Metabolic inflammation is associated with increased expression of saturated free fatty acids, proinflammatory cytokines, chemokines, and adipose oxidative stress. Macrophage inflammatory protein (MIP)-1α recruits the inflammatory cells such as monocytes, macrophages, and neutrophils in the adipose tissue; however, the mechanisms promoting the MIP-1α expression remain unclear. We hypothesized that MIP-1α co-induced by palmitate and tumor necrosis factor (TNF)-α in monocytic cells/macrophages could be further enhanced in the presence of reactive oxygen species (ROS)-mediated oxidative stress. To investigate this, THP-1 monocytic cells and primary human macrophages were co-stimulated with palmitate and TNF-α and mRNA and protein levels of MIP-1α were measured by using quantitative reverse transcription, polymerase chain reaction (qRT-PCR) and commercial enzyme-linked immunosorbent assays (ELISA), respectively. The cognate receptor of palmitate, toll-like receptor (TLR)-4, was blunted by genetic ablation, neutralization, and chemical inhibition. The involvement of TLR4-downstream pathways, interferon regulatory factor (IRF)-3 or myeloid differentiation (MyD)-88 factor, was determined using IRF3-siRNA or MyD88-deficient cells. Oxidative stress was induced in cells by hydrogen peroxide (H₂O₂) treatment and ROS induction was measured by dichloro-dihydro-fluorescein diacetate (DCFH-DA) assay. The data show that MIP-1α gene/protein expression was upregulated in cells co-stimulated with palmitate/TNF-α compared to those stimulated with either palmitate or TNF-α (P < 0.05). Further, TLR4-IRF3 pathway was implicated in the cooperative induction of MIP-1α in THP-1 cells, and this cooperativity between palmitate and TNF-α was clathrin-dependent and also required signaling through c-Jun and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB). Notably, ROS itself induced MIP-1α and could further promote MIP-1α secretion together with palmitate and TNF-α. In conclusion, palmitate and TNF-α co-induce MIP-1α in human monocytic cells via the TLR4-IRF3 pathway and signaling involving c-Jun/NF-κB. Importantly, oxidative stress leads to ROS-driven MIP-1α amplification, which may have significance for metabolic inflammation.

中文翻译：

---

棕榈酸酯和TNF-α共同刺激人单核细胞诱导的MIP-1α表达涉及TLR4-IRF3途径，并被氧化应激放大。

代谢性炎症与饱和游离脂肪酸，促炎细胞因子，趋化因子和脂肪氧化应激的表达增加有关。巨噬细胞炎性蛋白（MIP）-1α在脂肪组织中募集炎性细胞，例如单核细胞，巨噬细胞和中性粒细胞。但是，促进MIP-1α表达的机制尚不清楚。我们假设，在存在活性氧（ROS）介导的氧化应激的情况下，棕榈酸酯和肿瘤坏死因子（TNF）-α在单核细胞/巨噬细胞中共同诱导的MIP-1α可以进一步增强。为了研究这一点，将THP-1单核细胞和原代人巨噬细胞与棕榈酸酯共刺激，并通过定量逆转录测定TNF-α和MIP-1α的mRNA和蛋白水平，聚合酶链反应（qRT-PCR）和商业酶联免疫吸附测定（ELISA）。棕榈酸酯的同源受体，toll​​-like受体（TLR）-4，由于遗传消融，中和和化学抑制作用而减弱。使用IRF3-siRNA或MyD88缺陷型细胞确定了TLR4下游途径，干扰素调节因子（IRF）-3或骨髓分化（MyD）-88因子的参与。过氧化氢（H 使用IRF3-siRNA或MyD88缺陷细胞确定。过氧化氢（H 使用IRF3-siRNA或MyD88缺陷细胞确定。过氧化氢（H₂ O ₂）处理和ROS诱导通过二氯二氢荧光素二乙酸酯（DCFH-DA）测定来测量。数据显示，与棕榈酸酯或TNF-α刺激的细胞相比，与棕榈酸酯/TNF-α共同刺激的细胞中MIP-1α基因/蛋白表达上调（P<0.05）。此外，TLR4-IRF3途径与THP-1细胞中MIP-1α的协同诱导有关，并且棕榈酸酯和TNF-α之间的这种协同作用是网格蛋白依赖性的，并且还需要通过c-Jun和核因子kappa-light-激活的B细胞（NF-κB）的链增强剂。值得注意的是，ROS本身会诱导MIP-1α并与棕榈酸酯和TNF-α一起进一步促进MIP-1α的分泌。总之，棕榈酸酯和TNF-α通过TLR4-IRF3途径和涉及c-Jun /NF-κB的信号传导在人单核细胞中共诱导MIP-1α。重要的是，氧化应激会导致ROS驱动的MIP-1α扩增，这可能对代谢性炎症具有重要意义。