- Title
- The role of interferon epsilon in protection against chlamydial reproductive tract infections
- Creator
- Mayall, Jemma Rose
- Relation
- University of Newcastle Research Higher Degree Thesis
- Resource Type
- thesis
- Date
- 2017
- Description
- Research Doctorate - Doctor of Philosophy (PhD)
- Description
- Chlamydia trachomatis is the most common cause of sexually transmitted bacterial infections, affecting over 131 million individuals annually worldwide. In women, infection commonly ascends from the vagina into the upper reproductive tract (RT) and leads to a range of RT diseases such as infertility, pelvic inflammatory disease (PID), and ectopic pregnancy. While antibiotics are a safe and effective method for treating infection, 50-90% of Chlamydia RT infections (RTIs) are asymptomatic. This means that infections often progress for some time before treatment is initiated, leading to permanent immunopathology in the upper RT tissues, which causes disease. Significantly, there are no vaccines or immunotherapies currently available for the prevention of infection or the development of severe RT sequelae. This is largely due to the relatively limited understanding of the immunobiology of Chlamydia infections in the female RT. An improved understanding of the complex immune processes that are involved in both the clearance and immunopathology of Chlamydia infections is required in order to identify novel targets for the development of better therapeutic strategies for the prevention and treatment of Chlamydia infections and associated disease. Previous studies indicate that levels of the female sex hormones, progesterone and oestradiol, affect the susceptibility of an individual to Chlamydia RTIs. The pioneering studies and preliminary data collected by our research group and collaborators demonstrate that interferon (IFN)ε, a novel type I IFN that is exclusively and constitutively expressed in the female RT (predominantly uterine tissue), fluctuates throughout the oestrous cycle and, therefore, may be linked to the hormone-dependent susceptibility of the female RT to infection. In this thesis, I demonstrate the role of IFNε in protecting against Chlamydia RTIs, identify the key immune processes that underpin IFNε-mediated protection, and determine the potential of recombinant (r)IFNε as a therapeutic agent to prevent infection. Using wild-type (WT) and IFNε knockout (IFNε-/-), C57BL/6 mice, I investigated the effects of IFNε deficiency and exogenous IFNε (via administration of rIFNε) on Chlamydia infection and the effects of female sex hormones on these responses (Chapter 2). I demonstrate that IFNε protects against Chlamydia infections from the earliest stages of infection (3 days post infection [dpi]) in progesterone-pre-treated mice, but that, despite hormone pre-treatment having substantial effects on the expression IFNε in the female RT, hormone-mediated effects on IFNε expression may not fully account for the effects of female sex hormones on susceptibility to Chlamydia infections in the female RT. I also show that intravaginal (IVAG) rIFNε administration may be an effective preventative therapy for the treatment of Chlamydia RTIs. In order to determine the mechanisms that underpin IFNε-mediated protective responses during these early stages of infection, I then performed flow cytometry, and microarray and subsequent bioinformatical analyses, on uterine tissues from Chlamydia- and sham-infected, WT and IFNε-/- mice at 3dpi. The findings from these analyses were used to identify the key immune cells and molecular pathways and networks affected by IFNε and informed the more focussed studies that aimed to clarify potential roles of natural killer (NK) cell (Chapter 3) and inflammasome (Chapter 4) responses in IFNε-mediated protection against Chlamydia infection. I show, for the first time, that IFNε primes for and induces both local and systemic NK cell responses that protect against Chlamydia RTIs (Chapter 3). Specifically, I showed that IFNε-/- mice have reduced numbers of conventional (c)NK (CD45⁺ CD3⁻ NK1.1⁺) and uterine (u)NK (CD45⁺ CD3⁻ NK1.1⁻ CD49b⁻ CD122⁺) cells in their upper RTs during the early stages (3dpi) of Chlamydia infection. In particular, I show that the numbers of activated (CD69⁺) and IFNγ-producing NK cells are reduced in the RTs of IFNε-/- mice. I also show that NK cells are the predominant cellular source of IFNγ in the uterus at 3dpi. Interestingly, I show that the number of systemic (splenic) NK cells, as well as precursor and mature, but not immature, NK cells in the bone marrow, are reduced in both Chlamydia- and sham-infected IFNε-/- mice, compared to WT controls. These responses correspond with a reduction in interleukin (IL)-15, a cytokine important for the development, activation, and homing of NK cells, and C-X-C motif chemokine ligand (CXCL)10, a potent NK cell chemoattractant, in the uterine horns of IFNε-/- mice. This shows that IFNε, produced in the female RT, primes for both local and systemic NK cell responses, potentially by increasing IL-15 and CXCL10. Finally, by depleting NK cells in WT animals using anti-asialo ganglio-N-tetrasylceramide (ASGM1), I not only show for the first time that NK cells in the uterus are vital for restricting uterine Chlamydia growth, but that the depletion of systemic/infiltrating NK cells has little effect on reducing the number of activated NK cells in uterine tissue. These findings not only highlight the importance of IFNε in systemic NK cell mobilisation but, more importantly, also demonstrate the potent effects of IFNε in maintaining NK cell numbers and activity in the uterus in the absence of infiltrating NK cells and the importance of these responses in mediating protection against ascending Chlamydia RTI. I also show that IFNε regulates inflammasome signalling pathways during Chlamydia RTI and that IFNε-induced up-regulation of the nucleotide-binding oligomerisation domain (NOD)- and leucine-rich repeat (LRR)-containing receptor (NLR) family pyrin domain-containing 3 (NLRP3) inflammasome axis plays an important role in mediating protection against infection. Specifically, I show that IFNε-/- mice have reduced expression of IL-1β prior to and caspase-1 prior to and during infection, and reduced NLRP3-positive uterine epithelial cells during infection, compared to WT controls. These findings correspond with a reduction in the number of active caspase-1⁺ leukocytes present in their upper RTs early during Chlamydia infection (3dpi). Interestingly, a significant number of the active caspase-1⁺ leukocytes observed in the uterus during Chlamydia infection were NK cells. By inhibiting NLRP3 in WT animals using IVAG administration of MCC950, I show that NLRP3 inflammasome responses in the female RT play an important role in restricting uterine Chlamydia growth during the early stages of Chlamydia RTI but may not solely be responsible for IFNε-mediated caspase-1 activation. I also show that IFNε-/- mice have reduced expression of absent in melanoma 2 (AIM2) and caspase-4 in their upper RTs, compared to WT controls. These findings highlight the potential importance of IFNε-mediated NLRP3 and other inflammasome responses in protecting against ascending Chlamydia RTI. The investigations conducted during my PhD studies have identified several novel mechanisms by which IFNε may protect against Chlamydia RTIs from the earliest stages of infection. I have shown that IFNε plays an important role in the induction of NK cells responses, not only in the upper female RT but also systemically, during the early stages of Chlamydia infection and that these responses are important in protecting against infection. I also show that IFNε plays an important role in priming inflammasome-mediated responses in the upper female RT during infection and that these responses play an important role in protecting against Chlamydia infection. The findings from my thesis provide new insights into the effects of IFNε on mucosal immunology in the female RT and extend upon the current body of literature describing the immune processes that are important in mediating protection against Chlamydia infection. Significantly, these findings will now be used to guide future investigations that will aim to determine how the responses I have identified affect long-term sequelae and, therefore, may help inform new therapeutic targets for the development of improved treatment and prevention strategies for Chlamydia RTIs and associated disease.
- Subject
- interferon epsilon; chlamydial reproductive tract infections; <i>Chlamydia trachomatis</i>; interferon (IFN)ε
- Identifier
- http://hdl.handle.net/1959.13/1333764
- Identifier
- uon:27150
- Rights
- Copyright 2017 Jemma Rose Mayall
- Language
- eng
- Full Text
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