Bone-like Extracellular Matrix Enhances Osteoblastic Differentiation of BMSCs

An in-vitro generated, bone-like extracellular matrix (ECM) secreted by cells has been shown to significantly enhance mineralized matrix deposition by rat bone marrow stromal cells (BMSCs). Objectives: The aim of this study was to investigate the effect of a bone-like ECM on markers of osteoblast terminal differentiation at the level of mRNA expression. Methods: To generate bone-like ECM, rat BMSCs were seeded onto electrospun poly(e-caprolactone) (PCL) scaffolds and cultured for 12 days to allow ECM deposition (PCL-ECM). Fresh BMSCs cells were seeded onto plain PCL and decellularized PCL-ECM scaffolds, cultured for 16 days in standard osteogenic media, and harvested at various time points. Gene expression was analyzed by real-time PCR using the following primer sets for alpha-1 chain of type I collagen, alkaline phosphatase, bone sialoprotein and osteocalcin. Furthermore, cell distribution and spreading within the scaffold and ECM formation was assessed by histologic analysis. Results: Real-time PCR analysis revealed an earlier onset and increased expression levels of osteoblast marker genes in cells after 16 days on the PCL-ECM scaffolds. Cells on these PCL-ECM scaffolds exhibited more matrix deposition over the course of culture compared to that of plain PCL as demonstrated by methylene blue/basic fuchsin and von Kossa staining. Conclusion: Bone-like ECM secreted by cells enhances and accelerates the osteoblastic differentiation of BMSCs by upregulating various osteoblast-specific genes. This research was supported by NIH grant R01 AR42639 to AGM and R01-DE013368 to RDS.