HO-1修饰的BMMSCs联合常温机械灌注对肝移植急性排斥反应Brown Norway大鼠肠道屏障功能的影响  

Effect of heme oxygenase-1 modified bone marrow mesenchymal stem cells combined with normothermic machine perfusion on intestinal barrier function in Brown Norway rats with acute rejection of liver transplantation

作　　者：林玲[1] 袁梦淑 吴龙龙 田轩 郑卫萍[3] 曹欢 宋红丽[3] Lin Ling;Yuan Mengshu;Wu Longlong;Tian Xuan;Zheng Weiping;Cao Huan;Song Hongli(Tianjin Medical University,First Central Clinical College,Tianjin 300192,China;Nankai University,School of Medicine,Tianjin 300074,China;Department of Organ Transplantation,Tianjin First Central Hospital,Tianjin Key Laboratory of Organ Transplantation,Tianjin Organ Transplantation Clinical Medical Research Center,Tianjin 300192,China)

机构地区：[1]天津医科大学一中心临床学院,天津300192 [2]南开大学医学院,天津300074 [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,天津市器官移植临床医学研究中心,天津300192

出　　处：《中华肝胆外科杂志》2022年第3期210-215,共6页Chinese Journal of Hepatobiliary Surgery

基　　金：国家自然科学基金(82070639、81670574)。

摘　　要：目的分析血红素加氧酶-1(HO-1)修饰的骨髓间充质干细胞(BMMSCs)联合常温机械灌注(NMP)对肝移植急性排斥反应大鼠肠道屏障功能的影响。方法无特定病原体4~5周龄、40~60 g雄性Brown Norway(BN)大鼠2只提取BMMSCs,通过腺病毒转导HO-1;24只7~8周龄、200~220 g雄性Lewis大鼠为供体,30只8~9周龄、220~240 g雄性BN大鼠为受体,"二袖套"法建立原位肝移植急性排斥反应模型。BN大鼠随机分为5组:Sham组仅开腹关腹;NMP组供肝NMP 4 h;BMP组供肝NMP 4 h+门静脉灌注BMMSCs;HBP组供肝同BMP组,但灌注HO-1/BMMSCs;FK506组供肝NMP 4 h,肝移植术后灌胃他克莫司,每组6只。术后第14天取材检测并计算排斥活动指数(RAI),HE染色及透射电镜分别观察肠道病理改变和超微结构,Western印迹检测肠道组织紧密连接蛋白表达,酶联免疫吸附法检测血清脂多糖、D-乳酸、二胺氧化酶(DAO)。结果HBP组、FK506组RAI为(2.80±0.84)分、(2.20±0.84)分,显著低于NMP组(7.60±1.14)分、BMP组(6.00±1.58)分,差异均有统计学意义(均P<0.05)。肠道HE染色NMP组肠绒毛明显稀疏、皱缩,排列杂乱,BMP组、HBP组、FK506组较NMP组肠道损伤程度减轻。电镜观察HBP组肠上皮细胞微绒毛丰富且排列整齐,细胞间紧密连接结构完整。Western印迹检测闭锁小带-1相对表达,HBP组(0.87±0.06),高于NMP组(0.41±0.12)和FK506组(0.52±0.15);闭合蛋白相对表达HBP组(1.28±0.26),高于NMP组(0.27±0.18)和FK506组(0.63±0.22),差异均有统计学意义(均P<0.05)。HBP组血清脂多糖、D-乳酸和DAO浓度均低于NMP组、FK506组,差异均有统计学意义(均P<0.05)。结论HO-1/BMMSCs联合NMP可以保护肝移植术后急性排斥反应BN大鼠的肠道屏障功能。Objective To investigate the effect of heme oxygenase-1(HO-1)modified bone marrow mesenchymal stem cells(BMMSCs)combined with normothermic machine perfusion(NMP)on the intestinal barrier function in rats with acute rejection of liver transplantation.Methods Specific pathogen free 2 male Brown Norway(BN)rats(4-5 weeks,40-60 g)were used to isolat BMMSCs,and HO-1 was infected by adenovirus.Of 24 male Lewis rats(7-8 weeks old,200-220g)were used as donors,30 male BN rats(8-9 weeks old,220-240 g)were used as recipients.Acute rejection models of orthotopic liver transplantation were established in rats using two cuff technique.BN recipient rats were randomly divided into five groups:sham group,abdomen of the mice was open and closed within 30 min;NMP livers were simply mechanically perfused for 4 h;the BMP group were perfused with BMMSCs through the portal vein;the HBP group were perfused with HO-1/BMMSCs through the portal vein;the FK506 livers were mechanically perfused for 4 h and administered intragastrically of tacrolimus daily following surgery,6 per group,on days 14 after surgery,the relevant indicators were taken and the rejection activity index(RAI)changes were investigated.The changes of intestinal pathological were analyzed by HE staining and transmission electron microscope,the expression levels of zonula occludens-1(ZO-1)and occludin protein in intestinal tissue were detected by Western blotting,the concentrations of lipopolysaccharide,D-lactic acid and diamine oxidase(DAO)in serum were detected by ELISA.Results The RAI of HBP group(2.80±0.84)and FK506 group(2.20±0.84)were significantly lower than that of NMP group(7.60±1.14)and BMP group(6.00±1.58),the differences were statistically significant(all P<0.05).The intestinal villi in NMP group were significantly sparse,wrinkled and disorderly arranged while the degree of intestinal injury in BMP group,HBP group and FK506 group were more mitigated.Electron microscope observation showed that the microvilli of intestinal epithelial cells in HBP group were ric

关 键 词：肝移植 骨髓间充质干细胞 常温机械灌注 急性排斥反应 肠道屏障 

分 类 号：R657.3[医药卫生—外科学]