内质网应激相关蛋白1对衣霉素诱导HepG2细胞内质网应激的影响
내질망응격상관단백1대의매소유도HepG2세포내질망응격적영향
Effects of stress-associated endoplasmic reticulum protein 1 on endoplasmic stress induced by the ;tunicamycin in HepG2 cells
  • 万方数据
    中华诊断学电子杂志 중화진단학전자잡지

    2014年 1期 32-37 ,共6页

    肖元元%韩峻峰%毛月芹%王倩倩%魏美林%殷峻%黄金伟%魏丽

    초원원%한준봉%모월근%왕천천%위미림%은준%황금위%위려

    内质网应激%内质网应激相关蛋白1%葡萄糖调节蛋白78%C/EBP同源蛋白%钙联蛋白 內質網應激%內質網應激相關蛋白1%葡萄糖調節蛋白78%C/EBP同源蛋白%鈣聯蛋白
    내질망응격%내질망응격상관단백1%포도당조절단백78%C/EBP동원단백%개련단백
    Endopasmic reticulum stress%Stress-associated Endoplasmic Reticulum Protein 1%Glucose-regulated protein 78%C/EBP homologous protein%Calnexin
    目的：研究过表达内质网应激相关蛋白1（SERP1）对衣霉素诱导肝癌HepG2细胞内质网应激的影响。方法以衣霉素诱导HepG2细胞发生内质网应激，将细胞分为以下5组：正常对照组、衣霉素组、衣霉素＋0．25μg SERP1转染组、衣霉素＋0．5μg SERP1转染组和衣霉素＋1．0μg SERP1转染组，每组实验重复3次；采用MTT法检测不同浓度与作用时间的衣霉素对HepG2细胞存活率的影响，以吸光度（A ）值表示。Western blot法检测各组细胞内内质网应激标志蛋白葡萄糖调节蛋白（GRP78）、C／EBP同源蛋白（CHOP）以及钙联蛋白的表达水平。采用SPSS 15．0统计软件进行统计学分析，比较蛋白表达水平。结果与对照组相比，衣霉素处理组HepG2细胞中内质网应激标志性蛋白GRP78、CHOP及 Calnexin 蛋白表达量显著升高，分别为对照处理组的3．8倍（t ＝11．5， P ＜0．05）、1．3倍（t ＝3．498，P ＜0．05）和1．4倍（t ＝4．1，P ＜0．05），差异均有统计学意义；随着SERP1过表达量的逐渐升高，变化呈现剂量依赖性。随着SERP1转染剂量的增加，各组GRP78蛋白的表达较单独衣霉素处理组分别下降了12％[（1．83±0．29）A值，（1．61±0．13）A值，t ＝2．36， P ＞0．05]、24％和30％[（1．83±0．29）A值，（1．40±0．11）A值，（1．27±0．21）A值；F ＝50．56， P ＜0．05]，CHOP蛋白的表达水平分别下降了23％，29％和34％[（1．0±0．15）A值，（0．79±0．07）A值，（0．72±0．55）A值，（0．67±0．14）A值；F ＝9．532，P ＜0．05]，Calnexin蛋白的表达水平分别下降了5％[（1．20±0．18）A值，（1．15±0．13）A值；P ＞0．05]、24％和28％[（1．20±0．18） A值，（0．92±0．07）A值，（0．87±0．18）A值；F＝8．116，P＜0．05]。结论外源性过表达SERP1蛋白通过下调内质网应激蛋白的表达，降低HepG2细胞内质网应激水平，缓解内质网应激介导的细胞损伤。
    목적：연구과표체내질망응격상관단백1（SERP1）대의매소유도간암HepG2세포내질망응격적영향。방법이의매소유도HepG2세포발생내질망응격，장세포분위이하5조：정상대조조、의매소조、의매소＋0．25μg SERP1전염조、의매소＋0．5μg SERP1전염조화의매소＋1．0μg SERP1전염조，매조실험중복3차；채용MTT법검측불동농도여작용시간적의매소대HepG2세포존활솔적영향，이흡광도（A ）치표시。Western blot법검측각조세포내내질망응격표지단백포도당조절단백（GRP78）、C／EBP동원단백（CHOP）이급개련단백적표체수평。채용SPSS 15．0통계연건진행통계학분석，비교단백표체수평。결과여대조조상비，의매소처리조HepG2세포중내질망응격표지성단백GRP78、CHOP급 Calnexin 단백표체량현저승고，분별위대조처리조적3．8배（t ＝11．5， P ＜0．05）、1．3배（t ＝3．498，P ＜0．05）화1．4배（t ＝4．1，P ＜0．05），차이균유통계학의의；수착SERP1과표체량적축점승고，변화정현제량의뢰성。수착SERP1전염제량적증가，각조GRP78단백적표체교단독의매소처리조분별하강료12％[（1．83±0．29）A치，（1．61±0．13）A치，t ＝2．36， P ＞0．05]、24％화30％[（1．83±0．29）A치，（1．40±0．11）A치，（1．27±0．21）A치；F ＝50．56， P ＜0．05]，CHOP단백적표체수평분별하강료23％，29％화34％[（1．0±0．15）A치，（0．79±0．07）A치，（0．72±0．55）A치，（0．67±0．14）A치；F ＝9．532，P ＜0．05]，Calnexin단백적표체수평분별하강료5％[（1．20±0．18）A치，（1．15±0．13）A치；P ＞0．05]、24％화28％[（1．20±0．18） A치，（0．92±0．07）A치，（0．87±0．18）A치；F＝8．116，P＜0．05]。결론외원성과표체SERP1단백통과하조내질망응격단백적표체，강저HepG2세포내질망응격수평，완해내질망응격개도적세포손상。
    Backgroud Endoplasmic reticulum stress was induced by the accumulation and aggregation of unfolded proteins due to stresses that disturbed the cellular energy levels,the redox state,or Ca2+concentration,and leading to the unfolded protein response (UPR)pathway.The hepatic UPR was activated in several forms of liver disease.Recent data showed that the role of the UPR in hepatic cells have identified molecular mechanisms that may underlie the association between UPR activation and liver disease. SERP1 was known as ribosome-associated membrane protein 4 (RAMP4),was homologous to yeast suppressor of SecY 6 protein (YSY6p)which suggested a role in pathways controlling membrane protein biogenesis at&nbsp;the ER level.Expression of SERP1 was enhanced during cellular stress,causing accumulation of unfolded proteins in the ER.By interaction with the molecular chaperone calnexin,SERP1/RAMP4 could control biogenesis of membrane proteins and take participate in the endoplasmic reticulum stress.Objective To study the effects of stress-associated endoplasmic reticulum protein 1 (SERP1 )on the endoplasmic reticulum stress induced by the tunicamycin in HepG2 cells.Methods The tunicamycin was used to induce endoplasmic reticulum stress in the HepG2 cells.We divided the cells into 5 groups:normal control group, tunicamycin treated group,tunicamycin +0.25μg/μl SERP1 transfected group,tunicamycin +0.25μg/μl SERP1 transfected group,tunicamycin +0.5μg/μl SERP1 transfected group,tunicamycin +1μg/μl SERP1 transfected group.Each experiment was repeated three times.MTT was used to detect the effect on the survival rate of the HepG2 cells and selected the optimal concentration and time of tunicamycin treatment. Western blot was used to detect the standard of expression of endoplasmic reticulum stress spcific mark proteins,glucose-regulated protein 78(GRP78),C/EBP homologous protein (CHOP)and calnexin.Results Compared with the control group,the expression levels of GRP78,CHOP and calnexin were significantly increased in the tunicaymicin treated group,which were 3.8 times,1.3 times and 1.4 times respectively. With the increasing amount of transfection,SERP1 over expression was found to relieve the expression of GRP78 12%(1.838 ±0.29,1.6 ±0.132,P >0.05 ),24% and 30%(1.838 ±0.29,1.40 ±0.11,1.27 ±0.21,F =50.56,P <0.01 )compared with the tunicamycin group,the expression of CHOP were decreased by 23%,29% and 34%(1.0 ±0.15,0.79 ±0.07,0.72 ±0.55,0.67 ±0.14,F =9.532, P <0.01 )respectively and calnexin were decreased by 5%(1.20 ±0.18,1.15 ±0.13,P >0.05)、24% and 28%(1.20 ±0.18,0.92 ±0.07,0.87 ±0.18,F =8.116 ,P <0.01)respectively,which were induced by tunicamycin treatment.Conclusion SERP1 overexpression could attenuate the ER stress induced by tunicamycin,and may reduce the cell damage mediated by the ER stress.the ER level.Expression of SERP1 was enhanced during cellular stress,causing accumulation of unfolded proteins in the ER.By interaction with the molecular chaperone calnexin,SERP1/RAMP4 could control biogenesis of membrane proteins and take participate in the endoplasmic reticulum stress.Objective To study the effects of stress-associated endoplasmic reticulum protein 1 (SERP1 )on the endoplasmic reticulum stress induced by the tunicamycin in HepG2 cells.Methods The tunicamycin was used to induce endoplasmic reticulum stress in the HepG2 cells.We divided the cells into 5 groups:normal control group, tunicamycin treated group,tunicamycin +0.25μg/μl SERP1 transfected group,tunicamycin +0.25μg/μl SERP1 transfected group,tunicamycin +0.5μg/μl SERP1 transfected group,tunicamycin +1μg/μl SERP1 transfected group.Each experiment was repeated three times.MTT was used to detect the effect on the survival rate of the HepG2 cells and selected the optimal concentration and time of tunicamycin treatment. Western blot was used to detect the standard of expression of endoplasmic reticulum stress spcific mark proteins,glucose-regulated protein 78(GRP78),C/EBP homologous protein (CHOP)and calnexin.Results Compared with the control group,the expression levels of GRP78,CHOP and calnexin were significantly increased in the tunicaymicin treated group,which were 3.8 times,1.3 times and 1.4 times respectively. With the increasing amount of transfection,SERP1 over expression was found to relieve the expression of GRP78 12%(1.838 ±0.29,1.6 ±0.132,P >0.05 ),24% and 30%(1.838 ±0.29,1.40 ±0.11,1.27 ±0.21,F =50.56,P <0.01 )compared with the tunicamycin group,the expression of CHOP were decreased by 23%,29% and 34%(1.0 ±0.15,0.79 ±0.07,0.72 ±0.55,0.67 ±0.14,F =9.532, P <0.01 )respectively and calnexin were decreased by 5%(1.20 ±0.18,1.15 ±0.13,P >0.05)、24% and 28%(1.20 ±0.18,0.92 ±0.07,0.87 ±0.18,F =8.116 ,P <0.01)respectively,which were induced by tunicamycin treatment.Conclusion SERP1 overexpression could attenuate the ER stress induced by tunicamycin,and may reduce the cell damage mediated by the ER stress.
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