Characterisation of Aspergillus fumigatus Endocytic Trafficking within Airway Epithelial Cells Using High-Resolution Automated Quantitative Confocal Microscopy
Abstract
:1. Introduction
2. Materials and Methods
2.1. Fungal Strains
2.2. Cell Lines
2.3. Quantification of the Number of Lysosomes during A. fumigatus Infection of Lung Epithelial Cells
2.4. Quantification of the Rate of Lysosome—Phagosome Fusion during A. fumigatus Infection of Epithelial Cells
2.5. Determining A. fumigatus Viability within Phagolysosomes by Confocal Microscopy
2.6. Assessing the Impact of A. fumigatus Cell Wall Components on Phagosome-Lysosome Fusion Rates in the Lung Epithelium
2.7. Determining Synchronization of Phagosome Acidification and Phagosome Maturation upon A. fumigatus Infection of the Lung Epithelium
2.8. Fungal Killing Assays
2.9. siRNA Treatment
2.10. Image Acquisition and Data Analyses
2.11. Statistical Analyses
3. Results
3.1. Rate of Lysosome Recruitment to the Phagosome Determines A. fumigatus Fate within the Lung Alveolar Epithelial Cells
3.2. Bronchial Epithelial Cell Phagolysosomes Demonstrate Increased Fungal-Killing, Compared to Alveolar Cells
3.3. Phagolysosome Maturation Relies on A. fumigatus Cell Wall Integrity
3.4. Validation of High Content Live-Cell Imaging on vATPase Silenced Lung Epithelial Cells
4. Discussion
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Ben-Ghazzi, N.; Moreno-Velásquez, S.; Seidel, C.; Thomson, D.; Denning, D.W.; Read, N.D.; Bowyer, P.; Gago, S. Characterisation of Aspergillus fumigatus Endocytic Trafficking within Airway Epithelial Cells Using High-Resolution Automated Quantitative Confocal Microscopy. J. Fungi 2021, 7, 454. https://doi.org/10.3390/jof7060454
Ben-Ghazzi N, Moreno-Velásquez S, Seidel C, Thomson D, Denning DW, Read ND, Bowyer P, Gago S. Characterisation of Aspergillus fumigatus Endocytic Trafficking within Airway Epithelial Cells Using High-Resolution Automated Quantitative Confocal Microscopy. Journal of Fungi. 2021; 7(6):454. https://doi.org/10.3390/jof7060454
Chicago/Turabian StyleBen-Ghazzi, Nagwa, Sergio Moreno-Velásquez, Constanze Seidel, Darren Thomson, David W. Denning, Nick D. Read, Paul Bowyer, and Sara Gago. 2021. "Characterisation of Aspergillus fumigatus Endocytic Trafficking within Airway Epithelial Cells Using High-Resolution Automated Quantitative Confocal Microscopy" Journal of Fungi 7, no. 6: 454. https://doi.org/10.3390/jof7060454
APA StyleBen-Ghazzi, N., Moreno-Velásquez, S., Seidel, C., Thomson, D., Denning, D. W., Read, N. D., Bowyer, P., & Gago, S. (2021). Characterisation of Aspergillus fumigatus Endocytic Trafficking within Airway Epithelial Cells Using High-Resolution Automated Quantitative Confocal Microscopy. Journal of Fungi, 7(6), 454. https://doi.org/10.3390/jof7060454