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知母皂苷元对成骨细胞活性和破骨细胞分化及功能的影响

Effects of sarsasapogenin on the activity of osteoblasts and the differentiation and the function of osteoclasts

  • 摘要: 目的 : 考察知母皂苷元(SAR)对体外培养成骨细胞活性和破骨细胞分化及功能的影响。 方法 : 培养小鼠胚胎成骨细胞MC3T3-E1,分别采用MTT法、碱性磷酸酶(ALP)试剂盒检测及茜素红染色法观察SAR对MC3T3-E1细胞增殖、ALP活性及矿化结节形成的影响;分别采用皮质骨来源破骨细胞分离及骨髓间充质干细胞诱导破骨细胞形成两种 方法 培养破骨细胞,利用抗酒石酸酸性磷酸酶(TRAP)阳性细胞计数及骨吸收陷窝计数来观察不同浓度SAR对成熟破骨细胞活性及破骨细胞诱导分化的影响。 结果 : 与对照组相比,各质量浓度SAR(0.01,0.1,1 μg/mL)对MC3T3-E1细胞均具有明显的促增殖作用(P<0.05,P<0.01);处于快速增殖的MC3T3-E1细胞给药处理后各组间ALP活性未见明显差异,但对于增殖晚期MC3T3-E1细胞,不同浓度SAR均可使ALP活性增高,其中1 μg/mL组效应最强,与对照组相比差异非常显著(P<0.01);连续给药处理细胞15 d,SAR各组矿化结节形成数量与对照组相比均有增多趋势。此外,各浓度SAR对成熟破骨细胞数量及骨吸收能力均无明显影响,但均能抑制骨髓细胞分化形成破骨细胞。 结论 : SAR能促进体外培养的成骨细胞的增殖与分化成熟;SAR对分化成熟的破骨细胞无明显影响,但可抑制骨髓细胞向破骨细胞的分化,从而减少破骨细胞的产生。

     

    Abstract: Aim :To observe the effects of sarsasapogenin(SAR) on osteoblasts and osteoclasts cultured in vitro. Methods :Colonal murine calvarial osteoblast-like cell line MC3T3-E1 cells were cultured in vitro.MTT,p-nitropheneye phosphate and tinctorial method of alizarin Bordeaux were used to investigate the effects of SAR on the proliferation,ALP expression,and mineralization tuberculation of MC3T3-E1 cells.Mature osteoclasts were isolated from the long bone of one-day rat.Meanwhile,marrow cells of mouse bone were cultured with induction of 1,25(OH)2VitD3.During the culturing of osteoclasts or marrow cells,SAR of different concentrations was added into the medium.The number of osteoclasts was recognized as tartrate resistant acid phosphatase(TRAP)(+) multinucleate cells and the resorption lacuna on bone slice were examined with toluidine blue staining. Results :Comparing with the control group,SAR (0.01,0.1,1 μg/mL) significanthy increased the proliferation of MC3T3-E1 cells (P<0.05,P<0.01).There was no significant difference in the expression of ALP in early proliferating MC3T3-E1 cells exposed to SAR of 0.01,0.1,1 μg/mL,but in the differentiation phase MC3T3-E1 cells,SAR improved ALP activity very significantly if compared with the control group,of which SAR of 1 μg/mL had the most promotion effect(P<0.01).In addition,compared to the control group,there were, to various extents,increased in the number of mineral nodes in MC3T3-E1 cells after 15day incubation with SAR of different conentrations.Furthermore,no obvious effects of 0.01-1 μg/mL SAR on mature osteoclast were observed.But typical osteoclasts were formed when marrow cells were cultured with the induction of 1,25(OH)2D3 in medium for 7 days while little or no osteoclasts were induced from marrow cells in the presence of SAR. Conclusion :The results suggest that SAR can effectively promote the proliferation,differentiation and mineralization of osteoblasts cultured in vitro.Besides,SAR can inhibit the generation of osteoclasts from marrow cells.

     

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