龙牙楤木皂苷Ⅳ对原代成骨细胞分化和矿化的影响

Effect of Tarasaponin Ⅳ on differentiation and mineralization of primary osteoblasts

目的研究龙牙楤木皂苷Ⅳ(TarasaponinⅣ)对原代成骨细胞的分化及矿化的影响。方法通过消化SD乳鼠颅盖骨,获得原代前成骨细胞,体外诱导培养并鉴定;采用MTT比色法检测TarasaponinⅣ(10、20、30、40、50、60、70、80、90、100μg/mL)作用于前成骨细胞的适宜质量浓度范围;用低、中、高浓度(0.5、5、50μg/mL)TarasaponinⅣ对成骨细胞进行干预,培养4 d后,试剂盒法进行细胞上清中碱性磷酸酶(ALP)活性测定;培养14 d后,按茜素红S染色试剂说明对细胞进行钙化结节染色。结果与对照组比较,TarasaponinⅣ各浓度组均能明显提高前成骨细胞活性(P<0.05、0.01),0～60μg/mL内细胞生存率呈递增趋势。与对照组比较,TarasaponinⅣ低、中、高浓度组细胞上清ALP活性均显著下降(P<0.01),中、高浓度组细胞矿化结节数目显著增多(P<0.01),且均呈剂量相关性。结论 TarasaponinⅣ可以促进前成骨细胞生长,增强成骨细胞矿化过程。

Objectives To study the influence of TarasaponinⅣon the differentiation and mineralization of primary osteoblasts.Methods The craniums of male SD neonatal rats were digested to obtain primary pre-osteoblasts,and cells were then cultured in vitro and identified.The suitable concentration range of TarasaponinⅣ(10,20,30,40,50,60,70,80,90,100μg/mL)was determined by MTT colorimetry.The osteoblasts were respectively cultured with TarasaponinⅣ(0.5,5,50μg/mL).Alkaline phosphatase(ALP)activity in cell supernatant was determined by kit method after 4d of culture,and calcified nodules were stained by alizarin red S staining after 14 d of culture.Results Compared with the control group,all concentration groups of TarasaponinⅣcould significantly increase the activity of osteoblasts(P<0.05,0.01),and the cell survival rate showed an increasing trend in the concentration range of 0—60μg/mL.Compared with the control group,the ALP activities in the supernatant of TarasaponinⅣlow,medium,and high dose groups were decreased significantly(P<0.01),and the number of mineralized nodules in the medium and high concentration groups was increased significantly(P<0.01),in a dose-dependent manner.Conclusion TarasaponinⅣcan promote the proliferation of pre-osteoblast and enhance the mineralization process of osteoblasts.