Fundamental Steps for Mesenchymal Stem Cell Isolation for Experimental Research

Abstract

Background : Mesenchymal stem cells (MSCs) are undifferentiated cells that give rise to the mesodermal lineage. Adipose-derived MSCs are an easy and widely used source for MSCs isolation. In this study, adipose tissue was isolated and processed for MSCs isolation. MSCs’ proliferation, surface marker expression, in vitro differentiation potential, and polymerase chain reaction (PCR) results were evaluated by subculturing. Methods : Adipose tissue collected from a patient during spinal cord injury surgery was stored in PBS without shaking. First, the connective tissue was removed and the fat tissue was secured. Thereafter, the fat tissue was digested with collagenase type 1 at 37°C and 140 rpm for 1 h. After centrifugation, the remaining cell pellet was resuspended and filtered. Cells were cultured in Dulbecco’s modified Eagle’s medium supplemented with 10% FBS, fibroblast growth factor, and platelet-derived growth factor. Characterization analyses were performed to assess trilineage differentiation and marker expression using PCR and fluorescence-activated cell sorting. Results : Our results confirm that adipose-derived MSCs have a high proliferation rate. Additionally, marker gene expressions were confirmed by PCR. Evaluation of the surface marker expression of MSCs revealed positive expressions of CD73, CD90, and CD105, and negative expressions of CD14, CD34, and CD45. The MSCs showed differentiation potential into adipocytes, chondrocytes, and osteocytes in differentiation medium. Conclusion : s: MSCs can be isolated from the adipose tissue. Adipose-derived MSCs have adipose, chondrogenic, and osteogenic differentiation potential. The characterization and differentiation potential of MSCs are useful for evaluating their potential applications in various field of basic research, including pain research.

Keywords

• adipogenesis
• adipose tissue
• chondrogenesis
• mesenchymal stem cell
• osteogenesis