川芎嗪对药物性聋大鼠自噬相关蛋白表达水平影响

Effects of Ligustrazine on Autophagy Related Protein Expression in Drug-induced Deafness Rats

目的 通过观察川芎嗪对顺铂诱导大鼠耳蜗组织中自噬相关蛋白LC3-Ⅰ、LC3-Ⅱ和P62表达水平的影响,探讨川芎嗪对药物所致耳聋的大鼠耳蜗毛细胞自噬的干预及其作用机制。方法 选择110只健壮的听力正常的SD大鼠常规适应性养殖7 d后,将其分为2组,其中模型组(n=65)在腹腔内注入顺铂,对照组(n=45)在腹腔内注射相同计量的生理盐水。持续输注5 d后,经模式评估(每组各取5只大鼠)后,对照组随机分配为正常对照组和川芎嗪对照组各20只。模型组随机分配为模型对照组(n=20)、川芎嗪治疗组(n=20)和3-MA组(n=20),腹腔注射同等计量药剂,连续注射7 d。实验采用免疫组化和Western-blot的方法,观察实验中大鼠耳蜗组织中自噬蛋白LC3-Ⅰ、LC3-Ⅱ和P62表达水平的变化。结果 与3-MA组比较,空白对照组和川芎嗪对照组中LC3-Ⅱ/LC3-Ⅰ比值略微上升(P<0.05);与空白对照组耳蜗组织中P62蛋白表达水平比较,模型对照组和川芎嗪治疗组蛋白表达水平下降(P<0.01);与模型对照组耳蜗组织中P62蛋白表达水平比较,川芎嗪治疗组蛋白表达水平下降,3-MA组耳蜗组织中蛋白表达水平上升,差异均具有统计学意义(P<0.01);与空白对照组耳蜗组织中LC3-Ⅰ和LC3-Ⅱ蛋白表达水平比较,模型对照组和川芎嗪治疗组蛋白表达水平上升,差异均具有统计学意义(P<0.01);与模型对照组耳蜗组织中LC3-Ⅰ和LC3-Ⅱ蛋白表达水平比较,川芎嗪治疗组蛋白表达水平上升,而3-MA组蛋白表达水平下降,差异均具有统计学意义(P<0.01)。结论 川芎嗪能够上调顺铂所致药物性聋大鼠耳蜗组织中的自噬蛋白LC3-Ⅰ和LC3-Ⅱ的表达,下调自噬蛋白P62的表达,促进耳蜗保护性自噬的发生,从而抑制顺铂诱导的听力损伤作用。

Objective To observe the effect of ligustrazine on the expression levels of autophagy-related proteins LC3-Ⅰ,LC3-Ⅱand P62 in rat cochlea tissue induced by cisplatin.To investigate the effect of ligustrazine on cochlear hair cell autophagy in rats with drug-induced deafness and its mechanism.Methods A total of 110 healthy SD rats with normal hearing were divided into two groups after 7 days of conventional adaptive culture.The model group(n=65)was intraperitoneal injected with cisplatin.The control group(n=45)received intraperitoneal injection of the same amount of normal saline.After 5 days of continuous infusion,the control group was randomly divided into normal control group and ligustrazine control group with 20 rats in each group after pattern assessment(5 rats in each group).The model group was randomly assigned to model control group(n=20),tetramethylpyrazine treatment group(n=20)and 3-MA group(n=20)by intraperitoneal injection of the same dosage for 7 days.Immunohistochemistry and Western-blot were used to observe the expression of autophagy proteins LC3-Ⅰ,LC3-Ⅱand P62 in the cochlea of rats.Results Compared with 3-MA group,LC3-Ⅱ/LC3-Ⅰratio in blank control group and ligustrazine control group was slightly increased(P<0.05).Compared with the blank control group,the expression of P62 protein in the model control group and ligustrazine treatment group decreased(P<0.01).Compared with the model control group,the expression of P62 protein in the cochlea tissue was significantly decreased in the ligustrazine treatment group,and increased in the 3-MA group(P<0.01).Compared with the blank control group,the expression of LC3-Ⅰand LC3-Ⅱprotein in the cochlear tissue of the model control group and the ligustrazine treatment group increased,and the difference was statistically significant(P<0.01).Compared with the LC3-Ⅰand LC3-Ⅱprotein expression levels in the model control group,the histone expression level of ligustrazine treatment was increased,while the histone expression level of 3-MA was decreased,with significant statistical differences(P<0.01).Conclusion Ligustrazine can up-regulate the expression of autophagy proteins LC3-Ⅰand LC3-Ⅱand down-regulate the expression of autophagy protein P62 in the cochlea of cisplatin-induced drug-induced deafness rats,and promote the development of protective autophagy in the cochlea,thereby inhibiting the hearing damage induced by cisplatin-induced hearing loss.