Abstract: Objective　To study the effects of two kinds of new collagen membrane (fish collagen and porcine collagen) on the osteogenic differentiation of SD rat bone marrow mesenchymal stem cells (rBMSCs) and observe their repair effects on the cranial parietal defects of SD rat. Methods　The surface morphology and water contact angle of the two membranes were measured by scanning electron microscopy (SEM) and contact angle measuring instrument. rBMSCs were cultured on membranes, and cells were also seeded on blank well plates as a blank control group. The effects of the two membranes on the proliferation of rBMSCs at 1, 3, 5 and 7 days were examined by CCK-8 and the adhesion and extension of the cells were observed by laser confocal microscopy at 24h. The osteogenic properties in vitro of the two membranes were evaluated by alkaline phosphatase (ALP) staining and activity assay, alizarin red staining and semi-quantitative determination of calcium nodules and real-time fluorescent quantitative PCR. In vivo experiments, bone defects with diameter of 5mm were prepared on both sides of the cranial suture of SD rats. The left bone defect area was implanted with fish collagen membrane or porcine collagen membrane. The right bone defect area was used as a blank control group. After 3 months, micro-CT was used to detect the bone regeneration in the cranial parietal defect area. Results　SEM: The surface of the fish collagen membrane was dense, and the porcine collagen membrane had a loose porous surface. Contact angle measuring instrument: The hydrophilicity of porcine collagen membrane was better than fish collagen membrane (P<0.05). CCK-8 and laser confocal microscopy: The cells spread well at 24h and stably proliferated within 7 days on both membranes and blank well plates. Osteogenic properties in vitro: ALP activity of rBMSCs on porcine collagen membrane at 5d, extracellular matrix mineralization at 14d, and the expression of cell-associated osteogenic genes Bmp2, Col1, Runx2 at 7d were higher than fish collagen group and blank control group (P<0.05). In vivo osteogenic experiment (3 months): Porcine collagen membrane promoted bone regeneration significantly better than fish collagen membrane and blank control group (P<0.05). There was no significant difference between fish collagen membrane group and blank control group. Conclusion　The effects of porcine collagen membrane on promoting bone regeneration in vivo and in vitro are significantly better than fish collagen membrane, and it has the potential as a membrane for guiding bone tissue regeneration.

Key words: Bone marrow mesenchymal stem cells, Collagen, GBR, Osteogenic differentiation, Bone regeneration