转位分子蛋白经由Keap1/Nrf2/HO-1通路激活自噬缓解大鼠糖尿病神经病理性疼痛  

Translocator protein activates autophagy in diabetic neuropathic pain rats via regulation of the Keap1/Nrf2/HO-1 signaling

作　　者：高楠 郝璨 马冰洁[1] 靳天 马柯[1] 刘晓明[1] GAO Nan;HAO Gem;MA Bingjie;JIN Tian;MA Ke;LIU Xiaoming(Department of Pain Management,Xinhua Hospital,Shanghai Jiao Tong University School of Medicine,Shanghai 200082,China)

机构地区：[1]上海交通大学医学院附属新华医院疼痛科,上海200092

出　　处：《上海交通大学学报（医学版）》2023年第8期988-996,共9页Journal of Shanghai Jiao tong University:Medical Science

基　　金：国家自然科学基金(81771184)。

摘　　要：目的·探讨转位分子蛋白(translocator protein,TSPO)激动剂Ro5-4864对糖尿病神经病理性疼痛(diabetic neuropathic pain,DNP)大鼠坐骨神经自噬和Kelch样ECH关联蛋白(Kelch-like ECH-associated protein 1,Keap1)/核因子E2相关因子(nuclear factor erythroid-derived-2-like 2,Nrf2)/血红素加氧酶-1(heme oxygenase-1,HO-1)通路的影响。方法·通过高脂饮食和链脲佐菌素构建2型糖尿病模型,经行为学筛选获得DNP大鼠。24只大鼠根据数字表随机分配到假手术组(Sham组)、DNP模型组(DNP组)、TSPO激动剂Ro5-4864处理组(Ro组)和TSPO激动剂Ro5-4864合用Nrf2抑制剂ML385处理组(Ro+ML385组)。采用up-down法测量大鼠50%机械缩足反射阈值(paw 50%mechanical withdrawal threshold,50%PMWT),时间点选定为处理前(baseline),处理后第3(D3)、7(D7)、14(D14)、21(D21)和28(D28)天。在最后1天收集坐骨神经,使用免疫荧光和Western blotting分析鞘内注射Ro5-4864对DNP大鼠坐骨神经中自噬相关蛋白和Keap1/Nrf2/HO-1通路相关蛋白的影响。结果·与Sham组相比,DNP组大鼠的50%PMWT自第3天开始大幅下降,全程无改善(各时间点均P=0.000);另外,DNP大鼠坐骨神经中Bcl-2相互作用蛋白(Bcl-2 interacting coiled-coil protein 1,Beclin-1)、微管相关蛋白轻链3-Ⅱ(microtubule-associated protein light chain 3-Ⅱ,LC3-Ⅱ)、HO-1和核Nrf2蛋白的含量显著减少(均P=0.000),p62则显著增加(P=0.000)。鞘内注射Ro5-4864改善了以上损伤,大鼠50%PMWT与DNP组相比逐渐上升(D14 P=0.039,D21和D28均P=0.000),自噬和Keap1/Nrf2/HO-1通路的损伤也被修复,表现为Beclin-1、LC3-Ⅱ、HO-1和核Nrf2蛋白含量的升高(均P=0.000)以及p62含量的降低(P=0.001)。但是,Ro5-4864的效应被ML385完全抑制。结论·TSPO激动剂Ro5-4864缓解了DNP,其作用机制可能与通过对Keap1/Nrf2/HO-1通路的正向调节激活了DNP大鼠坐骨神经中的自噬相关。这可能为治疗DNP提供了一个新的策略。Objective·To study the effects of translocator protein(TSPO)agonist Ro5-4864 on autophagy and Kelch-like ECHassociated protein 1(Keap1)/nuclear factor erythroid-derived-2-like 2(Nrf2)/heme oxygenase-1(HO-1)signaling in diabetic neuropathic pain(DNP)rats.Methods·Type 2 diabetic rats were established by high-fat diet and streptozotocin(STZ),and DNP rats were filtered by behavioral assessment.Twenty-four rats were randomly assigned to the Sham group,DNP group,TSPO agonist Ro5-4864 group(Ro group),and TSPO agonist Ro5-4864 combined with Nrf2 inhibitor ML385 group(Ro+ML385 group).Up-Down method was used to measure paw 50%mechanical withdrawal threshold(50%PMWT)of the rats before high-fat diet(baseline),and on Day 3,7,14,21 and 28 after STZ.Sciatic nerves were collected on the last day to analyze the effects of Ro5-4864 on autophagy related proteins and Keap1/Nrf2/HO-1 signaling related proteins of DNP rats by using immunofluorescence and Western blotting.Results·The 50%PMWT in the DNP group decreased from D3 to D28(P=0.000 at all timing),and the expression of Bcl-2 interacting coiled-coil protein 1(Beclin-1),microtubule-associated protein light chain 3-Ⅱ(LC3-Ⅱ),HO-1,and nuclear Nrf2(P=0.000)were significantly reduced in the sciatic nerves of DNP rats(all P=0.000),compared with those in the sham group,but p62 was significantly increased(P=0.000).Administration of Ro5-4864 attenuated these changes in the rats of the Ro group.There was a gradual increase in the 50%PMWT,compared with that of the rats in the DNP group(D14 P=0.039,both D21 and D28 P=0.000),and the impairment of autophagy and the Keap1/Nrf2/HO-1 signaling was repaired,which was demonstrated by increases of Beclin-1,LC3-Ⅱ,HO-1,and nuclear Nrf2 protein contents(all P=0.000)and a decrease in p62 content(P=0.001).However,the beneficial effects of Ro5-4864 were totally abrogated by ML385 in rats of the Ro+ML385 group.Conclusion·TSPO alleviates DNP in rats,of which the mechanism involves activation of autophagy via upregulation of the Keap1/Nrf2/HO-1 sig

关 键 词：转位分子蛋白 神经病理性疼痛 糖尿病 自噬 Keap1/Nrf2/HO-1通路 

分 类 号：R614[医药卫生—麻醉学]