槲皮素对糖尿病大鼠神经病理性疼痛的改善作用及对PI3K/AKT信号通路的影响

Effect of Quercetin on Neuropathic Pain of Diabetic Rats and the Influence on PI3K/AKT Signaling Pathway

目的:探讨槲皮素对糖尿病大鼠神经病理性疼痛的改善作用及对磷脂酰肌醇-3激酶(PI3K)/蛋白激酶B(AKT)信号通路的影响。方法:腹腔注射45 mg/kg的链脲佐菌素溶液制备糖尿病神经病理性疼痛(DNP)大鼠模型,随机分为模型对照组、槲皮素25、50、100 mg/kg组、加巴喷丁40 mg/kg组(阳性对照),每组12只,另取12只大鼠设为正常对照组,造模成功后,每天灌胃相应药物或生理盐水,连续14 d。通过机械性疼痛刺激试验和热敏试验检测大鼠疼痛症状,分别得出缩爪阈值及热敏潜伏期;以多导生理记录仪测定大鼠神经传导速度;以TUNEL染色检测各组大鼠脊髓背角神经细胞凋亡情况;通过酶联免疫吸附(ELISA)法测定大鼠血清和脊髓背角组织中炎性因子白细胞介素-6(IL-6)和IL-18含量;通过蛋白免疫印迹试验检测大鼠脊髓背角组织中PI3K/AKT通路相关蛋白磷酸化PI3K(p-PI3K)/PI3K、磷酸化AKT(p-AKT)/AKT蛋白表达。结果:与正常对照组相比,模型对照组大鼠缩爪阈值、热敏潜伏期和神经传导速度明显降低(P<0.05),神经细胞凋亡率、大鼠血清和骨髓背角组织中IL-6和IL-18含量、大鼠脊髓背角组织p-PI3K/PI3K、p-AKT/AKT比值明显升高(P<0.05);与模型对照组比较,槲皮素25、50、100 mg/kg组和加巴喷丁40 mg/kg组大鼠缩爪阈值、热敏潜伏期、神经传导速度明显升高(P<0.05),神经细胞凋亡率、IL-6和IL-18含量、大鼠脊髓背角组织p-PI3K/PI3K、p-AKT/AKT比值明显降低(P<0.05),且槲皮素各组之间呈剂量依赖性(P<0.05)。结论:槲皮素可抑制糖尿病大鼠炎症反应,减少脊髓背角神经细胞凋亡,促进神经传导,减轻疼痛症状,可能是通过下调PI3K/AKT通路实现的。

Objective:To investigate the effect of quercetin on neuropathic pain of diabetic rats and the influence on phosphatidylinositol 3-kinase(PI3 K)/protein kinase B(AKT)signaling pathway.Methods:Diabetic neuropathic pain(DNP)was induced in rats with 45 mg/kg streptozotocin solution(intraperitoneal injection)and then they were randomized into model control group,quercetin(25 mg/kg,50 mg/kg,100 mg/kg)groups,and 40 mg/kg gabapentin group(positive control),12 in each group.Another 12 rats were set as normal control.After modeling,all rats were given corresponding drugs or normal saline,and the administration lasted 14 days.Mechanical stimulation test and thermal sensitivity test were performed to measure the paw withdrawal threshold and thermal latency,respectively.The nerve conduction velocity(NCV)was measured by a polygraph.The apoptosis of neurons in spinal dorsal horn was examined based on TUNEL staining and the levels of interleukin(IL)-6 and IL-18 in serum and spinal dorsal horn were measured by enzyme-linked immunosorbent assay(ELISA).The levels of phosphorylated-PI3 K(p-PI3 K)/PI3 K and phosphorylated-AKT(p-AKT)/AKT in PI3 K/AKT pathway in spinal dorsal horn were detected by Western blotting.Results:The paw withdrawal threshold,thermal latency,and nerve conduction velocity were lower(P<0.05),and the apoptosis rate of nerve cells,the levels of IL-6 and IL-18 in serum and spinal dorsal horn,and levels of p-PI3 K/PI3 K and p-AKT/AKT in spinal dorsal horn were higher(P<0.05)in the model control group than in the normal control group.The paw withdrawal threshold,thermal latency,and nerve conduction velocity were higher(P<0.05)and the apoptosis rate of nerve cells,levels of IL-6 and IL-18 in serum and spinal dorsal horn,and levels of p-PI3 K/PI3 K and p-AKT/AKT in spinal dorsal horn were lower(P<0.05)in quercetin groups(25 mg/kg,50 mg/kg,and 100 mg/kg)and 40 mg/kg gabapentin group than in the model control group,with a dose-dependent relationship in quercetin groups(P<0.05).Conclusion:Quercetin can inhibit the inflammatory reaction,reduce the apoptosis of neurons in spinal dorsal horn,promote the nerve conduction,and relieve pain symptoms in diabetic rats,which may be related to the down-regulation of PI3 K/AKT pathway.