Human infections with highly pathogenic avian influenza A (H5N1) virus are frequently fatal but the mechanisms of disease remain ill-defined. H5N1 infection is associated with intense production of proinflammatory cytokines, but whether this cytokine storm is the main cause of fatality or is a consequence of extensive virus replication that itself drives disease remains controversial. Conventional intratracheal inoculation of a liquid suspension of H5N1 influenza virus in nonhuman primates likely results in efficient clearance of virus within the upper respiratory tract and rarely produces severe disease. We reasoned that small particle aerosols of virus would penetrate the lower respiratory tract and blanket alveoli where target cells reside. We show that inhalation of aerosolized H5N1 influenza virus in cynomolgus macaques results in fulminant pneumonia that rapidly progresses to acute respiratory distress syndrome with a fatal outcome reminiscent of human disease. Molecular imaging revealed intense lung inflammation coincident with massive increases in proinflammatory proteins and IFN-α in distal airways. Aerosolized H5N1 exposure decimated alveolar macrophages, which were widely infected and caused marked influx of interstitial macrophages and neutrophils. Extensive infection of alveolar epithelial cells caused apoptosis and leakage of albumin into airways, reflecting loss of epithelial barrier function. These data establish inhalation of aerosolized virus as a critical source of exposure for fatal human infection and reveal that direct viral effects in alveoli mediate H5N1 disease. This new nonhuman primate model will advance vaccine and therapeutic approaches to prevent and treat human disease caused by highly pathogenic avian influenza viruses.
This work was supported by funds from the University of Pittsburgh (to S.M.B.-B.) and the University of Georgia (to D.R.P.), as well as by National Institutes of Health Grant AI111598 (to S.J.B. and C.A.W.) and National Institutes of Health Contract HHSN272201400008C (to D.R.P.).
The online version of this article contains supplemental material.
Abbreviations used in this article:
- alveolar epithelial cell
- bronchoalveolar lavage
- computed tomography
- dendritic cell
- Madin–Darby canine kidney
- positron emission tomography
- Received October 14, 2016.
- Accepted December 9, 2016.
- Copyright © 2017 by The American Association of Immunologists, Inc.