护骨胶囊对激素性股骨头坏死大鼠炎性因子的影响

Effect of HuGu capsule on inflammatory factors in rats with steroid induced osteonecrosis of femoral head

目的观察护骨胶囊对激素性股骨头坏死大鼠炎性因子的影响,探讨护骨胶囊作用于激素性股骨头坏死的可能机制。方法56只体质量200±20 g左右,不分雌雄SD大鼠随机分成4只空白对照组(CON组)和不同实验组52只。CON组大鼠正常饮食,余下52只大鼠进行脂多糖和甲泼尼龙琥珀酸钠(methylprednisolone,MET)注射造模。造模完成后,随机取其中4只大鼠作为模型组0天对照(MET-0 d组),与CON组同时处死取材;余下48只造模大鼠分成模型组(MET组),基础剂量护骨胶囊组(basal HuGu capsule group,HG1组)和双倍剂量护骨胶囊组(double HuGu capsule group,HG2组),每组大鼠随机分成2个时间组,分别进行42 d和60 d的灌胃处理后,取大鼠胸腺和脾脏计算免疫器官指数,酶联免疫吸附法(enzyme-linked immunosorbent assay,ELISA)测定大鼠股骨头关节囊内关节液的肿瘤坏死因子-α(tumor necrosis factorα,TNF-α)、白细胞介素-6(interleukin-6,IL-6)、白细胞介素-1β(interleukin-1β,IL-1β)、白细胞介素-33(interleukin-33,IL-33)的含量。结果(1)造模完成后检测MET-0 d大鼠发现,该组较空白组胸腺指数明显增加分别为(0.066±0.008)%vs.(0.106±0.017)%,P<0.01],脾指数下降分别为(0.160±0.014)%vs.(0.124±0.021)%,P<0.05]。与MET-0 d组比较,造模后生理盐水灌胃的模型组MET-42 d组和MET-60 d组的胸腺指数无变化,而脾指数明显上升分别为(0.169±0.018)%(P<0.01)和(0.177±0.020)%(P<0.01]。护骨胶囊给药一段时间后发现,与同时间点MET组比较,仅HG1-42 d组胸腺指数增加,为(0.143±0.019)%(P<0.05),而两剂量组60 d的胸腺指数无差异;与同时间点MET组比较,护骨胶囊给药组大鼠脾指数无差异。(2)ELISA检测股骨头关节液发现,与CON组相比,MET-0 d组关节液TNF-α、IL-6、IL-1β和IL-33均明显增加(均P<0.01)。与MET-0 d组比较,MET-42 d组和MET-60 d组关节液IL-6和IL-33均出现下降(均P<0.01)。护骨胶囊给药后发现,与同时间点MET组相比,HG1组和HG2组的关节液TNF-α、IL-6、IL-1β和IL-33均明显降低(均P<0.01),但不同剂量组同一时间点比较,以及同剂量给药不同时间点两组比较均无差异。结论护骨胶囊通过调节细胞免疫,减少炎性因子分泌,进而降低股骨头炎性损害,可有效防止激素性股骨头坏死进一步发展。

Objective To observe the effect of HuGu capsule on inflammatory factors in rats with steroid-induced osteonecrosis of the femoral head(ONFH),and to explore the possible mechanism of HuGu Capsule.MethodsFifty-six male and female SD rats weighing about 200±20 g were randomly divided into blank control group(CON group,n=4)and different experimental groups(n=52).The rats in CON group had normal diet,and the remaining 52 rats were injected with lipopolysaccharide and methylprednisolone(MET).After modeling,4 rats in the model group(named MET-0 d group)and CON group were killed at the same time;the remaining rats in the model group were divided into normal saline control group(MET group),basal dose HuGu capsule group(HG1 group),and double dose HuGu capsule group(HG2 group).Rats in each group were randomly divided into two time groups.After gavage for 42 and 60 days respectively,thymus and spleen were taken to calculate immune organ index,and enzyme linked immunosorbent assay(ELISA)was used to detect inflammatory factors in intra-articular fluid of rat femoral head joint,such as tumor necrosis factor-α(TNF-α),Interleukin-6(IL-6),interleukin-1β(IL-1β),and interleukin-33(IL-33).Results(1)after the completion of modeling,it was found that the thymus index increased significantly and the spleen index decreased in MET-0 d group compared with CON group.The thymus index and spleen index in the blank group were:(0.066±0.008)%and(0.160±0.014)%;The thymus index and spleen index in MET-0 d group were(0.106±0.017)%(P<0.01)and(0.124±0.021)%(P<0.05).Compared with MET-0 d group,the thymus index of MET-42 d group and MET-60 d group had no change,but the spleen index increased significantly.The spleen indexes of the model group,MET-42 d group and MET-60 d group were:(0.169±0.018)%(P<0.01)and(0.177±0.020)%(P<0.01),respectively.After administration of HuGu capsule for a period of time,compared with MET group at the same time point,only the thymus index in HG1-42 d group increased(0.143±0.019)%(P<0.05),but there was no difference between the two dose groups at 60 days.Compared with MET group at the same time point,there was no difference in spleen index of rats in HuGu capsule administration group.(2)ELISA detection of femoral head joint fluid showed that in intra-articular fluid of MET-0 d group,compared with CON group,TNF-α,IL-6,IL-1β,and IL-33 increased significantly(all P<0.01).Compared with MET-0 d group,IL-6 and IL-33 in intra-articular fluid of MET-42 d group and MET-60 d group were decreased(all P<0.01).After administration of HuGu capsule,it was found that compared with MET group at the same time point,TNF-α,IL-6,IL-1β,and IL-33 decreased significantly in intra-articular fluid of HG1 group and HG2 group(all P<0.01),but there were no differences between the two dose groups at the same time point and the same dose groups at different times.Conclusion HuGu capsule can effectively prevent the further development of steroid-induced ONFH by regulating cellular immunity,reducing the secretion of inflammatory factors,reducing the inflammatory damage of femoral head.