Senescent profile of cryopreserved adipose mesenchymal stem cells after multiple passages

Introduction: Human adipose derived mesenchymal stem cells have generated a great deal of excitement in regenerative medicine. The need for in vitro propagation to obtain therapeutic quantities of cells imposes a risk of impaired functionality due to cellular senescence. Therefore, this study analyzed the senescent profile of previously cryopreserved human adipose mesenchymal stem cells after multiple passages. Methods: Samples of this in vitro observational analytic study, which was performed from April–September 2016, were adipose mesenchymal stem cells that were cryopreserved in passage one in Stem Cell Integrated Medical Technology Unit RSCM-FKUI. After being recultured, the cells were cryopreserved for the second time. We compared cell size, viability, population doubling time (PDT) and senescence of once and twice cryopreserved cells. Cryopreservation groups were analyzed by independent t-test and ANOVA test for each passage analysis. Results: Senescent cells appeared at passage ten in the once- and passage nine in the twice-cryopreserved group, respectively. In terms of once and twice cryopreservation, there were significant differences in 30% confluence cell size, PDT, percentage of senescent cells, and the size of senescent negative cells at P10 (p < 0.001, p < 0.001, p < 0.001, p = 0.032), respectively. Meanwhile there were no differences in viability and the size of senescent positive cells at P10 (p = 0.98, p = 0.212), respectively. Conclusion: The adipose mesenchymal stem cells were senescent at the tenth passage and the ninth passage after once and twice cryopreservation, respectively.