基于Sirt1/PGC-1α/TFAM通路探讨电针治疗2型糖尿病周围神经病变的机制

Electroacupuncture improves peripheral neuropathy by up-regulating Sirt1/PGC-1α/TFAM pathway in type 2 diabetes rats with peripheral neuropathy

目的:观察电针治疗糖尿病周围神经病变(DPN)的效应并从沉默信息调节因子1(Sirt1)/过氧化物酶体增殖受体γ辅激活因子α(PGC-1α)/促进线粒体转录因子A(TFAM)通路探讨电针治疗DPN的可能机制。方法:SD大鼠随机分成空白组、模型组和电针组,每组8只。采取高脂饲料喂养联合小剂量链脲佐菌素(35 mg/kg)腹腔注射法复制2型糖尿病大鼠模型。电针组大鼠电针“天枢”,每次20 min,每天1次,每周6次,连续治疗6周。测量各组大鼠血糖、体质量、糖耐量曲线下面积(AUC);采用Von Frey法检测大鼠机械痛阈值,哈格里夫斯实验检测大鼠热痛阈值;免疫荧光染色法观察大鼠后足足垫表皮神经纤维密度(IENFD);神经电生理方法测量大鼠坐骨神经传导速度(MNCV);HE染色法观察坐骨神经病理形态改变;透射电子显微镜观察大鼠坐骨神经超微结构改变;Western blot法检测大鼠坐骨神经中Sirt1、PGC-1α、TFAM蛋白表达水平。结果:与空白组相比,模型组大鼠血糖和AUC升高(P<0.001),体质量、机械痛与热痛阈值、后足足垫表皮IENFD降低(P<0.001),MNCV减慢(P<0.01),坐骨神经横切面轴突分布较空白组零碎松散且无序,髓鞘与轴突广泛分离;坐骨神经中Sirt1、PGC-1α、TFAM蛋白表达下调(P<0.05,P<0.01)。与模型组相比,电针组大鼠血糖和AUC下降(P<0.05,P<0.01),机械痛和热痛阈值上升(P<0.01),IENFD增加(P<0.05),MNCV加快(P<0.05);坐骨神经形态和超微结构病变缓解;坐骨神经中Sirt1、PGC-1α、TFAM蛋白表达上调(P<0.05)。结论:电针可能是通过调控Sirt1/PGC-1α/TFAM通路,改善DPN的神经损伤,达到修复糖尿病周围神经病变的作用。

Objective To observe the effect of electroacupuncture(EA)on activation of silent information regulator 1(Sirt1)/peroxisome proliferator-activated receptorγcoactivator 1α(PGC-1α)/mitochondrial transcription factor A(TFAM)pathway in type 2 diabetes(T2DM)rats with peripheral neuropathy(DPN),so as to explore its possible mechanisms underlying improvement of DPN.Methods Thirty male SD rats were randomly divided into blank control group(n=8)and DPN model group(n=22)which were further divided into model group(n=8)and EA group(n=8)after successful modeling.The model of T2DM was established by high-fat diet and low-dose intraperitoneal injection of streptozocin(35 mg/kg).For rats of the EA group(anesthetized with isoflurane),EA stimulation(2 Hz/15 Hz,2 mA)was applied to“Tianshu”(ST25)for 20 min,once daily,6 times a week for 6 weeks.The blood glucose level,body weight,area under curve(AUC)of glucose tolerance test,and hind-paw mechanical pain threshold and thermal pain threshold were observed.The intra-epidermal nerve fiber density(IENFD)of the hind-foot pad was observed by immunofluorescence staining.The motor nerve conduction velocity(MNCV)of the sciatic nerve was measured by using electrophysiological method.H.E.staining was used to observe the histopathological changes of the sciatic nerve after modeling.Transmission electron microscopy(TEM)was used to observe the ultrastructural changes of the sciatic nerve.The protein expressions of energy-related Sirt1,PGC-1αand TFAM in the sciatic nerve was detected by Western blot.Results Compared with the blank control group,the model group had a higher blood glucose contents and AUC(P<0.001),a slower MNCV(P<0.01),and a decrease in the body weight and in the mechanical and thermal pain thresholds(P<0.001)and IENFD(P<0.001),and in the expression levels of Sirt1,PGC-1αand TFAM(P<0.05,P<0.01).In contrast to the model group,the EA group had a decrease in the blood glucose contents and AUC(P<0.05,P<0.01),and an increase in mechanical and thermal pain thresholds,MNCV,IENFD,and expression levels of Sirt1,PGC-1αand TFAM proteins(P<0.01,P<0.05).In addition,results of histopathological and ultrastructural changes of the sciatic nerve showed more fragmented and disordered distribution of axons on the transverse section,and extensive separation of myelin and axons,uneven myelin thickness,axonal degeneration and irregular shape in the model group,whereas in the EA group,the axons on the transverse section were relatively more dense and more complete,the myelin sheath of the sciatic nerve was relatively uniform,and the axonal shape was relatively regular with relatively milder lesions.Conclusion EA up-regulates the expressions of Sirt1,PGC-1α,TFAM in T2DM rats with DPN,which may be associated with its functions in improving and repairing the injured peripheral nerves in rats with DPN.