Investigating Cell Surface Markers and Differentiation Potential of Compact Bone-Derived Mesenchymal Stem Cells

Abstract

Background: The differentiation potential of mesenchymal stem cells (MSCs) derived from the bone-tissue to multiple lineages is not clear. Objective: This study was conducted to investigate the surface antigen expression and multilineage stem cell potential of the cells derived from culture of collagenase digested marrow-free compact bones of C57BL/6 mouse. Materials & Methods: Long bones of C57BL/6 mouse (n=6) were collected aseptically and bone marrow was flushed out. Collagenase-digested bone fragments were washed and cultured in plastic flasks. The plastic-adherent fibroblast-like spindle-shaped cells were cultured sequentially in multiple passages in low-glucose DMEM (Dulbecco's Modified Eagle's Medium) supplemented with 15% FBS (Foetal Bovine Serum) and antibiotics in a 37°C incubator with 5% CO 2 . Immu- nophenotyping for cell surface markers was done using flow cytometry. The cells were differentiated into the osteoblastic, adipogenic and chondrogenic lineages. Results: The culture of the adherent cells exhibited active proliferation and multiplication in consequent passages. The cultured cells revealed evidence of adipogenic and osteogenic differentiation confirmed by staining with oil red O and von Kossa stains. Under flow cytometry observation, a significant proportion of cultured cells expressed CD29 and stem cell antigen (Sca-1). Only 9.8% cells showed expression of CD105. These MSCs exhibited low ability in chondrogenic dif- ferentiation, which can potentially be attributed to their lack of CD105 expression. Lack of expression of CD45 showed evidence of absence of hematopoietic stem cells. Conclusion: This study showed that murine compact bone-chip culture can yield MSCs with significant proliferation ca- pacity. The cells displayed the ability to differentiate into osteoblast and adipocyte lineages.