The Influences Of Fluid Shear Stress Along With Growth Factor/Matrix Stiffness On The Differentiation Of Bone Marrow Mesenchymal Stem Cells

Bone marrow stromal cells(BMSCs) is a multi-directional differentiation potential stem cells with the potential to differentiate into osteoblasts, chondrocytes, fat cells, and so on. Research shows that mechanical stimulation, growth factors and the matrix hardness can regulate the differentiation of BMSCs, in which mechanical stimulation is proved to be an effective regulatory factor. Therefore, we adopted the mechanical loading belong with growth factor/matrix hardness on BMSCs at the same time, detected the distribution of osteogenic cell differentiation markers alkaline phosphatase(ALP) and chondrogenic differentiation markers glycosaminoglycans(GAG), to examine the influence of mechanical stimulation, growth factors and cell matrix hardness on the differentiation of BMSCs.The main research method, content and the results are as follows:(1) Used the whole bone marrow suspension adherence method to obtain BMSCs, second or third generation cells were used for experiments. The control group is static cultured BMSCs, the experimental group was vaccinated on two kind of matrixs for 24 h or 48 h, and later apply two increases linearly speed fluid shear stress(? SS) for 20 min. We made two different hardness polyacrylamide hydrogel(respectively 34 KPa and 80 KPa). Two kind of fluid shear stress were applied to BMSCs after inoculated 24 h or 48 h, ? SS is starting from 0 min after standing at 0 min and 2min from a linear velocity 0 dyn/cm2 to 10dyn/cm2(respectively 0-0'and 0-2'). After loading, BMSCs were cultured 48 h in a static state, detected ALP activity and GAG expression.Results of 24 h are as follows: ALP vitality value:(80,0-2') >(34,0-2') >(34,0-0') >(80,0-0'), in addition to last two groups, there are significant differences(P<0.05); GAG expressed quantity:(34,0-0') >(80,0-0') >(80,0-2') >(34,0-2'), and there are significant differences(P<0.05). As can be seen from the above results, fluid shear stress have a greater impact on BMSCs differentiation. Results of 48 h are as follows, ALP vitality value:(80,0-2') >(80,0-0') >(34,0-2') >(34,0-0'), in addition to the two groups behind, there were significant differences(P<0.05); GAG expressed quantity:(34,0-0') >(34,0-2') >(80,0-0') >(80,0-2'), we can see that a culture of 48 h on two kind of matrixs have a greater impact on BMSCs differentiation than FSS.(2) The control group is static cultured BMSCs, the experimental group with growth factor-induced differentiation for 3d, 5d or 7d, induction was divided into two directions, which are Osteogenic and Chondrogenic(referred to as O and C), later applied two increases linearly speed fluid shear stress(? SS) for 20 min. The results are as follows: after induction of 3d, ALP results are as following:(O,0-2') >(C,0-2') >(O,0-0') >(C,0-0'), GAG expression(C,0-0') >(O,0-0') >(C,0-2') >(O,0-2'), we can see that ALP vitality value(C,0-2') >(O,0-0'), GAG expression(O,0-0') >(C,0-2'); After 5 d differentiation after loading, the vigor value of ALP :(O,0-2') >(C,0-2') >(O,0-0') >(C,0-0'), in addition,(C,0-2') and(O,0-0') groups exist significant difference(P<0.05), GAG expression quantity:(C,0-0') >(C,0-2') >(O,0-0') >(O,0-2'), in addition to(C,0-2') and(O,0-0'), there were significant differences in the rest of group(P<0.05); ALP vitality values induction of 7d detected as follows:(O,0-2') >(O,0-0') >(C,0-2') >(C,0-0'), GAG expression in the following order(C,0-0') >(C,0-2') >(O,0-0') >(O,0-2'), two groups has the opposite effect ALP quantization values are as follows:(O,0-0') >(C,0-2'), GAG expression(C,0-2') >(O,0-0').Comprehensive the above results, we can know that: the fluid shear stress belong with matrix hardness or growth factor can regulate the bone and cartilage to differentiation of BMSCs, 20 min of different speed increase of fluid shear stress is an effective regulator.