自噬诱导、抑制的人三阴性乳腺癌细胞系MDA-MB-231中ATG2A和ATG9A表达观察  

Expression of ATG2A and ATG9A gene in human triple negative breast cancer cell line MDA-MB-231 of starvation induced autophagy and autophagy inhibition

作　　者：杨鑫娇 邹迪 熊喆 张海珠 夏从龙 赵红业 YANG Xinjiao;ZOU Di;XIONG Zhe;ZHANG Haizhu;XIA Conglong;ZHAO Hongye(College of Pharmacy,Dali University,Dali 671000,China;不详)

机构地区：[1]大理大学药学院,云南大理671000 [2]云南省动物基因编辑与体细胞克隆技术重点实验室 [3]云南省异种器官移植工程研究中心

出　　处：《山东医药》2021年第21期13-17,共5页Shandong Medical Journal

基　　金：国家自然科学基金面上项目(81872452)。

摘　　要：目的观察自噬诱导、抑制的人三阴性乳腺癌细胞系MDA-MB-231中ATG2A、ATG9A表达变化。方法人三阴性乳腺癌细胞系MDA-MB-231分为4组,对照组加入DMEM培养液正常培养,Baf-A1组在DMEM培养液中加100 nmol/L的巴弗洛霉素A1(Baf-A1)进行自噬阻断,EBSS组加入Earle's平衡盐溶液(EBSS)进行饥饿诱导细胞自噬,EBSS+Baf-A1组在EBSS培养液中加入100 nmol/L的Baf-A1,四组均培养4 h。采用实时荧光定量(Q-PCR)法检测各组细胞中ATG2A、ATG9A mRNA,采用WesternBlotting法检测各组细胞中ATG2A、ATG9A、P62、微管蛋白1轻链3(LC3)蛋白,计算LC3B-Ⅱ/LC3B-Ⅰ。结果与对照组、Baf-A1组相比,EBSS组、EBSS+Baf-A1组细胞中ATG2A、ATG9A mRNA相对表达量均显著升高(P均<0.05),且EBSS+Baf-A1组细胞中ATG9A mRNA相对表达量高于EBSS组(P均<0.05)。与对照组、Baf-A1组相比,EBSS组、EBSS+Baf-A1组细胞中ATG2A蛋白相对表达量均显著升高(P均<0.05);EBSS组细胞中ATG9A蛋白相对表达量均高于其余三组(P均<0.05);EBSS组细胞中P62蛋白相对表达量均低于其余三组(P均<0.05),EBSS+Baf-A1组P62蛋白相对表达量低于对照组(P均<0.05);EBSS组细胞中LC3B-Ⅱ/LC3B-Ⅰ均低于其余三组(P均<0.05),EBSS+Baf-A1组LC3B-Ⅱ/LC3B-Ⅰ均高于其余三组(P均<0.05)。结论EBSS成功诱导MDA-MB-231细胞发生自噬,饥饿诱导的MDA-MB-231细胞自噬过程中ATG2A、ATG9A mRNA和蛋白表达升高。Objective To investigate the expression changes of ATG2A gene and ATG9A gene in autophagy in-duced and inhibited human triple negative breast cancer cell line MDA-MB-231 of starvation induced autophagy and au-tophagy inhibition.Methods The inhibited human triple negative breast cancer cell line MDA-MB-231 cells were divid-ed into 4 groups.Cells in the control group was cultured in DMEM medium.Cells in the Baf-A1 group was treated with 100 nmol/L bafilomycin A1(Baf-A1)in DMEM culture medium for autophagy blocking.Cells in EBSS group were added with Earle's equilibrium salt solution(EBSS)to induce autophagy,and cells in the EBSS+Baf-A1 group were added 100 nmol/L Baf-A1 to EBSS medium.The above four groups were cultured for 4 h.The ATG2A and ATG9A of each group were detected by real-time fluorescent quantitative(Q-PCR),and the levels of ATG2A,ATG9A,P62 protein and tubulin 1 light chain 3(LC3)protein were detected by Western blotting.The gray values of LC3B-Ⅱand LC3B-Ⅰwere analyzed by image lab software,and the ratio of LC3B-Ⅱ/LC3B-Ⅰwas calculated.Results Compared with the control group and Baf-A1 group,the relative expression of ATG2A and ATG9A in cells of EBSS group and EBSS+Baf-A1 group were significantly increased(all P<0.05),and the relative expression of ATG9A in cells of EBSS+Baf-A1 group was higher than that in EBSS group(all P<0.05).Compared with the control group and Baf-A1 group,the relative expression of ATG2A in cells of EBSS group and EBSS+Baf-A1 group were significantly increased(all P<0.05);the relative expres-sion of ATG9A in cells of EBSS group was higher than that in the other three groups(all P<0.05);the relative expression of P62 protein in cells of EBSS group was lower than that in the other three groups(all P<0.05).The relative expression of P62 protein in cells of EBSS+Baf-A1 group was lower than that in the control group(all P<0.05);LC3B-Ⅱ/LC3B-Ⅰin cells of EBSS group were lower than those in other three groups(all P<0.05),and LC3B-Ⅱ/LC3B-Ⅰin cells of EBSS+Baf-A1 group was hi

关 键 词：乳腺肿瘤 三阴性乳腺癌 ATG2A基因 ATG9A基因 细胞自噬 MDA-MB-231细胞 

分 类 号：R966[医药卫生—药理学]