Sebti Yasmine, Ferri Lise, Zecchin Mathilde, Beauchamp Justine, Mogilenko Denis, Staels Bart, Duez Hélène, Pourcet Benoit
European Genomic Institute for Diabetes (E.G.I.D.), Lille, France.
UNIV LILLE, Lille, France.
Methods Mol Biol. 2019;1951:189-207. doi: 10.1007/978-1-4939-9130-3_15.
The NLRP3 inflammasome is a cellular sensor of danger signals such as extracellular ATP or abnormally accumulating molecules like crystals. Activation of NLRP3 by such compounds triggers a sterile inflammatory response that may be involved in numerous pathologies including rheumatoid arthritis, atherosclerosis, diabetes, and Alzheimer's disease. A better understanding of the mechanisms that govern NLRP3 inflammasome activation is an important step toward the development of novel therapeutic strategies to dampen over-activation of the immune system. Recent findings demonstrate that ligand-activated nuclear receptors regulate the NLRP3 inflammasome pathway, thus representing possible therapeutic targets. It is therefore important to assess the potential of these putative targets in the regulation of the NLRP3 inflammasome activation in the most appropriate pathophysiological models. Fulminant hepatitis (FH) results from massive hepatocyte apoptosis, hemorrhagic necrosis, and inflammation. Low doses of LPS in combination with the specific hepatotoxic agent D-galactosamine (D-GalN) promote liver injury in mice and induce the production of inflammatory cytokines associated with increased NLRP3 protein and caspase 1 activity, thus recapitulating the clinical picture of FH in humans. We provide a simple method to examine the involvement of nuclear receptors in NLRP3-driven fulminant hepatitis, consisting in the induction of FH, in the isolation of liver macrophages, and in the extraction and analysis of RNA content.
NLRP3炎性小体是一种细胞危险信号传感器，可感应细胞外ATP或晶体等异常累积的分子。此类化合物激活NLRP3会引发无菌性炎症反应，这可能与包括类风湿性关节炎、动脉粥样硬化、糖尿病和阿尔茨海默病在内的多种病理状况有关。更好地理解调控NLRP3炎性小体激活的机制，是开发新型治疗策略以抑制免疫系统过度激活的重要一步。最近的研究结果表明，配体激活的核受体调节NLRP3炎性小体途径，因此代表了可能的治疗靶点。因此，在最合适的病理生理模型中评估这些假定靶点在调控NLRP3炎性小体激活方面的潜力非常重要。暴发性肝炎（FH）是由大量肝细胞凋亡、出血性坏死和炎症引起的。低剂量脂多糖（LPS）与特异性肝毒性剂D-半乳糖胺（D-GalN）联合使用可促进小鼠肝损伤，并诱导与NLRP3蛋白增加和半胱天冬酶1活性增加相关的炎性细胞因子产生，从而重现人类FH的临床症状。我们提供了一种简单的方法来检测核受体在NLRP3驱动的暴发性肝炎中的作用，包括诱导FH、分离肝巨噬细胞以及提取和分析RNA含量。
