基于自噬与凋亡机制研究左归丸对化疗损伤颗粒细胞及膜细胞的影响  

The Effect of Zuogui Pill on Autophagy and Apoptosis in Chemotherapy-damaged Granular Cells and Theca Cells

作　　者：刘涛 孙晓峰[2] 曾贵荣 黄欣怡 曾文卓 龚力民 阳松威[5] 刘梦琳 LIU Tao;SUN Xiao-Feng;ZENG Gui-Rong;HUANG Xin-Yi;ZENGWen-Zhuo;GONG Li-Min;YANG Song-Wei;LIU Meng-Lin(School of Medicine,Hunan University of Chinese Medicine,Changsha 410208,China;Department of Histology and Embryology,Hunan University of Chinese Medicine,Changsha 410208,China;Hunan Center of Drug Safety Evaluation and Research of Drugs&Hunan Key Laboratory of Pharmacodynamics and Safety Evaluation of New Drugs,Changsha 410331,China;Department of Cardiovascular Medicine,The Third Xiangya Hospital,Central South University,Changsha 410006,China;School of Pharmacy,Hunan University of Chinese Medicine,Changsha 410208,China)

机构地区：[1]湖南中医药大学医学院,长沙410208 [2]湖南中医药大学医学院组织学与胚胎学教研室,长沙410208 [3]湖南省药物安全评价研究中心&新药药效与安全性评价湖南省重点实验室,长沙410331 [4]中南大学湘雅三医院心内科,长沙410006 [5]湖南中医药大学药学院,长沙410208

出　　处：《生物化学与生物物理进展》2022年第9期1741-1750,共10页Progress In Biochemistry and Biophysics

基　　金：supported by grants from The National Natural Science Foundation of China (81774362, 81303004);The Natural Science Foundation of Hunan Province (2018JJ2290)

摘　　要：目的探讨左归丸含药血清对化疗损伤性颗粒细胞和膜细胞的影响及作用机制。方法制备左归丸含药血清,培养大鼠卵巢颗粒细胞和膜细胞,使用磷酰胺氮芥造模分组后给药。CCK-8法测定颗粒细胞和膜细胞存活率,实时荧光定量PCR法(RT-PCR)及蛋白质免疫印迹法(Western blot)分别检测卵巢自噬启动因子Beclin-1、微管结合蛋白轻链3(LC3B)、自噬受体蛋白p62、凋亡蛋白Bax、Caspase3在转录水平和翻译水平上的表达。结果10%左归丸含药血清对于细胞存活的挽救率最高。Beclin-1、LC3B、Bax、Caspase3在磷酰胺氮芥作用的颗粒细胞和膜细胞中,相对于空白对照组有高表达(P<0.05),10%左归丸含药血清可下调上述蛋白质在模型组中的表达(P<0.05);然而受体蛋白p62较空白对照组升高(P<0.05),10%左归丸含药血清可上调模型组p62的表达(P<0.05)。此外,在颗粒细胞实验组中,激活或抑制自噬途径后,自噬相关蛋白的表达在发生相应改变的同时,凋亡相关蛋白的表达也会发生相应改变。结论磷酰胺氮芥可通过促进凋亡、激活自噬/溶酶体降解途径的机制损伤颗粒细胞和膜细胞。10%左归丸含药血清能缓解由此带来的损伤,同时影响了颗粒细胞和膜细胞自噬和凋亡过程。在磷酰胺氮芥损伤颗粒细胞的过程和10%左归丸含药血清缓解其损伤过程中均存在自噬与凋亡串流(cross-talk)。Objective To investigate the effect and mechanism of Zuogui pill-containing serum(ZGP-containing serum)on chemotherapy-damaged granular cells(GCs)and theca cells(TCs).Methods GCs and TCs were cultured respectively,the model group was established with phosphoramide mustard(PM),then treated by ZGP-containing serum.The survival rates of GCs and TCs was determined by CCK-8.Real-time fluorescent quantitative PCR(RT-PCR)and Western blot methods were used to detect the expression of Beclin-1,light chain 3(LC3B),p62,Bax and Caspase3.Results 10%ZGP-containing serum had the best effect on the recovery of cell survival rate.Compared with the blank control group,Beclin-1,LC3B,Bax and Caspase3 have higher expression in the model group(P<0.05),and 10%ZGP-containing serum can down-regulate the expression of them(P<0.05).Moreover,the expression of p62 is lower in the model group than the blank control group(P<0.05),and 10%ZGP-containing serum can upregulate it(P<0.05).In addition,in the groups of GCs,after activating or inhibiting the autophagy pathway,the expression of autophagy-related proteins and apoptosis-related proteins both changed correspondingly.Conclusion PM can damage GCs and TCs by promoting apoptosis and activating autophagy/lysosomal degradation pathways,10%ZGP-containing serum can alleviate the damage.There is a cross-talk between autophagy and apoptosis in the process of PM damaging GCs which can be alleviated by 10%ZGP containing serum.

关 键 词：左归丸 化疗 颗粒细胞 膜细胞 自噬 凋亡 

分 类 号：R285.5[医药卫生—中药学] R-332[医药卫生—中医学]