In vitro short term cartilage tissue engineering with oriented cartilage extracellular matrix scaffolds and bone marrow mesenchymal stem cells

姚军(Yao Jun，Orthop Research Inst，PLA Gen Hosp，Beijing 100853)…∥Chin J Orthop.－2010，30(12).－1217～1222

In vitro short term cartilage tissue engineering with oriented cartilage extracellular matrix scaffolds and bone marrow mesenchymal stem cells

姚军(Yao Jun，Orthop Research Inst，PLA Gen Hosp，Beijing 100853)…∥Chin J Orthop.－2010，30(12).－1217～1222

ObjectiveTo fabricate cartilage extracellular matrix(ECM)oriented scaffolds and investigate the attachment，proliferation，distribution and orientation of bone marrow mesenchymal stem cells(BMSCs)cultured within the scaffolds in vitro.MethodsCartilage slices were shattered in sterile phosphate-buffered saline(PBS)and the suspension were differentially centrifugated untill the micro-fiber of the cartilage extracellular matrix was disassociated from the residue cartilage fragments.At last the supernatant were centrifugated，the precipitation were collected and were made into 2% －3%suspension.U-sing unidirectional solidification as a freezing process and freeze-dried method，the cartilage extracellular matrix derived oriented scaffolds was fabricated.The scaffolds were then cross-linked by exposure to ultraviolet radiation and immersion in a carbodiimide solution.By light microscope and scan electron microscope(SEM)observation，histological staining，and biomechanical test，the traits of scaffolds were studied.After being labelled with PKH26 fluorescent dye，rabbit BMSCs were seeded onto the scaffolds.The attachment，proliferation and differentiation of the cells were analyzed using inverted fluorescent microscope.ResultsThe histological staining showed that toluidine blue，safranin O，alcian blue and anti-collagenⅡimmunohistochemistry staining of the scaffolds were positive.A perpendicular pore-channel structures which has a diameter of 100 μm were verified by light microscope and SEM analysis.The cell-free scaffolds showed the compression moduli were(2.02 ± 0.02)MPa in the mechanical testing.Inverted fluorescent microscope showed that most of the cells attached to the scaffold.Cells were found to be widely distributed within the scaffold，which acted as a columnar arrangement.The formation of a surface cells layer was found on the surface of the scaffolds which resembled natural cartilage.ConclusionThe cartilage extracellular matrix derived oriented scaffolds have promising biological，structural，and mechanical properties.27 refs，5 figs.

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