Chlamydia Muridarum Genital Infection Research Articles

Evaluation of a multisubunit recombinant polymorphic membrane protein and major outer membrane protein T cell vaccine against Chlamydia muridarum genital infection in three strains of mice

An efficacious vaccine is needed to control Chlamydia trachomatis infection. In the murine model of Chlamydia muridarum genital infection, multifunctional mucosal CD4 T cells are the foundation for protective immunity, with antibody playing a secondary role. We previously identified four Chlamydia outer membrane proteins (PmpE, PmpF, PmpG and PmpH) as CD4 T cell vaccine candidates using a dendritic cell-based immunoproteomic approach. We also demonstrated that these four polymorphic membrane proteins (Pmps) individually conferred protection as measured by accelerated clearance of Chlamydia infection in the C57BL/6 murine genital tract model. The major outer membrane protein, MOMP is also a well-studied protective vaccine antigen in this system. In the current study, we tested immunogenicity and protection of a multisubunit recombinant protein vaccine consisting of the four Pmps (PmpEFGH) with or without the major outer membrane protein (MOMP) formulated with a Th1 polarizing adjuvant in C57BL/6, Balb/c and C3H mice. We found that C57BL/6 mice vaccinated with PmpEFGH+MOMP elicited more robust cellular immune responses than mice immunized with individual protein antigens. Pmps elicited more variable cellular immune responses than MOMP among the three strains of mice. The combination vaccine accelerated clearance in the three strains of mice although at different rates. We conclude that the recombinant outer membrane protein combination constitutes a promising first generation Chlamydia vaccine construct that should provide broad immunogenicity in an outbred population.

Usefulness of 11C-Choline Positron Emission Tomography for Genital Chlamydial Infection Assessment in a Balb/c Murine Model

The aim of this study is to explore the feasibility of 11C-Choline PET in the assessment of the degree of inflammation in the Chlamydia muridarum genital infection model. Forty female Balb/c mice received 2.5 mg of medroxyprogesterone acetate i.m. 9 and 2 days prior to the infection: 21 mice were infected by C. muridarum into the vaginal vault, 12 mice were treated with inactivated chlamydiae, and 7 mice were SPG buffer-treated as negative controls. Three healthy control mice were not treated with progesterone. Mice in each category were randomly subdivided in two groups: (1) sacrificed at 5, 10, 15, and 20 days for histological analysis and (2) undergoing 11C-Choline PET at days 5, 10, and 20 post-infection (20 MBq of 11C-Choline, uptake time of 10 min, acquisition through a small-animal PET tomograph for 15 min). Infected animals showed a significantly higher standardized uptake value than both controls and animals inoculated with heat-inactivated chlamydiae in each PET scan (P<0.05). All organs of the infected animals had scores of inflammation ranging between 2 and 3 at day 5, decreasing to 1-2 at day 20. This preliminary result demonstrated that 11C-Choline PET can highlight a specific proliferation mechanism of inflammatory cells induced by C. muridarum, thanks to a very high sensitivity in detecting very small amounts of tracer in inflammatory cells.

Interruption of CXCL13-CXCR5 axis increases upper genital tract pathology and activation of NKT cells following chlamydial genital infection.

BackgroundRegulation of immune responses is critical for controlling inflammation and disruption of this process can lead to tissue damage. We reported that CXCL13 was induced in fallopian tube tissue following C. trachomatis infection. Here, we examined the influence of the CXCL13-CXCR5 axis in chlamydial genital infection.Methodology and Principal FindingsDisruption of the CXCL13-CXCR5 axis by injecting anti-CXCL13 Ab to BALB/c mice or using Cxcr5−/− mice increased chronic inflammation in the upper genital tract (UGT; uterine horns and oviducts) after Chlamydia muridarum genital infection (GT). Further studies in Cxcr5−/− mice showed an elevation in bacterial burden in the GT and increased numbers of neutrophils, activated DCs and activated NKT cells early after infection. After resolution, we noted increased fibrosis and the accumulation of a variety of T cells subsets (CD4-IFNγ, CD4-IL-17, CD4-IL-10 & CD8-TNFα) in the oviducts. NKT cell depletion in vitro reduced IL-17α and various cytokines and chemokines, suggesting that activated NKT cells modulate neutrophils and DCs through cytokine/chemokine secretion. Further, chlamydial glycolipids directly activated two distinct types of NKT cell hybridomas in a cell-free CD1d presentation assay and genital infection of Cd1d−/− mice showed reduced oviduct inflammation compared to WT mice. CXCR5 involvement in pathology was also noted using single-nucleotide polymorphism analysis in C. trachomatis infected women attending a sub-fertility clinic. Women who developed tubal pathology after a C. trachomatis infection had a decrease in the frequency of CXCR5 SNP +10950 T>C (rs3922).Conclusions/SignificanceThese experiments indicate that disruption of the CXCL13-CXCR5 axis permits increased activation of NKT cells by type I and type II glycolipids of Chlamydia muridarum and results in UGT pathology potentially through increased numbers of neutrophils and T cell subsets associated with UGT pathology. In addition, CXCR5 appears to contribute to inter-individual differences in human tubal pathology following C. trachomatis infection.

Significant Role of IL-1 Signaling, but Limited Role of Inflammasome Activation, in Oviduct Pathology during Chlamydia muridarum Genital Infection

IL-1β has been implicated in the development of oviduct pathology during Chlamydia muridarum genital infection in the mouse model. The goal of this study was to characterize the role of IL-1 signaling and the inflammasome-activation pathways during genital chlamydial infection. Compared with control mice, IL-1R-deficient mice displayed delayed clearance and increased chlamydial colonization. Consistent with the role for IL-1 signaling in infection clearance, mice deficient for the IL-1R antagonist cleared infection at a faster rate. Despite increased infection, IL-1R-deficient mice had significantly reduced oviduct pathology, which was associated with decreased numbers of neutrophils, but more macrophages, in the genital tract. IL-1β secretion is dependent on caspase-1 and apoptosis-associated speck-like protein containing caspase recruitment domain (ASC) inflammasome during in vitro infection of primed macrophages with C. muridarum. To investigate the role of inflammasome components during in vivo genital infection, mice lacking NLRP3, NLRC4, and ASC were tested and found to display no reduction in oviduct pathology compared with control mice. Mice deficient for ASC displayed a prolonged course of infection, which was associated with reduced T cell recruitment and proliferation. Further, ASC-deficient mice displayed normal levels of IL-1β in genital secretions. However, a significant decrease in caspase-1-dependent IL-18 was observed in both ASC- and NLRP3-deficient mice. These data demonstrate a major role for IL-1 signaling, but a limited role for the inflammasome pathway, in IL-1β secretion and development of oviduct pathology during genital chlamydial infection. The data also suggest an IL-1-independent role for ASC in adaptive immunity during genital chlamydial infection.

Interferon Regulatory Transcription Factor 3 Protects Mice from Uterine Horn Pathology during Chlamydia muridarum Genital Infection

Mice with the type I interferon (IFN) receptor gene knocked out (IFNAR KO mice) or deficient for alpha/beta IFN (IFN-α/β) signaling clear chlamydial infection earlier than control mice and develop less oviduct pathology. Initiation of host IFN-β transcription during an in vitro chlamydial infection requires interferon regulatory transcription factor 3 (IRF3). The goal of the present study was to characterize the influence of IRF3 on chlamydial genital infection and its relationship to IFN-β expression in the mouse model. IRF3 KO mice were able to resolve infection as well as control mice, overcoming increased chlamydial colonization and tissue burden early during infection. As previously observed for IFNAR KO mice, IRF3 KO mice generated a potent antigen-specific T cell response. However, in contrast to IFNAR KO mice, IRF3 KO mice exhibited unusually severe dilatation and pathology in the uterine horns but normal oviduct pathology after infection. Although IFN-β expression in vivo was dependent on the presence of IRF3 early in infection (before day 4), the IFN-independent function of IRF3 was likely driving this phenotype. Specifically, early during infection, the number of apoptotic cells and the number of inflammatory cells were significantly less in uterine horns from IRF3 KO mice than in those from control mice, despite an increased chlamydial burden. To delineate the effects of IFN-β versus IRF3, neutralizing IFN-β antibody was administered to wild-type (WT) mice during chlamydial infection. IFN-β depletion in WT mice mimicked that in IFNΑR KO mice but not that in IRF3 KO mice with respect to both chlamydial clearance and reduced oviduct pathology. These data suggest that IRF3 has a role in protection from uterine horn pathology that is independent of its function in IFN-β expression.

Significant role of IL-1 signaling, but limited role of inflammasome activation in oviduct pathology during Chlamydia muridarum genital infection (110.6)

Abstract IL-1β has been implicated in the development of oviduct pathology during Chlamydia muridarum genital infection in the mouse model. The goal of this study was to characterize the role of IL-1 signaling and the inflammasome activation pathways during C. muridarum genital infection. Compared to control mice, IL-1 receptor KO (knockout) mice display delayed clearance and increased infection burden, but very few incidences of oviduct hydrosalpinx. Consistent with this observation, IL-1R antagonist KO mice display an opposite phenotype of rapid infection clearance. Flow cytometry analysis at day 10 and day 20 post infection reveal significantly more macrophages in the oviducts of IL-1R KO mice in comparison to control mice. Despite increased infection and recruited macrophages, IL-1R-KO mice have less tissue fibrosis and dilation, suggesting an anti-inflammatory role for these cells in the absence of IL-1 signaling. To investigate the role of inflammasome components during in vivo genital infection, mice lacking NLRP3, NLRC4 and ASC were tested. Although IL-1β secretion is dependent on caspase-1 during in vitro infection of primed macrophages and requires ASC, it is independent of NLRP3 and NLRC4 activation. Surprisingly, during in vivo genital infection, mice lacking NLRP3, NLRC4, as well as ASC display no reduction in oviduct pathology. These data suggest a major role for IL-1 signaling but a limited role for inflammasome pathway in development of oviduct pathology.

Expression library immunization confers partial protection against Chlamydia muridarum genital infection

Protective sequences of Chlamydia muridarum were identified as potential vaccine candidates by screening a genomic DNA expression library and assessing the immune responses of mice immunized with individual library clones following vaginal challenge with live Chlamydia. Groups of female BALB/c mice were immunized intra-abdominally by gene gun delivery of DNA three times at three-weekly intervals with individual library clones expressing chlamydial protein fragments and humoral and cell-mediated immune responses were evaluated. Chlamydia-specific cytokines including tumour necrosis factor-α (TNF-α) interleukin-10 (IL-10), interleukin-4 (IL-4), interleukin-12 (IL-12) and interferon-γ (IFN-γ) were detected in mice immunized either with selected DNA clones in spleen cells (0.2–135.2 pg/mL) or lymph nodes (0.15–84.9 pg/mL). The most protective antigen identified was TC0512, a putative outer membrane protein (OMP). Immunization of mice with this clone elicited T-helper type-1 (Th-1) and T-helper type-2 (Th-2) cytokines as well as and IgG1 and IgG2a in sera of these animals. Ten days after the last immunization, animals were challenged intra-vaginally with 5 × 10 4 inclusion-forming units (IFUs) of C. muridarum. At 9 days following challenge TC0512 showed a 73% reduction in the number of recoverable Chlamydia compared with vector only immunized controls. Six additional clones were identified that also conferred varying degrees of protection against live chlamydial challenge. Significant protection against the initial stages of infection was shown by two DNA clones (encoding hypothetical proteins) and five clones showed enhanced clearance of chlamydial infection following DNA immunization and live chlamydial challenge. These results demonstrate that the C. muridarum genome can be screened for individual vaccine candidates by genetic immunization and that the screen produces novel and partially protective vaccine candidates.