Carlos Daniela, Costa Frederico R C, Pereira Camila A, Rocha Fernanda A, Yaochite Juliana N U, Oliveira Gabriela G, Carneiro Fernando S, Tostes Rita C, Ramos Simone G, Zamboni Dario S, Camara Niels O S, Ryffel Bernhard, Silva João S
Department of Biochemistry and Immunology, Ribeirão Preto Medical School, University of São Paulo , Ribeirao Preto , Brazil.
Department of Pharmacology, Ribeirão Preto Medical School, University of São Paulo , Ribeirao Preto , Brazil.
Front Immunol. 2017 Feb 27;8:164. doi: 10.3389/fimmu.2017.00164. eCollection 2017.
Although a correlation between polymorphisms of NOD-like receptor family-pyrin domain containing 3 (NLRP3) and predisposition to type 1 diabetes (T1D) has been identified, the potential function and activation of the NLRP3 inflammasome in T1D have not been clarified. The present study shows that non-obese diabetic mice exhibited increased NLRP3, and pro-IL-1β gene expression in pancreatic lymph nodes (PLNs). Similar increases in gene expression of NLRP3, apoptosis associated speck like protein (ASC) and pro-IL-1β were induced by multiple low doses of streptozotocin (STZ) in C57BL/6 mice. In addition, diabetic C57BL/6 mice also exhibited increased IL-1β protein expression in the pancreatic tissue at day 7, which remained elevated until day 15. Diabetic mice also showed increased positive caspase-1 macrophages in the PLNs, which were decreased in NLRP3 mice, but not in ASC mice, after STZ treatment. NLRP3- and IL-1R-deficient mice, but not ASC-deficient mice, showed reduced incidence of diabetes, less insulitis, lower hyperglycemia, and normal insulin levels compared to wild-type (WT) diabetic mice. Notably, these mice also displayed a decrease in IL-17-producing CD4 and CD8 T cells (Th17 and Tc17) and IFN-γ-producing CD4 and CD8 T cells (Th1 and Tc1) in the PLNs. Following STZ treatment to induce T1D, NLRP3-deficient mice also exhibited an increase in myeloid-derived suppressor cell and mast cell numbers in the PLNs along with a significant increase in IL-6, IL-10, and IL-4 expression in the pancreatic tissue. Interestingly, diabetic mice revealed increased circulating expression of genes related to mitochondrial DNA, such as cytochrome and cytochrome , but not NADH dehydrogenase subunit 6 (NADH). Mitochondrial DNA (mDNA) from diabetic mice, but not from non-diabetic mice, induced significant IL-1β production and caspase-1 activation by WT macrophages, which was reduced in NLRP3 macrophages. Finally, mDNA administration increased Th17/Tc17/Th1/Tc1 cells in the PLNs and precipitated T1D onset, which was abolished in NLRP3 mice. Overall, our results demonstrate that mDNA-mediated NLRP3 activation triggers caspase-1-dependent IL-1β production and contributes to pathogenic cellular responses during the development of STZ-induced T1D.
尽管已确定含NOD样受体家族吡啉结构域3（NLRP3）的多态性与1型糖尿病（T1D）易感性之间存在关联，但NLRP3炎性小体在T1D中的潜在功能和激活机制尚未阐明。本研究表明，非肥胖糖尿病小鼠胰腺淋巴结（PLN）中NLRP3和前白细胞介素-1β（pro-IL-1β）基因表达增加。在C57BL/6小鼠中，多次低剂量链脲佐菌素（STZ）诱导了NLRP3、凋亡相关斑点样蛋白（ASC）和pro-IL-1β基因表达的类似增加。此外，糖尿病C57BL/6小鼠在第7天时胰腺组织中白细胞介素-1β蛋白表达也增加，直至第15天一直保持升高。糖尿病小鼠PLN中活化的半胱天冬酶-1阳性巨噬细胞也增加，STZ处理后，NLRP3基因敲除小鼠中的该细胞减少，而ASC基因敲除小鼠中的未减少。与野生型（WT）糖尿病小鼠相比，NLRP3和白细胞介素-1受体（IL-1R）缺陷小鼠的糖尿病发病率降低、胰岛炎减轻、高血糖水平降低且胰岛素水平正常。值得注意的是，这些小鼠PLN中产生白细胞介素-17的CD4和CD8 T细胞（Th17和Tc17）以及产生干扰素-γ的CD4和CD8 T细胞（Th1和Tc1）也减少。在STZ处理诱导T1D后，NLRP3缺陷小鼠PLN中髓系来源的抑制细胞和肥大细胞数量也增加，同时胰腺组织中白细胞介素-6、白细胞介素-10和白细胞介素-4表达显著增加。有趣的是，糖尿病小鼠循环中与线粒体DNA相关的基因表达增加，如细胞色素 和细胞色素 ，但烟酰胺腺嘌呤二核苷酸（NADH）脱氢酶亚基6（NADH）未增加。糖尿病小鼠而非非糖尿病小鼠的线粒体DNA（mDNA）可诱导WT巨噬细胞产生显著的白细胞介素-1β并激活半胱天冬酶-1，而在NLRP3巨噬细胞中这种作用减弱。最后，给予mDNA增加了PLN中的Th17/Tc17/Th1/Tc1细胞并加速了T1D发病，而在NLRP3基因敲除小鼠中这种作用消失。总体而言，我们的结果表明，mDNA介导的NLRP3激活触发了半胱天冬酶-1依赖性白细胞介素-1β的产生，并在STZ诱导的T1D发展过程中促成了致病性细胞反应。
