Culture of adipose-derived stem cells: a critical analysis

Abstract

Introduction: adipose-derived stem cells (ADSCs) are pluripotent and present little invasive collection and high cell yield in culture, besides the fact that the number of liposuctions, the main source of these cells, is increasing around the world. Plastic Surgery has found the benefits of the use of those cells in several areas, which increases the importance of critically analyzing the protocols currently used for cell cultivation. Objective: to describe in detail the cultivation protocol adopted by our laboratory in order to perform a critical analysis of the culture of ADSCs. Method: surpluses of adipose tissue were donated to the laboratory with research purposes for 5 patients of both genders and age between 20 and 45 years. From each patient, 30 mL of fatty material was collected in a sterile flask and transported to the laboratory for cell culture, where they were subjected during 45 min to enzymatic digestion by 30 mg of type IA collagenase diluted in 30mL of Dulbecco’s Modified Eagle Medium (DMEM) and then centrifuged to isolate the cells from the extracellular matrix. The amplification of the cultures was made with 0.05% trypsin solution + EDTA 0.02%. For cryopreservation was used freezing media consisting of 60% DMEM, 10% dimethylsulfoxide (DMSO) and 30% FBS. Results: The above protocol for growing ADSCs obtained a satisfactory cell culture, completing three cell passages in all samples, with subsequent freezing and thawing. Conclusions: The scientific literature indicates that collagenase get the best results in the extraction and isolation of ADSCs, paying attention to the type used. Regarding freezing, given the cytotoxicity of DMSO, new protocols with large samples replacing this cryoprotectant by trehalose should be tested, since the latter has produced good results and showed no toxic effects in published studies.