| Objective:To research the influence of ALN and ICA on the ability to proliferate and to differentiate into osteoblast of Bone mesenchymal stem cells(BMSCs),observe the the influence of ALN and ICA on the expression of transforming growth factor-betal(TGF-β1) and bone morphogenetic protein-2(BMP-2) in the progress to promote MSCs differentiate into osteoblast and analyze the appoach of ALN and ICA to go into effect can provide experimental evidence to explain the mechanism of ALN and ICA in preventing and treating OP,feasible experimental exploration in establishing a platform of TGF-β1 and BMP-2 to screen the medicine of preventing and treating OP and a new thought of preventing and treating OP by traditional Chinese and western medicine.Methods:1.BMSCs from rabbits were isolated and fostered by using differential time adherent method method;2.To observe the growth,proliferate and morphologic characteristics of the original cells and their descendant by inverted phase contrast microscope day by day and darw their auxodrome.3.To get the well growing cells of third generation,and identify BMSCs by detecting surface markers and cell life cycle by morphology and flow cytometer;4.MTT method was used to confirm the most effective concentration of the ALN and ICA on the proliferation of BMSCs;5.The most effective concentration of the ALN and ICA on the differentiation of BMSCs into osteoblast was jurged by alkaline phosphatase(ALP);6.According to the different induced condition,MSCs were divided into 3 groups: control group,classic group(induced by the classic osteoblast-induced system) and drugs group(induced by the most effective concentration of drugs on differentiation into osteoblast).ALP and calcium nodes in each group were detected to indicate the osteoblast-formation function of different group; 7.ELISA was used to detect the expression of TGF-β1,BMP-2 of each induced group in the progress of BMSCs differenting into osteoblast to explain the mechanism of the drugs in inducing BMSCs differenting into osteoblast;8.To reach the conclusion by contrasting and analyzing the statistics methods.Result:1.The results of the surface antigens of BMSCs detected by flow cytometer showed the positive expressions of CD29 and CD44 and the negative expressions of CD34 and CD45;MSCs isolated from rabbit could be induced successfully into osteoblasts,chondrocytes and adipocytes;2.The most effective concentration of the ALN and ICA on the proliferation of BMSCs were 1×10-8mol/L and 1×10-9mol/L;and the most effective concentration of them on the differentiations of BMSCs into osteoblast were 1×10-8 mol/L and 1×10-9mol/L;the ALN(1×10-8mol/L) and ICA(1×10-9mol/L) could both increase the expression of osteoblast-indices which follwed the increase of the secretion of TGF-β1 and BMP-2;3.ALN and ICA which had been proved to possess the ability to induce BMSCs differenting into osteoblast in former study could both increase the expression of TGF-β1 and BMP-2.Conelution:1.The stable and uniformal BMSCs of rabbit could be obtained by differential time adherent method;2.The ALN and ICA could both promote the proliferation of the BMSCs and the differentiation of BMSCs into osteoblast;3.In the osteoblast-differentiati on progress,the induced groups of the drugs in this expriment all could increase the expression of TGF-β1 and BMP-2,which may be one of the mechanisms that could promote the differentiation of BMSC into osteoblast. |
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