鼠特异性Fas抗体对人鼠嵌合肝中人肝细胞增殖的促进作用

Repopulation of human fetal hepatocytes in nude mouse model with chimeric human liver using mouse-specific anti-Fas antibody

目的:探讨人胎肝细胞移植联合使用JO2抗体的策略,促进人胎肝细胞在小鼠肝内存活和增殖.方法:裸鼠经脾移植1×106人胎肝细胞,移植后第1天ipJO2抗体,剂量为0.2mg/kg,1次/wk,持续12wk为实验组,裸鼠经人胎肝细胞移植后未给予JO2抗体为对照组,建立人鼠嵌合肝动物模型.采用HE染色、原位末端标记方法(TUNEL染色)观察未经人胎肝细胞移植而给予JO2抗体24h后处死的裸鼠肝脏组织.免疫组化、逆转录聚合酶链反应(RT-PCR)检测不同时相点实验组和对照组嵌合肝中肝组织人白蛋白、特异人增殖细胞核抗原(PCNA)和人白蛋白mRNA表达.结果:未经人胎肝细胞移植而给予JO2抗体的裸鼠病理组织切片发现肝组织出血、坏死和细胞凋亡.实验组和对照组裸鼠均能存活至24wk.移植后嵌合鼠肝组织表达人白蛋白和人PCNA阳性细胞的时间:实验组2-20wk,对照组2-12wk;白蛋白mRNA表达:实验组4-16wk,对照组4-8wk;实验组与对照组移植后8、12wkPCNA表达差异有显著性(25.7%±8.5%vs13.4%±7.8%,29.4%±5.0%vs8.5%±2.3%,均P<0.05).结论:人肝细胞异种移植于裸鼠体内能够存活,经小剂量JO2抗体ip裸鼠,使人鼠嵌合肝中人肝细胞得以增殖,存活时间延长.

AIM：To investigate repopulation of human fetal hepatocytes in an animal model of nude mice with chimeric human liver following induction of mouse hepatocyte apoptosis using a mouse-specific anti-Fas monoclonal antibody（Jo2 mAb） that does not engage xenogeneic fas. METHODS：For experiment group,nude mice were transplanted with human fetal hepatocytes intrasplenically and treated with 0.2 mg/kg Jo2 mAb intraperitoneally once a week for 12 weeks consistently.Nude mice in the control group were transplanted with human fetal hepatocytes but not administrated with Jo2 mAb.Liver section from non-transplanted nude mice administered with Jo2 mAb were analyzed using hematoxylin and eosin staining and terminal uridine deoxynucleotidyl transferase dUTP nick end labeling（TUNEL）staining.Reverse transcription-polymerase chain reaction（RT-PCR）and S-P immunohistochemistry were used to detect human albumin mRNA,human albumin and specific proliferating cell nuclear antigen（PCNA）in chimeric liver tissues. RESULTS：Liver sections from non-transplanted nude mice administered with Jo2 mAb showed hepatocyte death,massive apoptosis and hemorrhage.Nude mice in both experiment group and control group survived 24 weeks after transplantation.Human albumin and specific human PCNA were detected from the week 2 to week 20 after transplantation,but they could only be detected from the week 2 to week 12 in the controls.Human albumin mRNA（356 bp）was detected in mice livers from the week 4 to week 16 after transplantation,but they could only be detected from the week 4 to week 8 in the controls.The number of PCNA in experiment group is significantly higher than in the control group at 8,12 wk（25.7%±8.5%vs 13.4%±7.8%,29.4% ±5.0%vs 8.5%±2.3%,both P〈0.05）. CONCLUSION：Human fetal hepatocytes of xenogeneic graft can survive in nude mice.The repopulation of human fetal hepatocytes can be promoted and prolonged in nude mouse model with chimeric human liver using mouse-specific anti-Fas antibody intraperitoneally.