The Study On Osteogenic Differentiation Capacity Of Mixed Culture Of Adipose-derived Mesenchymal Stem Cells And Bone Marrow Mesenchymal Stem Cells

Objective : To assess the osteogenic differentiation on the condition of co-culture of BMSC and ADSCMethod: Flow cytometry was used for identifying ADSC and BMSC obtained by adherent screening method and enzymatic digestion method. Oil red O was used to identify two cells after being induced into fat. Alkaline phosphatase and alizarin red staining were used in osteogenic induction group; toluidine blue staining was used in Chondrogenic induction group,SOX9 and type II collagen were tested in this group. Osteogenic induction group was divided into four sub-groups, group A: uninduced stem cells group; group B: BMSC osteogenic induction group; group C: ADSC osteogenic induction group; group D: mixed BMSC and ADSC osteogenic induction group(cell density wass 1.0×104/cm2 per group,BMSC vs ADSC=1:1 in group D).OCN and RUNX2 expression were tested in osteogenic induction group.Twenty Wista rats which were divided into experimental group and control group were used to make a model of bone defects. Experimental group: the above-mentioned osteogenic induction cells which implanted into support material were planted into bone defects in rats; Control group: nothing was planted into bone defects.RESULT: ADSC and BMSC cells could be induced into adipose cells under adipogenic induction conditions, oil red O stain demonstrated red, into osteoblast under condition of osteogenic induction with alkaline phosphatase(ALP) and Alizarin red staining showing flaky red, and into cartilage cells under chondrogenic induction conditions with toluidine blue staining showing in blue. In the osteogenic induction group, OCN in group b and c and the expression of RUNX2 are significantly higher in group a(P<0.05); OCN and RUNX2 expression were higher in group B than that of group C(P<0.05);OCN and RUNX2 expression in group d were significantly high compared with the other three groups(P<0.01). SOX9, type II collagen gene expression in two stem cells were significantly increased under the condition of chondrogenic induction.The implanted materials were histologically examined after planted into bone defects in rats one month and two months later respectively, Experimental group showed the generation of bone trabecular in the first month and bone laminar in the second month.The order of osteoblastic reaction was co-culture group,BMSC group and ADSC group.Conclusion: The ability of osteogenic potential strengthened to some extent under the condition of co-culturing. Osteoblasts which implanted into support material can promote bone healing by forming bone trabecular when planted into bone defects in rats.Compared with seperated induction, the osteogenic capacity was stronger by mixed induction.