改构型酸性成纤维细胞生长因子对体外培养肝细胞增殖活性的影响

Effects of modified acidic fibroblast growth factor on the proliferation activity cultured hepatocytes in vitro

目的探讨改构型酸性成纤维细胞生长因子(MaFGF)对体外培养大鼠肝细胞增殖的影响。方法采用"0.25%胰酶+0.1%胶原酶"消化法分离培养大鼠肝细胞并传代纯化扩增,取第3代细胞鉴定后实验:(1)形态学法观察在不同浓度MaFGF组中细胞的形态变化;(2)将实验随机分为对照组和MaFGF组,MaFGF组加入一系列浓度的MaFGF,MTT法测定不同浓度MaFGF组作用于肝细胞24、48、72h的促增殖活性;(3)MTT法筛选出最适的MaFGF浓度,用此浓度和不加生长因子2种方式培养细胞,并绘制细胞生长曲线。结果(1)MaFGF对体外培养的肝细胞有一定的促增殖作用,但是与浓度不成正比;且实验表明24h的增殖率较其它时间点显著增高(P<0.05);比较对照组和各组(4.68×10^(-3)~7.80×10^(-3)mg/LMaFGF组)均差异有统计学意义(P<0.05),但各组间对肝细胞的促增殖作用的差异无统计学意义;(2)最适浓度(6.24×10^(-3)mg/L)MaFGF的第4天细胞数量较对照组明显增加(P<0.05);MaFGF组的第10天细胞数(12.4×10~4)是对照组(6×10~4)的2.1倍(P<0.05)。结论在适宜浓度和时间内,MaFGF对肝细胞具有一定的促增殖作用且不具有浓度依赖性。

Abstract Objective To explore effects of modified acidic fibroblast growth factor （MaFGF） on the growth of hep- atocytes cultured in vitro. Methods Hepatocytes of SD rats were isolated, cultured in vitro and subculture for three generations. After being identified, the cells were used in the experiment： Hepatocytes was observed by inverted phase contrast microscope; The experiment were randomly divided into control and experimental group, MaFGF with different concentration were added into the culture medium of hepatocytes, after 24,48172 h, the mitogenic effects were measured by the MTr test; The MTT test was used to detect the effects of MaFGF on the prolifera- tion of the hepatocytes in 24 h. The growth curve was plotted. Results MaFGF potentiated the proliferation of hepatocytes, which was no significant effect with dose increased. The proliferative rate of 24 h were higher than that of the other time points （P 〈 0.05 ）. there was statistical difference between the MaFGF group （4.68 x 10- 3 x 7.80 x 10-3 mg/L） and the control group （P 〈 0. 05 ）. However, the mitogeoic activities of each MaFGF group were in- significant; Four days after MaFGF（6. 24 x 10-3 rag/L） added into the medium, the population of the hepato- cytes was larger than that of the control （ P 〈 0.05 ）. The number of the hepatocytes incubated with MaFGF （ 12.4 x 104） was 2. 1 times as large as the control（6 x 104 ） in the 10th day （ P 〈 0. 05 ）. Conclusion MaFGF has a certain role in promoting the proliferation of the hepatocytes at the appropriate concentration and time. However, the effects don＇t have concentration dependent manner.