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为研究胎肝中造血和肝上皮发育的关系,建立了小鼠胎肝高增殖潜能集落形成细胞(HPP-CFC)培养体系,并进行了单克隆培养以及诱导分化实验.在造血和肝诱导因子的共同作用下,对单克隆来源的HPP集落细胞向造血和肝上皮细胞进行诱导分化,采用透射电镜(TEM)、巢式RT-PCR、细胞免疫荧光检测,从细胞形态、超微结构、上皮细胞分化标志等方面对分化后的细胞进行检测.检测结果显示,诱导后的部分细胞具有肝细胞特异性的超微结构,并不同程度地表达白蛋白(ALB)、甲胎蛋白(AFP)、细胞角蛋白(CK8,CK18)等肝上皮分化标志,同时还表达间质标志α-SMA和血管内皮细胞标志Flk-1.免疫磁珠分选表明:胎肝来源的HPP-CFC主要来自于CD45+细胞,CD45-细胞不具有形成造血细胞克隆的能力.在肝上皮细胞分化潜能上,流式分选获得的CD49f+/Sca-1+细胞与未分选细胞无明显差异.该模型的克隆源性通过细胞混合实验进行证明.研究结果表明,改进的胎肝来源的HPP-CFC可能代表了一个新的造血细胞向肝上皮细胞分化的单克隆模型,为研究胎肝中造血和非造血细胞的发育关系提供了一个新的切入点.
To study the relationship between hematopoietic and hepatic epithelial development in fetal liver, a culture system of mouse fetal liver hyperproliferative potential colony-forming cells (HPP-CFCs) was established and cultured in vitro and in vitro.Hematopoietic and hepatic induction factors , The monoclonal antibody derived HPC cells were induced to differentiate into hematopoietic and hepatic epithelial cells. Transmission electron microscopy (TEM), nested RT-PCR and immunofluorescence were used to detect the cell morphology, ultrastructure, Cell differentiation markers, etc. The results showed that some of the induced cells had hepatocyte-specific ultrastructure and expressed ALB, AFP, Cytokeratin (CK8, CK18) and other markers of liver epithelial differentiation, but also express interstitial markers α-SMA and vascular endothelial cell marker Flk-1. Immunomagnetic bead sorting showed that: fetal liver derived HPP-CFC mainly from CD45 + Cells, CD45- cells do not have the ability to form hematopoietic cells cloning .In the differentiation potential of liver epithelial cells, CD49f + / Sca-1 + cells obtained by flow sorting had no significant difference with unsorted cells.The clonality by Cell hybridization experiments.The results show that the improved fetal liver derived HPP-CFC may represent a new monoclonal model of hematopoietic cell differentiation to liver epithelial cells, in order to study the development of fetal liver hematopoietic and non-hematopoietic cells Provides a new entry point.