人骨髓单个核细胞向肝细胞诱导分化的体外研究

The differentiation of human bone marrow mononuclear cell into hepatocyte in vitro

目的 探讨在HGF、FGF_4诱导下骨髓单个核细胞能否向肝细胞分化。方法 找健康忐愿者,年龄4～50岁,胸骨抽取骨髓,用淋巴分离液分离出骨髓单个核细胞,用含2％胎牛血清和1×ITS的DMEM培养,分别用单独HGF、单独FGF_4、HGF和FGF_4联合刺激、无生长因子4种处理因素进行诱导培养,分别于诱导培养0、7、14、21、28天时,留取细胞,用免疫细胞化学法检测CK18、AFP、白蛋白,用PAS法进行糖元染色试验以验证诱导分化的结果。结果 0天时,AFP、白蛋白少数细胞阴性,CK18阴性,糖元未见阳性;加生长因子的三组AFP在7、14天时表达逐渐增高,21、28天减弱;糖元、CK18、白蛋白表达逐渐增高。无生长因子组,在7、14、21、28天时,这此指标均未见表达。结论 在HGF、FGR_4诱导下,骨髓单个核细胞可分化为具肝细胞表型和功能的细胞。

Objective To observe whether the bone marrow mononuclear cell (BMMC) can differentale into the cells with hepatocytic phenotype and funcation in some conditions. Methods Bone marrow cells from the sternum of healthy volun-leers between 4 to 50 years old. And BMMCs were separated by gradient centrifugation on Lymophoeyte Separating Liquid (density 1 .077) .The cells were plated in DMEM supplemented with 2% FBS, 1× ITS( insulin/transferrin/selenium) . And single HGF, single FGF4,HGF + FGF4,no growth factor was added respectively. Cells were collected on day 7,14,21,28.CK18, AFP,Albumin were tested by immunoeytoehemistry, glycogen by PAS( Periodic: acid-schiff) .Results Albumin ,AFP were positive In lew new isolated BMMCs,and CK18 , Glycogen were negative in BMMCs. The expression of AFP increased progressively on day 7 and 14, then decresed on day 21 and 28. The expressions of CK18 刧lycogen were stronger with the cultured time longer. Conclusion Bone marrow mononuclear cell can differentiate into the cells with hepatic phenotype and funtion in the presence of HGF and(or) FGF4 .