Effect of holding time, temperature and different parenteral solutions on viability and functionality of adult bone marrow-derived mesenchymal stem cells before transplantation

Abstract

Mesenchymal stem cells (MSCs) have the ability to proliferate and differentiate into various lineages, given the appropriate microenvironment, thus making MSCs promising candidates for cell transplantation. For clinical applications, MSCs need to be stored in optimal conditions so that they may be transported and made available as an off-the-shelf product for companies to market. Freshly harvested and cultured or frozen-thawed bone marrow-derived MSCs were prepared for cell transplantation. Both freshly cultured or frozen-thawed MSCs were washed and resuspended in parenteral solutions, either 0.9% saline, Dulbecco's phosphate-buffered saline (DPBS), plasmalyte A or 5%dextrose and held for 2, 4, 6 and 8 h at 4 degrees C, 37 degrees C and RT (22 degrees C). The viability of the cells, differentiation capability and expression of cell surface markers were analysed. MSCs harvested from fresh cultures, resuspended in the parenteral solutions and maintained at 4 degrees C for 6 h showed more than 90% viability, and the viability was appreciably better when suspended in 5% dextrose at 4 degrees C for 8 h. In contrast, frozen-thawed cells can be held for a maximum of 2 h after thawing before losing their viability significantly below permissible limits for transplantation. We are reporting for the first time the effect of various parenteral solutions, holding times and temperatures on the viability and functionality of bone marrow-derived freshly cultured or frozen-thawed MSCs for transplantation. Our results suggested that freshly harvested MSCs can be held for 8 h at 4 degrees C in 5% dextrose or for up to 6 h at 4 degrees C in saline, DPBS or plasmalyte A. Freeze-thawed MSCs can be held for a maximum of 2 h in plasmalyte A before transplantation without affecting their viability and ability to differentiate.