含笑内酯通过抑制NF-κB/NLRP3轴改善单侧输尿管梗阻模型小鼠的肾脏病变

Micheliolide ameliorates renal lesion of unilateral ureteral obstruction mice by inhibiting NF-κB/NLRP3 axis

目的探究含笑内酯(MCL)对单侧输尿管梗阻(UUO)模型小鼠肾脏病变的干预作用及机制。方法将雄性C57BL/6J小鼠随机分为假手术(Sham)组、UUO组、UUO+MCL组,UUO+MCL组建立UUO模型前1 d以25 mg/kg MCL灌胃。术后8 d采集肾组织标本进行HE、Masson、TUNEL染色;蛋白免疫印迹(Western blot)和免疫组化检测核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)、胱天蛋白酶1(caspase-1)、白细胞介素(IL)-1β、肿瘤坏死因子(TNF)-α、核因子(NF)-κB p65蛋白表达情况。结果HE染色结果显示,Sham组小鼠肾组织无明显病变;UUO组肾小管进行性扩张,间质水肿伴有少量的炎性细胞浸润;UUO+MCL组肾脏病变较UUO组明显改善。Masson染色结果显示,与Sham组相比,UUO组胶原容积分数明显增加,而UUO+MCL组胶原容积分数较UUO组明显减少。TUNEL染色结果显示,与Sham组相比,UUO组细胞凋亡率升高,UUO+MCL组较UUO组细胞凋亡率明显下降。Western blot和免疫组化结果显示,与Sham组相比,UUO组肾组织中NLRP3、caspase-1、IL-1β、TNF-α、p-NF-κB p65的表达水平均升高;与UUO组相比,UUO+MCL组上述因子均显著下降。结论UUO小鼠肾组织NLRP3炎症小体及相关炎性因子表达增加,MCL通过抑制NF-κB通路及NLRP3炎症小体的活化,减轻肾小管间质的炎症反应,从而改善肾纤维化。

Objective To investigate the effect and mechanism of micheliolide on renal lesions of unilateral ureteral obstruction(UUO)mice.Methods Male C57BL/6J mice were randomly divided into the sham group,the UUO(model)group and the UUO+micheliolide(administration)group.Mice in the UUO+micheliolide group were given micheliolide by gavage at 25 mg/kg body weight after the establishment of the animal model.Renal tissue samples were collected 8 days after operation,and the renal tissue was stained with HE,Masson and TUNEL staining.The protein expressions of nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3),caspase-1,interleukin(IL)-1β,tumor necrosis factor(TNF)-αand(phosphorylation)-nuclear transcription factor(NF)-κB p65 were detected by immunohistochemistry and Western blot assay.Results HE staining showed that there were no pathological changes in renal tissue in the sham group.Mice in the UUO group had progressive renal tubule dilation and interstitial edema with a small amount of inflammatory cell infiltration.The renal lesions were significantly improved in the UUO+MCL group compared with those of the UUO group.Masson staining showed that compared with the sham group,the collagen volume fraction(CVF)was significantly increased in the UUO group,while CVF was significantly reduced in the UUO+MCL group than that in the UUO group.TUNEL staining showed that compared with the sham group,the apoptosis rate was increased in the UUO group,and the apoptosis rate was significantly decreased in the UUO+MCL group than that of the UUO group.Western blot assay and immunohistochemistry result showed that the expression levels of NLRP3,caspase-1,IL-1β,TNF-αand p-NF-κB p65 were increased in kidney tissue of the UUO group compared with those of the sham group.Compared with the UUO group,NLRP3,caspase-1,IL-1β,TNF-αand p-NF-κB p65 were significantly decreased in the UUO+MCL group.Conclusion The expression of NLRP3 inflammasome and related inflammatory factors in renal tissue of mice with unilateral ureteral obstruction are increased.Micheliolide can reduce the inflammatory response of renal tubule interstitium by inhibiting NFκB pathway and NLRP3 inflammasome activation,thereby improving renal fibrosis.