萝卜硫素抑制糖尿病小鼠耳蜗毛细胞损伤和Trx、Nrf2及p-Erk1/2表达下调

Sulforaphane Inhibits cochlear hair cell damage and down-regulation of Trx,Nrf2 and p-Erk1/2 expression in the diabetic mice

目的探讨萝卜硫素(sulforaphane,SF)对糖尿病引起的耳蜗毛细胞退行性病变的保护作用及机制,为临床糖尿病耳聋防治寻找新的靶点。方法选取20只1月龄雄性Balb/c小鼠,随机分为正常(NC)组,糖尿病(DM)组,DM+SF组,DM+SF+PX-12组。DM组给予高脂饲料喂养同时腹腔注射链脲佐菌素(streptozotocin,STZ)。采用HE染色、琥珀酸脱氢酶(SDH)活性染色及免疫荧光染色观察耳蜗毛细胞形态学改变及硫氧还蛋白(thioredoxin,Trx)表达变化;Western blot检测耳蜗组织中Trx、NF-E2-相关因子2(NF-E2-related factor-2,Nrf2)及p-Erk1/2蛋白表达水平变化。结果与NC组相比,DM组内外毛细胞数量均减少且排列不规则,SF处理可延缓毛细胞的丢失,且其排列规则;Trx抑制剂PX-12处理则阻断SF对耳蜗毛细胞损伤的保护作用。Western blot检测显示,与NC组相比,DM组Trx、Nrf2及p-Erk1/2表达明显下降,而给予SF后,Trx表达则明显回升,Nrf2及p-Erk1/2水平上调;而Trx抑制剂PX-12处理抑制SF对Nrf2及p-Erk1/2表达的上调作用。结论SF可能通过p-Erk1/2/Nrf2抗氧化信号通路可有效上调小鼠耳蜗组织中Trx表达,从而延缓DM小鼠耳蜗毛细胞损伤。

Objective To investigate the protective effect of Sulforaphane(SF)on the degeneration of cochlear hair cells induced by diabetes and to elucidate its mechanism aiming to find a new therapeutic target in clinic.Methods Twenty 1-month age male Balb/c mice were randomly divided into 4 groups:normal(NC),diabetes(DM),DM+SF,DM+SF+PX12.DM group was fed with high-fat diet while streptozotocin(STZ)was administered intraperitoneally.The morphological changes of cochlear hair cells were observed by HE staining,succinate dehydrogenase(SDH)staining and the expression of Trx was detected by immunofluorescence.The expression of Trx,Nrf2 and p-Erk1/2 was detected by Western blot.Results Compared with NC group,inner and outer hair cells in DM group were reduced in number and irregular in arrangement.SF treatment could delay the loss of hair cells,and make the arrangement of hair cells regular.Treatment with Trx inhibitor PX-12 blocked the protective effect of SF on cochlear hair cell injury.The protective effect of SF on cochlear hair cell injury was inhibited after treated with PX-12 which is the inhibitor of Trx.Western blot results showed that the expression of Trx,Nrf2 and p-Erk1/2 in DM group was significantly decreased compared with NC group.The expression of Trx,Nrf2 and p-Erk1/2 was up-regulated after SF treatment,while the treatment with Trx inhibitor,PX-12,inhibited the up-regulation of Nrf2 and p-Erk1/2 expression caused by SF.Conclusion SF can delay the degeneration of cochlear hair cells in DM mice through p-Erk1/2/Nrf2 antioxidant signaling pathway to up-regulate the expression of Trx.