乳酸调控大鼠肠黏膜微血管内皮细胞的NF-κB信号通路

Lactic acid inhibits the NF-κB signaling pathways in intestinal mucosal microvascular endothelial cells of rats

目的探讨内毒素(LPS)刺激大鼠肠黏膜微血管内皮细胞(RIMMVECs)后,乳酸(LA)调控NF-κB信号通路中磷酸化IκBα和NF-κB p65蛋白表达情况,肿瘤坏死因子α(TNF-α)和白细胞介素6(IL-6)mRNA表达情况,阐明乳酸发挥作用的最佳时间及其调控NF-κB信号通路的部位。方法提取RIMMVECs总蛋白和总RNA,用Western blotting检测NF-κB p65、IκBα及p-IκBα蛋白表达水平,用real-time PCR对TNF-α和IL-6 mRNA进行定量检测。结果乳酸能降低LPS诱导RIMMVECs分泌的TNF-α和IL-6 mRNA表达水平,并分别于24 h和3 h下调效果最明显;乳酸能抑制IκBα磷酸化及NF-κB转录活性,并于4～8 h达到最佳效果;乳酸发挥作用部位是抑制信号通路中IκBα磷酸化。结论乳酸通过抑制IκBα磷酸化而阻断NF-κB的激活,抑制下游炎性因子表达,进而发挥出很好的预防炎症效果。

Objective The aim of this study was to explore the effects over time of lactic acid（LA） on IκBα phosphorylation and expressions of nuclear factor-kappa B p65 protein,tumor necrosis factor α（TNF-α） and interleukin-6（IL-6） mRNA in the NF-κB signaling pathway in rat intestinal mucosal microvascular endothelial cells stimulated by lipopolysaccharide（LPS）,and then to assess the optimal action time of LA and its regulating site in the NF-κB signaling pathways.Methods IκBα,phosphorylated IκBα（p-IκBα） and NF-κB p65 proteins were monitored by Western blot analysis,and the TNF-α and IL-6 mRNA was analyzed using real-time PCR.Results LA treatment reduced TNF-α and IL-6 mRNA expression in LPS-stimulated RIMMVECs,with the greatest effect after 24 h and 3 h.The highest inhibitory effect of LA on IκBα phosphorylation and NF-κB transcriptional activity was during 4 to 8 h.The site of action of LA was IκBα phosphorylation in the signaling pathways.Conclusions The results of this study suggest that LA inhibits downstream inflammatory cytokine expression through inhibiting IκBα phosphorylation and blocking activation of NF-κB,thus playing a key anti-inflammatory role in the intestinal mucosal microvascular endothelial cells in rats.