Functionalization of porous BCP scaffold by generating cell-derived extracellular matrix from rat bone marrow stem cells culture for bone tissue engineering

Abstract

The potential of decellularized cell-derived extracellular matrix (ECM) deposited on biphasic calcium phosphate (BCP) scaffold for bone tissue engineering was investigated. Rat derived bone marrow mesenchymal stem cells were cultured on porous BCP scaffolds for 3 weeks and decellularized with two different methods (freeze-thaw [F/T] or sodium dodecyl sulfate [SDS]). The decellularized ECM deposited scaffolds (dECM-BCP) were characterized through scanning electron microscopy, energy dispersive X-ray spectrometer, and confocal microscopy. The efficiency of decellularization was evaluated by quantifying remaining DNA, sulfated glycosaminoglycans, and collagens. Results revealed that F/T method was more effective procedure for removing cellular components of cultured cells (95.21% DNA reduction) than SDS treatment (92.49%). Although significant loss of collagen was observed after decellularization with both F/T (56.68%) and SDS (70.87%) methods, F/T treated sample showed higher retaining amount of sulfated glycosaminoglycans content (75.64%) than SDS (33.28%). In addition, we investigated the cell biocompatibility and osteogenic effect of dECM-BCP scaffolds using preosteoblasts (MC3T3-E1). Compared to bare BCP scaffolds, dECM-BCP_F/T scaffolds showed improved cell attachment and proliferation based on immunofluorescence staining and water soluble tetrazolium salts assay (p < .001). Moreover, dECM-BCP scaffolds showed increased osteoblastic differentiation of newly seeded preosteoblasts by up-regulating three types of osteoblastic genes (osteopontin, alkaline phosphatase, and bone morphogenic protein-2). This study demonstrated that functionalization of BCP scaffold using cell-derived ECM could be useful for improving the bioactivity of materials and providing suitable microenvironment, especially for osteogenesis. Further study is needed to determine the potential of dECM-BCP scaffold for bone formation and regeneration in vivo.

Keywords: BCP; SDS; decellularization; extracellular matrix; freeze-thaw; rat bone marrow derived mesenchymal stem cells.