@article{c7b1fe72dff94507a9c140f86ba7817c,
title = "ATP exposure stimulates glutathione efflux as a necessary switch for NLRP3 inflammasome activation",
abstract = "The NLRP3 inflammasome is a multiprotein complex responsible for the maturation of precursor forms of interleukin (IL)-1β and IL-18 into active proinflammatory cytokines. Increasing evidence suggests that modulation of redox homeostasis contributes to the activation of the NLRP3 inflammasome. However, specific mechanistic details remain unclear. We demonstrate here that ATP exposure evoked a sharp decrease in glutathione (GSH) levels in macrophages, which led to NLRP3 inflammasome activation. We detected an increase in GSH levels in culture supernatants that was comparable to the GSH decrease in macrophages, which suggests that exposure to ATP stimulated GSH efflux. Exogenous addition of P2X7 receptor antagonist, GSH, or the oxidized form GSSG attenuated this efflux. Also, exogenous GSH or GSSG strongly inhibited NLRP3 inflammasome activation in vitro and in vivo. These data suggest that GSH efflux controls NLRP3 inflammasome activation, which may lead to development of novel therapeutic strategies for NLRP3 inflammasome-associated disorders.",
keywords = "ATP, GSH efflux, Glutathione, NLRP3 inflammasome, Reactive oxygen species, Redox regulation",
author = "Tianli Zhang and Hiroyasu Tsutsuki and Waliul Islam and Katsuhiko Ono and Kohsuke Takeda and Takaaki Akaike and Tomohiro Sawa",
note = "Funding Information: We thank J.B. Gandy for her editing of the manuscript. This work was supported in part by Grants-in-Aid for Scientific Research ([B], [C], Challenging Exploratory Research) from the Ministry of Education, Science, Sports, and Technology ( MEXT ), Japan ( 18H02098 , 19K22258 ), the Japan Society for the Promotion of Science ( 20F20408 ), and the Amabie Research Grant from Kumamoto University to T.S.; and by the JSPS International Research Fellow and a grant from the Otsuka Toshimi Scholarship Foundation to T.Z. Funding Information: We next evaluated the previous result implicating GSH transporters as possible candidates inducing GSH efflux. We added MK571, an inhibitor of multidrug resistance proteins (common GSH transporters), to cells during activation of the NLRP3 inflammasome. However, we observed no marked difference in GSH efflux after ATP treatment between cells either treated with MK571 or not treated, as Fig. S5A demonstrates. We also found that both p-aminohippuric acid (PAH) [inhibitor of organic anion transporters (OATs)] and rifamycin SV (inhibitor of OAT polypeptides) failed to prevent ATP-induced GSH efflux (Figs. S5B and S5C). Similar results were observed when cells were incubated with inhibitors of other ATP-binding cassette (ABC) transporters, including glibenclamide (cystic fibrosis transmembrane conductance regulator inhibitor) and Ko143 (ABCG2 inhibitor) (Figs. S5D and S5E). Moreover, two possible channel blockers—phloretin (volume-regulated anion channel blocker) and 4,4ʹ-diisothiocyanatostilbene-2,2ʹ-disulfonic acid disodium salt hydrate (DIDS) (maxi-channel blocker)—did not affect GSH efflux in NLRP3 inflammasome-activated J774.1 cells (Figs. S5F and S5G). ATP-induced GSH efflux was not associated with the endoplasmic reticulum (ER)-Golgi transport system, as supported by the finding of no substantial alteration of GSH efflux when cells were treated with ATP in the presence of brefeldin A (BFA) (ER-Golgi protein trafficking inhibitor) (Fig. S5H).We thank J.B. Gandy for her editing of the manuscript. This work was supported in part by Grants-in-Aid for Scientific Research ([B], [C], Challenging Exploratory Research) from the Ministry of Education, Science, Sports, and Technology (MEXT), Japan (18H02098, 19K22258), the Japan Society for the Promotion of Science (20F20408), and the Amabie Research Grant from Kumamoto University to T.S.; and by the JSPS International Research Fellow and a grant from the Otsuka Toshimi Scholarship Foundation to T.Z. Publisher Copyright: {\textcopyright} 2021 The Author(s)",
year = "2021",
month = may,
doi = "10.1016/j.redox.2021.101930",
language = "English",
volume = "41",
journal = "Redox Biology",
issn = "2213-2317",
publisher = "Elsevier BV",
}