内质网应激参与介导脂多糖诱导的大鼠肝细胞凋亡-论文.pdf

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1、法医学杂志2014年2月第30卷第1期·13·内质网应激参与介导脂多糖诱导的大鼠肝细胞凋亡季英磊，闰骏，王艳莎，刘夷嫦，谷振勇(南通大学医学院法医学系，江苏南通226001)摘要：目的探讨内质网应激在脂多糖(1ip0p0lysaccharide，LPS)诱导的肝细胞凋亡中的作用。方法肝细胞系BRL细胞培养．用LPS、内质网应激诱导剂毒胡萝卜素(thapsigargin，TG)、内质网应激抑制剂4一苯基丁酸(4一phenylbutyricacid，4一PBA)分别或组合处理肝细胞。用M1Tr比色法检测细胞活力的变化，Heochst33258荧光染色检测细胞凋亡形态的变化，Annexin

2、V—FITC／PI双染流式细胞术检测细胞凋亡率，Western印迹法检测内质网应激标志蛋白GRP78和内质网应激相关的凋亡蛋白CHOP、caspase一12和激活型casDase一3的表达。结果LPS引起肝细胞活力降低、细胞凋亡率明显增加，并具有量效和时效关系，LPS诱导GRP78、CIIOP,caspase-12和激活型caspase一3的表达上调；TG可引起肝细胞活力降低和细胞凋亡增加．并能加重LPS引起的肝细胞损伤；4一PBA明显抑制LPS引起的肝细胞凋亡增加。结论内质网应激参与介导了LPS引起的肝细胞凋亡．提示内质网应激是LPS诱导肝细胞损伤的重要发病途径。关键词：法医病理学

3、；内质网；脂多糖类；肝细胞；细胞凋亡；大鼠中图分类号：DF795．1文献标志码：Adoi：10．3969~．issn．1004-5619．2014．01．003文章编号：1004—5619(2014)01—0013—06EndoplasmicReticulumStressMediatesLipOpOlysaccharide—inducedApoptosisinRatHepatocyte1lYing-leiYANlun，WANGYan—sha,LIUYi-chang,GUZhen-yong(DepartmentofForensicMedicine，MedicalCollegeofNan

4、tongUniversity,Nantong226001，China)Abstract：ObjectiveToinvestigatetheroleofendoplasmicreticulumstress(ERS)inlipopolysaccharide(LPS)．inducedhepatocyteapoptosis．MethodsCellsoftherathepatocyte1ineBRLwerecultured．ThehepatocytesweretreatedwithLPS，ERSinducerthapsigargin(TG)，andERSinhibitor4-phenylbut

5、yricacid(4．PBA)．respectivelyorintheirdifferentcombination．Thecel1viabilitywasmeasuredbyMTTassay．Thecyto-nuclearmorphologicalchangesofapoptosiscellsweredetectedbythefluorescentdyeHoechst33258．TheapoptosisratewasassessedbyflowcytometrywithAnnexinV-FITC／PIdouble—staining．Ex-pressionsofGRP78asERSma

6、rkerprotein，CHOP，caspase一12andcleaved—caspase-3asERSrelatedproteinweredetectedbyWesternblotting．ResultsLPScouldcauseadecreaseincellviabilityandanincreaseinapoptosisrateinadose—andtime．dependentmanner．TheexpressionofGRP78，CHOP，cas-pase一12andcleaved—caspase．3proteinsweresignificantlyincreasedwith

7、LPStreatment．TG1edtoamarkeddecreaseincellviabilityandanincreaseinapoptosisrate，whichaggravatedthehepatocytein—iuryinducedbyLPS；whereas4．PBAalleviatedLPS—inducedapoptosis．ConclusionERSmediatesLPS-inducedhepatocyteinjuries，indicatin