基于Wnt/β-catenin信号通路的电针穴位刺激改善神经炎性反应初步机制探讨

Preliminary mechanism of electrical acupoint stimulation in improving neuroinflammatory response based on Wnt/β-catenin signaling pathway

目的基于Wnt/β-catenin分子信号通路,探讨穴位电刺激改善神经病理性疼痛神经炎性反应的可能机制。方法将30只大鼠按照随机数表法分为M组(模型组即CCI模型)、S组(假手术组)及D组(穴位电刺激组),每组各10只。其中CCI模型参照国际公认方法建立,即坐骨神经慢性压迫损伤模型。观察各组术T_(0)(术前1 d)、T_(1)(术后3 d)、T_(2)(术后1周)、T_(3)(术后2周)各时间点每组大鼠行为学指标机械性缩足反应阈值(MWT)和热缩足潜伏期(TWL)。行为测试结束后处死大鼠,取L_(4~5)节段脊髓组织,采用组织病理学免疫组化染色检测各组星形胶质细胞活化程度及标志物胶质纤维酸性蛋白(GFAP)表达水平。采用Western-blot技术检测脊髓组织(L_(4~5)节段)Wnt 3a和β-catenin蛋白表达水平以及脊髓组织白介素-18(IL-18)浓度水平。结果T_(0)、T_(1)时S组、M组和D组的MWT和TWL比较,差异无统计学意义(P>0.05)。T_(2)、T_(3)时M组的MWT和TWL小于S组,D组的MWT和TWL大于M组,差异有统计学意义(P<0.05)。T_(3)时间点后处死每组大鼠,免疫组化结果显示,M组大鼠的L_(4~5)节段脊髓组织的星形胶质细胞的活化情况及GFAP表达水平高于S组,D组L_(4~5)节段脊髓组织星形胶质细胞活化情况及GFAP表达水平低于M组,差异有统计学意义(P<0.05)。Western-blot结果显示,M组L_(4~5)节段脊髓组织Wnt3a、β-catenin蛋白表达水平和脊髓组织IL-18浓度水平高于S组,D组L_(4~5)节段脊髓组织Wnt3a、β-catenin蛋白表达水平和脊髓组织IL-18浓度水平低于M组,差异有统计学意义(P<0.05)。结论电针穴位刺激通过抑制Wnt/β-catenin信号通路,使星形胶质细胞活化程度降低,抑制神经病理性疼痛神经炎性反应发生。

Objective To investigate the role of Wnt/β-catenin signaling pathway in neuropathic pain,so as to identify the possible mechanism of electrical acupoint stimulation(EAS)in improving neuroinflammatory response in neuropathic pain.Methods A total of 30 rats were randomly divided into group M(CCI model group),group S(sham operation group)and group D(acupoint electrical stimulation group)according to the random number table method,with 10 rats in each group.The CCI model was established according to the internationally recognized method,that is,the chronic compression injury model of the sciatic nerve.The mechanical paw withdrawal threshold(MWT)and thermal paw withdrawal latency(TWL)were observed at T_(0)(1 day before surgery),T_(1)(3 days after surgery),T_(2)(1 week after surgery)and T_(3)(2 weeks after surgery).After the behavioral test,the rats were sacrificed and their L_(4-5) spinal cord tissues were removed.Immunohistochemical staining was used to detect the activation of astrocytes and the expression of glial fibrillary acidic protein(GFAP).The expression levels of Wnt3a andβ-catenin proteins in the L_(4-5) spinal cord tissues and the interleukin-18(IL-18)concentration in the spinal cord tissues were detected by Western-blot.Results There was no significant difference in mechanical paw withdrawal threshold and thermal paw withdrawal latency at T_(0) and T_(1) among S,M and D groups(P>0.05).The MWT and TWL of group M were lower than those of group S at T_(2) and T_(3),while those of group D were higher than those of group M,with statistically significant differences(both P<0.05).After time point T3,the rats in each group were sacrificed.The immunohistochemical results showed that the activation of astrocytes and the expression level of GFAP in L_(4-5) spinal cord tissues of group M were higher than those of group S,while the activation of astrocytes and the expression level of GFAP in L_(4-5) spinal cord tissues of group D were lower than those of group M,with statistically significant differences(P<0.05).Western-blot data showed that the expression levels of Wnt3a andβ-catenin in the L_(4-5) spinal cord tissues and IL-18 concentration in the spinal cord tissues in group M were higher than those in group S,while the expression levels of Wnt3a andβ-catenin in the L_(4-5) spinal cord tissues and IL-18 concentration in the spinal cord tissues in group D were lower than those in group M,with statistically significant differences(P<0.05).Conclusion EAS can reduce the activation of astrocytes by inhibiting the Wnt/β-catenin signaling pathway,and thus improve the neuroinflammatory response in neuropathic pain.