NLRP3 Inflammasome Activation in Lung Vascular Endothelial Cells Contributes to Intestinal Ischemia/Reperfusion-Induced Acute Lung Injury This information is current as of September 28, 2021. Homare Ito, Hiroaki Kimura, Tadayoshi Karasawa, Shu Hisata, Ai Sadatomo, Yoshiyuki Inoue, Naoya Yamada, Emi Aizawa, Erika Hishida, Ryo Kamata, Takanori Komada, Sachiko Watanabe, Tadashi Kasahara, Takuji Suzuki, Hisanaga Horie, Joji Kitayama, Naohiro Sata, Kazuyo Yamaji-Kegan and Masafumi Takahashi Downloaded from J Immunol published online 29 July 2020 http://www.jimmunol.org/content/early/2020/07/28/jimmun ol.2000217 http://www.jimmunol.org/ Supplementary http://www.jimmunol.org/content/suppl/2020/07/28/jimmunol.200021 Material 7.DCSupplemental Why The JI? Submit online. by guest on September 28, 2021 • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2020 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. Published July 29, 2020, doi:10.4049/jimmunol.2000217 The Journal of Immunology NLRP3 Inflammasome Activation in Lung Vascular Endothelial Cells Contributes to Intestinal Ischemia/ Reperfusion-Induced Acute Lung Injury Homare Ito,*,† Hiroaki Kimura,* Tadayoshi Karasawa,* Shu Hisata,‡ Ai Sadatomo,*,† Yoshiyuki Inoue,*,† Naoya Yamada,* Emi Aizawa,* Erika Hishida,* Ryo Kamata,* Takanori Komada,* Sachiko Watanabe,* Tadashi Kasahara,* Takuji Suzuki,‡ Hisanaga Horie,† Joji Kitayama,† Naohiro Sata,† Kazuyo Yamaji-Kegan,x and Masafumi Takahashi* Intestinal ischemia/reperfusion (I/R) injury is a life-threatening complication that leads to inflammation and remote organ damage. Downloaded from The NLRP3 inflammasome regulates the caspase-1–dependent release of IL-1b, an early mediator of inflammation after I/R injury. In this study, we investigated the role of the NLRP3 inflammasome in mice with intestinal I/R injury. Deficiency of NLRP3, ASC, caspase-1/11, or IL-1b prolonged survival after intestinal I/R injury, but neither NLRP3 nor caspase-1/11 deficiency affected intestinal inflammation. Intestinal I/R injury caused acute lung injury (ALI) characterized by inflammation, reactive oxygen species generation, and vascular permeability, which was markedly improved by NLRP3 deficiency. Bone marrow chimeric experiments showed that NLRP3 in non–bone marrow–derived cells was the main contributor to development of intestinal http://www.jimmunol.org/ I/R-induced ALI. The NLRP3 inflammasome in lung vascular endothelial cells is thought to be important to lung vascular permeability. Using mass spectrometry, we identified intestinal I/R-derived lipid mediators that enhanced NLRP3 inflammasome activation in lung vascular endothelial cells. Finally, we confirmed that serum levels of these lipid mediators were elevated in patients with intestinal ischemia. To our knowledge, these findings provide new insights into the mechanism underlying intestinal I/R-induced ALI and suggest that endothelial NLRP3 inflammasome–driven IL-1b is a novel potential target for treating and preventing this disorder. The Journal of Immunology, 2020, 205: 000–000. ntestinal ischemia/reperfusion (I/R) injury is a common life- endotoxins from ischemic intestine (4, 5). However, the mecha- by guest on September 28, 2021 threatening complication that can occur as a result of many nism by which intestinal I/R induces inflammation in the intestine I clinical conditions, such as mesenteric arterial thrombosis, and distant organs remains to be elucidated. strangulated ileus, trauma, abdominal aortic aneurysm surgery, Increasing evidence indicates that the nucleotide-binding olig- and intestinal transplantation (1). Intestinal I/R not only injures the omerization domain-like receptor (NLR) family pyrin domain intestine itself but also can severely damage distant organs. containing 3 inflammasome initiates an inflammatory response in In particular, the lung is susceptible to developing acute injury the development of various diseases (6–9). The NLR family pyrin after intestinal I/R (2, 3). One prominent feature of intestinal domain containing 3 (NLRP3) inflammasome is a large, intra- I/R-induced acute lung injury (ALI) is an excessive inflammatory cellular, multiprotein complex that consists of NLRP3, the adaptor response characterized by the release of proinflammatory cyto- protein apoptosis speck-like protein containing a caspase recruit- kines, the infiltration of inflammatory cells, and bacteria-derived ing domain (ASC), and cysteine protease caspase-1. In response to *Division of Inflammation Research, Center for Molecular Medicine, Jichi Medical discussed the data and provided scientific advice. H.I. and M.T. wrote the manuscript. University, Tochigi 329-0498, Japan; †Department of Surgery, Jichi Medical Univer- All authors approved this study. sity, Tochigi 329-0498, Japan; ‡Division of Pulmonary Medicine, Department of x Address correspondence and reprint requests to Prof. Masafumi Takahashi, Division Medicine, Jichi Medical University, Tochigi 329-0498, Japan; and Department of of Inflammation Research, Center for Molecular Medicine, Jichi Medical University, 3311- Anesthesiology, University of Maryland School of Medicine, Baltimore, MD 21201 1 Yakushiji, Shimotsuke, Tochigi 329-0498, Japan. E-mail address: [email protected] ORCIDs: 0000-0001-9523-1335 (H.I.); 0000-0001-5721-2619 (A.S.); 0000-0001- The online version of this article contains supplemental material. 6970-9657 (R.K.); 0000-0003-3360-3185 (T.Komada); 0000-0001-7687- 5477 (T.Kasahara); 0000-0002-5907-4050 (H.H.); 0000-0002-5708-7130 (J.K.); Abbreviations used in this article: AA, arachidonic acid; ALI, acute lung injury; ALT, 0000-0002-8500-972X (K.Y.-K.); 0000-0003-2716-7532 (M.T.). alanine aminotransferase; ASC, adaptor protein apoptosis speck-like protein contain- ing a caspase recruiting domain; AST, aspartate aminotransferase; BALF, bronchoal- Received for publication February 27, 2020. Accepted for publication July 6, 2020. veolar lavage fluid; BMT, bone marrow transplantation; BUN, blood urea nitrogen; This work was supported by Japan Society for the Promotion of Science Grant-in-Aid EBD, Evans blue dye; Gr-1, granulocyte receptor 1; HETE, hydroxyeicosatetraenoic for Scientific Research 18K08112 (to M.T.), a Private University Research Branding acid; HMGB1, high-mobility group box 1; 4-HNE, 4-hydroxy-2-nonenal; HODE, Project (to M.T.), Japan Agency for Medical Research and Development Core Re- hydroxyoctadecadienoic acid; I/R, ischemia/reperfusion; KIM-1, kidney injury mol- search for Evolutional Science and Technology Grant 18gm0610012h0105 (to M.T.), ecule 1; LA, linoleic acid; LC/MS, liquid chromatography/mass spectrometry; LDH, the Takeda Science Foundation (to M.T.), a Jichi Medical University Graduate Stu- lactate dehydrogenase; LVEC, lung vascular endothelial cell; NLR, nucleotide- dent Start-up Award (to H.I.), and a Jichi Medical University Graduate Student binding oligomerization domain-like receptor; NLRP3, NLR family pyrin domain Research Award (to H.I.). containing 3; ROS, reactive oxygen species; SMA, superior mesenteric artery; SMV, superior mesenteric vein; VE, vascular endothelial; WT, wild-type. H.I. and M.T. designed the study concept and experiments. H.I., H.K., T. Karasawa, S.H., A.S., Y.I., N.Y., E.A., E.H., R.K., T. Komada, and S.W. performed the exper- Ó iments and analyzed the data. T. Kasahara, T.S., H.H., J.K., N.S., and K.Y.-K. Copyright 2020 by The American Association of Immunologists, Inc. 0022-1767/20/$37.50 www.jimmunol.org/cgi/doi/10.4049/jimmunol.2000217 2 NLRP3 INFLAMMASOME IN INTESTINAL I/R-INDUCED ALI danger signals, NLRP3 inflammasome components are assembled the liver, heart, and kidney were assessed as described previously (29–31). to activate caspase-1 with subsequent maturation of IL-1b (pro- The severity of lung injury was graded according to the following pa- cessing to its active form), resulting in inflammation and tissue rameters (4): 1) periluminal infiltrates (around the airway/vessels), 2) pneumonitis (alveolar/interstitial), and 3) percentage of affected lung tis- damage. Our group has recently shown that the NLRP3 inflam- sue. Immunohistochemical analysis was performed as described previously masome is involved in the pathogenesis of several disease con- (32, 33) with Abs to the following proteins: Ly-6G (BioLegend, San ditions (10–15). In terms of I/R injury, we and other investigators Diego, CA), CD45 (BD Biosciences, Franklin Lakes, NJ), F4/80 (clone showed that the NLRP3 inflammasome mediates the initial in- A3-1; AbD Serotec), granulocyte receptor 1 (Gr-1; eBioscience, San Diego, CA), kidney injury molecule 1 (KIM-1; R&D Systems, Minneap- flammatory response after I/R in the heart, liver, kidney, and in- olis, MN), cleaved caspase-3 (Cell Signaling Technology, Boston, MA), testine (10, 16–18). Furthermore, several reports have shown that and 4-hydroxy-2-nonenal (4-HNE;