中性粒细胞胞外诱捕网(NETs)是机体对抗外源性细菌的重要手段。然而,研究表明,过量的NETs释放会损害其他细胞。越来越多的证据表明丁酸可以减轻细胞的炎症反应。然而,丁酸对金黄色葡萄球菌诱导的NETs形成的影响及其潜在机制尚不清楚。在这项研究中,西方印迹,免疫荧光和CCK-8测定用于检查丁酸钠(NaB)对NETs形成的影响。结果表明,NaB抑制了金黄色葡萄球菌诱导的NETs形成的释放,正如DNA水平下降所表明的那样,组蛋白,髓过氧化物酶,和中性粒细胞弹性蛋白酶.金黄色葡萄球菌可以诱导自噬,自噬在NETs的形成中起关键作用。我们的数据显示,NaB激活哺乳动物雷帕霉素靶蛋白(mTOR)和Ser757处的激酶蛋白激酶B(AKT)和unc-51样激酶1(ULK1),并抑制AMP激活的蛋白激酶(AMPK)。探讨NaB是否通过抑制自噬抑制NETs的形成,我们添加了3-甲基腺嘌呤(自噬抑制剂)(3-MA,5mM)对牛中性粒细胞,结果表明,3-MA显著抑制NETs的释放。此外,我们发现NETs及其组蛋白对奶牛乳腺上皮细胞(BMECs)的细胞毒作用显著增强,表明NETs及其组蛋白在BMEC损伤中起关键作用。总之,NaB可以通过抑制自噬减少NETs的过度形成,从而减少NET对BMEC的破坏作用。
Neutrophil extracellular traps (NETs) are an important means by which the body fights against exogenous bacteria. However, studies have shown that excessive NETs release can damage other cells. Accumulating evidence has shown that butyric acid can alleviate the inflammatory response of cells. However, the effect of butyric acid on Staphylococcus aureus-induced NETs formation and its underlying mechanism are still unclear. In this study, western blotting, immunofluorescence and CCK-8 assays were used to examine the effect of NETs formation by sodium butyrate (
NaB). The results showed that
NaB suppressed the release of S. aureus-induced NETs formation, as indicated by decreases in the levels of DNA, histones, myeloperoxidase, and neutrophil elastase. S. aureus can induce autophagy, and autophagy plays a key role in the formation of NETs. Our data showed that
NaB activated mammalian target of rapamycin (mTOR) and the kinases protein kinase B (AKT) and unc-51 like kinase 1 (ULK1) at Ser757 and inhibited AMP-activated protein kinase (AMPK). To explore whether
NaB inhibited the formation of NETs by inhibiting autophagy, we added 3-methyladenine (autophagy inhibitor) (3-MA, 5 mM) to bovine neutrophils, and the results showed that 3-MA significantly inhibited NETs release. Furthermore, we found that NETs and their component histones exhibited significantly increased the cytotoxic effects on bovine mammary epithelial cells (BMECs), indicating that NETs and their component histones play a key role in BMEC damage. In conclusion,
NaB can reduce the excessive formation of NETs by inhibiting autophagy, thus reducing the damaging effect of NETs on BMECs.