title: NLRP3 activation induces ASC-dependent programmed necrotic cell death, which leads to neutrophilic inflammation.
abstract: NLR family pyrin domain containing 3 (NLRP3) is a cytoplasmic pattern recognition receptor that regulates innate immune responses by forming a protein complex, the inflammasome. It leads to production of proinflammatory cytokine productions such as interleukin 1β (IL-1β). We and others demonstrated that an induction of activated NLRP3 also induced cell death. However, little is known about the characteristics and mechanisms of the cell death and its involvement in the pathogenesis of inflammatory conditions. In this study, we established cell lines in which NLRP3 was induced by doxycycline using a tetracycline-inducible expression (Tet-on) system. Using this system, the expression of NLRP3 mutants in cryopyrin-associated periodic syndrome (CAPS) patients was sufficient for the induction of necrotic cell death without lipopolysaccharide stimulation or generation of mature IL-1β. We also found that CA074-Me, a cathepsin B inhibitor, blocked cell death before oligomerization of apoptosis-associated speck-like protein containing a CARD (ASC), whereas Z-VAD-fmk, a pan-caspase inhibitor, blocked the cell death after the oligomerization. Silencing of the ASC gene (Pycard) by small hairpin RNA treatment inhibited the NLRP3 mutant-induced cell death, but silencing of the caspase-1 gene (Casp1) did not. Taken together, these results indicated that ASC was indispensable for NLRP3-mediated programmed necrotic cell death, and that this type of cell death was distinct from 'pyroptosis', which requires caspase-1. Finally, we demonstrated in an in vivo model that the programmed necrotic cell death induced by activated NLRP3 could cause neutrophil infiltration, indicating a possible role of cell death in neutrophil infiltration of skin lesions in CAPS patients.
authors.affiliation: Department of Dermatology, Chiba University Graduate School of Medicine, Chiba, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Satoh
authors.forename: N
authors.initials: N
authors.lastname: Kambe
authors.forename: H
authors.initials: H
authors.lastname: Matsue
mesh_terms: D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D051017:Apoptosis Regulatory Proteins; D053468:CARD Signaling Adaptor Proteins; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D020170:Caspase 1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D061945:Caspase Inhibitors / Q000494:pharmacology; D003598:Cytoskeletal Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D004151:Dipeptides / Q000494:pharmacology; D004318:Doxycycline / Q000494:pharmacology; D006801:Humans; D007249:Inflammation; D053583:Interleukin-1beta / Q000378:metabolism; D051379:Mice; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; D009336:Necrosis; D009504:Neutrophils / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism
chemical_list: D000590:Amino Acid Chloromethyl Ketones; D051017:Apoptosis Regulatory Proteins; C400541:CA 074 methyl ester; D053468:CARD Signaling Adaptor Proteins; D002352:Carrier Proteins; D061945:Caspase Inhibitors; D003598:Cytoskeletal Proteins; D004151:Dipeptides; D053583:Interleukin-1beta; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; C492716:Nlrp3 protein, mouse; C498534:Pycard protein, mouse; D034741:RNA, Small Interfering; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D020170:Caspase 1; D004318:Doxycycline