Increasing osteogenic differentiation capacity of bone marrow mesenchymal stem cells by design of osteo-mimetic culture environment

Trauma, fractures, infectious diseases or tumors can cause bone defects that are difficult to heal. The use of biocompatible artificial bone scaffolds with high porosity is a preferred method in tissue engineering studies for the treatment of various bone injuries. Three dimensional (3D) artificial bone scaffolds allow mimicking natural bone structure by providing a suitable support environment for osteogenic differentiation and tissueization of cells. Development of collagen/hyaluronic acid/chondroitin sulfate-hydroxyapatite COL/HYA/CS-HA polymer-ceramic composite scaffolds as a result of coating 1% collagen, 1% hyaluronic acid and 0.1% chondroitin sulfate polymer composites with hydroxyapatite mineral in SBF solution with biomimetic approach and bone marrow using mesenchymal stem cells (MSC). It was aimed to determine the ability to create texture. The scaffolds were activated with rat bone marrow-derived mesenchymal stem cells to determine the ability to form bone tissue under osteogenic induction with Dulbecco's Modified Eagle's Medium-Low Glucose (DMEM-LG) supplemented with bone morphogenetic protein (BMP). Samples were taken from the culture at weekly intervals to assess bone formation. Biocompatibility was tested by MTT kit analysis. Alkaline phosphatase activity was evaluated by alkaline phosphatase (ALP) analysis. In addition, bone tissue was followed by immunohistochemistry, histology and SEM analysis. The surface morphology of the hollow scaffolds and the mineralized extracellular matrix (ECM) structure after culture with MSC were clearly seen in the SEM analysis. According to the results of MTT analysis, mitochondrial activity increased partially from the 7th day and reached the highest level on the 28th day. Alizarin red staining demonstrated increased calcium deposition during MSC cultures on Col/HYA/CS-HA scaffolds. Alsian and Safranin-O staining showed proteoglycans and collagen fibers, respectively, in the ECM of bone-like tissue. Immunohistochemistry staining showed that bone-like tissue formation occurred at the end of culture and the MSC-initiated tissueization process proceeded in accordance with natural osteogenesis.