Effects Of Major Components In Yunnan Baiyao On Bone Formation

Objectives To investigate the effects of Yunnan Baiyao on bone formation and tissue repair, comprehend the regulation of Yunnan Baiyao on osseous tissue mineralization, bone growth factor, pick out the obvious bone-formating element without poisonousness, originate new compatibility.Methods 1: SD rats marrow stroma cells (MSCs) isolation and cultivation: MSCs were isolated from the bone marow of SD rats (12 weeks old) sacrificed by decapitation, and then were cultured in at 37℃, 5%CO₂ humidified incubator in DMEM medium supplemented with 10% FBS. MSCs were digested by Trypsin and serial subcultivation were carried out. The third generetion were used in experiments. 2: The third generation MSCs inoculated in 6-well culture plates (2.5×10⁴,cm²) were cultured in LG-DMEM media supplemented with 5ug constituent of Yunnan Baiyao and the cells were harvested on 1, 3, 5, 7, 14 days after being induced. The proliferation of the stromal cells was observed to estimate drug toxicity, and alkaline phosphatase (ALP) staining and examination were done to check the differentiation ability of bone marrow cells. Alizarin red staining was performed to evaluate the mineralization of the osteoblasts in culture. Semiquantitative RT-PCR was carried out to detective the mRNA expression of BMP-2, BMP-4, BMP-7, ON, OPN, osterix, TGF-β1, BSP in the cells. Results Yunnan Baiyao inhibited the proliferation of bone marrow stromal cells compared with control group. Yunnan Baiyao enhanced the ALP activity of stromal cells. On 7th to 15th day, ALP activity was stronger in groups E, G, H, I and J than in control group ( P< 0. 05). Bone nodule formation was very obvious at 14th day in E, G, H, I and J groups. OPN mRNA and osterix mRNA in A group, BMP-2 mRNA and osteocalcin mRNA in B group, BMP-2 mRNA in C group, BMP-2 mRNA and BSP mRNA and osteocalcin mRNA in D group , BMP-2 mRNA, BSP mRNA in G group, TGF mRNA and BMP-2 mRNA and OPN mRNA in H group, TGF mRNA and BMP-2 mRNA and BSP in I group, TGF mRNA and BSP mRNA and BMP-2 mRNA in J group were higher than in the contrlo group.Conclusion This study suggests that Yunnan Baiyao may inhibit the proliferation, but stimulate the differentiation of osteoblasts derived from bone marrow stromal dells, accelerate the mineralization of osteoblasts and promote the maturation of bone nodules, and promote significantly mRNA expression of TGF-β1,BMP-2 and etc..