山萘酚联合戊巴比妥钠缓解骨关节炎大鼠炎性反应和神经根疼痛的效果观察

Combination of Kaempferol and Pentobarbital Sodium Inhibits Inflammatory Response and Relieves Radiculopathic Pain in Osteoarthritis Rats

目的探讨山萘酚联合戊巴比妥钠对骨关节炎(osteoarthritis,OA)大鼠炎性反应和神经根疼痛的影响。方法选择56只SD大鼠随机分为健康对照组10只、山萘酚组10只、OA模型组12只、戊巴比妥钠组12只、联合治疗组12只。利用4%木瓜蛋白酶和0. 3 mol/L半胱氨酸诱导OA模型。造模成功后,山萘酚组大鼠腹腔注射山萘酚20 mg/kg每日1次;戊巴比妥钠组大鼠腹腔注射戊巴比妥钠30 mg/kg每日1次;联合治疗组大鼠腹腔注射戊巴比妥钠30 mg/kg每日1次和山萘酚20 mg/kg每日1次;健康对照组和OA模型组则注射0. 9%氯化钠注射液每日1次。苏木素伊红染色观察各组大鼠膝关节软骨组织病理变化;采用蛋白印记分析检测各组大鼠MMP-13、Caspase-3、Ki67、P-p65/p65水平;采用酶联免疫吸附实验检测各组大鼠白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、诱导型一氧化氮合酶(i NOS)和白细胞介素-10(IL-10)的表达;给药25 d后检测各组大鼠机械性撤足阈值。结果山萘酚联合戊巴比妥钠可减轻OA大鼠膝关节软骨组织表面增生,过渡层细胞排列紊乱等病理改变。与健康对照组相比,OA模型组MMP-13、Caspase-3、IL-6、TNF-α、i NOS及P-p65/p65比值升高,Ki67、IL-10和机械性撤足阈值下降(P <0. 01)。山萘酚组MMP-13、Caspase-3、Ki67、IL-6、TNF-α、i NOS、IL-10水平、P-p65/p65比值及机械性撤足阈值与健康对照组比较无明显差异(P> 0. 05)。与OA模型组相比,戊巴比妥钠组MMP-13、Caspase-3、IL-6、TNF-α、i NOS水平及P-p65/p65比值降低,Ki67、IL-10水平和机械性撤足阈值升高(P <0. 01)。与戊巴比妥钠组相比,联合治疗组MMP-13、Caspase-3、IL-6、TNF-α、i NOS及P-p65/p65比值下降,Ki67、IL-10水平和机械性撤足阈值升高(P <0. 01)。结论山萘酚联合戊巴比妥钠可减轻OA大鼠膝关节软骨组织病变,缓解炎性反应和神经根疼痛。

Objective To explore the effect of combination of kaempferol and pentobarbital sodium on the inflammatory response and radiculopathic pain in osteoarthritis （OA） rats. Methods Fif-y-six rats were randomly divided into five groups, including healthy control （Ctrl） group （n = 10）, kaempferol （KF） group （n = 10）, OA model group （ n = 12 ）, pentobarbital sodium treatment （ OA ＋ PS ） group （ n = 12 ） and combination treatment （ OA ＋ PS ＋ KF） group （n = 12）. OA model was induced by 4% papain and 0.3 mol/L cysteine. After successful modeling, the rats in KF group were intraperitoneally injected with kaempferol 20 mg/kg once a day; the rats in OA ＋ PS group were intraperitoneally injected with pentobarbital sodium 30 mg/kg once a day. Rats in combination treatment group were intraperitoneally injected with pentobarbital sodium 30 mg/kg once daily and kaempferol 20 mg/kg once daily; Ctrl group and OA model group were injected with 0.9% sodium chloride once daily. Patho- logic changes of articular cartilage tissues in each group were observed by hematoxylin-eosin （HE） staining. The prorein levels of MMP-13, Caspase-3, Ki67, and P-p65/p65 were detected by western blot. The expressions of interleu- kin （IL） -6, tumor necrosis factor-or （ TNF-ot）, inducible nitric oxide synthase （iNOS） and IL-10 were measured by enzyme-linked immunosorbentassay （ELISA） in all groups. Mechanical withdrawal threshold was tested 25 days after treatment. Results The pathologic changes of articular cartilage tissues were alleviated by combination of kaempferol and pentobarbital sodium. Compared with Ctrl group, the levels of MMP-13, Caspase-3, IL-6, TNF-ot, iNOS and the ratio of P-p65/p65 in OA model group were increased, with decreased levels of Ki67, IL-10 and the nlechanical withdrawal threshold （P 〈0.01）. The levels of MMP-13, caspase-3, Ki67, IL-6, TNF-ot, iNOS, IL-10, the ratio of P-p65/p65 and the mechanical withdrawal threshold in the KF group were not significantly different from the Ctrl group （P 〉 0.05）. Compared with OA model group, levels of MMP-13, Caspase-3, IL-6, TNF-ot, iNOS and ratio of P-p65/p65 in OA ＋ PS group were reduced with elevated levels of Ki67, IL-10 and the mechanical withdrawal threshold （P 〈 0.01）. As compared with OA ＋ PS group, levels of MMP-13, Caspase-3, IL-6, TNF-ot, iNOS and ratio of P-p65/p65 in OA ＋ PS ＋ KF group were decreased with increased levels of Ki67, IL-10 and mechanical with drawal threshold （P 〈 0.01 ）. Conclusion Combination of kaempferol and pentobarbital sodium alleviates pathologic changes of articular cartilage tissues and relieves inflammatory response and radiculopathic pain in OA rats.