Hvdrostatic Pressure And TGF-β3Regulate The Chondrogenesis Of Rat Bone-marrow Derived Chondrogenesis Of Rat Bone-marrow Derived Mesenchymal Stem Cells In Alginate Beads

Objectives:Bioactive factors, such as TGF β3as well as mechanical factors i.e. hydraulicpressure, have been shown to induce and/or modulate chondrogenesis of bone marrow derived mesenchymal stromal cells(BMSCs). Since these factors are intracellularly transduced through different mechanisms, it is hypothesized that TGF β3and hydraulic pressure may act synergistically on chondrogenic differentiation of BMSCs. The purpose of this study was to investigate the effects of Hydrostatic pressure and TGF-β3on chondrogenesis employing rat Bone-Marrow Derived Mesenchymal Stem Cells(BM-MSCs).METHODS:BM-MSCsderived from the tibias and femurs of rats, were grown in α-modified Eagle medium supplemented with10%fetal calf serum, which were characterized through flow cytometry using specific antibodies. BM-MSCs encapsulated in alginate beads, were divided into four groups:control; transforming growth factor (TGF-β3)(with TGF-P3treatment); loading (with stimulation of Hydrostatic pressure loading); and TGF-β31oading (with TGF-β3treatment and loading stimulation) groups. In the loading experiment, specimens were subjected to100kPaby an in-house designed pressure chamber for4hours a day.Experiments were conducted for1,3, and7consecutive days. Cell differentiation was verified through histochemistry, quantitativereverse transcription-polymerase chain reaction (RT-PCR) analysis.RESULT:MSCs of passage2.3,4were positive(≥80%、≥98%、>98%)for CD29, CD90and negative(≤2.0%、≤0.4%、≤0.2%)for CD11and CD45. Staining of the extracellular matrix was positive. TGF-(33, loading, and TGF-β3loading exhibitedsignificantly higher levels of expressions of chondrogenicmarkers (collagen II and AGG) at the third and seventh day.TGF-β3loading exhibited higher collagen type II and AGG than medium with TGF—β3alone or loading at the third day.CONCLUSION:The used concentrations of TGF-β3and parameters for hydraulicpressure were able to induce and modulatechondrogenesis of rat BMSCs in some extend, but the co-effect of TGF-β3and hydraulicpressure is complicated, there are no synergistic effects were observed.