Proliferation and differentiation potential of cryopreserved human skin-derived precursors

Abstract

Skin-derived precursors are recognized to be a potentially autologous and accessible source of neural precursor cells for drug screening or cell-based treatments, in many neurological disorders. Thus, it is necessary to investigate appropriate methods for cryopreservation of such human skin-derived precursors (hSKPs). The aim of this study was to evaluate different cryopreservation techniques for retention of hSKPs to discover an optimized protocol. We cryopreserved hSKPs treated with 0%, 10%, 20%, 30% and 40% foetal bovine serum (FBS) and three concentrations of dimethylsulphoxide (DMSO) 5%, 10% and 15%, with two different storage periods in liquid nitrogen (2 days: short-term storage; and 2 months: long-term storage). Then, we assessed survival and proliferation levels of the cells after freeze-thaw processes, by viability measurement and colony-forming assay. For detecting hSKPs, we used immunocytochemistry and RT-PCR assessments. Our findings indicated that hSKPs cryopreserved in 5% DMSO without FBS, had better survival and proliferation potentials compared to other working formulations. With various concentrations of cryoprotectants over different time periods, hSKPs retained their differentiation potentiality and were able to differentiate into neurons (NFM and βΙΙΙ tubulin-positive), glial cells (GFAP-positive) and smooth muscle cells (SMA-positive). Results revealed that in only 5% DMSO, hSKPs could be cryopreserved for long-term storage with considerable survival and proliferation levels, without losing multipotency.