Molecular Cell Biology Unit, Laboratory of Molecular Immunology, National Institute of Allergy and Infectious Diseases and National Institutes of Health, Bethesda, Maryland, United States of America.
PLoS One. 2012;7(2):e30404. doi: 10.1371/journal.pone.0030404. Epub 2012 Feb 9.
Toll-like receptors (TLRs) that recognize pathogen associated molecular patterns and chemoattractant receptors (CKRs) that orchestrate leukocyte migration to infected tissue are two arms of host innate immunity. Although TLR signaling induces synthesis and secretion of proinflammatory cytokines and chemokines, which recruit leukocytes, many studies have reported the paradoxical observation that TLR stimulation inhibits leukocyte chemotaxis in vitro and impairs their recruitment to tissues during sepsis. There is consensus that physical loss of chemokine receptor (CKR) at the RNA or protein level or receptor usage switching are the mechanisms underlying this effect. We show here that a brief (<15 min) stimulation with LPS (lipopolysaccharide) at ~0.2 ng/ml inhibited chemotactic response from CCR2, CXCR4 and FPR receptors in monocytes without downmodulation of receptors. A 3 min LPS pre-treatment abolished the polarized accumulation of F-actin, integrins and PIP(3) (phosphatidylinositol-3,4,5-trisphosphate) in response to chemokines in monocytes, but not in polymorphonuclear neutrophils (PMNs). If chemoattractants were added before or simultaneously with LPS, chemotactic polarization was preserved. LPS did not alter the initial G-protein signaling, or endocytosis kinetics of agonist-occupied chemoattractant receptors (CKRs). The chemotaxis arrest did not result from downmodulation of receptors or from inordinate increase in adhesion. LPS induced rapid p38 MAPK activation, global redistribution of activated Rap1 (Ras-proximate-1 or Ras-related protein 1) GTPase and Rap1GEF (guanylate exchange factor) Epac1 (exchange proteins activated by cyclic AMP) and disruption of intracellular gradient. Co-inhibition of p38 MAPK and Rap1 GTPase reversed the LPS induced breakdown of chemotaxis suggesting that LPS effect requires the combined function of p38 MAPK and Rap1 GTPase.
模式识别受体（TLRs）识别病原体相关分子模式，趋化因子受体（CKRs）协调白细胞迁移到感染组织，它们是宿主固有免疫的两个组成部分。虽然 TLR 信号诱导合成和分泌前炎症细胞因子和趋化因子，从而募集白细胞，但许多研究报告了一个矛盾的观察结果，即 TLR 刺激在体外抑制白细胞趋化性，并损害其在败血症期间向组织的募集。人们普遍认为，趋化因子受体（CKR）在 RNA 或蛋白质水平上的物理丧失或受体使用转换是产生这种效应的机制。我们在这里表明，用 LPS（脂多糖）短暂（<15 分钟）刺激约 0.2ng/ml 抑制了单核细胞中 CCR2、CXCR4 和 FPR 受体的趋化反应，而不降低受体水平。3 分钟的 LPS 预处理可消除单核细胞中对趋化因子的 F-肌动蛋白、整合素和 PIP(3)（磷脂酰肌醇-3,4,5-三磷酸）的极化积累，但不能消除多形核白细胞（PMN）中的极化积累。如果趋化因子在 LPS 之前或同时添加，则保留趋化性极化。LPS 不改变初始 G 蛋白信号或激动剂占据的趋化因子受体（CKRs）的内吞动力学。趋化性阻滞不是由于受体下调或过度增加粘附引起的。LPS 诱导快速的 p38 MAPK 激活，激活的 Rap1（Ras-proximate-1 或 Ras-related protein 1）GTPase 的全球重分布和 Rap1GEF（环 AMP 激活的交换蛋白）以及细胞内梯度的破坏。p38 MAPK 和 Rap1 GTPase 的共同抑制逆转了 LPS 诱导的趋化性破坏，表明 LPS 效应需要 p38 MAPK 和 Rap1 GTPase 的联合功能。
