Objective: 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.
Methods: 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.
Results: 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).
Conclusion: 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.