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Isolation and In Vitro Culture of Murine and Human Alveolar Macrophages
Published on: April 20, 2018
Alveolar macrophages support interferon gamma-mediated viral clearance in RSV-infected neonatal mice
Katherine M Eichinger1, Loreto Egaña1, Jacob G Orend1
1Department of Pharmacy and Therapeutics, Center for Clinical Pharmaceutical Sciences, University of Pittsburgh School of Pharmacy, Pittsburgh, PA, USA.
Background:
Poor interferon gamma (IFNγ) production during respiratory syncytial virus (RSV) is associated with prolonged viral clearance and increased disease severity in neonatal mice and humans. We previously showed that intra-nasal delivery of IFNγ significantly enhances RSV clearance from neonatal lungs prior to observed T-lymphocyte recruitment or activation, suggesting an innate immune mechanism of viral clearance. We further showed that alveolar macrophages dominate the RSV-infected neonatal airways relative to adults, consistent with human neonatal autopsy data. Therefore, the goal of this work was to determine the role of neonatal alveolar macrophages in IFNγ-mediated RSV clearance.
Methods:
Clodronate liposomes, flow cytometry, viral plaque assays, and histology were used to examine the role of alveolar macrophages (AMs) and the effects of intra-nasal IFNγ in RSV infected neonatal Balb/c mice. The functional outcomes of AM depletion were determined quantitatively by viral titers using plaque assay. Illness was assessed by measuring reduced weight gain.
Results:
AM activation during RSV infection was age-dependent and correlated tightly with IFNγ exposure. Higher doses of IFNγ more efficiently stimulated AM activation and expedited RSV clearance without significantly affecting weight gain. The presence of AMs were independently associated with improved RSV clearance, whereas AM depletion but not IFNγ exposure, significantly impaired weight gain in RSV-infected neonates.
Conclusion:
We show here for the first time, that IFNγ is critical for neonatal RSV clearance and that it depends, in part, on alveolar macrophages (AMs) for efficient viral clearing effects. Early reductions in viral burden are likely to have profound short- and long-term immune effects in the vulnerable post-natally developing lung environment. Studies are ongoing to elucidate the pathologic effects associated with early versus delayed RSV clearance in developing neonatal airways.
Insights
Interferon gamma (IFNγ) is crucial for clearing respiratory syncytial virus (RSV) in neonates, working partly through alveolar macrophages (AMs). This finding is key for understanding neonatal lung immunity and RSV infection.
Area of Science:
- Immunology
- Neonatal Respiratory Infections
- Innate Immunity
Background:
- Poor interferon gamma (IFNγ) production in respiratory syncytial virus (RSV) infection correlates with severe disease and delayed viral clearance in neonates.
- Previous studies indicated intra-nasal IFNγ enhances RSV clearance in neonatal lungs via innate immune mechanisms, preceding adaptive immune responses.
- Alveolar macrophages (AMs) are the predominant immune cells in neonatal airways during RSV infection, as supported by human autopsy data.
Purpose of the Study:
- To investigate the specific role of neonatal alveolar macrophages (AMs) in IFNγ-mediated clearance of respiratory syncytial virus (RSV).
- To determine how IFNγ influences AM function and RSV clearance in the context of neonatal infection.
Main Methods:
- Utilized clodronate liposomes for AM depletion, flow cytometry, viral plaque assays for viral quantification, and histology in RSV-infected neonatal Balb/c mice.
- Assessed functional outcomes of AM depletion by measuring viral titers and monitored illness severity through weight gain.
Main Results:
- Alveolar macrophage (AM) activation during RSV infection was age-dependent and directly correlated with IFNγ exposure levels.
- Higher doses of IFNγ enhanced AM activation and accelerated RSV clearance, with no significant impact on weight gain.
- AMs were independently associated with improved RSV clearance; however, AM depletion, but not IFNγ administration, significantly impaired weight gain in neonates.
Conclusions:
- Interferon gamma (IFNγ) is critical for neonatal respiratory syncytial virus (RSV) clearance, functioning partly through alveolar macrophages (AMs).
- Early reduction of viral load in neonates is likely to have significant short- and long-term impacts on the developing lung immune environment.
- Further research is underway to understand the pathological effects of early versus delayed RSV clearance in neonatal airways.
