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磷酸甘油酸变位酶5调控细胞焦亡在肝脏缺血-再灌注损伤中的作用

乔兵兵 李世朋 宋浩森 季敏 赵龙栓

乔兵兵, 李世朋, 宋浩森, 等. 磷酸甘油酸变位酶5调控细胞焦亡在肝脏缺血-再灌注损伤中的作用[J]. 器官移植, 2021, 12(4): 412-420. doi: 10.3969/j.issn.1674-7445.2021.04.007
引用本文: 乔兵兵, 李世朋, 宋浩森, 等. 磷酸甘油酸变位酶5调控细胞焦亡在肝脏缺血-再灌注损伤中的作用[J]. 器官移植, 2021, 12(4): 412-420. doi: 10.3969/j.issn.1674-7445.2021.04.007
Qiao Bingbing, Li Shipeng, Song Haosen, et al. Effect of phosphoglycerate mutase 5 mediated pyroptosis on liver ischemia-reperfusion injury[J]. ORGAN TRANSPLANTATION, 2021, 12(4): 412-420. doi: 10.3969/j.issn.1674-7445.2021.04.007
Citation: Qiao Bingbing, Li Shipeng, Song Haosen, et al. Effect of phosphoglycerate mutase 5 mediated pyroptosis on liver ischemia-reperfusion injury[J]. ORGAN TRANSPLANTATION, 2021, 12(4): 412-420. doi: 10.3969/j.issn.1674-7445.2021.04.007

磷酸甘油酸变位酶5调控细胞焦亡在肝脏缺血-再灌注损伤中的作用

doi: 10.3969/j.issn.1674-7445.2021.04.007
基金项目: 

河南省高等学校重点科研项目计划 20A320077

河南省医学科技攻关计划省部共建项目 SB201902007

详细信息
    通讯作者:

    乔兵兵,男,1982年生,博士,副主任医师,研究方向为肝胆胰外科基础与临床研究,Email: popzxcbb@126.com

  • 中图分类号: R617, R364.5

Effect of phosphoglycerate mutase 5 mediated pyroptosis on liver ischemia-reperfusion injury

More Information
  • 摘要:   目的  研究磷酸甘油酸变位酶5(PGAM5)调控细胞焦亡在肝脏缺血-再灌注损伤(IRI)中的作用及分子机制。  方法  建立C57小鼠肝脏IRI模型,随机分别予再灌注6 h(6 h组)与12 h(12 h组),并设立假手术组(Sham组),每组10只。分析IRI对小鼠肝组织及血清学指标的影响; 分析小鼠肝脏IRI过程中PGAM5、半胱氨酸天冬氨酸蛋白酶(Caspase)-1的表达情况。建立肝细胞IRI模型(IRI组),采用Caspase-1抑制剂Z-YVAD-FMK预处理后再建立肝细胞IRI模型(抑制剂组),将未处理的AML12细胞作为对照组,分析抑制Caspase-1活性对细胞焦亡的影响。采用脂质体3000将PGAM5小干扰核糖核酸(siRNA)(siRNA组)和siRNA阴性对照(siRNA-NC)(siRNA-NC组)转染至AML12细胞,再建立肝细胞IRI模型,将未处理的AML12细胞作为对照组,分析PGAM5调控细胞焦亡对肝细胞IRI的影响。  结果  6 h组和12 h组小鼠肝组织中部分肝细胞水肿,肝窦区变窄,中央静脉充血,偶见点灶状坏死等,且12 h组较6 h组病变加重。与Sham组小鼠比较,6 h组和12 h组小鼠血清丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)水平升高,且12 h组高于6 h组; 6 h组和12 h组肿瘤坏死因子(TNF)-α、白细胞介素(IL)-1β水平升高,12 h组低于6 h组; 6 h组和12 h组小鼠肝组织IL-1β信使核糖核酸(mRNA)相对表达量升高,12 h组低于6 h组; 6 h组和12 h组肝组织细胞凋亡率升高,12 h组低于6 h组(P < 0.01~0.05)。与Sham组小鼠比较,6 h组和12 h组小鼠肝组织PGAM5 mRNA相对表达量和蛋白相对表达量均升高,且12 h组高于6 h组(P < 0.01~0.05);6 h组和12 h组肝组织PGAM5、Caspase-1蛋白表达增多。IRI组细胞NOD样受体蛋白3(NLRP3)、裂解Caspase (cleaved Caspase)-1及Gasdermin D(GSDMD)蛋白相对表达量较对照组升高,GSDMD荧光强度较对照组增强; 抑制剂组NLRP3、cleaved Caspase-1及GSDMD蛋白相对表达量较IRI组下降,GSDMD荧光强度较IRI组减弱(P < 0.01~0.05)。与对照组比较,siRNA-NC组细胞存活率下降,PGAM5、NLRP3、cleaved Caspase-1、GSDMD蛋白相对表达量升高(P < 0.01~0.05);与siRNA-NC组比较,siRNA组细胞存活率升高,PGAM5、NLRP3、cleaved Caspase-1、GSDMD蛋白相对表达量下降(P < 0.01~0.05)。  结论  PGAM5可加重小鼠肝脏IRI,其机制可能为通过PGAM5/Caspase-1/GSDMD信号通路调控细胞焦亡,加速肝细胞损伤。

     

  • 图  1  各组小鼠肝组织和血清学指标的表现

    注:A图示各组小鼠肝组织病理学表现(HE,×200);B图示各组小鼠血清ALT水平; C图示各组小鼠血清AST水平; D图示各组小鼠血清TNF-α水平; E图示各组小鼠血清IL-1β水平; F图示各组小鼠肝组织IL-1β mRNA相对表达量; G、H图示各组小鼠肝组织中细胞凋亡水平(TUNEL,×200)。与Sham组比较,aP < 0.01;与6 h组比较,bP < 0.05,cP < 0.01。

    Figure  1.  Manifestations of the indexes of liver tissues and serology of mice in each group

    图  2  各组小鼠肝组织PGAM5和Caspase-1的表达情况

    注:A图示各组小鼠肝组织中PGAM5 mRNA相对表达量; B图示各组小鼠肝组织PGAM5蛋白相对表达量; C图示各组小鼠肝组织PGAM5、Caspase-1表达与分布情况(免疫组化,×200)。与Sham组比较,aP < 0.05,bP < 0.01;与6 h组比较,cP < 0.05,dP < 0.01。

    Figure  2.  Expression of PGAM5 and Caspase-1 in liver tissues of mice in each group

    图  3  各组肝细胞焦亡相关蛋白的表达情况

    注:A图示各组细胞NLRP3、cleaved Caspase-1及GSDMD蛋白相对表达量; B图示各组细胞GSDMD蛋白的表达与分布(免疫荧光,×200)。与对照组比较,aP < 0.05,bP < 0.01;与IRI组比较,cP < 0.05,dP < 0.01。

    Figure  3.  Expression of pyroptosis related proteins of hepatocytes in each group

    图  4  各组肝细胞存活及PGAM5/Caspase-1/GSDMD信号通路蛋白表达情况

    注:A图示各组细胞存活率; B图示各组细胞PGAM5、NLRP3、cleaved Caspase-1及GSDMD蛋白相对表达量; C图示各组细胞GSDMD蛋白的表达与分布(免疫荧光,×200)。与对照组比较,aP < 0.05,bP < 0.01;与siRNA-NC组比较,cP < 0.05,dP < 0.01。

    Figure  4.  Survival and protein expression of PGAM5/Caspase-1/GSDMD signaling pathway in hepatocytes in each group

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出版历程
  • 收稿日期:  2021-03-27
  • 网络出版日期:  2021-07-13
  • 刊出日期:  2021-07-15

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