PLCε通过GLS/p-mTOR途径促进膀胱癌细胞T24生存  

PLCε Can Promote Bladder Cancer Cell T24 Survival via GLS/p-mTOR Pathway

作　　者：袁鸿玲 范佳鑫 杨锦潇 李婷[1] 何镇廷 吴小候[2] 陈琪 欧俐苹[1] 罗春丽[1] YUAN Hongling;FAN Jiaxin;YANG Jinxiao;LI Ting;HE Zhenting;WU Xiaohou;CHEN Qi;OU Liping;LUO Chunli(Key Laboratory of Diagnostics Medicine of Ministry of Education,Chongqing Medical University,Chongqing 400016,China;Department of Urinary Surgery,First Affiliated Hospital of Chongqing Medical University,Chongqing 400016,China;Jiajia Central Health Center,Chengdu 610000,China)

机构地区：[1]重庆医科大学检验医学院,临床检验诊断学教育部重点实验室,重庆400016 [2]重庆医科大学附属第一医院泌尿外科,重庆400016 [3]贾家中心卫生院,成都610000

出　　处：《中国细胞生物学学报》2020年第9期1588-1597,共10页Chinese Journal of Cell Biology

基　　金：国家自然科学基金(批准号:81072086)资助的课题。

摘　　要：谷氨酰胺对于细胞的代谢和生长十分重要,也是血液中含量最丰富的氨基酸,且肿瘤的代谢特征之一就是谷氨酰胺成瘾。该研究探讨磷脂酰肌醇特异性磷脂酶PLC epsilon(phospholipase C epsilon,PLCε)是否通过谷氨酰胺酶(glutaminase,GLS),调节膀胱癌细胞T24自噬,促进膀胱癌细胞生存。首先通过数据库Su Multi-cancer Statistics、Sanchez-Carbayo Bladder 2和细胞实验分析PLCε在膀胱癌中表达情况。结果表明,PLCε在膀胱癌中高表达。并通过LV-shPLCε转染膀胱癌细胞T24后,q-PCR和Western blot检测PLCε在膀胱癌细胞T24中的表达情况以及对凋亡和自噬的影响,同时免疫荧光检测细胞内自噬斑点(LC3)的变化。结果显示,敲低PLCε后,Caspase-3/Caspase-8/LC3-Ⅱ表达增加,p62表达降低;流式细胞术结果显示凋亡率增高;免疫荧光发现自噬斑点LC3均增多;GLS和p-mTOR的表达受到抑制。在shPLCε组中添加过表达GLS质粒后,p-mTOR和p62表达增加,LC3-Ⅱ表达降低并且免疫荧光自噬斑点LC3减少;加入敲低GLS质粒后出现相反结果。该研究得出,PLCε通过GLS/p-mTOR抑制膀胱癌细胞T24自噬,促进膀胱癌细胞T24的生存。Glutamine which is the most abundant amino acid in the blood,plays a particularly important role in cell growth and metabolism.One of the characteristics of tumor metabolism is glutamine addiction.This study investigated whether PLCεcan regulate autophagy and promote the survival of bladder cancer cells T24 targeted by GLS(glutaminase).Firstly,PLCεexpression in bladder cancer was analyzed using Su Multi-cancer Statistics,Sanchez-Carbayo Bladder 2 database and cell culture experiment.Results showed that PLCεhighly expressed in bladder cancer.Then,shPLCεcell lines were established by transfecting LV-shPLCεinto bladder cancer cell T24.The expression of PLCεin bladder cancer cell T24 and its effects on apoptosis and autophagy were detected by q-PCR and Western blot.Meanwhile,the changes of autophagy spot(LC3)were detected by immunofluorescence.The results illustrated that PLCεknockdown increased the expression of Caspase-3/Caspase-8/LC3-Ⅱ,apoptosis cell ratio and decreased p62;meanwhile increased the number of green autophagy spots LC3 in the cytoplasm by immunofluorescence and inhibited the expression of GLS and p-mTOR.Adding GLS over-expression plasmid into shPLCεgroup decreased the expression of LC3-Ⅱand the number of LC3 autophagy spots,and improved p-mTOR/p62 expression.However the results were reversed after adding GLS knockdown plasmids.This study showed that PLCεcould inhibit autophagy in bladder cancer cell T24 via GLS/p-mTOR,thus promoting the survival of tumor cells.

关 键 词：膀胱癌 谷氨酰胺代谢 PLCε GLS 自噬 

分 类 号：R737.14[医药卫生—肿瘤]