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Performance study of two new collagen membranes for guiding bone tissue regeneration in vivo and in vitro
Author(s): WANG Lili, YAN Jia, LI Dongsheng, MO Xiumei, HU Xiaokun, ZHANG Feimin, LIU Mei, Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Department of Prosthodontics, Affiliated Hospital of Stomatology, Nanjing Medical University
Pages: 481-
487
Year: 2019
Issue:
6
Journal: Stomatology
Keyword: bone marrow mesenchymal stem cells; collagen; GBR; osteogenic differentiation; bone regeneration; SD rats;
Abstract: Objective To study the effects of two kinds of new collagen membranes(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 24 h. 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 5 mm 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 24 h and stably proliferated within 7 days on both membranes and blank well plates. Osteogenic properties in vitro: ALP activity of rB MSCs on porcine collagen membrane at 5 d,extracellular matrix mineralization at 14 d,and the expression of cell-associated osteogenic genes Bmp2,Col1,Runx2 at 7 d were higher than fish collagen membrane 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.
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