Satisfactory repair of damaged articular cartilage is still a challenge, while tissue engineering provides a promising strategy. Collagen-based hydrogels have been widely applied in cartilage tissue engineering due to their biocompatibility. In this study, type I collagen and type II collagen were selected to prepare physically crosslinked composite hydrogels by self-assembly of collagen, and the effects of their physicochemical properties on chondrocyte phenotype maintenance and extracellular matrix (ECM) secretion were investigated. First, the microstructure of hydrogels was observed by a scanning electron microscope, and the compressive modulus was measured by a dynamic mechanical analyzer. Then, chondrocytes were encapsulated in hydrogels and detected by Live/Dead staining. The secretion of ECM was qualitatively estimated by histological staining and quantitatively analyzed by sulfated glycosaminoglycans and DNA content detection. Finally, cartilage-specific gene expression was analyzed by quantitative real-time polymerase chain reaction analysis. The results showed that the microstructure and mechanical property of hydrogels were relevant to the composition of composite hydrogels. The compressive modulus of hydrogels improved with the increase of type I collagen content in the hydrogels. Chondrocytes could maintain their round or oval morphology and secrete cartilage-specific ECM in the four groups of hydrogels, but higher the compressive modulus of composite hydrogels, the more ECM secretion of chondrocytes.

Porous calcium phosphate ceramics (CaP ceramics) could induce ectopic bone formation which was regulated by various signal molecules. In this work, bone marrow mesenchymal stem cells (MSCs) were cultured on the surface of osteoinductive hydroxyapatite (HA) and biphasic calcium phosphate (BCP) ceramics in comparison with control (culture plate) for up to 14 days to detect the signal molecules which might be affected by the CaP ceramics. Without adding osteogenic factors, MSCs cultured on HA and BCP both expressed higher Runx2, Osterix, collagen type I, osteopontin, bone sialoprotein, and osteocalcin at various stages compared with control, thus confirmed the osteoblastic differentiation of MSCs. Later study demonstrated the messenger RNA level of bone morphogenetic protein 2 (BMP2) and BMP4 were also significantly enhanced by HA and BCP. Furthermore, Smad1, 4, 5, and Dlx5, the main molecules in the BMP/Smads signaling pathway, were upregulated by HA and BCP. Moreover, the higher expression of Smads and BMP2, 4 in BCP over HA, corresponded to the better performance of BCP in stimulating in vitro osteoblastic differentiation of MSCs. This was in accordance with the better osteoinductivity of BCP over HA in vivo. Altogether, these results implied that the CaP ceramics may initiate the osteoblastic differentiation of MSCs by influencing the expression of molecules in BMP/Smads pathway.

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