Banashree Das

Cytokine expression pattern in the genital tract of Chlamydia trachomatis positive infertile women – implication for T-cell responses

Human genital infection caused by Chlamydia trachomatis is thought to be immunologically mediated, resulting in local recruitment of lymphocyte subsets and inducing the production of cytokines. Little information is available about the role of lymphocyte recruitment and the regulation of cytokine production in the genital tract of C. trachomatis positive infertile women. We have evaluated the recruitment of lymphocyte subsets in the genital tract and production of Th1/Th2 cytokines in cervical secretions and laparoscopic specimens from the fallopian tubes of C. trachomatis positive infertile women (n = 17) and compared them with controls, viz. C. trachomatis negative infertile women (n = 20) using ELISA and flow cytometry. None of these patients were found to be infected either with Candida sps., bacterial vaginosis, Trichomonas vaginalis, Neisseria gonorrhoeae, Mycoplasma hominis or Ureaplasma urealyticum in the cervix. Flow cytometric analysis of cervical secretions in Chlamydia positive women revealed recruitment of both CD4 and CD8 lymphocytes to the genital tract was up‐regulated and a variation in the production rates of different cytokines in cervical secretions and fallopian tube was observed. We found that the immune responses in cervical secretions were of Th0 type, since all the analysed cytokines, viz. IFN‐γ, TNF‐α, IL‐10 and IL‐12 were up‐regulated. As, both CD4 and CD8 cells contribute to the production of IFN‐γ and IL‐10, these results suggest that along with CD4 cells, CD8 lymphocytes  also  may  be  important  for  local  regulation  of  Th1/Th2  responses  in  the genital  tract  during C. trachomatis infection.

Predominance of Chlamydia trachomatis Serovars Associated with Urogenital Infections in Females in New Delhi, India

ABSTRACT Screening for Chlamydia trachomatis was done for 280 endocervical swab samples by PCR specific for endogenous plasmid. Age dependency was seen in symptomatic patients, with a high chlamydial prevalence rate (28%) found in younger women. Genotyping by restriction fragment length polymorphism analysis of omp1 PCR-positive samples showed serovars D, E, and F to be the most prevalent.

Effect of treatment for Chlamydia trachomatis during pregnancy

Objectives: To screen and treat chlamydial infection in pregnant women in order to assess the effects of therapeutic intervention on the outcome of Chlamydia trachomatis‐infected pregnancy. Methods: Enrolled in the study were 350 women in the first to third trimesters of pregnancy. Endocervical swabs were collected for C. trachomatis diagnosis by DFA and PCR. A few STD infections, viz.: Neisseria gonorrhoeae, Candida spp., bacterial vaginosis, Trichomonas vaginalis and Treponema pallidum were ruled out. After excluding 22 patients infected with other sexually transmitted infections, a cohort of 328 pregnant women comprised the study population. Anti‐chlamydial treatment (viz.: oral therapy with erythromycin stearate, 500 mg 4 times daily for 7 days) was given to 17 women (group I) and their partners. Fifteen patients of group I were retested by DFA and PCR assay for C. trachomatis infection 2 weeks following therapy. Other Chlamydia‐positive patients (n=42) who were lost to follow‐up were classified as untreated positive cases (group II) while group III included C. trachomatis negative cases (n=269). Data on obstetric outcome were recorded in a total of 164 cases. Statistical comparison of the data were done using the χ2‐test and means were compared using Students t‐test. Results: Among the 350 pregnant women enrolled initially for the study, C. trachomatis positivity was found to be 18.8% (n=66) in the endocervix by DFA and PCR assay. Co‐infection with Candida spp., bacterial vaginosis, T. vaginalis and T. pallidum was 2.0%, 1.7%, 1.7% and 0.5%, respectively. None of the pregnant women was infected with N. gonorrhoeae. Pregnant women at an increased risk of chlamydial infection included those who had multiple births and were in second trimester of pregnancy. Fifteen patients of group I became Chlamydia‐negative following treatment. Data on obstetric outcome were recorded in 11, 26 and 127 patients of groups I, II and III, respectively. The mean duration of gestation for premature deliveries was found to be significantly higher in group I in comparison with group II [35.5 vs. 33.1 weeks (P<0.05)], thereby showing an improved effect of treatment on pregnancy outcome. The mean of low birth weight births was higher in group I compared with group II (2200.0 vs. 2113.3 g), however, this was statistically non‐significant. Stillbirths were significantly higher among group II in comparison with group III [11.5% vs. 4.7% (χ2=1.79; P<0.5)]. No stillbirths were recorded in patients who had taken anti‐chlamydial treatment. Conclusions: Our findings suggest that routine screening and treatment of C. trachomatis infection in pregnant women, especially those in high risk groups, should be mandatory to reduce the adverse effects on obstetric outcome.

A severe case of pubic symphysis diastasis in pregnancy.

Pubic symphysis diastasis in pregnancy is a rare occurrence. We report a case of severe pubic symphysis diastasis in pregnancy in a primigravida.

Serum IgM to Chlamydia trachomatis in pregnancy: its usefulness for screening.

Abstract Asymptomatic infection with Chlamydia trachomatis represents an important health problem. A non-invasive diagnostic test to screen pregnant women is needed that is cost effective and can be used widely, especially in developing countries. In this setting, quantitation of antichlamydial IgM antibodies may offer an additional strategy for the control of C. trachomatis infection. The aim of this prospective study is to evaluate the quantitation of serum antichlamydial IgM antibodies, based on absorbance (A) values, in pregnant women for the prediction of C. trachomatis infection. Serum samples from a cohort of 148 pregnant women (first to third trimesters; age range: 18-35 years) presenting to the antenatal department at Safdarjang Hospital were tested for IgM antibodies specific to C. trachomatis by an enzyme-linked immunosorbent assay (ELISA) kit (Novum Diagnostics, Germany). Co-infection with other STD pathogens was ruled out. In this cohort, 85 (57.4%) pregnant women were found to be positive for IgM antibodies to C. trachomatis. Based on the cut-off value of the ELISA test (calculated as 0.558), pregnant women with an A value between 0.558 and 0.999 and those with a value >1.000 were categorised as low positive (LP, n=41) and high positive (HP, n=44), respectively. The differences in mean A values for the LP versus negative groups (0.7504 versus 0.2249, P<0.05) and the HP versus negative groups (1.5353 versus 0.2249, P<0.05) were statistically significant. Maximum seroprevalence (44.4%, P<0.05) was found among those in the HP group in the first trimester of pregnancy. Multigravidae (34.4%, P<0.5) and multiparous (34.9%, P<0.5) pregnant women in the HP group were at an increased risk of chlamydial infection. As overall results indicated that pregnant women in the HP group were at higher risk, we stress the importance of large-scale screening of pregnant women for C. trachomatis infection, particularly in developing countries where sophisticated techniques for collection/diagnosis are as yet unavailable.

Does aberrant expression of cyclooxygenase-2 and prostaglandin-E2 receptor genes lead to abortion in Chlamydia trachomatis-infected women

Abstract Objective: The aim of the study was to characterize the expression of cyclooxygenase-2 (COX-2) and prostaglandin-E2 (PGE2) genes in spontaneously aborted tissues from women infected with Chlamydia trachomatis. Methods: A total of 135 spontaneous aborters (Group I) and 45 induced aborters (controls; Group II) attending Obstetrics and Gynaecology Department at Safdarjung hospital (New Delhi, India), were enrolled. Polymerase chain reaction (PCR) assay was performed to detect C. trachomatis DNA in endometrial curettage tissue (ECT). Differential expression of COX-2 and PGE2 receptors at mRNA level was analysed in ECT using reverse transcription PCR and real-time PCR. Results: In total, 14.8% patients were diagnosed as C. trachomatis-positive in Group I whereas all control patients were C. trachomatis-negative. Qualitative expression of COX-2 (p < 0.05) and PGE2 (p < 0.0001) receptors was found increased in C. trachomatis-positive patients (Group I) in comparison to controls. Quantitative real-time PCR analysis also showed upregulation in transcript levels of both COX-2 (p < 0.002) and PGE2 (p < 0.0001) receptors in infected patients (Group I) versus Group II. COX-2 and PGE2 expression was higher (p < 0.002) in recurrent spontaneous aborters in comparison to sporadic spontaneous aborters. Conclusions: Results suggest that chlamydial infection leads to upregulation of COX-2 in C. trachomatis-positive recurrent spontaneous aborters, which probably mediates increased prostaglandin synthesis.

Expression of prostaglandin receptors in Chlamydia trachomatis-infected recurrent spontaneous aborters.

A study was undertaken to quantify the expression of prostaglandin (PG) receptors and find the effect of gestational age on expression of PG receptor genes in Chlamydia trachomatis-infected recurrent spontaneous aborters (RSA). Endometrial curettage tissue (ECT) was collected from 130 RSA (Group I) and 100 age-matched controls (Group II) at the Department of Obstetrics and Gynecology, Safdarjung Hospital, New Delhi (India). PCR was performed for diagnosis of C. trachomatis cryptic plasmid; mRNA expression of PG receptor genes was assessed by real-time PCR (q-PCR), while serum progesterone/estrogen levels were determined by respective commercial kits. Data were evaluated statistically. A total of 15.4 % RSA (GroupI) were diagnosed as C. trachomatis-positive (200 bp), whereas controls were uninfected. q-PCR showed significant upregulation (P<0.0001) of PGE2 (EP-1, EP-2, EP-3, EP-4), PGF2α (FP) and PGI2 (IP) receptors in Group I versus Group II. The expression of PG receptors increased significantly with advanced gestational age (P<0.002); however, only contractile receptors, EP-1, EP-3 and FP, were positively correlated with gestational age in Group-I. In infected RSA, mean serum progesterone level was significantly low (P<0.0001) while serum oestrogen was high (P<0.0001). Overall, the data suggest that increased expression of PG receptors, particularly contractile gene receptors (EP-1, EP-3, FP), with advanced gestational age and altered steroid levels could be a possible risk factor for abortion in Chlamydia-infected RSA.

Involvement of matrix metalloproteinases and their inhibitors in endometrial extracellular matrix turnover in Chlamydia trachomatis-infected recurrent spontaneous aborters

&NA; Mechanism underlying Chlamydia trachomatis disease pathology is not completely understood. It was hypothesised that dysregulation of extracellular matrix by matrix metalloproteinases (MMPs) might be involved in sequelae of events leading to abortion in C. trachomatis‐infected women. Hence, this study was undertaken to elucidate the expression of MMP‐2/MMP‐9/TIMP‐1/TIMP‐3 in infected recurrent spontaneous aborters (RSA) and simultaneously ascertain effect of few cytokines (TNF‐&agr;, TGF‐&bgr;1, TGF‐&bgr;2)/cyclooxygenase‐2 (Cox‐2) on MMPs. Endometrial curettage tissue was collected from 135 RSA (group I) and 120 age‐matched controls (group II) at Department of Obstetrics and Gynecology, SJH, New Delhi (India). PCR was performed for diagnosis of C. trachomatis MOMP/plasmid gene. Expression of MMP‐2/MMP‐9/TIMP‐1/TIMP‐3 mRNA was assessed by quantitative real‐time PCR. Data were statistically evaluated. 15.6% (n = 21) RSA (Group‐I) were diagnosed as C. trachomatis positive. Significant upregulation of MMP‐2/MMP‐9 and decreased TIMP‐1/TIMP‐3 were found in C. trachomatis‐positive RSA (group I) versus control women (group II). MMP‐2 was found positively correlated with cytokines/Cox‐2. Significant positive correlation was observed between MMP‐2/MMP‐9 and C. trachomatis copy load. Overall data suggested that dysregulated MMPs/TIMPs promoted excessive endometrial matrix degradation which, in turn, affected pregnancy leading to spontaneous abortion in infected women. &NA; Graphical Abstract Figure. Dysregulated MMPs and TIMPs expression promoted endometrial extracellular matrix degradation and affected early pregnancy eventually leading to spontaneous abortion in Chlamydia trachomatis‐infected recurrent aborters.

Cytokine-induced expression of nitric oxide synthases in Chlamydia trachomatis-infected spontaneous aborters

Abstract Purpose: The aim of study was to evaluate expression of inducible nitric oxide synthase (iNOS), endothelial nitric oxide synthase (eNOS), interferon-gamma (IFN-γ), and tumor necrosis factor-alpha (TNF-α) in Chlamydia trachomatis (CT)-infected spontaneous aborters (SA). Materials and methods: Endometrial curettage tissue was collected from 140 SA (sporadic SA- 70; recurrent SA- 70) (Group I) and 140 age-matched controls (Group II) from Department of Obstetrics and Gynecology, Safdarjung Hospital, New Delhi, India. Polymerase chain reaction was performed for diagnosis of CT. The expression of iNOS/ eNOS/ IFN-γ/ TNF-α was assessed by real-time polymerase chain reaction (PCR). Results: 15.7% SA were CT-positive (Group I); none in controls. Sporadic spontaneous aborters (SSA) (n = 8/70), recurrent spontaneous aborters (RSA) (n = 14/70) diagnosed as CT-positive (Group-I). Significant upregulation of iNOS/ eNOS was found in CT-positive SSA/RSA compared with CT-negative SSA/RSA and healthy controls. TNF-α and IFN-γ were expressed in CT-positive SSA/RSA compared with negative SSA/controls. iNOS showed a significant strong positive correlation with TNF-α and IFN-γ in CT-infected SA. eNOS showed a significant positive correlation with TNF-α and no correlation with IFN-γ in CT-infected SA. TNF-α was positively correlated with IFN-γ. Conclusions: Significantly high expression of iNOS/ eNOS and proinflammatory cytokines affected pregnancy in CT-infected RSA, thereby implying that there occurs cytokine-induced expression of nitric oxide synthase (NOS).

In situ localization of Chlamydia trachomatis and Chlamydial Heat Shock Protein 60 in Endometrial Curettage Tissue of Recurrent Spontaneous Aborters

Sangita Rastogi1*, Chanchal Yadav1 and Banashree Das2 1Microbiology Laboratory, National Institute of Pathology (ICMR), Sriramachari Bhawan, Safdarjung Hospital Campus, Post Box No. 4909, New Delhi-110029, India 2Department of Obstetrics and Gynecology, Vardhman Mahavir Medical College and Safdarjung Hospital, New Delhi-110029, India *Corresponding author: Dr. Sangita Rastogi, Microbiology Laboratory, National Institute of Pathology (ICMR), Sriramachari Bhawan, Safdarjung Hospital Campus, Post Box No. 4909, New Delhi-110029, India, Tel: +91-09810217260; E-mail: rastogi_sangita@rediffmail.com