天冬酰胺合成酶对移植黑色素瘤APP/PS1小鼠骨骼肌减少的影响  

Effect of asparagine synthetase on reduction of skeletal muscle in APP/PS1 mice transplanted with melanoma

作　　者：董营[1] 吕航 郭家女 郑鸿 郭丹 王力可 王丹[1] 陈昌捷[2] 孙连坤[3] 张勇[3] 于春艳[1] DONG Ying;LYU Hang;GUO Jianv;ZHENG Hong;GUO Dan;WANG Like;WANG Dan;CHEN Changjie;SUN Liankun;ZHANG Yong;YU Chunyan(Department of Pathology,College of Basic Medical Sciences,Beihua University,Jilin 132013,China;Department of Laboratory,Affiliated Hospital,Beihua University,Jilin 132013,China;Department of Pathophysiology,School of Basic Medical Sciences,Jilin University,Changchun 130021,China)

机构地区：[1]北华大学基础医学院病理教研室,吉林吉林132013 [2]北华大学附属医院检验科,吉林吉林132013 [3]吉林大学基础医学院病理生理学系,吉林长春130021

出　　处：《吉林大学学报（医学版）》2021年第4期849-856,共8页Journal of Jilin University：Medicine Edition

基　　金：国家自然科学基金项目(81541148);吉林省科技厅科技发展计划项目(YDZJ202101ZYTS090,20180623018TC);吉林省教育厅“十三五”科学技术项目(JJKH20200057KJ);吉林省卫健委卫生与健康技术创新项目(2019J040);北华大学省级大学生创新创业训练计划项目(202010201073);吉林省国家级大学生创新创业训练计划项目(202010201002);吉林省吉林市科技局医疗卫生指导性计划项目(20200404101)。

摘　　要：目的:探讨天冬酰胺合成酶(ASNS)在移植黑色素瘤模型淀粉样蛋白前体蛋白/早老素1(APP/PS1)小鼠骨骼肌减少中的作用,并阐明其作用机制。方法:C57BL/6J(C57)小鼠和APP/PS1小鼠分别移植黑色素瘤B16细胞(C57移植瘤组和APP/PS1移植瘤组),同时设C57对照组和APP/PS1对照组,每组7只。取小鼠腓肠肌组织,分析天平称量小鼠腓肠肌质量,计算小鼠腓肠肌质量与体质量比值即肌肉减少症指数(SI),采用HE染色观察小鼠腓肠肌组织形态表现,实时荧光定量PCR(RT-qPCR)法检测小鼠腓肠肌组织中肌肉组织特异萎缩相关基因E3泛素连接酶1(Atrogin-1)、肌肉环指蛋白1(MuRF1)和ASNS mRNA表达水平,Western blotting法检测小鼠腓肠肌组织中ASNS及内质网应激途径相关蛋白葡萄糖调节蛋白78(GRP78)、活化转录因子4(ATF4)、真核细胞起始因子2α(eIF2α)和磷酸化eIF2α(p-eIF2α)蛋白表达水平。结果:与C57对照组比较,C57移植瘤组小鼠腓肠肌SI降低(P<0.05),腓肠肌细胞呈玻璃样变性,肌浆蓝染,部分细胞核可见核内移现象,小鼠腓肠肌组织中Atrogin-1、MuRF1和ASNS mRNA表达水平升高(P<0.05),ASNS蛋白表达水平升高(P<0.05),GRP78、ATF4蛋白表达水平和p-eIF2α/eIF2α蛋白表达水平比值明显升高(P<0.05);与APP/PS1对照组比较,APP/PS1移植瘤组小鼠腓肠肌SI降低(P<0.05),腓肠肌部分肌纤维肿胀,部分肌纤维变细,细胞核内移现象多见,肌浆颜色以红色为主,腓肠肌组织中Atrogin-1、MuRF1和ASNS mRNA表达水平升高(P<0.05),ASNS蛋白表达水平升高(P<0.05),GRP78、ATF4蛋白表达水平和p-eIF2α/eIF2α蛋白表达水平比值明显升高(P<0.05);与C57移植瘤组比较,APP/PS1移植瘤组小鼠腓肠肌SI降低(P<0.05),腓肠肌组织中Atrogin-1 mRNA表达水平升高(P<0.05),ASNS mRNA水平降低(P<0.05),ASNS蛋白表达水平差异无统计学意义(P>0.05),GRP78和ATF4蛋白表达水平及p-eIF2α/eIF2α蛋白表达水平比值明显降低(P<0.05)。结论:APP/PS1移植Objective:To investigate the role of asparagine synthetase(ASNS)on reduction of skeletal muscle in the amyloid precursor protein/presenilin 1(APP/PS1)mice transplanted with melanoma,and to clarify its mechanism.Methods:The C57BL/6J(C57)and APP/PS1 mice were transplarted with melanoma cells(C57 transplanted tumor group and APP/PS1 transplanted tumor group),at the same time,C57 control group and APP/PS1 control group were set up.The mass of gastrocnemius was measured and the sarcopenia index(SI)was calculated with the gastrocnemius mass to body weight ratio.The morphology of gastrocnemius tissue of the mice was determined by HE staining.The mRNA expression levels of muscle specific atrophy gene E3 ubiquitin protein ligase 1(Atrogin-1),muscle ring finger protein 1(MuRF1)and ASNS in gastrocnemius tissue of the mice were measured by Real-time fluorescence quantitative PCR(RT-qPCR).The expression levels of ASNS and endoplasmic stress related glucose regulated protein 78(GRP78),activating transcription factor-4(ATF4),eukaryotic cell initiation factor 2α(eIF2α),phosphorylated eIF2α(p-eIF2α)in gastrocnemius tissue of the mice were measured by Western blotting method.Results:Compared with C57 control group,the SI of the mice in C57 transplanted tumor group was decreased(P<0.05),gastrocnemius cells had hyaline degeneration,the sarcoplasm was stained blue,and some muscle fiber showed intranuclear migration;the expression levels of Atrogin-1,MuRF1 and ASNS mRNA in gastrocnemius tissue of the mice were increased(P<0.05),and the expression level of ASNS protein was increased(P<0.05);the expression levels of GRP78 and ATF4 and the ratio of p-eIF2α/eIF2αwere increased(P<0.05).Compared with APP/PS1 control group,the SI of the mice in APP/PS1 transplanted tumor group was decreased(P<0.05),some muscle fibers of gastrocnemius were swelling,and some were thin;intranuclear migration was easily seen,and the sarcoplasm was red;the expression levels of Atrogin-1,MuRF1 and ASNS mRNA in gastrocnemius tissue were increased(P<0.05),and

关 键 词：天冬酰胺合成酶 肌肉减少 淀粉样蛋白前体蛋白/早老素1 移植瘤 黑色素瘤B16细胞 

分 类 号：R739.5[医药卫生—肿瘤] R746.4[医药卫生—临床医学]