不同浓度肝细胞生长因子及表皮细胞生长因子体外联合诱导兔骨髓间充质干细胞向肝细胞分化

背景：研究证实在多种微环境下可以将骨髓间充质干细胞诱导分化为成熟肝细胞,但诱导条件及诱导分化率尚无定论,选择合适的诱导剂及诱导剂浓度尤为重要。 目的：观察肝细胞生长因子和表皮细胞生长因子体外联合诱导兔骨髓间充质干细胞向肝细胞分化的最适浓度。 方法：不同浓度肝细胞生长因子、表皮细胞生长因子联合诱导培养原代兔骨髓间充质干细胞,观察细胞形态学变化,并检测肝细胞表面标志物甲胎蛋白、白蛋白表达及肝细胞合成功能。 结果与结论：随诱导时间延长,可观察到多极性的肝细胞样细胞。7 d时甲胎蛋白表达阳性,14 d达最大值后即开始下降(P < 0.05),此后各浓度组间表达无差别(P  > 0.05)。14 d时白蛋白表达阳性,随时间延长,阳性细胞数递增(P  < 0.05),且细胞生长因子浓度越高,阳性细胞数越高(P < 0.05)。诱导早期(9~15 d) 白蛋白上清液中白蛋白水平与诱导剂浓度成正比(P < 0.05)。18 d或21 d达高峰后下降,此时各浓度组间含量无差别(P  > 0.05)。结果显示高浓度的肝细胞生长因子及表皮细胞生长因子可提高骨髓间充质干细胞向肝细胞的分化率,诱导剂最适浓度为肝细胞生长因子60 μg/L、表皮细胞生长因子4.5 mg/L。

In vitro combination of different-concentration hepatocyte growth factor and epidermal growth factor induces differentiation of rabbit bone marrow mesenchymal stem cells into hepatocytes

BACKGROUND:Studies have confirmed that in a variety of micro-environments, bone marrow mesenchymal stem cells (BMSCs) can be induced to differentiate into mature hepatocytes, but the inducible condition and differentiating rate are inconclusive. To select the appropriate inducer and its concentration is particularly important. OBJECTIVE:Through study on the differentiation of rabbit BMSCs into hepatocytes in vitro induced by different concentrations of hepatocyte growth factor (HGF) and epidermal cell growth factor (EGF), to find the best concentration combination of HGF and EGF. METHODS:BMSCs were induced with different concentrations of HGF and HGF. The morphological changes of cells were observed. In different culture periods, the hepatic surface phenotype including alpha fetoprotein (AFP), albumin (ALB) and synthesis function of hepatocytes were determined. RESULTS AND CONCLUSION:After prolonged culture, hepatocyte-like cells were seen. Until day 7, AFP was positive, but reduced in the following days (P < 0.05), and after that, there were no difference among all groups (P > 0.05). Until day 14, ALB expression was positive, and increased with the culture time (P < 0.05). The higher the concentration of cell growth factors, the higher the number of positive cells (P< 0.05). The concentration of ALB in the supernatant in the early stage (9-15 days) showed a concentration-dependent manner, and reached the peak at 18 or 21 days. Then the concentration began to decreased, and there were no difference (P > 0.05). The high concentrations of HGF and EGF may enhance the rate of differentiation, and the best combination is HGF 60 ug/L and EGF 4.5 mg/L.