Wang Yan, Zhu Ying-qian, Lin Jing-ming, Zhang Zhong-yi, Gao Yi
The Southern Medical University Affiliated Zhujiang Hospital, Guangzhou 510282, China.
Zhong Yao Cai. 2007 Dec;30(12):1551-4.
To observe the influence of Astragalus membranaceus Extraction (AE) on the primary-cultured human fetal hepatocytes stored in liquid nitrogen and explore a new method for the cryopreservation of human hepatocytes with improved function.
Human fetal hepatocytes were harvested by two-step collagenase perfusion, and then stored in a liquid nitrogen for one month with five different cryoprotectants (I: 10% DMSO, II: 5% DMSO + 2 mg/L AE, III: 5% DMSO + 20 mg/L AE, IV: 5% DMSO + 60 mg/L AE, V: 5% DMSO + 100 mg/L AE). One month later, the cells were thawed rapidly and the viability, morphology and basic function of them were tested.
The human fetal hepatocytes in different groups showed various levels of viability, morphological manifestation and cell function respectively. After thawing, the viability rate and flash adhering rate in group IV and V had no significant difference with group I (P > 0.05), but were higher than group II and III (P < 0.05); the cell function analysis in the group IV, the results of ALB and AST level determination, NH4Cl transformation test, were the best among the groups (P < 0.05).
AE can provide protection for human fetal hepatocytes in cryopreservation, and the best performance concentration level of its is 60 mg/L; the preservation dosage of DMSO can be reduced when combined with AE in the preservation solution, which shows that AE has a synergistic effect with DMSO.
观察黄芪提取物（AE）对液氮保存的原代培养人胎肝细胞的影响，探索一种能改善功能的人肝细胞冷冻保存新方法。
采用两步胶原酶灌注法获取人胎肝细胞，然后用五种不同的冻存保护剂（I：10%二甲基亚砜，II：5%二甲基亚砜+2mg/L AE，III：5%二甲基亚砜+20mg/L AE，IV：5%二甲基亚砜+60mg/L AE，V：5%二甲基亚砜+100mg/L AE）在液氮中保存1个月。1个月后，迅速解冻细胞并检测其活力、形态和基本功能。
不同组的人胎肝细胞分别表现出不同程度的活力、形态表现和细胞功能。解冻后，IV组和V组的活率和贴壁率与I组无显著差异（P>0.05），但高于II组和III组（P<0.05）；IV组的细胞功能分析，即白蛋白和谷草转氨酶水平测定、氯化铵转化试验结果在各组中最佳（P<0.05）。
AE能在冷冻保存中对人胎肝细胞提供保护，其最佳作用浓度水平为60mg/L；在保存液中与AE联合使用时可降低二甲基亚砜的保存用量，表明AE与二甲基亚砜具有协同作用。
