Effect of a cryopreservation protocol on the in vitro yield of adipose-derived stem cells

Background: Adipose tissue obtained by liposuction may be an accessible source of stem cells for future clinical application in tissue regeneration. Cryopreservation maintains stem cells in a live state for long periods of time, without impairing their function. The aim of this study was to assess the effect of a cryopreservation protocol on the proliferation of adipose-derived stem cells. Methods: Fragments of mouse adipose tissue were subjected to enzymatic digestion in order to isolate cells that were then cultured in minimum essential medium (α-MEM) supplemented with 10% fetal bovine serum (FBS). Cells were main tained at 37°C in 5% carbon dioxide (CO2). At the first passage, an aliquot of 1 × 10 6 cells was cryopreserved in FBS with 10% dimethylsulfoxide (DMSO) for 30 days, whereas the remaining cells were seeded and maintained in culture. When these cells reached the third passage, the 2 groups of cells (cryopreserved and non-cryopreserved) were seeded for experiments. Cell viability and proliferation curves were established at intervals of 24, 48, and 72 hours by counting cells with a hemocytometer and MTT assay. Nuclear morphology was assessed by DAPI staining. The data were statistically analyzed, with a significance level of 5%. Results: Increasing cellular proliferation was observed in both groups, with no significant difference throughout the experiment (P > 0.05). There was no significant change in cell viability and signs of nuclear damage were not detected in both the groups studied. Conclusions: The cryopreservation protocol analyzed was effective for maintaining the integrity of adipose-derived stem cells, allowing their storage for later use.