BMMSCs联合NMP减轻大鼠心脏死亡供肝移植中肝细胞凋亡的机制研究  

Mechanisms of bone marrow mesenchymal stem cells plus normothermic mechanical perfusion alleviating hepatocyte apoptosis after donation after cardiac death transplantation in rats

作　　者：吴龙龙 袁梦淑 林玲 田轩 郑卫萍 曹欢 宋红丽 Wu Longlong;Yuan Mengshu;Lin Ling;Tian Xuan;Zheng Weiping;Cao Huan;Song Hongli(School of Medicine,Nankai University,Tianjin 300074,China;First Central Clinical College,Tianjin Medical University,Tianjin 300190,China;Department of Organ Transplantation,Tianjin First Central Hospital,Tianjin Key Laboratory of Organ Transplantation,School of Medicine,Nankai University,Tianjin 300192,China)

机构地区：[1]南开大学医学院,天津300074 [2]天津医科大学一中心临床学院,天津300190 [3]天津市第一中心医院器官移植科,天津市器官移植重点实验室,南开大学医学院,天津300192

出　　处：《中华器官移植杂志》2022年第4期228-234,共7页Chinese Journal of Organ Transplantation

基　　金：国家自然科学基金(82070639、81670574)。

摘　　要：目的研究骨髓间充质干细胞(BMMSCs)联合常温机械灌注(NMP)对大鼠心脏死亡供肝移植术后早期肝细胞凋亡的影响及其机制。方法贴壁法分离培养BMMSCs并鉴定。SD大鼠随机数字表法分为5组:假手术组(Sham组)、冷保存组(SCS组)、NMP组、BMMSC组和BMMSCs联合NMP组(BP组),每组每个时间点6只大鼠。分别在术后1 d和7 d取各组大鼠肝组织和血清,血生化检测肝脏酶学;HE染色观察肝组织病理变化;TUNNEL染色检测肝细胞凋亡;免疫组化检测肝细胞转录因子C/EBP同源蛋白(CHOP)表达;Western Blot检测内质网应激相关蛋白葡萄糖调节蛋白78(GRP-78)、磷酸化蛋白激酶R样内质网激酶(p-PERK)、活化转录因子4(ATF4)、CHOP、半胱氨酸天冬氨酸蛋白酶-3剪切体(Cleaved caspase-3)的表达。结果与SCS组相比,NMP组、BMMSC组和BP组肝脏损伤和炎症明显减轻,BP组损伤改善最明显(P<0.05)。BP组肝细胞凋亡在1 d和7 d时较SCS组显著减少,差异有统计学意义(P<0.05)。SCS组中GRP-78、p-PERK和ATF4表达增加,并且促凋亡蛋白CHOP和Cleaved caspase-3表达显著升高,而在BP组中均明显降低,差异有统计学意义(P<0.05)。结论BMMSCs联合NMP能够显著改善DCD供肝移植术后早期肝细胞凋亡和炎症反应,其机制可能与抑制移植肝的内质网应激有关。Objective To explore the effect and possible mechanism of normal temperature mechanical perfusion(NMP)plus bone marrow mesenchymal stem cells(BMMSCs)on early endoplasmic reticulum stress(ERs)and cellular apoptosis after donation after cardiac death(DCD)donor liver transplantation.Methods BMMSCs were isolated and cultured by adherence method.Sixty Sprague-Dawley(SD)rats were randomly divided into five groups of sham operation(sham),static cold storage(SCS),NMP,BMMSC and NMP plus BMMSCs(BP)(n=12 each).Liver tissue and serum sample of each group were harvested at Day 1/7 post-operation.Hematoxylin-eosin(HE)staining was employed for observing pathological changes of liver tissue;TdT-mediated dUTP nick end labeling(TUNNEL)staining for detecting cellular apoptosis;immunohistochemistry for detecting the expression of hepatocyte transcription factor C/EBP homologous protein(CHOP);Western blot for detecting the expressions of GRP-78,p-PERK,ATF4,CHOP and cleaved caspase-3.Results Compared with SCS group,hepatic injury and inflammation significantly declined in NMP,BMMSC and BP groups and improvement of hepatic injury was the most pronounced in BP group.Cellular apoptosis lessened markedly in BP group at Day 1/7 as compared with SCS group and the difference was statistically significant.The expressions of ERs-related proteins GRP-78,p-PERK and ATF4 spiked in SCS group and the expressions of pro-apoptotic proteins CHOP and cleaved caspase-3 were significantly elevated and declined markedly in BP group.And the difference was statistically significant.Conclusions BMMSCs plus NMP can significantly improve hepatocyte apoptosis and inflammatory response after DCD donor liver transplantation.And its mechanism may be correlated with suppressing early endoplasmic reticulum stress of hepatocytes.

关 键 词：肝移植 骨髓间充质干细胞 常温机械灌注 

分 类 号：R657.3[医药卫生—外科学]