLL were better than those of HE and FLL alone(P<0.05,P<0.01).In addition,the compatible group could up-regulate the expression of caspase-3 protein(P<0.05)and down-regulate the expression of p53 protein(P<0.01);However,there was no significant effect on the mTOR,p-mTOR,Bax and Bcl-2 protein.Conclusion The compatibility of HE and FLL could suppress the excessive autophagy/apoptosis level of RAP-induced ASMCs,restore the cell viability and proliferation activity of ASMCs,and avoid excessive damage of ASMCs.

关 键 词：气道平滑肌细胞 淫羊藿/女贞子 自噬 凋亡 增殖 

分 类 号：R285.5[医药卫生—中药学]