大鼠胎肝干细胞的体外分离、培养与鉴定

Isolation and identification of hepatic stem cells from rat fetal liver in vitro

目的:体外分离、培养原代大鼠胎肝干细胞(HSCs),并对其生物学特性进行鉴定。方法:采用胶原酶灌注法及剪碎消化法分离大鼠胎肝干细胞,并用含10%优等胎牛血清的H-DMEM培养液培养。利用免疫细胞化学方法在激光共聚焦扫描显微镜下观察该细胞表面抗原的表达情况,进行鉴定。结果:分离的大鼠胎肝干细胞体外培养24 h贴壁,5 d左右可长成单层,光镜下为致密圆形细胞,边缘清楚。8 d后细胞铺展,呈上皮样,表达甲胎蛋白(AFP)抗原、细胞角蛋白(CK)18、细胞角蛋白(CK)19。结论:分离并鉴定为大鼠胎肝干细胞可在体外培养条件下迅速扩增。

Objective To isolate and cultivate hepatic stem cells （HSCs） from rat fetal liver in vitro and identify their biological features. Methods Collagenase perfusion method and mechanical cutting method were used to isolate HSCs from rat fetal liver which were then cultivated by H-DMEM containing 10% fetal bovine serum. The cell surface antigen expression of HSCs was observed with immunocytochemical method under confocal laser scanning microscope. Results The isolated HSCs from rat fetal liver grew to monolayer 5 d after cultivation in vitro. They presented pykno-round cells and distinct borderline under the light microscope. After 8 d the cells grew like epithelium. These cells expressed AFP antigen, CK 18 and CK19. Conclusion The cultivated cells are proved to be HSCs and can proliferate quickly in vitro.