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Investigation of the molecular and cellular mechanisms of in vitro phage therapy in human cells and the innate immune response against Escherichia coli K1 and bacteriophage K1F
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Møller-Olsen, Christian (2021) Investigation of the molecular and cellular mechanisms of in vitro phage therapy in human cells and the innate immune response against Escherichia coli K1 and bacteriophage K1F. PhD thesis, University of Warwick.
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Official URL: http://webcat.warwick.ac.uk/record=b3736673~S15
Abstract
The last few decades have seen bacteria becoming increasingly resistant to antibiotic
treatment. This is considered a threat to human health and financial stability by
global non-governmental organisations (NGOs) and governments alike. As a result,
multiple technologies are being pursued to mitigate the effects of antimicrobial
resistance, one of which is phage therapy. Phage therapy is a re-emerging technology
that exploits the natural enemy of bacteria, the bacteriophage (phage), to treat
bacterial infection in humans. Although phages are used extensively in biotechnology
and food security industries, research is needed to advance the clinical potential of
the technology. This study uses clinically relevant tripartite in vitro model systems to
investigate the cellular and molecular processes underpinning phage therapy and the
interactions between phages and human cells.
It is demonstrated that phage K1F effectively can locate and infect intracellular E. coli
EV36 in both epithelial (T24) and endothelial (hCMEC) cell lines. Microscopy and flow
cytometry experiments further demonstrated that phage K1F is taken up by T24 cells
and hCMECs by the endocytic machinery and processed by non-selective and LC3-
assisted phagocytosis. In contrast, E. coli EV36 uptake activated galectin-8 and
ubiquitin-dependent autophagy. The inflammatory responses of hCMEC cultures
were analysed by RT qPCR. The analyses demonstrated that phage K1F does not elicit
inflammatory measures, while E. coli induced expression of cytokines TNFa, IL-6, IL-
8, IL-10, or IFNb. Phage intervention of bacteria-infected hCMEC cultures resulted in
reduced expression of TNFa and increased expression of IL-6 and IL-8. Temporal
measurements of the barrier integrity of hCMEC cultures, using xCELLigence,
demonstrated that phages K1F and T7 decrease the impendence of hCMEC cultures,
indicating an increase in barrier permeability. This function has the potential to
promote immune cell extravasation through capillaries to a site of infection.
Collectively, this study presents valuable experimental in vitro data to further the
advancement of phage therapy. The data suggests that while human cell lines
recognise phages through PRR activation, the response is non-inflammatory with
potentially additional clinical benefits.
Item Type: | Thesis (PhD) | ||||
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Subjects: | Q Science > QR Microbiology | ||||
Library of Congress Subject Headings (LCSH): | Bacteriophages, Bacteriophages -- Therapeutic use, Escherichia coli, Immune response -- Molecular aspects | ||||
Official Date: | May 2021 | ||||
Dates: |
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Institution: | University of Warwick | ||||
Theses Department: | School of Life Sciences | ||||
Thesis Type: | PhD | ||||
Publication Status: | Unpublished | ||||
Supervisor(s)/Advisor: | Sagona, Antonia ; Leppard, K. N. (Keith N.) | ||||
Extent: | xiv, 194 leaves : illustrations, charts | ||||
Language: | eng |
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