Biological Characteristics Of Rabbit-derived Bone Marrow Mesenchymal Stem Cells And Subcutaneous Bone Formation By Autologous Implantation

Objective To study biological characteristics of rabbit-derived bone marrow mesenchymal stem cells (BMSCs), osteoinductive characteristics of dexamethasone(DEX) for BMSCs and the feasibility of tissue engineered bone which combined BMSCs with 10% coral hydroxyapatite (CHA) become one of the materials to repair the bone defects.1. The study of biological characteristics and osteoinductive of rabbit-derived bone marrow mesenchymal stem cellsMethods BMSCs was isolated from the rabbit marrow using density gradient centrifugation with 1.073g/ml lymphocyte separation media, then plastic adherence in culture. The second subculture BMSCs was treated with different DEX density from 10^-6mol/L, 10^-7 mol/L, 10^-8 mol/L to 10^-9mol/L, no DEX would be control group (n=5) , the proliferation of BMSCs were detected with MTT methods after 1, 3, 5 and 7 days. After 1, 2, 3 and 4 weeks, the osteoinductive symbols of BMSCs were detected, the expression of ALP with quantificational method, collagen-I with immuno-histochemistry method and mineralized nodes with acheomycin sign. BMSCs were cultured for 1, 2, 3, 4, 5, 6 and 7 days in no-serum low-alimentation environment, the proliferation results were gotten with MTT method after been recultured for 3 days (n=4) and 7 days (n=3) in 13% serum DMEM, no hunger would be control group.Results BMSCs was spindly which isolated from bone marrow after density gradient centrifugation and plastic adherence in culture. The results of BMSCs'proliferation, which was the highest after treated with density 10"6mol/L of DEX after one day, there were prominence differences with any one of other groups. After 3 and 5 days, the results of the control group was the lowest, there were prominence differences with any one of other groups.After 7 days, the results with density 10"9mol/L of DEX was the highest, there were prominence differences with any one of other groups. The ALP expressions were prominence hoisted after 4 weeks, the groups with density lO^mol/L and 10"9mol/L of DEX were the most prominence. The electropositive cells of collagen-I with immuno-histochemistry method would be seen in the groups with density 10'8mol/L and 10⁹mol/L of DEX after 4 weeks, which was brown in cytoplasm. After 5 weeks, mineralized nodes can be seen, which was golden in acheomycin sign. After recultured for 3 days, there was P<0.05 between control group and any one of trial groups too, P<0.05 between hunger one day group and other groups. After recultured for 7 days, there was P>0.05 between groups with exception of between group a and group h.Conclusions (1) Bone marrow is one of good sources of mesenchymal stem cells. BMSCs can be isolated from rabbit bone marrow with density gradient centrifugation, and have great ability of proliferation and multipotent. (2) BMSCs can be accelerated to osteoinductive under DEX combined with sodium 6-glycerophosphate lOmmol/L and L-ascorbic acid 50mg/L, the action are most in DEX density 10"9mol/L and 10'8mol/L. (3) DEX can protect and accelerate BMSCs to proliferation. (4) BMSCs can be able to endure low-nutrition and still have great ability of proliferation by went into inhibiting state low-consumedly.