温阳固本汤对糖尿病神经病变大鼠NGF及BDNF表达、脂代谢和坐骨神经超微结构的影响

Effect of Wenyang Guben decoction on lipid metabolism,expression of NGF and BDNF and ultrastructure of sciatic nerve in rats with diabetic peripheral neuropathy

目的探讨温阳固本汤对糖尿病周围神经病变(DPN)大鼠NGF及BDNF表达、脂代谢和坐骨神经超微结构的影响。方法5-6周SD大鼠60只被随机分为6组,每组10只。除对照组外,其余组腹腔注射STZ溶液制备糖尿病大鼠模型,造模成功后饲养4周,给予糖尿病大鼠缺血再灌注制备DPN大鼠模型。模型成功后,温阳固本汤低、中、高剂量组分别给予灌胃温阳固本汤9.17 g/kg、18.33 g/kg及36.67 g/kg,1次/d,连续56 d,对照组和模型组等体积生理盐水灌胃。干预结束后,ELISA法测定血清TC、TG;免疫比浊法测定LDL-C、HDL-C;免疫组化法检测大鼠坐骨神经NGF、BDNF表达;RT-PCR法检测大鼠坐骨神经NGF mRNA、BDNF mRNA表达水平;电子透射显微镜观察大鼠坐骨神经超微结构。结果与模型组对比,温阳固本汤高、中剂量组TG、TC、LDL-C、HDL-C含量低于模型组(P<0.05);温阳固本汤中、高剂量组NGF、BDNF表达水平明显高于模型组,且随剂量的增加表达水平不断升高;温阳固本汤低、中、高剂量组NGF mRNA、BDNF mRNA水平均明显高于模型组(P<0.01)。电镜显示,模型组坐骨神经横切面髓鞘板层分离严重,层间排列紊乱,可见较多空泡;温阳固本汤各剂量组髓鞘均有改善,但低、中剂量组未恢复正常结构,高剂量组恢复正常髓鞘结构,仅存在部分线粒体、微丝及微管结构异常。结论温阳固本汤可降低DPN大鼠血脂、血清游离脂肪酸含量,可提高DPN大鼠坐骨神经NGF、BDNF及基因的表达,同时对神经髓鞘、微丝等具有修复作用。

Objective To explore the effects of Wenyang Guben decoction on the lipid metabolism,NGF and BDNF expression and ultrastructure of sciatic nerve of diabetic peripheral neuropathy rats.Methods Sixty adult SD rats aged 5-6 weeks were selected.Ten rats were chosen as control group,and the other 50 rats were intraperitoneally injected with STZ solution to prepare diabetic rat model.At 4 week after the successful modeling,the DPN model was induced by ischemic reperfusion in diabetic rats.After successful model,the model rats were randomly divided into model group,positive control group,low,medium and high dose groups of Wenyang Guben decoction according to the random number table.The model rats were gavaged with 9.17 g/kg,18.33 g/kg,36.67 g/kg Wenyang Guben decoction once a day for 56 d in low,medium and high dose groups,respectively.The rats were given equal volume of normal saline in control group and model group,and 20 mg/kgα-lipoic acid in positive control group.After the intervention,ELISA was used to detect the levels of TC and TG,and immunoturbidimetric assay was used to measure LDL-C and HDL-C.Immunohistochemistry was used to detect the expression of NGF and BDNF in rat sciatic nerve.RT-PCR was used to detect the expression of NGF mRNA and BDNF mRNA in rat sciatic nerve.The ultrastructure of rat sciatic nerve was observed by electron projection microscope.Results Compared with model group,the contents of TG,TC,LDL-C and HDL-C in high and medium dose groups of Wenyang Guben decoction were decreased(P<0.05).The expression of BDNF was significantly higher in high and medium dose groups of Wenyang Guben decoction than in model group(P<0.01),and the expression level increased with the increase of dose.The expression levels of NGF mRNA and BDNF mRNA in low,medium and high dose groups of Wenyang Guben decoction were significantly higher than those in model group(P<0.01).The expression levels of NGF mRNA and BDNF mRNA in low,medium and high dose groups of Wenyang Guben decoction were significantly higher than those in model group(P<0.01).Electron microscopy results showed that the myelin layer of the sciatic nerve in model group was severely separated,and the layers were disordered with more vacuoles.The myelin sheath was improved to normal level in high dose group of Wenyang Guben decoction,and only some mitochondria,microfilaments and microtubules were abnormal,but it was not returned to normal structure in low and medium dose groups.Conclusion Wenyang Guben decoction can decrease the blood lipid level and the serum free fatty acid level of DPN rats,promote the expression of NGF,BDNF mRNA in sciatic nerve of DPN rats,and have a repairing effect on the ultrastructure of sciatic nerve.