槐角苷及染料木素对成骨细胞生物学特性的影响

Effects of sophoricoside and genistein on biological characteristics of osteoblasts

目的:比较槐角苷及染料木素对成骨细胞生物学特性的影响。方法:成骨细胞由新生24hSD大鼠头盖骨分离得到。通过细胞增殖率测定、碱性磷酸酶活性测定、茜素红染色矿化结节形态计量方法,对比槐角苷和染料木素在体外对成骨细胞骨形成功能的影响。利用基因克隆和报告基因方法,检测槐角苷和染料木素对人骨保护素(osteoprotegerin,OPG)启动子活性的作用,观察两者对骨保护素基因转录活性的调节。结果:体外培养的成骨细胞在1和0.1μmol/L槐角苷的干预下,均显示增殖刺激作用(与对照组比较P<0.05)。10μmol/L染料木素使细胞增殖受抑制,但0.1μmol/L染料木素则显示增殖刺激作用(与对照组比较P<0.05)。槐角苷在实验的各种浓度中均能提高ALP活性,而染料木素只在0.1μmol/L浓度才能提高ALP活性。槐角苷在10、1和0.1μmol/L浓度对成骨细胞的矿化总面积较对照组分别提高了73%、138.6%和114.3%,而染料木素各个浓度下成骨细胞矿化总面积均减少。10、1和0.1μmol/L浓度槐角苷干预后检测报告β-半乳糖苷酶基因(LacZ)活性均增加,但染料木素仅在0.1μmol/L浓度对LacZ活性有增强作用(与对照组比较P<0.05)。结论:槐角苷能够促进成骨细胞的增殖、分化和矿化功能,与染料木素比较,不出现对成骨细胞成骨功能的抑制作用。而对于成骨细胞产生的破骨细胞抑制因子骨保护素,较染料木素有更强的上调作用。

Objective. To observe the effects of sophoricoside and genistein on the biological characteristics of osteoblasts. Methods： Osteoblasts were isolated from parietal bone of the newborn SD rats （less than 24 hours） and cultured in minimum essential medium （MEM）. The effects of different concentrations of sophoricoside and genistein （10, 1 and 0. 1 μmol/L） on the bone formation of osteoblasts in vitro were observed by cell proliferation rate. Differentiation of osteoblasts was evaluated by detecting the activity of alkaline phosphatase （ALP） using 4-nitrophenyl phosphate disodium salt （PNPP） method and by measuring the mineralized nodiform dyed by alizarin bordeaux. And the activity of osteoprotegerin （OPG） promoters of human beings was tested by using gene cloning and reporter gene. Results： Sophoricoside at 1 and 0.1μmol/L could promote the proliferation of osteoblasts （ vs control group, P〈0.05）. Genistein at 10 μmol/L could inhibit the cell proliferation, while genistein at 0.1μmol/L showed promotion to cell proliferation （ vs control group, P〈0.05）. The activity of ALP of osteoblasts was increased by sophoricoside at concentrations from 0. 1 μmol/L to 10 μmol/L, but genistein only at concentration of 0.1 μmol/L could make it. The total areas of mineralization of the osteoblasts in 10, 1 and 0.1μmol/L sophoricoside groups increased by 73%, 138. 6% and 114. 3% as compared with the control group, respectively, but those in different concentrations of genistein groups decreased. The activity of LacZ was increased by sophoriceside at concentrations from 0. 1μmol/L to 10μmol/L, as well as by genistein at concentration of 0.1 μmol/L. Conclusion： Sophoricoside can stimulate the proliferation, differentiation and mineralization of osteoblasts. And it shows no inhibition to bone formation of osteoblasts compared with genistein. Sophoricoside can also increase the activity of osteoprotegerin promoter and is more effective than genistein.