无血清培养基培养乳猪肝细胞的效果  

Serum-free medium for culture of suckling pig hepatocytes

作　　者：陈钟[1] 丁义涛[1] 张鹤云[2] 

机构地区：[1]南京大学医学院附属鼓楼医院肝胆外科,江苏省南京市210008 [2]南京大学生化系,江苏省南京市210093

出　　处：《世界华人消化杂志》2002年第3期320-323,共4页World Chinese Journal of Digestology

基　　金：江苏省卫生厅重点发展项目基金;No.BQ200020

摘　　要：目的:比较无血清培养基和含血清培养基培养乳猪肝细胞的效果,以寻找一种用于生物人工肝猪肝细胞无血清培养的良好培养基。 方法:采用改良原位两步胶原酶灌注法分离乳猪肝细胞。将肝细胞以5×10~8·L^(-1)分别培养在无血清培养基和含血清培养基中,动态观察培养7d中肝细胞活率、蛋白质和葡萄糖合成功能、G-6-Pase活性、安定转化功能及LDH漏出量。 结果:无血清培养的猪肝细胞各项指标除G-6-Pase活性在0,1,2d较低、葡萄糖合成功能在2,7d较低外,其余与含血清培养基培养的肝细胞无显著差异。肝细胞活率随着培养时间的延长而下降,但均高于88％;无血清培养的肝细胞的蛋白质合成功能在培养7d中保持稳定。安定转化功能在培养2,3d最强(安定浓度2d时为75±4pg·cell^(-1),3d时为77±5pg·cell^(-1);葡萄糖合成功能从1d时1.00±0.15nmol·cell^(-1)下降到2d时0.71±0.02nmol·cell^(-1);G-6-Pase活性从0d时1290±30zmol·cell^(-1)下降到1d时306±38zmol·cell^(-1),然后维持在较低水平;LDH漏出量在2,3d较高。AIM: To compare the effect of serum-free medium with serum-contained medium for culture of sucking pig hepatocytes and to search for a kind of better serum-free medium for bioartificial liver.METHODS: Suckling pig hepatocytes were isolated by modified two-step in situ collagenase perfusion method and cultured at 5 × 108·L-1 both in serum-free medium and serum-contained medium, respectively. Viability, protein and glucose syntheses, G-6-Pase activity, diazepam transformation of hepatocytes and LDH release were evaluated during 7 d of culture.RESULTS: There were no significant differences in above indices except for lower G-6-Pase activity on 0, 1, 2 d and glucose synthesis on 2,7 d. High viability ( > 88 %) was obtained though it declined with time. Protein synthesis of hepatocytes cultured in serum-free medium was stable over the course of 7 d. Diazepam transformation of hepatocytes was stronger on 2d and 3 d (diazepam concentration was 75 ×4 pg · cell-1 on 2 d and 77 × 5 pg · cell-1 on 3 d). Glucose synthesis of hepatocytes declined from 1.00 × 0.15 nmol· cell-1 on 1 d to 0. 71× 0. 02 nmol · cell-1 on 2 d. G-6-Pase activity of the hepatocytes declined apparently from 1290 × 30 zmol·cell-1 on 0 d to 306 × 38 zmol · cell-1 on 1 d, and it was maintained at low level for 7d. LDH release was higher on 2 d and 3d.CONCLUSION:The serum-free medium may be applied in the culture of porcine hepatocytes of bioartificial liver.

关 键 词：无血清培养基 培养 乳猪肝细胞 效果 

分 类 号：R329-33[医药卫生—人体解剖和组织胚胎学]