右归饮对激素性股骨头坏死大鼠体外诱导培养成骨细胞作用的研究

In vitro induction studies of YouGui drink(右归饮) on culture of steroid induced necrosis of femoral head rat osteoblast

目的:在右归饮对体外正常大鼠成骨细胞的增殖和分化作用研究基础上,进一步观察右归饮对激素性股骨头坏死大鼠体外诱导培育成骨细胞的增殖和分化以及基因表达的影响。方法:取体重为100～120g雄性SD大鼠20只,臀肌注射醋酸强的松龙49mg/kg·d,连用6d(造模),7d后处死,无菌条件下取造模大鼠骨髓基质细胞,诱导培养成骨细胞,然后随机分为4组:高、中、低浓度右归饮含药血清组(R1、R2、R3)和对照组(R4),分别添加高、中、低浓度右归饮含药血清及对照血清,继续培养72h后,进行成骨细胞增殖、碱性磷酸酶活性、骨保护素(OPG)和破骨细胞分化因子(RANKL)表达水平检测(实时荧光定量PCR检测),并进行统计学分析。结果:高、中浓度右归饮含药血清组(R1、R2)与对照组(R4)相比,成骨细胞增殖率显著升高(P<0.01),碱性磷酸酶活性显著升高(P<0.01),成骨细胞OPGmRNA的相对表达量显著升高(P<0.01),且与浓度剂量呈一定的正相关;成骨细胞RANKLmRNA显著降低(P<0.01)。低浓度右归饮含药血清组(R3)与对照组(R4)相比,成骨细胞增殖率、碱性磷酸酶活性以及OPG、RANKLmRNA相对表达量无统计学差异。结论:高、中浓度的右归饮含药血清对体外培养的SINFH大鼠成骨细胞的增殖和分化具有明显的促进作用,并可提高成骨细胞OPGmRNA的相对表达量而对其RANKLmRNA有抑制作用。右归饮含药血清对体外诱导培养成骨细胞的作用与剂量有一定相关性,在一定的浓度范围内促进成骨细胞的分化、增殖。

Objective ： On the basis of YouGui drink （右归饮） to act on the normal rats in vitro osteoblast proliferation and differentiation, further observe YouGui drink （右归饮 ） inductive effect for steroid induced necrosis of femoral head rats in vitro on cultivation of osteoblast proliferation and differentiation. Methods ：Prednisolone acetate 49 mg/kg-d were injected into gluteal of 20 male SD rats （body weight in 100-120 g）, continuously 6 days for model. After 7 days, under sterile conditions from the model rats bone marrow stromal cells were induced and cultured to osteoblast, and then randomly divided into 4 groups, respectively, add the high, medium and low concentration YouGui drink （右归饮 ） containing serum （group R 1, R2, R3 ） and control serum （group R4）. After 72 h, osteoblast proliferation, alkaline phosphatase activities （APA） were detected, and expression level of osteoprotegerin （OPG） and osteoclast differentiation factor （RANKL）were detected by real-time fluo- rescence quantitative PCR. Results：The high and medium concentration YouGui drink （右归饮） containing serum group compared with the control group （R 1, R2 vs R4）, the rate of osteoblast proliferation was significantly high （P〈0.01 ）, APA significantly increased （P〈0.01）,relative expression level of osteoblast OPG mRNA significantly increased （P〈0.01）, and with the concentration of a certain dose was positively correlated; RANKL mRNA significantly was lower （P〈0.01）. The low concentra- tion YouGui drink （右归饮） containing serum group compared with the control group （R3 vs R4）, there was no significantly different in the rate of osteoblast proliferation, ALP and relative expression level of osteoblast OPG mRNA, RANKL mRNA. Conclusion ： The high concentration of YouGui drink （右归饮） containing serum can obviously promote the proliferation and differentiation of steroid induced necrosis of femoral head rats in vitro osteoblast,and plays a clear role in promoting and enhance the relative expression of osteoblast OPG mRNA and an inhibitory effect on RANKL mRNA. The effect of YouGui drink （右归饮 ） containing serum acting on induced in vitro cultured osteoblasts have relevance with dose, at a certain concentration range it can promote osteoblast differentiation and proliferation.