Repairing Rabbit Thyroid Cartilage Defects With Transgenic Cell Sheets Of Cartilage-derived Morphogenetic Protein1

ObjectiveStructure transgenic cell sheets of cartilage-derived morphogenetic protein1, identifying the biological activity. Combined cell sheet technology and tissueengineering technology to explore the repairing effect of the new novelscaffold-free tissue engineering cell sheets on rabbit thyroid cartilage defects,hope to offer a new idea or method on cartilage reconstruction and regenerativemedicine.MethodsRabbit bone mesenchymal stem cells(BMSCs) were cultured andexpanded in vitro, isolated from bone marrow of1-month-old rabbit throughoptimizations (based on preliminary study).The4th generation cells weredivided into3groups: A group was transfected byadenovirus(Ad)-cytomegalovirus(CMV)-human CDMP1(hCDMP1)-internalribosome entry site(IRES)-enhanced green fluorescent protein(eGFP); B groupwas transfected by Ad-CMV-EGFP; C group was untransfected BMSCs. UsingMTT assay to detect the cell proliferation, and obtain the cell sheets withtemperature-responsive culture dish after14days’ continuous culturing.Detecting the express of CDMP1, glycosaminoglycan (GAG) and collagen typeΠ through histological staining, immunohistochemistry, Real time PCR andWestern blot.Manufacturing the model of rabbit thyroid cartilage defects undergeneral anesthesia, implanted into the three group cell sheets.In regularfeeding, make gross and histological observation after4or8weeks. ResultsThe BMSCs had an active proliferative ability in vitro with no change afterhCDMP1adenovirus-mediated gene transfer.The three-dimensional transgenicBMSCs cell sheets can be successfully harvested by temperature-responsiveculture dish with certain toughness and operability, can express collagen Π andGAG. Transplanted into the animal body, the BMSCs cell sheets transfected byCDMP1can promote and repair thyroid cartilage defects.In group A, thecartilage defect area had chondrocyte-like changes with a smooth surface, Alcian blue staining and collagen Π immunohistochemistry tested positive, wassignificantly higher in group A than that in groups B and C (P<0.05).Conclusions1. CDMP1transgenic cell sheet can be successfully harvested in vitro.2. The cell sheets can promote and repair rabbit thyroid cartilage defects.