College of Pharmacy, Chonnam National University, Gwangju, Republic of Korea.
Department of Bioengineering and Biotechnology, College of Engineering, Chonnam National University, Gwangju 61186, Republic of Korea.
Metabolism. 2023 May;142:155527. doi: 10.1016/j.metabol.2023.155527. Epub 2023 Mar 3.
Obesity is a state of chronic low-grade systemic inflammation. Recent studies showed that NLRP3 inflammasome initiates metabolic dysregulation in adipose tissues, primarily through activation of adipose tissue infiltrated macrophages. However, the mechanism of NLRP3 activation and its role in adipocytes remains elusive. Therefore, we aimed to examine the activation of TNFα-induced NLRP3 inflammasome in adipocytes and its role on adipocyte metabolism and crosstalk with macrophages.
The effect of TNFα on adipocyte NLRP3 inflammasome activation was measured. Caspase-1 inhibitor (Ac-YVAD-cmk) and primary adipocytes from NLRP3 and caspase-1 knockout mice were utilized to block NLRP3 inflammasome activation. Biomarkers were measured by using real-time PCR, western blotting, immunofluorescence staining, and enzyme assay kits. Conditioned media from TNFα-stimulated adipocytes was used to establish the adipocyte-macrophage crosstalk. Chromatin immunoprecipitation assay was used to identify the role of NLRP3 as a transcription factor. Mouse and human adipose tissues were collected for correlation analysis.
TNFα treatment induced NLRP3 expression and caspase-1 activity in adipocytes, partly through autophagy dysregulation. The activated adipocyte NLRP3 inflammasome participated in mitochondrial dysfunction and insulin resistance, as evidenced by the amelioration of these effects in Ac-YVAD-cmk treated 3T3-L1 cells or primary adipocytes isolated from NLRP3 and caspase-1 knockout mice. Particularly, the adipocyte NLRP3 inflammasome was involved in glucose uptake regulation. Also, TNFα induced expression and secretion of lipocalin 2 (Lcn2) in a NLRP3-dependent manner. NLRP3 could bind to the promoter and transcriptionally regulate Lcn2 in adipocytes. Treatment with adipocyte conditioned media revealed that adipocyte-derived Lcn2 was responsible for macrophage NLRP3 inflammasome activation, working as a second signal. Adipocytes isolated from high-fat diet mice and adipose tissue from obese individuals showed a positive correlation between NLRP3 and Lcn2 gene expression.
This study highlights the importance of adipocyte NLRP3 inflammasome activation and novel role of TNFα-NLRP3-Lcn2 axis in adipose tissue. It adds rational for the current development of NLRP3 inhibitors for treating obesity-induced metabolic diseases.
肥胖是一种慢性低度全身炎症状态。最近的研究表明，NLRP3 炎性小体在脂肪组织中引发代谢失调，主要通过激活脂肪组织浸润的巨噬细胞。然而，NLRP3 的激活机制及其在脂肪细胞中的作用仍不清楚。因此，我们旨在研究 TNFα 诱导的脂肪细胞 NLRP3 炎性小体的激活及其对脂肪细胞代谢和与巨噬细胞相互作用的影响。
测定 TNFα 对脂肪细胞 NLRP3 炎性小体激活的影响。使用 caspase-1 抑制剂（Ac-YVAD-cmk）和 NLRP3 和 caspase-1 敲除小鼠的原代脂肪细胞来阻断 NLRP3 炎性小体的激活。使用实时 PCR、western blot、免疫荧光染色和酶试剂盒测量生物标志物。使用 TNFα 刺激的脂肪细胞的条件培养基建立脂肪细胞-巨噬细胞串扰。染色质免疫沉淀测定用于确定 NLRP3 作为转录因子的作用。收集小鼠和人脂肪组织进行相关性分析。
TNFα 处理诱导脂肪细胞中 NLRP3 的表达和 caspase-1 的活性，部分通过自噬失调。激活的脂肪细胞 NLRP3 炎性小体参与线粒体功能障碍和胰岛素抵抗，这可通过 Ac-YVAD-cmk 处理的 3T3-L1 细胞或从 NLRP3 和 caspase-1 敲除小鼠分离的原代脂肪细胞中的这些作用得到改善。特别是，脂肪细胞 NLRP3 炎性小体参与葡萄糖摄取调节。此外，TNFα 以 NLRP3 依赖性方式诱导脂联素 2（Lcn2）的表达和分泌。NLRP3 可以与启动子结合并在脂肪细胞中转录调节 Lcn2。用脂肪细胞条件培养基处理显示，脂肪细胞衍生的 Lcn2 负责巨噬细胞 NLRP3 炎性小体的激活，作为第二信号。从高脂肪饮食小鼠分离的脂肪细胞和肥胖个体的脂肪组织中 NLRP3 和 Lcn2 基因表达之间存在正相关。
本研究强调了脂肪细胞 NLRP3 炎性小体激活的重要性以及 TNFα-NLRP3-Lcn2 轴在脂肪组织中的新作用。这为当前开发 NLRP3 抑制剂治疗肥胖引起的代谢性疾病提供了合理依据。
