桑白皮总黄酮抑制分泌性中耳炎大鼠耳黏膜的自噬作用

Inhibition of autophagy of ear mucosa by total flavonoids of mulberry bark in otitis media with effusion

目的探讨桑白皮总黄酮对分泌性中耳炎(OME)大鼠耳黏膜水肿的改善以及对自噬的影响。方法26只大鼠腹腔注射卵清蛋白致敏后耳内激发建立OME模型,建立模型成功后,将26只大鼠随机分为模型组和治疗组,每组13只,另取10只大鼠作为正常组。治疗组大鼠灌胃桑白皮总黄酮25 mg·kg^(-1),正常组和模型组大鼠灌胃等剂量生理盐水,连续给药2周。ELISA检测血清肿瘤坏死因子α(TNF-α)、白细胞介素(IL-6)和白细胞介素-1β(IL-1β)表达,HE染色观察中耳组织病理学变化,TUNEL染色检测细胞凋亡率,蛋白印迹法检测Beclin1、LC3Ⅱ/LC3Ⅰ和p62蛋白相对表达量。结果模型组大鼠耳黏膜细胞肿胀,黏膜厚度增加,炎症细胞浸润,可见多处细胞脱落;治疗组耳黏膜细胞肿胀情况好转,少量黏膜细胞脱落,炎症细胞浸润减少。与正常组比较,模型组TNF-α、IL-6和IL-1β含量、细胞凋亡率、Beclin1以及LC3Ⅱ/LC3Ⅰ蛋白相对表达量升高,p62蛋白相对表达量降低(P<0.05);与模型组比较,治疗组TNF-α、IL-6和IL-1β含量、细胞凋亡率、Beclin1以及LC3Ⅱ/LC3Ⅰ蛋白相对表达量降低,p62蛋白相对表达量升高(P<0.05)。结论桑白皮总黄酮可改善OME大鼠耳黏膜水肿及炎症反应,可能是通过抑制自噬发挥作用。

Objective To investigate the effect of total flavonoids of mulberry bark on the improvement and anti-inflammatory effect of ear mucosa edema in rats with otitis media with effusion(OME),and the effect on autophagy.Methods A rat model of OME was established by intraperitoneal injection of ovalbumin sensitization and intra aural stimulation in 26 rats.After successful preparation of the model,the rats were randomly divided into model group and treatment group,with 13 rats in each group,and another 10 rats were selected as normal group.Rats in the treatment group were intragastrically given 25 mg·kg^(-1) of total flavonoids of mulberry,and rats in the normal group and model group were given equal amount of normal saline for 2 weeks.The expression of TNF-α,IL-6 and IL-1βin serum were detected by ELISA,the histopathological changes of middle ear were observed by HE staining,the apoptosis rate of cells was detected by TUNEL staining,and the relative expression levels of Beclin1,LC3Ⅱ/LC3Ⅰand p62 protein were detected by Western blot.Results In model group,the ear mucosa cells were swollen,the mucosa thickness increased,the inflammatory cells were infiltrated,and many cells were shed.In treatment group,the swelling of ear mucosa cells was improved,a few mucosa cells were exfoliated,and the infiltration of inflammatory cells was reduced.Compared with normal group,the contents of TNF-α,IL-6,IL-1β,apoptosis rate,relative expression levels of Beclin1,LC3Ⅱ/LC3Ⅰprotein in model group were increased,and the relative expression level of p62 protein was decreased(P<0.05).Compared with model group,the contents of TNF-α,IL-6,IL-1β,apoptosis rate,relative expression levels of Beclin1,LC3Ⅱ/LC3Ⅰprotein in treatment group were decreased,and the relative expression level of p62 protein was increased(P<0.05).Conclusion The total flavonoids of mulberry can improve the edema and inflammatory response of ear mucosa in OME rats,which may play a role in inhibiting autophagy.