Failure of Y-27632 to improve the culture of adult human adipose-derived stem cells

Abstract

Nuno Jorge Lamas,1–3 Sofia C Serra,1,2 António J Salgado,1,2 Nuno Sousa1,2 1Life and Health Sciences Research Institute (ICVS), School of Health Sciences (ECS), University of Minho, Braga, Portugal; 2ICVS/3B’s-PT Government Associate Laboratory, Braga/Guimarães, Portugal; 3Clinical Pathology Department, Centro Hospitalar do Alto Ave (CHAA), EPE, Guimarães, Portugal Abstract: Y-27632 is a well-known inhibitor of the Rho-associated coiled kinase (ROCK) and has been shown to significantly improve the culture of a variety of multipotent stem cell types. However, the effects of Y-27632 on the expansion of adult human adipose-derived stem cell (hADSC) cultures remain to be established. Here, we aimed to characterize the effects of Y-27632 on the culture of hADSCs. Adult hADSCs were isolated from subjects submitted to elective plastic surgery procedures and cultivated in vitro under optimized conditions. Our results show that the continuous supplementation of hADSC cultures with Y-27632 led to decreased numbers of cells and decreased global metabolic viability of hADSC cultures when compared with control conditions. This effect appeared to be dependent on the continuous presence of the drug and was shown to be concentration-dependent and significant for 10 µM and 20 µM of Y-27632. Moreover, the Y-27632-induced decrease in hADSC numbers was not linked to a block in global cell proliferation, as cell numbers consistently increased from the moment of plating until passaging. In addition, Y-27632 was not able to increase the number of hADSCs present in culture 24 hours after passaging. Taken together, our results suggest that, in contrast to other stem cell types, Y-27632 supplementation is not a suitable strategy to enhance hADSC culture expansion. Keywords: human mesenchymal stem cells, human multipotent stromal cells (hMSCs), ROCK inhibitor, MTS assay