The Impact of Ammonium Chloride-Based Erythrocyte Lysis Process on Banked Adipose-Derived Stem Cells

The safety of banked human adipose-derived stem cells (hADSCs) purified by 155 mM ammonium chloride (NH4Cl)-based erythrocyte lysis has not been evaluated. This study was conducted to determine the impact of NH4Cl-based erythrocyte lysis on the biological characteristics of cryopreserved hADSCs. Stromal vascular fractions (SVFs) were obtained from lipoaspirates and purified with NH4Cl-based erythrocyte lysis (lysis group) or without (nonlysis group). The hADSCs were freshly isolated (fresh group) from SVFs and/or cryopreserved for 2 weeks (cryo group). The morphologies, immunophenotypes, viability, apoptosis, and growth kinetics of each group were compared. The cell cycle and differentiation capacity assays were performed in both cryopreserved groups. All groups showed similar cell morphology, immunological phenotypes, and viability. However, the main effect of lysis and its interaction with cryopreservation were observed when early apoptosis was regarded as a dependent variable in two-way repeated-measures analysis of variance. After cryopreservation, significant growth retardation and S-phase fraction reduction were observed in lytic hADSCs compared with those in nonlytic hADSCs. No significant differences in the adipogenic and osteogenic differentiation capacities were found between the two groups. Although NH4Cl-based erythrocyte lysis did not affect the cell morphology, immunological phenotypes, viability, and adipogenic and osteogenic differentiation capacities of cryopreserved hADSCs, exposure to NH4Cl-based erythrocyte lysis or its synergistic action with cryopreservation may induce apoptosis and inhibit the proliferation and mitosis of cryopreserved hADSCs. These results indicate that NH4Cl-based erythrocyte lysis is not suitable for high-quality banked collection of hADSCs for future clinical applications. Further development of safe, convenient, and cost-effective purification methods of hADSCs is warranted.