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目的:提高骨髓间充质干细胞(mesenchymal stem cell,MSC)移植治疗急性肝衰竭时在肝脏的定植率及疗效.方法:采用慢病毒转染使MSC过表达CXCR4基因,RT-PCR和细胞流式检测CXCR4表达.ELISA实验检测肝衰竭裸鼠肝脏MSC向基质衍生因子-1α(stromal cell-derived factor-1α,SDF-1α)水平.迁移实验检测转染的SDF-1α趋化能力.20%(V/V)四氯化碳(CCl4)橄榄油溶液8μL/g腹腔注射诱导裸鼠急性肝衰竭模型,24 h后尾静脉注射CXCR4转染的间充质干细胞(CXCR4-MSC)和未转染的间充质干细胞(Null-MSC).于注射后各时间点活体示踪MSC分布情况、裸鼠肝功能、生存率、组织病理学改变和肝细胞增殖情况.结果:CXCR4-MSC高表达CXCR4基因和CXCR4蛋白,肝衰竭裸鼠肝脏SDF-1α水平升高,与正常裸鼠相比有统计学差异.体外迁移实验证实CXCR4-MSC比Null-MSC具有向SDF-1α更好的迁移能力.体内活体成像显示移植后5 d CXCR4-MSC主要定植在肝脏,而Null-MSC在肝脏和脾脏都有定植.高定植率使得转染组裸鼠比未转染组裸鼠肝细胞受损更轻且具有更长的生存期,同时转染组裸鼠肝细胞增殖明显多于未转染组,两者有统计学差异.我们的实验证实转染CXCR4基因的骨髓间充质干细胞能够更好地促进肝再生.结论:基因修饰使骨髓间充质干细胞过表达CXCR4基因能够促进干细胞在肝脏的定植,改善肝功能,提高肝细胞的再生和裸鼠的生存率,为干细胞移植治疗肝功能衰竭提供了更广阔的应用途径.
OBJECTIVE: To improve the colonization rate and therapeutic effect of mesenchymal stem cell (MSCs) transplantation in the treatment of acute liver failure.Methods: CXCR4 gene was overexpressed by lentivirus and transfected into MSC by RT-PCR and flow cytometry The expression of CXCR4 was detected by ELISA, the level of stromal cell-derived factor-1alpha (SDF-1alpha) was detected in MSC of nude mice with liver failure and the chemotactic ability of transfected SDF- (CCl4) olive oil solution was intraperitoneally injected into the model of acute hepatic failure in nude mice. After 24 h, CXCR4-transfected mesenchymal stem cells (CXCR4-MSCs) were injected into tail vein and untransfected (Null-MSCs) .The distribution of MSCs, liver function, survival rate, histopathological changes and hepatocyte proliferation in nude mice were observed at different time points after injection.Results: CXCR4-MSC highly expressed CXCR4 The level of SDF-1α in the liver and hepatic failure nude mice were significantly higher than those in normal nude mice, and migration in vitro showed that CXCR4-MSCs migrated better than Null-MSC to SDF-1α. In vivo biopsy showed that CXCR4-MSCs colonized the liver primarily 5 days after transplantation, whereas Nul l-MSC colonization in the liver and spleen.High colonization rate makes the transfection group of nude mice than untransfected group of liver cells lighter damage and have a longer survival, while transfection of nude mice liver cell proliferation Significantly more than the untransfected group, there is a significant difference between the two.Our experiments confirmed that the transfection of CXCR4 gene in bone marrow mesenchymal stem cells can better promote liver regeneration.Conclusion: gene modification of bone marrow mesenchymal stem cells overexpression of CXCR4 Genes can promote stem cell colonization in the liver, improve liver function, improve liver cell regeneration and survival of nude mice, and provide a broader application of stem cell transplantation for liver failure.