Myxobacteria-Derived Outer Membrane Vesicles: Potential Applicability Against Intracellular Infections
Abstract
:1. Introduction
2. Materials and Methods
2.1. Myxobacterial Culture
2.2. Isolation and Purification of Outer Membrane Vesicles
2.3. Liquid-Chromatography Coupled Mass Spectrometry
2.3.1. OMV Preparation
2.3.2. UHPLC MS Conditions
2.3.3. LC-MS Data Bucketing and Annotation Using Metaboscape Software
2.3.4. Conditions for MS² Analysis
2.3.5. GNPS Clustering Parameters
2.4. Cryogenic Electron Microscopy
2.5. Cell Culture
2.6. Cell Viability and Cytotoxicity
2.7. Cytokine Detection of OMV-Treated PBMCs
2.8. Antimicrobial Effect upon Storage
2.9. Bacteriomimetic Liposome Preparation
2.10. Flow Cytometry and Confocal Laser Scanning Microscopy to Assess OMV Uptake into Mammalian Cells
2.11. Bacterial Culture
2.12. Intracellular Infection
2.13. Statistical Analysis
3. Results
3.1. OMVs are Successfully Isolated by Ultracentrifugation and SEC
3.2. Myxobacterial OMVs Neither Affect Viability of Mammalian Cells nor Induce Cytotoxicity
3.3. Proinflammatory Cytokine Detection by Flow Cytometry
3.4. Myxobacterial OMVs Are Able to Kill Planktonic S. aureus
3.5. OMVs Are Taken Up by Mammalian Cells
3.6. OMVs Are Able to Kill Intracellular Pathogens
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Goes, A.; Lapuhs, P.; Kuhn, T.; Schulz, E.; Richter, R.; Panter, F.; Dahlem, C.; Koch, M.; Garcia, R.; Kiemer, A.K.; et al. Myxobacteria-Derived Outer Membrane Vesicles: Potential Applicability Against Intracellular Infections. Cells 2020, 9, 194. https://doi.org/10.3390/cells9010194
Goes A, Lapuhs P, Kuhn T, Schulz E, Richter R, Panter F, Dahlem C, Koch M, Garcia R, Kiemer AK, et al. Myxobacteria-Derived Outer Membrane Vesicles: Potential Applicability Against Intracellular Infections. Cells. 2020; 9(1):194. https://doi.org/10.3390/cells9010194
Chicago/Turabian StyleGoes, Adriely, Philipp Lapuhs, Thomas Kuhn, Eilien Schulz, Robert Richter, Fabian Panter, Charlotte Dahlem, Marcus Koch, Ronald Garcia, Alexandra K. Kiemer, and et al. 2020. "Myxobacteria-Derived Outer Membrane Vesicles: Potential Applicability Against Intracellular Infections" Cells 9, no. 1: 194. https://doi.org/10.3390/cells9010194