Autophagy Inhibition Contributes to ROS-Producing NLRP3-Dependent Inflammasome Activation and Cytokine Secretion in High Glucose-Induced Macrophages
Autophagy Inhibition Contributes to ROS-Producing NLRP3-Dependent Inflammasome Activation and Cytokine Secretion in High Glucose-Induced Macrophages
复制标题
自噬抑制有助于高糖诱导的巨噬细胞中 ROS 产生的 NLRP3 依赖性炎症小体激活和细胞因子分泌
DOI：
10.1159/000480367
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发表时间：
2017-01-01
影响因子：
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通讯作者：
Chai, Yimin
中科院分区：
文献类型：
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作者：
Dai, Jiezhi;Zhang, Xiaotian;Chai, Yimin
背景：2型糖尿病是一种损害愈合过程的持续性炎症反应。我们假设，在促炎信号之后用高糖刺激会导致自噬抑制、活性氧(ROS)的产生，并最终激活NOD样受体蛋白(NLRP)-3。方法：从人糖尿病创面分离巨噬细胞。用免疫印迹和实时荧光定量聚合酶链式反应检测NLRP3、caspase1和IL-1β的表达，用流式细胞仪检测细胞表面标志。将THP-1诱导的巨噬细胞暴露于高糖环境中，研究自噬、ROS和NLRP3激活之间的关系。免疫印迹和实时定量聚合酶链式反应检测Lc3-II、P62、NLRP3炎症反应和IL-1β的表达。用细胞内活性氧检测试剂盒检测ROS的产生。结果：从糖尿病创面分离的巨噬细胞表现出促炎表型，包括持续的NLRP3炎症体活性与IL-1β的分泌有关。我们的数据显示，高糖抑制了THP-1来源的巨噬细胞的自噬，诱导了ROS的产生，并激活了NLRP3炎症体和细胞因子的分泌。为了研究高糖诱导的NLRP3炎症小体信号，我们使用自噬诱导剂、ROS抑制剂和NLRP3抑制剂进行研究，发现所有这些都减少了NLRP3炎症小体的激活和细胞因子的分泌。结论：创面巨噬细胞中持续的NLRP3炎性小体活性参与了糖尿病创面的高炎症反应。自噬抑制和ROS的产生在高糖诱导的NLRP3炎症小体激活和巨噬细胞分泌细胞因子中起重要作用。(C)2017年作者(S)由S.Karger AG，巴塞尔出版
Background: Type 2 diabetes is a persistent inflammatory response that impairs the healing process. We hypothesized that stimulation with high glucose following a pro-inflammatory signal would lead to autophagy inhibition, reactive oxygen species (ROS) production and eventually to the activation of the Nod-like receptor protein (NLRP) -3. Methods: Macrophages were isolated from human diabetic wound. We measured the expression of NLRP3, caspase1 and interleukin-1 beta (IL-1 beta) by western blot and real-time PCR, and the surface markers on cells by flow cytometry. THP-1-derived macrophages exposed to high glucose were applied to study the link between autophagy, ROS and NLRP3 activation. LC3-II, P62, NLRP3 inflammation and IL-1 beta expression were measured by western blot and real-time PCR. ROS production was measured with a Cellular Reactive Oxygen Species Detection Assay Kit. Results: Macrophages isolated from diabetic wounds exhibited a pro-inflammatory phenotype, including sustained NLRP3 inflammasome activity associated with IL-1 beta secretion. Our data showed that high glucose inhibited autophagy, induced ROS production, and activated NLRP3 inflammasome and cytokine secretion in THP-1-derived macrophages. To study high glucose-induced NLRP3 inflammasome signalling, we performed studies using an autophagy inducer, a ROS inhibitor and a NLRP3 inhibitor and found that all reduced the NLRP3 inflammasome activation and cytokine secretion. Conclusion: Sustained NLRP3 inflammasome activity in wound-derived macrophages contributes to the hyper-inflammation in human diabetic wounds. Autophagy inhibition and ROS generation play an essential role in high glucose-induced NLRP3 inflammasome activation and cytokine secretion in macrophages. (C) 2017 The Author(s) Published by S. Karger AG, Basel