大鼠胚胎内注射h UCMSCs在肝内存活和肝分化研究

Intrahepatic survival and hepatic differentiation in rat embryos injected with hUCMSCs

目的观察大鼠胚胎期注射绿色荧光蛋白(GFP)标记的人脐带间充质干细胞(GFP-hUCMSCs)在SD大鼠肝中存活和肝分化,探索在胚胎发育期建立人-大鼠嵌合体肝的可行性。方法运用携带绿色荧光蛋白的慢病毒转染标记人脐带间充质干细胞,然后注射到实验组大鼠胚胎。胎鼠出生后,制作冰冻切片观察GFPh UCMSCs在肝组织的迁移和分布,同时免疫组织化学法检测人相关蛋白在鼠肝内的表达。对照组注射等体积生理盐水。结果实验组胎鼠出生45 d、75 d、120 d的肝组织内检测到GFP信号的分布,对应肝组织内可检测到人肝细胞相关蛋白ALB、HNF3β和HNF4α的表达,但120 d大鼠含量极少、部分甚至检测不到。对照组为阴性。结论胚胎期注射h UCMSCs的胎鼠出生75 d的肝内仍可检测到GFP信号的存在和人肝细胞相关蛋白ALB、HNF3β和HNF4α的表达,且随时间延长而呈现逐渐减少的趋势。

Objective To observe the survival and liver differentiation of Green Fluorescent Protein(GFP labeled human umbilical cord mesenchymal stem cells(GFP-hUCMSCs)in SD rats during the embryonic stage,to investigate the feasibility of establishing a human-rat chimeric liver in the embryonic stage of rats.Methods Lentiviral containing GFP gene was used to transfect and to label human umbilical cord mesenchymal stem cells,and then GFPh UCMSCs were injected into the experimental group rat embryos.After embryonic rats birth,the Frozen liver sections were used,and the migration and distribution of the GFP-hUCMSCs were observed in the liver tissue.While the expression of human-related proteins in the rat liver were detected by immunohistochemical method.The control group was injected with an equal volume of normal saline.Results GFP signal were detected in liver tissue from the experimental group at 45 d,75 d,and 120 d after birth.Correspondingly,expression of the human hepatocyte-related proteins ALB,HNF3β,and HNF4αwas detected in the liver tissue,but these proteins content at 120 d was extremely small,and partly not detected.The control group was negative.Conclusions In embryonic rats injected with h UCMSCs,the presence of GFP signal and the expression of the human hepatocyte-related proteins ALB,HNF3β,and HNF4αcan be detected in the liver at 75 d after birth,and gradually decrease over time.