感觉神经肽P物质对成纤维细胞核因子-κB激活作用的信号机制研究

Signal transduction mechanisms of substance P on the activation of nuclear factor-κB of granulation tissue fibroblasts

目的研究感觉神经肽P物质(SP)对肉芽组织成纤维细胞核因子-κB(NF-κB)的激活作用的信号机制,探讨SP在创伤修复中的作用。方法采用甲醛注射的方法造成Wistar大鼠局部无菌性炎症反应,提取肉芽组织进行成纤维细胞原代培养;采用凝胶迁移阻滞实验(EMSA)的方法,检测SP对肉芽组织成纤维细胞NF-κB的激活作用,观察NK-1受体,Ca2+,PKC以及氧自由基等在NF-κB活化中的作用,并采用抗体超迁移的方法观察NF-κB的亚基组成。结果SP可有效激活NF-κB,这一作用可被NK-1受体阻断剂、Ca2+拮抗剂以及抗氧化剂阻断且呈显著的剂量依赖性,而与PKC无关;活化的NF-κB是由p50和p65亚基组成的。结论SP可有效激活肉芽组织成纤维细胞NF-κB,这一作用是通过NK-1受体介导,并与氧自由基、Ca2+的激活有关,而与PKC无关。

AIM: To study the signal transduction mechanisms of neuropeptide substance P (SP) on the activation of nuclear factor κB (NF κB) of granulation tissue fibroblasts, in order to explore the effect of SP for repairing trauma.METHODS: A local aseptic inflammation was induced by injection of formaladehyde in Wistar rats, and its granulation tissue was undertaken the culture of fibroblast. Electriphoretic mobility shift assay was employed to observe the activation of NF κB after inducement of SP.The conformation of NF κB subunits was observed with antibody super shift assay, and the effect of neurokinin 1 receptor, Ca2+, PKC and oxygen derived free radicals were also observed. RESULTS:SP could activate NF κB.The activation effect could be blocked by NK 1 receptor, Ca2+antagonists and PDTC in a marked dosage dependent manner but not related with PKC. The activated NF κB consisted of p50 and p65.CONCLUSION:SP can activate NF κB,which is mediated by NK 1 receptor and correlated with activation of intercellular calcium and formation of oxygen derived free radicals as second messengers,but not with PKC .