艾灸对内脏痛镇痛作用的MAPKs信号转导机制研究

Pain is one of the most common symptoms in clinical practice, and moxibustion can produce a satisfactory result in managing or killing pain, especially the visceral pain. It has been proved that MAPKs are involved in the transduction of pain signals, and it has been confirmed that the activation of several members from MAPKs, p38MAPK, ERK and JNK, has played a crucial role in the production and maintenance of inflammatory hyperalgesia. Based on the previous work, this study was to investigate the intervention of moxibustion on the visceral hyperalgesia due to inflammatory bowel diseases by using immunohistochemistry techniques, Western blot, protein chips, and real-time fluorescence quantitative PCR. Focused on ERK, P38MAPK, and JNK, which are the signaling pathways closely related to the visceral hyperalgesia, the study also aimed to discover the changes of the expression of pERK, p38MAPK, NK-1, VR-1, NGF, COX-2, pJNK, and c-Jun in spinal dorsal horn and colon of the rats with post-inflammatory visceral hyperalgesia, as well as the regulation effect of moxibustion. From the angle of pathways ERK, P38 MAPK, and JNK, and by studying the signal transduction in the analgesic effect of moxibustion in the rat models of post-inflammatory visceral hyperalgesia, this study was going to make clear the action pathways and the key signaling molecules in the regulation of moxibustion on visceral hyperalgesia, and to provide scientific evidences for the explanation of the analgesia mechanism of moxibustion.

疼痛是临床最常见的症状之一,艾灸在缓解和消除疼痛尤其是内脏痛方面具有良好疗效。研究证实MAPKs参与疼痛信息传导,目前已明确MAPKs家族中p38MAPK、ERK和JNK信号分子的激活在炎性痛敏的产生和维持中具有重要作用。基于前期研究成果,本项目应用免疫组织化学、Western blot、蛋白芯片、荧光定量PCR等技术,研究艾灸对大鼠炎症性肠病炎症后慢性内脏痛敏的干预效应;并选择与内脏痛敏关系密切的ERK通路、P38MAPK通路和JNK通路,探讨炎症后内脏痛敏大鼠脊髓背角和结肠pERK、p38MAPK、NK-1、VR-1、NGF、COX-2、pJNK、c-Jun等表达的变化及艾灸的调节作用,从ERK通路、P38 MAPK通路、JNK通路调控角度研究艾灸对炎症后内脏痛敏大鼠镇痛作用的信号转导机制,拟阐释艾灸调节内脏痛敏信号通路的作用途径及关键信息分子,为艾灸镇痛机制的阐明提供科学实验资料。

艾灸在缓解和消除内脏痛方面具有良好疗效。MAPKs参与疼痛信息传导，在炎性痛敏的产生和维持中具有重要作用。本项目建立炎症性肠病慢性内脏痛大鼠模型，从与内脏痛敏关系密切的MAPKs信号通路（ERK通路、p38 MAPK通路、JNK通路）调控角度研究艾灸对慢性炎性内脏痛大鼠镇痛作用的信号转导机制。结果发现：隔药灸对慢性炎性内脏痛大鼠痛行为和痛情绪均有改善作用；对大鼠脊髓TRPV1、c-fos、SP有下调作用；能够降低下丘脑SP、5-HT、c-fos、TNF-α、CRF、CRFR1的表达，提高β-内啡肽、脑啡肽、阿黑皮素原的含量。慢性炎性内脏痛大鼠脊髓MAPK信号通路激活，隔药灸能使激活的MAPK信号通路受到抑制；芯片结果提示隔药灸调节脊髓的蛋白位点共有14个，包括ERK/MAPK通路：MEK1、Tau、STMN1、CREB、ELK-1、c-Myc；JNK/MAPK通路：MKK4、c-JUN、JunD、ELK-1；p38 MAPK通路：ASK1、MKK3、p53、CREB。免疫组化/Western blot、荧光定量PCR检测结果提示，隔药灸能降低慢性炎性内脏痛大鼠脊髓MAPK信号通路中MEK、ERK、CREB、p38 MAPK、ELK-1、MSK1、JNK、c-jun、ATF-2蛋白磷酸化及mRNA表达，对DRG中p38 MAPK、ERK、JNK、c-jun、SP、NK-1、c-fos、MCP-1、iNOS、COX-2、VR1蛋白和mRNA表达有抑制作用，提示隔药灸对脊髓和DRG的ERK、p38 MAPK和JNK通路均有一定的调节作用。对结肠组织的检测发现，隔药灸能降低结肠p38MAPK、JNK、NGF、MCP-1、c-fos、iNOS、COX2的表达，提示隔药灸对结肠p38MAPK、JNK通路有一定调节作用。研究结果从MAPKs通路角度阐释了艾灸治疗慢性炎性内脏痛的信号转导机制和关键信息分子，有助于促进艾灸疗法在慢性炎性内脏痛中的进一步应用。