三根汤含药大鼠血清对颗粒诱导的破骨细胞形成及其骨吸收功能的影响

Effects of rat serum containing Chinese herbal medicine Sangen Decoction on osteoclastogenesis and bone resorption of osteoclasts induced by polymethylmethacrylate particles

目的:研究三根汤含药大鼠血清在体外实验中对颗粒诱导的破骨细胞形成及其骨吸收功能的影响。方法:利用核转录因子-κB受体活化因子配体和巨噬细胞集落刺激因子共同诱导小鼠骨髓巨噬细胞分化成破骨细胞。将骨髓巨噬细胞同骨水泥即聚甲基丙烯酸甲酯颗粒按照1:3的比例分别加入至24孔板及装有骨片的96孔板中。50只SD雄性大鼠,随机分成5组,每组10只,分别给予不同药物后,同等条件下提取空白血清、西药(伊班膦酸钠)血清及高、中、低浓度中药(三根汤)血清并分别加入至相应组的培养液中。抗酒石酸酸性磷酸酶染色进行破骨细胞分化鉴定并计数,计算机图像分析技术测定骨片上骨吸收陷窝的面积。结果:加入含药血清组破骨细胞数量低于空白组(P<0.05),西药组与三根汤高剂量组破骨细胞数量低于三根汤中、低剂量组(P<0.05),且西药组与中药高剂量组两组间差异无统计学意义(P>0.05)。骨吸收陷窝实验显示空白组骨吸收陷窝面积大于其他组(P<0.05),西药组与三根汤高剂量组陷窝吸收面积小于三根汤中、低剂量组(P<0.05),且两组间差异无统计学意义(P>0.05)。结论:三根汤含药血清可以抑制颗粒诱导的破骨细胞的形成及其骨吸收功能。

Objective：To investigate the effects of Sangen Decoction,a compound Chinese herbal medicine,on osteoclastogenesis and bone resorption function of osteoclasts induced by polymethylmethacrylate particles in vitro.Methods：Macrophage colony stimulating factor （M-CSF） and receptor activator of nuclear factor-κB ligand （RANKL） were used to induce differentiation of bone marrow-derived macrophages （BMMs） towards osteoclasts. BMMs and polymethylmethacrylate particles with ratio of 1︰3 were added to the 24-well plate and 96-well plate with bone slices respectively. A total of 50 male SD rats were divided into 5 groups randomly with each group containing 10 rats. After being treated with different drugs,serum samples of rats in each group were extracted,i.e.,the blank serum,Western medicine （ibandronate） serum and high-,medium-,and low-dose Sangen Decoction serum and were added to the medium respectively. The tartrate-resistant acid phosphatase （TRAP） staining was used to identify the differentiation of BMMs and for counting of osteoclasts. Area of lacuna induced by osteoclast bone resorption on the bone slices was measured by computer image processing.Results：Numbers of osteoclasts of treatment groups were less than that of blank group by TRAP staining （P〈0.05）; numbers of osteoclasts of positive control group and high-dose Sangen Decoction group were much lower than those of medium-and low-dose Sangen Decoction groups （P〈0.05）,and no difference was found between Western medicine group and high-dose Sangen Decoction group （P〉0.05）. In bone resorption assay,area of lacuna of blank group was larger than those of treatment groups （P〈0.05）; areas of lacuna of Western medicine group and high-dose Sangen Decoction group were much smaller than those of medium-and low-dose Sangen Decoction groups （P〈0.05）,and no difference was found between Western medicine group and high-dose Sangen Decoction group （P〉0.05）.Conclusion：Sangen Decoction can inhibit osteoclastogenesis induced by polymethylmethacrylate particles as well as bone resorption function of osteoclasts