辛伐他汀对大鼠骨髓基质细胞成骨分化的影响

Effect of simvastatin on osteoblastic differentiation of bone marrow stromal cells in rats

目的研究辛伐他汀体内给药后对大鼠骨髓基质细胞在体外培养过程中成骨分化和增殖的影响,探讨其刺激成骨的作用机制.方法30只SD雌性大鼠,随机分为3组,每组10只.G1组(C)为对照组;G2组(SIM-10)给予辛伐他汀10mg/(kg.d);G3组(SIM-20)给予辛伐他汀20mg/(kg.d).连续给药28d后,取大鼠的骨髓细胞体外培养,诱导14d后用RT-PCR和Westernblot分别检测cbfa1mRNA和蛋白表达的变化;收集上清液,检测细胞碱性磷酸酶;应用cellcountingkit(CCK-8)测定细胞增殖情况.结果辛伐他汀体内干扰28d后,再经过骨髓细胞体外培养诱导14d后,cbfa1因子的mRNA及蛋白表达水平均增高,呈剂量依赖关系.上清液碱性磷酸酶表达增高,呈剂量依赖关系.实验组与对照组比较,G2组(10mg)组促进细胞增殖,而G3组(20mg组)未见显著性差异.结论辛伐他汀可促进骨髓基质细胞中cbfa1mRNA和蛋白的表达,碱性磷酸酶活性增高,辛伐他汀刺激成骨的作用机制可能与此有关.

AIM： To study the effect on osteoblastic differentiation and proliferation of bone marrow stromal cells in vitro by administration of simvastatin in vivo and to elucidate the mechanism of the anabolic osteogenetic effect of simvastatin. METHODS： Thirty 3-month-old virgin female SD rats, weighting 250 -300 g were randomly divided into 3 groups： Group 1, control, 10 rats; Group 2, given simvastatin 10 mg/（kg · d）, 10 rats; Group 3, given simvastatin 20 mg/（ kg · d） , 10 rats. All of the animals were given the agents through gastric tube for 28 d. At 29th day all the rats were sacrificed. Bone marrow stromal cells in femur and tibia were cultured in vitro. After treated with same condition for 14 d ALP activity of bone marrow stromal cells in supernatant was determined. The mRNA level of cbfal was detected by RT-PCR, and cbfal protein expression was analyzed by Western blot. Cell count kit （ CCK-8 ） was used to examine the cell proliferation. RESULTS： After the rats were adminis-trated with different-dose simvastatin in vivo for 28 d, and then the bone marrow stromal cells were cultured for 14 d in vitro, the level of cbfal mRNA was increased, and the expression of cbfal protein also increased in a dose-dependent manner, and supernatant ALP activity increased in a dose-dependent manner. T-test showed that the proliferation of bone marrow stromal cells in Group 2 had significant difference, but no significant difference in Group 3, when compared with control group. CONCLUSION： Simvastatin leads to the high expression of cbfal in bone marrow stromal cells and increased ALP activity, which may be parts of the mechanisms underlying the anabolic osteogenetic effect of simvastatin.