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. 2018 Apr 5;73(8):758–768. doi: 10.1136/thoraxjnl-2017-210326

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© Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/

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Aspergillus fumigatus (Af)-induced nucleotide-binding domain, leucine-rich-containing family, pyrin-domain-containing-3 (NLRP3) inflammasome assembly/activation is regulated by phosphoinositide 3-kinase (PI3K)-δ in epithelial cells (EpCs). (A and B) Representative confocal images of tracheal EpCs show the localisation of NLRP3 (red), caspase-1 (Casp-1; green) and apoptosis-associated speck-like protein containing a carboxy-terminal caspase-recruitment domain (ASC; green) in the control (no treatment), Af-stimulated cells (Af), Af-stimulated cells administered drug vehicle (Af+VEH), and Af-stimulated cells administered IC87114 (Af+IC). 4',6-Diamidino-2-phenylindole (DAPI) stain was used for nuclear localisation. Bars indicate 20 µm. (C) Representative immunoblot of ASC monomers and high order complexes in cross-linked cytosolic pellets from Af-stimulated EpCs in the presence and absence of IC87114. (D) Representative confocal images of tracheal EpCs show the localisation of NLRP3 (red) and Casp-1 (green) in the control (no treatment), Af-stimulated cells (Af), Af-stimulated cells administered scrambled siRNA (Af+Scram), or Af-stimulated cells administered PI3K-δ siRNA (Af+PI3K-δ). Bars indicate 20 µm. (E) Representative confocal images show the localisation of mitochondrial ROS (mtROS) in bronchoalveolar lavage (BAL) cells from saline-exposed mice administered drug vehicle (SAL+VEH), Af-exposed mice administered drug vehicle (Af+VEH), or Af-exposed mice administered NecroX-5 (Af+NX-5). Bars indicate 20 µm. (F) Cellular changes in BAL fluids of Af-exposed mice. Arrows and arrowheads indicate macrophages and eosinophils, respectively. Bars indicate 20 µm. (G) Representative immunoblots and densitometric analyses of IL-1β in lung tissues. Bars represent mean±SEM from seven mice per group. ^#P<0.05 versus SAL+VEH; *P < 0.05 versus Af+VEH. (H) Airway responsiveness assessed by invasive measurements from SAL+VEH, Af+VEH or Af+NX-5. Bars represent mean±SEM from six mice per group. (I) Cellular changes in bronchoalveolar lavage fluids. Bars represent mean±SEM from six mice per group. ^#P<0.05 versus SAL+VEH; *P < 0.05 versus Af+VEH. (J–L) Representative H&E stained sections of the lung from SAL+VEH (J), Af+VEH (K) and Af+NX-5 (L). Bars indicate 50 µm. (M) Representative confocal images show the localisation of mtROS in BAL cells from saline-exposed mice administered drug vehicle (SAL+VEH), Af-exposed mice administered drug vehicle (Af+VEH), Af-exposed mice administered 0.1 mg/kg IC87114 (Af+IC 0.1) or Af-exposed mice administered 1.0 mg/kg IC87114 (Af+IC 1.0). Bars indicate 20 µm. (N) Representative confocal images of tracheal EpCs show the localisation of mtROS in the control, Af, Af+VEH and Af+IC. Bars indicate 10 µm.