Mechanism of immune modulation in macrophages by Enterococcus faecalis
Enterococcus faecalis is a member of the human gastrointestinal microbiota and is a gram-positive, opportunistic pathogen associated with hospital acquired wound, bloodstream, and urinary tract infections. E. faecalis can suppress or evade immune-mediated clearance by macrophages which promotes pers...
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| Format: | Thesis-Doctor of Philosophy |
| Language: | English |
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Nanyang Technological University
2020
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| Online Access: | https://hdl.handle.net/10356/138251 |
| _version_ | 1811693722177372160 |
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| author | Tien, Brenda Yin Qi |
| author2 | Kimberly Kline |
| author_facet | Kimberly Kline Tien, Brenda Yin Qi |
| author_sort | Tien, Brenda Yin Qi |
| collection | NTU |
| description | Enterococcus faecalis is a member of the human gastrointestinal microbiota and is a gram-positive, opportunistic pathogen associated with hospital acquired wound, bloodstream, and urinary tract infections. E. faecalis can suppress or evade immune-mediated clearance by macrophages which promotes persistent infection. However, the mechanism by which E. faecalis modulates host immune function is not well understood. The overarching aim of this thesis is to fully characterize the interaction between E. faecalis and macrophages. I hypothesised that E. faecalis can modulate macrophage activation during infection thus affecting the host innate immune response in vivo. I established an in vitro model to study the immune-modulatory interaction of E. faecalis and macrophages. I observed that E. faecalis actively prevents NF-κB signalling in mouse RAW-Blue macrophages in the presence of Toll-like receptor agonists and during polymicrobial infection with Escherichia coli in a dose-dependent manner, resulting in global downregulation of cytokines and chemokines during enterococcal infection. To elucidate the bacterial factor(s) that are involved in this macrophage suppression, various E. faecalis virulence mutants were tested. I showed that lactic acid, secreted by E. faecalis, is essential for macrophage suppression. Moreover, I showed that lactate dehydrogenase-mediated immune suppression promotes E. coli survival during polymicrobial wound infection. Finally, to identify the pathways which E. faecalis modulates to suppress macrophage activation, I performed western blot analysis and RNA analysis. I observed that E. faecalis subverts macrophage activation by suppressing IκBα and ERK level, preventing p65 phosphorylation and potentially activating Hif-1α. In addition, E. faecalis infection at high doses also prevents polarisation of macrophages towards a M1 and M2 phenotype. Taken together, these results suggest that E. faecalis requires a high bacterial density to subvert the innate immune response, that E. faecalis-derived lactic acid is involved in this subversion, resulting in altered pathogenic outcomes. Understanding the interaction between E. faecalis and the host immune response may be helpful in developing alternative approaches or improved treatment strategies to prevent recalcitrant E. faecalis infection. |
| first_indexed | 2024-10-01T06:56:12Z |
| format | Thesis-Doctor of Philosophy |
| id | ntu-10356/138251 |
| institution | Nanyang Technological University |
| language | English |
| last_indexed | 2024-10-01T06:56:12Z |
| publishDate | 2020 |
| publisher | Nanyang Technological University |
| record_format | dspace |
| spelling | ntu-10356/1382512020-11-01T04:58:17Z Mechanism of immune modulation in macrophages by Enterococcus faecalis Tien, Brenda Yin Qi Kimberly Kline Interdisciplinary Graduate School (IGS) Singapore Centre for Environmental Life Sciences and Engineering kkline@ntu.edu.sg Science::Biological sciences Enterococcus faecalis is a member of the human gastrointestinal microbiota and is a gram-positive, opportunistic pathogen associated with hospital acquired wound, bloodstream, and urinary tract infections. E. faecalis can suppress or evade immune-mediated clearance by macrophages which promotes persistent infection. However, the mechanism by which E. faecalis modulates host immune function is not well understood. The overarching aim of this thesis is to fully characterize the interaction between E. faecalis and macrophages. I hypothesised that E. faecalis can modulate macrophage activation during infection thus affecting the host innate immune response in vivo. I established an in vitro model to study the immune-modulatory interaction of E. faecalis and macrophages. I observed that E. faecalis actively prevents NF-κB signalling in mouse RAW-Blue macrophages in the presence of Toll-like receptor agonists and during polymicrobial infection with Escherichia coli in a dose-dependent manner, resulting in global downregulation of cytokines and chemokines during enterococcal infection. To elucidate the bacterial factor(s) that are involved in this macrophage suppression, various E. faecalis virulence mutants were tested. I showed that lactic acid, secreted by E. faecalis, is essential for macrophage suppression. Moreover, I showed that lactate dehydrogenase-mediated immune suppression promotes E. coli survival during polymicrobial wound infection. Finally, to identify the pathways which E. faecalis modulates to suppress macrophage activation, I performed western blot analysis and RNA analysis. I observed that E. faecalis subverts macrophage activation by suppressing IκBα and ERK level, preventing p65 phosphorylation and potentially activating Hif-1α. In addition, E. faecalis infection at high doses also prevents polarisation of macrophages towards a M1 and M2 phenotype. Taken together, these results suggest that E. faecalis requires a high bacterial density to subvert the innate immune response, that E. faecalis-derived lactic acid is involved in this subversion, resulting in altered pathogenic outcomes. Understanding the interaction between E. faecalis and the host immune response may be helpful in developing alternative approaches or improved treatment strategies to prevent recalcitrant E. faecalis infection. Doctor of Philosophy 2020-04-29T12:41:23Z 2020-04-29T12:41:23Z 2020 Thesis-Doctor of Philosophy Tien, B. Y. Q. (2020). Mechanism of immune modulation in macrophages by Enterococcus faecalis. Doctoral thesis, Nanyang Technological University, Singapore. https://hdl.handle.net/10356/138251 10.32657/10356/138251 en This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License (CC BY-NC 4.0). application/pdf Nanyang Technological University |
| spellingShingle | Science::Biological sciences Tien, Brenda Yin Qi Mechanism of immune modulation in macrophages by Enterococcus faecalis |
| title | Mechanism of immune modulation in macrophages by Enterococcus faecalis |
| title_full | Mechanism of immune modulation in macrophages by Enterococcus faecalis |
| title_fullStr | Mechanism of immune modulation in macrophages by Enterococcus faecalis |
| title_full_unstemmed | Mechanism of immune modulation in macrophages by Enterococcus faecalis |
| title_short | Mechanism of immune modulation in macrophages by Enterococcus faecalis |
| title_sort | mechanism of immune modulation in macrophages by enterococcus faecalis |
| topic | Science::Biological sciences |
| url | https://hdl.handle.net/10356/138251 |
| work_keys_str_mv | AT tienbrendayinqi mechanismofimmunemodulationinmacrophagesbyenterococcusfaecalis |