Epithelial uptake ofAspergillus fumigatusdrives efficient fungal clearancein vivoand is aberrant in Chronic Obstructive Pulmonary Disease (COPD)

Abstract

AbstractHundreds of spores of the common mouldAspergillusfumigatus (Af)are inhaled daily by human beings, representing a constant, often fatal, threat to our respiratory health. The small size ofAfspores suggest that interactions withAirwayEpithelialCells (AECs) are frequent and we and others have previously demonstrated that AECs are able to internaliseAfspores. We thus hypothesised thatAfspore uptake and killing by AECs is important for driving efficient fungal clearancein vivoand that defective spore uptake and killing would represent major risk factors forAspergillus-related diseases. In order to test this, we utilised single-cell approaches based onImagingFlowCytometry (IFC) and live-cell microfluidic imaging to measure spore uptake and outcomesin vitro,in vivoand using primary human AECs.In vitro, viability of immortalised AECs was largely unaffected byAfuptake and AECs were able to significantly curtail the growth of internalised spores. Applying our approach directly to infected mouse lungs we demonstrated, for the first time, thatAfspores are internalised and killed by AECs during whole animal infection, whereby only ~3% of internalised spores remained viable after 8 hours of co-incubation with murine AECs. Finally,in vitroanalysis of primary human AECs from healthy and at-risk donors revealed significant alterations in the uptake and consequent outcomes in Chronic Obstructive Pulmonary Disease (COPD), whereby gorging COPD-derived AECs were unable to quell intracellularAfas efficiently as healthy primary AECs. We have thus demonstrated that AECs efficiently killAfspores upon uptakein vivoand that this process is altered in COPD, a well-known risk factor for debilitating fungal lung disease, thereby suggesting that AECs critically contribute to the efficient clearance of inhaledAfspores and that dysregulation of curative AEC responses represents a potent driver ofAspergillus-related diseases.