地黄活性成分梓醇对小鼠成骨细胞MC3T3-E1增殖、分化和矿化的影响

Effects of catalpol from Radix rehmanniae on proliferation,differentiation and matrix mineralization of MC3T3-E1 cells

目的检测地黄活性成分梓醇对成骨细胞株MC3T3-E1细胞增殖、分化和矿化的影响。方法制备不同浓度地黄活性成分梓醇提取液。以小鼠成骨细胞株MC3T3-E1作为药物筛选的细胞模型;用MTT法测定不同浓度的梓醇溶液的促细胞增殖作用;采用ALP活性和骨钙素定量检测分别观察不同浓度的梓醇溶液的促细胞分化作用;以Vonkos-sa钙化染色法了解不同浓度的梓醇溶液的促细胞钙化作用。结果梓醇在1×10-7～1×10-9mol·L-1浓度范围内培养24及48h促进成骨细胞株MC3T3-E1细胞增殖。梓醇在浓度1×10-5～1×10-6mol·L-148及72h提高成骨细胞株MC3T3-E1细胞内碱性磷酸酶的活性。梓醇在浓度1×10-5～1×10-6mol·L-1培养8及12d时能明显促进成骨细胞MC3T3-E1骨钙素合成和分泌。梓醇在浓度1×10-5～1×10-6mol·L-1培养19d时成骨细胞株MC3T3-E1细胞的矿化结节(mineralized bone nodular structure,MBNS)数目增多。结论梓醇可以提高成骨细胞株MC3T3-E1增殖和分化能力,梓醇可能是地黄治疗骨质疏松作用的活性成分之一。

Aim To investigate the effect of catalpol from Radix rehmanniae on the proliferation,differentiation and matrix mineralization of MC3T3-E1 cells in vitro.Methods Catalpol from Radix rehmanniae of different concentration preparations were extracted with ethanol（catalpol ethanol-extract）,respectively.A mouse osteoblast cell line MC3T3-E1 was used to determine the potency of catalpol.MTT were applied to determine proliferation of the cell treated by catalpol at different concentrations.Differentiating effects of the catalpol with different concentrations in the cell were evaluated through the examinations of alkaline phosphatase（ALP）activities and bone gla protein（BGP）levels.Von kossa staining method was used to demonstrate the effects of the catalpol on calcification of the cells.Results Catalpol at concentration from 1×10^-7 to 1×10^-9 mol·L^-1 for 24 hours and 48 hours effective promoted the proliferation of osteoblasts of MC3T3-E1 cells line.Catalpol at concentration from 1×10-5 to 1×10^-6 mol·L^-1 for 48 hours and 72 hours effective stimulated the activity of ALP of osteoblasts of MC3T3-E1 cells line.Catalpol at concentration from 1×10^-5 to 1×10^-6 mol·L^-1 for 8 days and 12 days effective increased the synthesis and secretion of bone gla protein（BGP） of osteoblasts of MC3T3-E1 cells line.Catalpol at concentration from 1×10^-5 to 1×10^-6 mol·L^-1 for 19 days effective increased the mineralized bone nodular structure number of osteoblasts of MC3T3-E1 cells line.Conclusion Catalpol could promote proliferation and differentiation of MC3T3-E1 cells in vitro.Catalpol may be one of effective monomer of Radix rehmanniae on treatment of osteoporosis.