血清浓度对高原体外肝细胞培养的影响

目的了解在高原环境条件下血清对培养肝细胞生长的影响,初步确定进行体外肝细胞培养的适宜血清浓度。方法采用已适应本地环境的成年家兔肝细胞进行体外单层培养,在常规RPMI1640培养基的基础上添加不同浓度的新生牛血清(0%、2%、10%及20%),观察培养细胞形态、数量及培养上清中白蛋白和尿素水平变化。结果无血清组始终未见明显细胞贴壁及增殖,其他组细胞均于接种4 h开始贴壁,8 h开始增殖,10%血清组和20%血清组细胞均于6 d生长满瓶,并形成均匀的单层,而2%血清组细胞则于8 d生长满瓶;2%血清组的细胞数量峰值明显低于10%血清组和20%血清组;除无血清组外,培养上清中白蛋白和尿素水平均呈现显著升高,其中10%血清组和20%血清组的白蛋白及尿素峰值水平明显高于2%血清组。结论在高原环境条件下,体外培养肝细胞的生长、增殖及其生物学功能维持明显依赖于常规基础培养基中添加的血清浓度;对于普通肝细胞培养实验而言,10%血清浓度较为适宜。

Effects of Serum Concentration on Hepatocyte Culture.at High Altitude Setting

Objective To understand the effects of serum concentration on hepatocyte growth and proliferation at high altitude setting and to establish an optimal serum concentration for experimental use. Methods The hepatocytes isolated from rabbit liver were cultured with RPMI1640 culture medium appended with bovine serum in different concentrations.The changes of cell shape and amount and the biochemical substance levels in culture supernatants of the cultured rabbit hepatocytes were monitored regularly. Results The hepatocytes of 0% serum group didn't proliferate.After cultured for 4 hours,the hepatocytes of the other 3 groups began to adherent to the walls of plant boards;after 8 hours,the hepatocytes began to proliferate,some of them were binuclear;after 6 days,the bottoms of plant boards had been suffused with growing hepatocytes in the 10% serum group and the 20% serum group,and after 8 days,2% serum group,too.The peak value of cell number of 2% serum group was lower than that of 10% serum group and 20% serum group.Except the 0% serum group,the levels of albumin and urea in culture supernatants increased remarkably,and their peak values of the 10% serum group and the 20% serum groups were higher than that of 2% serum group. Conclusion Hepatocyte growth,proliferation and biological function depend on the serum concentration appending basic medium at high altitude setting;and 10% serum appending basic medium might be optimal for hepatocyte culture.