Experimental Study On The Effect Of Simulated Microgravity On Repairing Articular Cartilage Defects By Chondrocyt E-Improved Fibrin Glue Bracket In Rabbits

Objective:To study the effects of using simulated microgravity by Rotating cell culture system(RCCS)on repairing defects of articular cartilage.The stabilized fibrin-chondrOcyte constructs were fibrin glue which were added in aprotinin and tranexamic acid.The stabilized fibrin -chondrocyte constructs were cultivated in vitro and proliferation of the cells on the bracket or in RCCS and degradation of bracket and secretion of extracellular matrix were observed.Methods 1.The chondrocytes from articular cartilage of 3 weeks old New Zealand rabbit were isolated and monolayer cultured,then the proliferation seed cells were seeded on the improved FG scaffold in RCCS or not.The stabilized fibrin-chondrocyte constructs were culltured and amplinedfor2 weeks in four-dimensional spatial in vitro.Gross and histological appearance,biochemical content(GAG),typeⅡcollagen immunohistochemical text of both group sampIes were assessed during two weeks culture.The degradated speeds of fibrin-chondrocyte constructs were evaluated in different periods.2.Two defects of articular cartilage were made in the trochlear groove of each new zealand rabbit.The defect of group A was filled with stabilized fibrin-chondrocyte constructs in RCCS and the defect of group B group was filled with stabilized fibrin-chondrocyte constructs without RCCS.The defect of group C was filled only with stabilized FG scaffold.The group D was left empty.The repaired defects were periodically examined grossly and histologically.Result:1.Using improved scaffold,the constructs can basically maintain the original size and shape after two weeks of culture.2.The cell multiplication time of the normal group spends 5.1 days.The cell multiplication time of the simulated microgravity group 4.5.days.2.In vitro,two weeks post cultured,there was significantly statistical difference of Glycosaminoglycan between two groups.3.Two weeks postoperatively,implanted cartilage blocks could be seen in all experimental groups and all blocks stayed in the defect of articular cartilage,but the borders were clearly identified and none of implanted blocks had connected with circumference.A little extravasated blood could be found in the group D.4.Six weeks postoperatively,untypical hyaline cartilage could be found in the bone morphogenic simulated microgravity group,and there were many cartilage-like cells, which arrayed regularly.The newly born cartilage has plenty toluidine blue staining male matrix,and few lymphocyte cells could be seen in some areas.The depth of all blocks reach as2/3—3/4 high as normal articular cartilage.Their surface were smooth.Such phenomena could also be seen in the group B,but the depth of all blocks only reach the high of 1/2—2/3 to normal articular cartilage.And the newly born cartilage of the group B had less toluidine blue staining male matrix than that of the group A;and the cartilage cells of the group B arrayed less regularly.And its surface looked smooth.In the group C,fibrous tissue covered defected areas,the surface of which were uneven and the borders were identified clearly.In the group D,the defected areas were covered by a thin sheet tissue mainly composed of fibrous scar and imflammation tissue.5.Twelve weeks postoperatively,the implanted blocks in the group A had tightly integration with vicinity cartilage,the defects of which were almost as high as normal cartilage.The newly born cartilage had nearly the same texture as the normal cartilage.The superficial layer cartilage cells had oval shape and the cartilage cells'major axis parallel with articular surface.The depth of the group B blocks reach the high of 2/3—1/2 to normal articular cartilage.There were some newly born cartilages.The newly born cartilage cells of the group B had more toluidine blue staining male matrix than before,and the defect surface looked smooth.In the group C,there were fibrous tissue covered defected areas,the surface of which were uneven and the borders were identified clearly.The defected areas the group D were covered by a thin sheet tissue mainly composed of fibrous scar and imflammation tissue.6.According to the Wakitani' s standard,the group A has the best plerosis effect on the repairing of articular cartilage defects compared with the other groups.It had been proved to have the significantly difference among all the control groups by statistics disposal.Conclusion:1.The results of this study demonstrated that the simulated microgravity by RCCS can improve the quality of tissue-engineered cartilage formed in vitro.2.The study showed that degradative speed of modified fibrin gel scaffold were kept up with cartilaginous tissue formed in vitro for a long duration,and it fitted for rotating culture,which maybe the suitable scaffold for cartilage tissue engineering.3.Cells can form big aggregates, because this condition simulated to the parent tissue condition in vitro,.with the optimization of the culture conditions,RCCS may be employed for the production of tissue-engineered cartilage and become a promising means for human cartilage reconstruction.