Inhibitory effects of berberine on proinflammatory M1 macrophage polarization through interfering with the interaction between TLR4 and MyD88
Inhibitory effects of berberine on proinflammatory M1 macrophage polarization through interfering with the interaction between TLR4 and MyD88
复制标题
小檗碱通过干扰 TLR4 和 MyD88 之间的相互作用对促炎性 M1 巨噬细胞极化的抑制作用
DOI：
10.1186/s12906-019-2710-6
复制
发表时间：
2019-11-19
影响因子：
--
通讯作者：
Lu, Fuer
中科院分区：
文献类型：
--
作者：
Gong, Jing;Li, Jingbin;Lu, Fuer
研究背景炎症被认为是2型糖尿病的关键病理机制。小檗碱(BBR)的降血糖作用与抑制炎症反应有关，但其机制尚不完全清楚。方法采用LPS诱导Raw264.7细胞和原代腹腔巨噬细胞炎症极化，探讨BBR的作用及潜在机制。采用不同浓度LPS处理不同时间建立炎症模型。采用ELISA法检测TNF-α的分泌情况。 RT-PCR检测M1炎症因子。流式细胞术测定原代腹腔巨噬细胞F4/80+比值和CD11c+比值。 Western blot检测p-AMPK和TLR4的表达。通过共聚焦显微镜观察NFκB p65的细胞质和核分布。 CoIP检测TLR4与MyD88的结合，分子对接评估BBR对TLR4的亲和力。结果LPS暴露后，巨噬细胞TNF-α分泌和炎症因子转录增加，细胞形态改变并逐渐出现突起，F4/80+CD11c+M1巨噬细胞比例增加，NFκB p65核分布增加。 BBR预处理部分抑制了上述变化。然而，LPS干预3小时后，TLR4和p-AMPK的表达没有显着变化。同时，CoIP显示TLR4和MyD88之间的相互作用增加，而BBR抑制了这种结合。分子对接提示BBR可能与TLR4相互作用。结论LPS刺激3h后巨噬细胞发生炎症变化，BBR预处理可抑制炎症极化。 BBR可能与TLR4相互作用，干扰TLR4/MyD88/NFκB信号通路，这可能是BBR早期减轻炎症的机制。
BackgroundsInflammation is recognized as the key pathological mechanism of type 2 diabetes. The hypoglyceamic effects of berberine (BBR) are related to the inhibition of the inflammatory response, but the mechanism is not completely clear.MethodsThe inflammatory polarization of Raw264.7 cells and primary peritoneal macrophages were induced by LPS, and then effects and underlying mechanisms of BBR were explored. An inflammatory model was established by LPS treatment at different concentrations for different treatment time. An ELISA assay was used to detect the secretions of TNF-α. RT-PCR was applied to detect M1 inflammatory factors. The F4/80+ratio and CD11c+ratio of primary peritoneal macrophages were determined by flow cytometry. The expressions of p-AMPK and TLR4 were detected by Western blot. The cytoplasmic and nuclear distributions of NFκB p65 were observed by confocal microscopy. The binding of TLR4 to MyD88 was tested by CoIP, and the affinity of BBR for TLR4 was assessed by molecular docking.ResultsUpon exposure to LPS, the secretion of TNF-α and transcription of inflammatory factors in macrophages increased, cell morphology changed and protrusions appeared gradually, the proportion of F4/80+CD11c+M1 macrophages increased, and the nuclear distribution of NFκB p65 increased. BBR pretreatment partially inhibited the changes mentioned above. However, the expression of TLR4 and p-AMPK did not change significantly after LPS intervention for 3 h. Meanwhile, CoIP showed that the interaction between TLR4 and MyD88 increased, and BBR inhibited the binding. Molecular docking suggested that BBR might interact with TLR4.ConclusionsInflammatory changes were induced in macrophages after LPS stimulation for 3 h, and BBR pretreatment inhibited inflammatory polarization. BBR might interact with TLR4 and disturb TLR4/MyD88/NFκB signalling pathway, and it might be the mechanism by which BBR attenuated inflammation in the early phase.