Host immune responses to chlamydial inclusion membrane proteins B and C in Chlamydia trachomatis infected women with or without fertility disorders

Rishein Gupta,Pragya Srivastava,Harsh Vardhan,Sudha Salhan,Aruna Mittal

doi:10.1186/1477-7827-7-38

Rishein Gupta, Pragya Srivastava + Show 3 more

Open Access

https://doi.org/10.1186/1477-7827-7-38

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Abstract

BackgroundWith an increase in the number of putative inclusion membrane proteins (incs) in chlamydial genomes, there is a need for understanding their contribution in host-pathogen interactions. Thus in this study we determined the host mucosal and peripheral immune responses to incs (IncB and IncC) of Chlamydia trachomatis (CT).MethodsFemale patients (n = 296) attending the gynaecology out patient department of Safdarjung hospital, New Delhi were enrolled for the study and were clinically characterized into two groups; CT-positive fertile women (n = 38) and CT-positive women with fertility disorders (n = 29). Uninfected healthy fertile women were enrolled as controls (n = 31). Gene specific PCRs were used for detection of incB and incC genes in endocervical samples of CT-positive patients. ELISA and Western blot assay were used for detection of IgA and IgG antibodies to IncB and IncC in cervical washes and sera. Effect of IncB and IncC stimulation of cervical cells and PBMCs on cellular proliferation and cytotoxity was determined using MTT assay and Lactate dehydrogenase (LDH)-cytotoxicity assay respectively. Modulation of cytokines (Interleukin (IL)-1 Beta, IL-4, IL-5, IL-6, IL-10, Interferon-gamma, IL-12, Tumor Necrosis Factor-alpha and Granulocyte macrophage colony-stimulating factor (GM-CSF)) in cervical cells and PBMCs upon stimulation with IncB and IncC was determined by real-time reverse-transcriptase (RT)-PCR and ELISA. Further, CD4 positive T cells were purified from cervical cells and peripheral blood mononuclear cells (PBMCs) and secreted cytokines (Interferon-gamma and IL-4) were evaluated by ELISPOT and real-time RT-PCR.ResultsUsing MTT assay, significantly high proliferative responses (P < 0.05) were observed in inc-stimulated cervical cells and PBMCs from CT-positive fertile women compared to CT-positive women with fertility disorders and controls. Interferon-gamma, IL-12 and GM-CSF were found to be elevated in inc-stimulated cervical cells and PBMCs of CT-positive fertile women compared to CT-positive women with fertility disorders and controls (P < 0.05). In contrast, IL-1 Beta, IL-4, IL-5, IL-6 and IL-10 levels were found to be higher in CT-positive women with fertility disorders compared to CT-positive fertile women and controls (P < 0.05). Interferon-gamma secreting cells and mRNA expression in inc-stimulated cervical and peripheral CD4 positive T cells were significantly higher (P < 0.05) in CT positive fertile women compared to CT-positive women with fertility disorders.ConclusionOur data overall suggests that CT incs, IncB and IncC modulate host immune responses and may have a role in protection/pathogenesis of genital chlamydial infection in women.

Highlights

With an increase in the number of putative inclusion membrane proteins in chlamydial genomes, there is a need for understanding their contribution in host-pathogen interactions
Seven CTpositive patients and two controls were excluded as the count of cervical cells was less than 2 million cells
All healthy controls tested negative for a current Chlamydia trachomatis (CT) infection as revealed by absence of CT IgM and IgG antibodies in their sera

Summary

Introduction

With an increase in the number of putative inclusion membrane proteins (incs) in chlamydial genomes, there is a need for understanding their contribution in host-pathogen interactions. In this study we determined the host mucosal and peripheral immune responses to incs (IncB and IncC) of Chlamydia trachomatis (CT). Studies on the involvement of CT IncA in homotypic membrane fusion via N-terminal SNARE-like motifs [8,21] and IncA mutant stains have been instrumental in elucidating the role of incs in disease pathogenesis and inducing modified hostcell transcriptional responses [22]. CT IncB and IncC with homologues in C. pneumoniae [23], C. psittaci [16], C. muridarum [24] and C. abortus [25] may be involved in processes like inclusion formation, transportation to perinuclear space and evasion of early lysosomal fusion as their corresponding genes are expressed within 0.5 hours of the infection cycle and coincides with these events

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