Experimental Of Three-dimensional Culture With Adipose-derived Stem Cells/Fibrin Gel Complex In Skeletally Mature Rabbit

Part I: Adult rabbit adipose-derived mesenchymal stem cell separation, Culture.Identification and chondrogenic inductionObjective Research the culturing characteristics of adipose-derived stem cells in vitiroto establish the method of isolation and culture and to explore can be induced to differentiateinto chondrocytesMethods Adipose was collected from the inguinal region of News Zealand rabbits andthe ADSCs were isolated with adherent method and lagans method and cultured iultured andpassages with high glucose DMEM medium containing9-10%fetal bovine serum. The formof cell growth observed under an inverted microscope.3.7cell line MTT determination oftheir added value. Line cartilage cells induced to differentiate. Observed inverted the growthof cells under the microscope. One week after induction of differentiation, cell Safranin Ostaining and RT-PCR of chondro-cyte type II collagen mRNA expression.Conclusion Primary fat-derived mesenchymal stem cells cultured2days after theadherent growth, showing the growth of spindle and polygonal cells,7or9days after cellfusion was almost a swirling arrangement. Cultured fat-derived mesenchymal stem cells invitro value-added faster growth is relatively stable.2to3days after induction of thechondrogenic cell aggregate. High-density clumps of cells positive Safranin O staining, notaggregate cells were weakly positive or negative. RT-PCR tests showed that the cells expressthe11collagen mRNA. Part II: Effect of different cell seeding concentrations on chondrogenicdifferentiation of fibrin gel with rabbit adipose-derived stem cellsin three-dimensional cultureObjective Mesenchymal stem cells in fibrin three-dimensional colloidal outsidedimensional culture of different planting density of adipose fat cells into cartilagedifferentiation capacityMethods Culture of adipose mesenchymal stem cells were2×10⁹L^-1,2×10^-10L^-1,2×10^-11L-1, grown in96-well plates containing fibrin glue in an inverted microscope andscanning electron microscopy. Chondrogenic induction3weeks after sampling, stained byhematoxylin-eosin staining, collagen immunohistochemistry, RT-PCR detectionConclusion Cells in each group three weeks after the induction of fibrin glue onadhesion is better, and the high density of2×10^-11L^-1Group of tightly packed cells, matrixformation of more cell adhesion to the growth of more fibrin within the scanning electronmicroscope, high-density group, The type II collagen-positive, a gradually increasing trendwith increasing planting density. The results showed that the RT-PCR, with the cell seedingdensity increased, type II collagen mRNA and proteoglycan expression are enhanced.