ACSS2通过PI3K/AKT信号通路调控食管鳞癌细胞的顺铂敏感性  

ACSS2 mediates the chemosensitivity of human esophageal squamous cell carcinoma via PI3K/AKT pathway

作　　者：高星宇 王景芝 凌锐[1] 周月鹏[2] 毛朝明[3] 陈德玉[1] GAO Xing-yu;WANG Jing-zhi;LING Rui;ZHOU Yue-peng;MAO Chao-ming;CHEN De-yu(Clinical Center of Tumor Therapy,Affiliated Hospital of Jiangsu University,Zhenjiang Jiangsu 212001,China;Oncological Laboratory,Affiliated Hospital of Jiangsu University,Zhenjiang Jiangsu 212001,China;Department of Nuclear Medicine, Affiliated Hospital of Jiangsu University, Zhenjiang Jiangsu 212001,China)

机构地区：[1]江苏大学附属医院肿瘤治疗中心,江苏镇江212001 [2]江苏大学附属医院肿瘤学实验室,江苏镇江212001 [3]江苏大学附属医院核医学科,江苏镇江212001

出　　处：《江苏大学学报（医学版）》2020年第6期486-492,共7页Journal of Jiangsu University：Medicine Edition

基　　金：国家自然科学基金面上项目(81572956);江苏省社会发展基金资助项目(BE2017696);江苏省医学创新团队(CXTDC2016009)。

摘　　要：目的:探讨乙酰辅酶A合成酶2(acetyl-CoA synthetase 2,ACSS2)表达对食管鳞癌细胞顺铂(cisplatin,DDP)敏感性的影响和可能机制。方法:通过免疫组化和蛋白质印迹法检测40对食管鳞癌瘤体和癌旁组织中ACSS2蛋白的表达。蛋白质印迹法验证食管鳞癌细胞株TE-1、ECA-109和KY-SE150与正常食管鳞状细胞Het-1A中ACSS2的表达差异;利用脂质体转染siRNA-ACSS2或者慢病毒转染LV-ACSS2至TE-1细胞构建ACSS2下调或过表达细胞株;CCK8实验检测下调ACSS2对顺铂IC50值的改变;顺铂处理(5μg/mL)前后,流式细胞术检测ACSS2干扰处理对TE-1细胞凋亡水平的影响;蛋白质印迹检测PI3K/AKT信号通路及凋亡相关蛋白cleaved-Caspase-3的表达。结果:免疫组化及蛋白质印迹结果显示,食管鳞癌组织中ACSS2蛋白表达显著高于癌旁组织(P<0.001),食管鳞癌细胞株中ACSS2表达水平明显高于正常鳞状上皮细胞Het-1A。CCK8结果显示siRNA-ACSS2处理显著下调TE-1细胞对顺铂的IC50值(P值均<0.05);流式细胞术结果证实,抑制ACSS2表达可显著上调食管鳞癌细胞凋亡率(P<0.01);结合5μg/mL顺铂处理,ACSS2干扰后TE-1细胞凋亡率明显上升(P值均<0.001);蛋白质印迹结果表明,顺铂处理上调TE-1细胞中ACSS2及p-PI3K、p-AKT的表达;顺铂作用下,靶向抑制ACSS2后可见p-PI3K、p-AKT的显著降低而凋亡相关蛋白cleaved-Caspase-3表达量明显升高,过表达ACSS2在维持PI3K/AKT信号活化的同时可有效逆转cleaved-Caspase-3的形成。结论:ACSS2通过调控PI3K/AKT信号通路活化以及cleaved-Caspase-3表达参与食管鳞癌细胞顺铂敏感性调节。Objective:To explore the effect of the expression change of acetyl-CoA synthetase 2(ACSS2)on the sensitivity of esophageal squamous cell carcinoma(ESCC)to cisplatin(DDP)and relevant mechanism.Methods:ACSS2 expression was detected in tumor and paracancerous tissues collected from 40 cases of ESCC by immunohistochemistry and western blotting.In vitro verification of ACSS2 expression difference was performed by Western blotting in ESCC cell line and normal esophageal squamous Het-1A cell line.ACSS2 downregulated and overexpressed cell strains were constructed by liposome transfection of siRNA-ACSS2 or lentivirus transfection of LV-ACSS2 into TE-1 cells.CCK8 assay was used to detect the change of IC50 value of DDP by downregulating the expression of ACSS2.Meanwhile,the effect of ACSS2 interference on TE-1 cell apoptosis was tested by flow cytometry before and after DDP treatment(5μg/mL).In addition,Western blotting was carried out to detect the expression of proteins related to PI3K/AKT signaling pathway and apoptosis-related protein cleaved-Caspase-3.Results:Immunohistochemistry and Western blotting results showed that the expression intensity of ACSS2 in ESCC tissue was significantly higher than that in paracancerous normal tissue(P<0.001).In vitro test indicated that the expression of ACSS2 in ESCC cell line was obviously higher than that of normal esophageal squamous Het-1A cell line.CCK8 results showed that IC50 of TE-1 cells was remarkedly downregulated after siRNA-ACSS2 treatment(all P<0.05).Flow cytometry results showed that the apoptosis rate of ESCC cells was obviously decreased by inhibiting ACSS2 expression(P<0.01),which,however,increased significantly under the combined treatment of 5μg/mL DDP and ACSS2 interference(all P<0.001).Furthermore,according to the results of Western blotting,expression levels of ACSS2,p-PI3K and p-AKT were upregulated in TE-1 cells after DDP treatment.Simultaneously,under DDP treatment,targeted inhibition of ACSS2 expression resulted in significant decrease of p-PI3K and p-AK

关 键 词：乙酰辅酶A合成酶2 顺铂 食管鳞癌 化疗敏感性 PI3K/AKT 

分 类 号：R735.1[医药卫生—肿瘤]