Abstract
Background Cigarette smokers are at increased risk of acquiring influenza, developing severe disease and requiring hospitalisation/intensive care unit admission following infection. However, immune mechanisms underlying this predisposition are incompletely understood, and therapeutic strategies for influenza are limited.
Methods We used a mouse model of concurrent cigarette smoke exposure and H1N1 influenza infection, colony-stimulating factor (CSF)3 supplementation/receptor (CSF3R) blockade and single-cell RNA sequencing (scRNAseq) to investigate this relationship.
Results Cigarette smoke exposure exacerbated features of viral pneumonia such as oedema, hypoxaemia and pulmonary neutrophilia. Smoke-exposed infected mice demonstrated an increase in viral (v)RNA, but not replication-competent viral particles, relative to infection-only controls. Interstitial rather than airspace neutrophilia positively predicted morbidity in smoke-exposed infected mice. Screening of pulmonary cytokines using a novel dysregulation score identified an exacerbated expression of CSF3 and interleukin-6 in the context of smoke exposure and influenza. Recombinant (r)CSF3 supplementation during influenza aggravated morbidity, hypothermia and oedema, while anti-CSF3R treatment of smoke-exposed infected mice improved alveolar–capillary barrier function. scRNAseq delineated a shift in the distribution of Csf3+ cells towards neutrophils in the context of cigarette smoke and influenza. However, although smoke-exposed lungs were enriched for infected, highly activated neutrophils, gene signatures of these cells largely reflected an exacerbated form of typical influenza with select unique regulatory features.
Conclusion This work provides novel insight into the mechanisms by which cigarette smoke exacerbates influenza infection, unveiling potential therapeutic targets (e.g. excess vRNA accumulation, oedematous CSF3R signalling) for use in this context, and potential limitations for clinical rCSF3 therapy during viral infectious disease.
Abstract
Cigarette smoke exacerbates PFU-independent viral RNA accumulation, morbidity-associated interstitial neutrophilia, Csf3-expressing neutrophil recruitment and CSF3-mediated alveolar–capillary barrier dysregulation during influenza infection in mice https://bit.ly/3HVOuiw
Footnotes
Published in volume 60, issue 2 of the European Respiratory Journal on 25 August 2022; republished 8 September 2022 to correct typographical errors in figures 6 and 7.
Single-cell RNA sequencing data have been uploaded to the Gene Expression Omnibus (GEO) database (GSE186976).
Author contributions: G. Vanderstocken, J.J.C. McGrath and M.R. Stämpfli conceived the study and designed the experiments. J.J.C. McGrath, G. Vanderstocken, S.P. Cass, S. Afkhami, M.F. Fantauzzi, D. Thayaparan, P. Wang, A. Beaulieu, R. Jiménez-Saiz, P. Shen, M. Morissette, S.D. Revill, A. Tabuchi and D. Zabini conducted experiments. A. Dvorkin-Gheva conducted bioinformatic analysis of scRNAseq data, while J.J.C. McGrath compiled scRNAseq data. A. Reihani and S.D. Revill performed HALO analysis. W.L. Lee, C.D. Richards, M.S. Miller, K. Ask, W.M. Kuebler and J.A. Simpson provided technical insights for experimental design. J.J.C. McGrath and M.R. Stämpfli wrote the manuscript with feedback from all authors. M.R. Stämpfli secured funding for the study.
Conflict of interest: M.S. Miller reports consulting fees for scientific advisory board participation from Seqirus and Medicago (influenza vaccines); lecture honoraria for lunch-and-learn on COVID-19 vaccines from Boehringer-Ingelheim; outside the submitted work. K. Ask reports grants from Alkermes, Prometic, GSK, Canadian Institute for Health Research, Pharmaxis, Indalo, Unity Biotechnology, Canadian Pulmonary Fibrosis Association, Collaborative Health Research Projects, Pieris Pharmaceuticals, Bold Therapeutics, Pliant, CSL Behring and Boehringer Ingelheim; lecture honoraria from Boehringer Ingelheim; travel support from Boehringer Ingelheim; outside the submitted work. W.M. Kuebler reports grant-in-aid from Canadian Institutes of Health Research (CIHR), during the course of the present study; grants-in-aid from German Research Foundation (DFG), German Ministry for Research and Education (BMBF), Berlin Institute of Health (BIH), German Centre for Cardiovascular Research (DZHK); outside the submitted work. M.R. Stämpfli reports support for the present manuscript from Canadian Institutes of Health Research; grants from RespiVert Pharmaceuticals, Canadian Heart and Stroke Foundation, and Ontario Thoracic Society; consulting fees from Boehringer Ingelheim; lecture honoraria from AstraZeneca; MTA for monoclonal antibody to block G-CSF from CSL Behring and is employed by CSL Behring since January 2020; outside the submitted work. All other authors have nothing to disclose.
Support statement: Funding for this study was provided by the Canadian Institutes for Health Research (CIHR; MOP-64390, MOP-142353, PJT-159792; M.R. Stämpfli) and the Deutsche Forschungsgemeinschaft (DFG; SFB TR84, SFB 1449; W.M. Kuebler). G. Vanderstocken was supported by the Canadian Respiratory Research Network (CRRN). M.S. Miller was supported, in part, by a CIHR New Investigator Award and an Ontario Early Research Award. Funding information for this article has been deposited with the Crossref Funder Registry.
- Received July 22, 2021.
- Accepted December 29, 2021.
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