Immunological response of fallopian tube epithelial cells to spermatozoa through modulating cytokines and chemokines.

BACKGROUND: Spermatozoa interactions with fallopian tubes may influence fertilization. The purpose was to investigate cytokines, chemokines and growth factors expression from human fallopian tube epithelial cells (OE-E6/E7) exposed to spermatozoa. METHODS: Fresh semen samples were obtained from 10 healthy normozoospermic men. Sperms were prepared and co-cultured with OE-E6/E7. The cell line without spermatozoa was considered as the control group. Afterwards, Expression of 84 cytokines from OE-E6/E7 cell line in the presence and absence of spermatozoa were measured using PCR-array. Quantitative PCR was performed on seven genes to confirm the results of PCR-array analysis. Differentially expressed genes were subjected to www.geneontology.org and www.pantherdb.org to perform GO enrichment and panther pathway analysis. The concentration of IL-8, IL-10, IL-1B and BMP-4 in culture medium were analyzed by ELISA. RESULTS: Sperm interaction with the epithelial cells resulted in a significant increase in expression of TGF-ß2, BMP-4, IL-10, IL-9, and CD40LG markers. Moreover, expression of IL-16, IL-17F, SPP-1, CXCL-13, MSTN, IL-1A, IL-1B, IL-8, BMP-7, CSF-2, CSF-3, VEGF-A, OSM, LTA, TNF, TNFRSF11B, TNFSF11, CCL-11, CCL-20, CCL-24, CCL-3, CCL-8, CX3CL1 and CXCL-9 were considerably reduced in presence of spermatozoa. Panther pathway analysis discovered 3 pathways for upregulated genes including gonadotropin-releasing hormone receptor, TGF-beta and interleukin signaling pathways. Furthermore, 9 pathways were detected for down-regulated genes. Inflammation signaling pathway which is mediated by chemokine and cytokine contains the most number of genes. CONCLUSION: This study indicates that sperm modifies expression of cytokines, chemokines and growth factors from OE-E6/E7. Moreover, altered genes expression are toward higher survival chance of the spermatozoa.

Toll-like receptor 2 mediates the immune response of the bovine oviductal ampulla to sperm binding.

We previously reported that sperm binding to cultured bovine oviduct epithelial cells induces an anti-inflammatory immune response. Now we have developed a differentiated explant model to focus on the oviductal ampulla, where fertilization occurs, and to study the effect of sperm capacitation on the immune response. We used heparin to stimulate bovine sperm capacitation. Fluorescence imaging showed that 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolyl-carbocyanine iodide-labeled sperm pretreated with (Hep(+) ) or without (Hep(-) ) heparin rapidly attached to the explant ciliated epithelium in similar numbers. However, only Hep(+) sperm upregulated explant messenger RNA (mRNA) transcription of TLR2, IL8, TGFB1, and PGES, without changes in TNFA and IL-10 expression, while Hep(-) sperm only upregulated PGES. The responses were primarily anti-inflammatory, with a greater response produced by Hep(+) sperm, which also produced a substantial increase in TLR2 protein expression in the epithelium. The addition of TLR1/2 (toll-like receptor 1/2) antagonist to the Hep(+) and (Hep(-) ) sperm-explant coincubations reduced sperm attachment to the epithelium and inhibited TLR2 protein expression and some of the Hep(+) sperm-induced mRNA transcription. Our observations suggest that the ampullar epithelium immunologically reacts more strongly to sperm that have undergone heparin stimulation of capacitation. This anti-inflammatory response could serve to protect capacitated sperm as they approach the oocyte in the ampulla.

Adrenomedullin insufficiency alters macrophage activities in fallopian tube: a pathophysiologic explanation of tubal ectopic pregnancy.

Ectopic pregnancy is the major cause of maternal morbidity and mortality in the first trimester of pregnancy. Tubal ectopic pregnancy (TEP) accounts for nearly 98% of all ectopic pregnancies. TEP is usually associated with salpingitis but the underlying mechanism in salpingitis leading to TEP remains unclear. Adrenomedullin (ADM) is a peptide hormone abundantly expressed in the fallopian tube with potent anti-inflammatory activities. Its expression peaks at the early luteal phase when the developing embryo is being transported through the fallopian tube. In the present study, we demonstrated reduced expression of ADM in fallopian tubes of patients with salpingitis and TEP. Using macrophages isolated from the fallopian tubes of these women, our data revealed that the salpingistis-associated ADM reduction contributed to aggravated pro-inflammatory responses of the tubal macrophages resulting in production of pro-inflammatory and pro-implantation cytokines IL-6 and IL-8. These cytokines activated the expression of implantation-associated molecules and Wnt signaling pathway predisposing the tubal epithelium to an adhesive and receptive state for embryo implantation. In conclusion, this study provided evidence for the role of ADM in the pathogenesis of TEP through regulating the functions of tubal macrophages.

Roadmap to pregnancy in the first 7 days post-insemination in the cow: Immune crosstalk in the corpus luteum, oviduct, and uterus.

The first 7 days post-insemination are critical for establishment of pregnancy. The pre-ovulatory luteinizing hormone (LH) surge induces ovulation through disruption of the follicle structure that elucidates pro-inflammatory (Th1) responses. Various types of immune cells are recruited into the corpus luteum (CL) to regulate luteal angiogenesis and progesterone (P4) secretion into the circulation to establish pregnancy. The active sperm-uterine crosstalk also induces Th1 responses, mainly via Toll-like receptor (TLR) 2/4 signaling pathway in vitro. The endometrial glands serve as sensors for sperm signals, which trigger Th1 responses. Conversely, the sperm-oviduct binding generates anti-inflammatory (Th2) responses to support sperm survival until fertilization. It is well-established that embryo-maternal crosstalk starts after the embryo hatches out from the zona pellucida (ZP). However most recently, it was shown that the 16-cell stage bovine embryo starts to secrete interferon-tau (IFNT) that induces Th2 immune responses in the oviduct. Once developing embryos descend into the uterine horn, they induce Th2 responses with interferon-stimulated genes (ISGs) expression in the uterine epithelium and local immune cells mainly via IFNT release. Likewise, multiple embryos in the uterus of superovulated donor cows on D7 post-insemination induce Th2 immune responses with ISGs expressions in circulating immune cells. These findings strongly suggest that the maternal immune system reacts to the embryo during the first 7 days post-insemination to induce fetal tolerance. It became evident that the innate immunity of the developing CL, oviduct, and uterus works together to provide optimal conditions for fertilization and early embryonic development during the first 7 days post-insemination.

The Risk of Ovarian Cancer Increases with an Increase in the Lifetime Number of Ovulatory Cycles: An Analysis from the Ovarian Cancer Cohort Consortium (OC3).

Repeated exposure to the acute proinflammatory environment that follows ovulation at the ovarian surface and distal fallopian tube over a woman's reproductive years may increase ovarian cancer risk. To address this, analyses included individual-level data from 558,709 naturally menopausal women across 20 prospective cohorts, among whom 3,246 developed invasive epithelial ovarian cancer (2,045 serous, 319 endometrioid, 184 mucinous, 121 clear cell, 577 other/unknown). Cox models were used to estimate multivariable-adjusted HRs between lifetime ovulatory cycles (LOC) and its components and ovarian cancer risk overall and by histotype. Women in the 90th percentile of LOC (>514 cycles) were almost twice as likely to be diagnosed with ovarian cancer than women in the 10th percentile (<294) [HR (95% confidence interval): 1.92 (1.60-2.30)]. Risk increased 14% per 5-year increase in LOC (60 cycles) [(1.10-1.17)]; this association remained after adjustment for LOC components: number of pregnancies and oral contraceptive use [1.08 (1.04-1.12)]. The association varied by histotype, with increased risk of serous [1.13 (1.09-1.17)], endometrioid [1.20 (1.10-1.32)], and clear cell [1.37 (1.18-1.58)], but not mucinous [0.99 (0.88-1.10), P-heterogeneity = 0.01] tumors. Heterogeneity across histotypes was reduced [P-heterogeneity = 0.15] with adjustment for LOC components [1.08 serous, 1.11 endometrioid, 1.26 clear cell, 0.94 mucinous]. Although the 10-year absolute risk of ovarian cancer is small, it roughly doubles as the number of LOC rises from approximately 300 to 500. The consistency and linearity of effects strongly support the hypothesis that each ovulation leads to small increases in the risk of most ovarian cancers, a risk that cumulates through life, suggesting this as an important area for identifying intervention strategies. SIGNIFICANCE: Although ovarian cancer is rare, risk of most ovarian cancers doubles as the number of lifetime ovulatory cycles increases from approximately 300 to 500. Thus, identifying an important area for cancer prevention research.

Toll-Like Receptor 3 Deficiency Leads to Altered Immune Responses to Chlamydia trachomatis Infection in Human Oviduct Epithelial Cells.

Reproductive tract pathology caused by Chlamydia trachomatis infection is an important global cause of human infertility. To better understand the mechanisms associated with Chlamydia-induced genital tract pathogenesis in humans, we used CRISPR genome editing to disrupt Toll-like receptor 3 (TLR3) function in the human oviduct epithelial (hOE) cell line OE-E6/E7 in order to investigate the possible role(s) of TLR3 signaling in the immune response to Chlamydia Disruption of TLR3 function in these cells significantly diminished the Chlamydia-induced synthesis of several inflammation biomarkers, including interferon beta (IFN-ß), interleukin-6 (IL-6), interleukin-6 receptor alpha (IL-6Rα), soluble interleukin-6 receptor beta (sIL-6Rß, or gp130), IL-8, IL-20, IL-26, IL-34, soluble tumor necrosis factor receptor 1 (sTNF-R1), tumor necrosis factor ligand superfamily member 13B (TNFSF13B), matrix metalloproteinase 1 (MMP-1), MMP-2, and MMP-3. In contrast, the Chlamydia-induced synthesis of CCL5, IL-29 (IFN-λ1), and IL-28A (IFN-λ2) was significantly increased in TLR3-deficient hOE cells compared to their wild-type counterparts. Our results indicate a role for TLR3 signaling in limiting the genital tract fibrosis, scarring, and chronic inflammation often associated with human chlamydial disease. Interestingly, we saw that Chlamydia infection induced the production of biomarkers associated with persistence, tumor metastasis, and autoimmunity, such as soluble CD163 (sCD163), chitinase-3-like protein 1, osteopontin, and pentraxin-3, in hOE cells; however, their expression levels were significantly dysregulated in TLR3-deficient hOE cells. Finally, we demonstrate using hOE cells that TLR3 deficiency resulted in an increased amount of chlamydial lipopolysaccharide (LPS) within Chlamydia inclusions, which is suggestive that TLR3 deficiency leads to enhanced chlamydial replication and possibly increased genital tract pathogenesis during human infection.

Use of Human Fallopian Tube Organ in Culture (FTOC) and Primary Fallopian Tube Epithelial Cells (FTEC) to Study the Biology of Neisseria gonorrhoeae Infection.

Epithelial cells represent one of the most important physical barriers to many bacterial pathogens. In the case of Neisseria gonorrhoeae, the epithelial cell response is critical because they are the main target of the tissue damage triggered by the pathogen, particularly when the organism reaches the Fallopian tube (FT). Although the irreversible damage triggered by N. gonorrhoeae in the FT has been previously reported (ectopic pregnancy, pelvic inflammatory disease and infertility), the mechanisms of gonococcal-induced tissue damage are not fully understood. In addition, the lack of animal models that efficiently mimic the human disease and the complexity of gonococcus-host interactions make studying gonococcal pathogenesis particularly difficult. The use of human immortalized cells is also limited, since a variety of commercial FT cell lines is not yet available. Finally, the phase and antigenic variation of many gonococcal surface molecules involved in attachment and invasion of epithelial tissues leads to a failure to reproduce results using different human cells lines used in previous studies. The FT organ in culture (FTOC) and primary human fallopian tube epithelial cell (FTEC) represent the closest ex vivo cell models to explore the biology of Neisseria gonorrhoeae during infection of the FT, since it is a natural host target of the gonococcus. In this chapter, we describe protocols to process human FT samples to obtain FTOC and FTEC and assess their response to infection with Neisseria gonorrhoeae.

How many cell types form the epithelial lining of the human uterine tubes? Revision of the histological nomenclature of the human tubal epithelium.

INTRODUCTION: Many widely used international histological textbooks claim that the epithelium of the human uterine tube consists of two, three, and, eventually, four types of cells. Most discrepancies among these textbooks relate to debates regarding the presence or absence of basal cells, whether the peg/intercalary cells and secretory cells are the same or distinct cell populations, and if the epithelium contains a population of immunologically active cells (T- and B-lymphocytes, NK cells, macrophages and dendritic cells) or dispersed endocrine cells. METHODS: Uterine tubes were obtained from 22 women (average age: 46.73 y) undergoing gynecological surgery. The women were in fertile age, mostly in the middle of the menstrual cycle (ovulation phase). Tissue samples were processed for immunohistochemistry using primary antibodies against proliferation markers (Ki67 and PCNA), immune system cells (CD1a, CD3, CD4, CD8, CD20, CD45RO, CD56, CD68, granzyme B and S100) and disperse endocrine cells (chromogranin A and synaptophysin). RESULTS: Most of the mature tubal epithelial cells, ciliated cells, and secretory cells were mitotically active (PCNA+), a population of basal undifferentiated cells was not identified. The dividing cells had a narrow-shaped nucleus (Ki67 positive). These cells were morphologically identical to - by the terminology mentioned - intercalary cells, assuming they represented actually dividing cells (epitheliocytus tubarius mitoticus). The tubal "basal cells" displayed small, hyperchromatic nuclei and very pale cytoplasm (clear cytoplasmic halo). They were located in the epithelium adjacent to the basement membrane, were non-mitotically active and their immunophenotype corresponded to intraepithelial regulatory T-lymphocytes (CD3+, CD8+, CD45RO+, CD4-, CD20-, CD56- and granzyme B-). Intraepithelial B-lymphocytes were only rarely identified. Intraepithelial NK cells, dendritic cells, macrophages and dispersed endocrine cells were not identified. CONCLUSIONS: We recommend replacing the term "epitheliocytus tubarius basalis" in the Terminologia Histologica with the term "lymphocytus T intraepithelialis tubarius", which represents intraepithelial regulatory T-cells (CD8+, CD45RO+) of the uterine tube. Additionally, we propose that intercalary/peg cells are actively dividing cells, instead of effete or degenerating cells. Finally, the histological nomenclature should be corrected in a way that peg/intercalary cells are not considered synonymous terms for secretory cells.

Chronic Chlamydia infection in human organoids increases stemness and promotes age-dependent CpG methylation.

Chronic infections of the fallopian tubes with Chlamydia trachomatis (Ctr) cause scarring and can lead to infertility. Here we use human fallopian tube organoids and genital Ctr serovars D, K and E for long-term in vitro analysis. The epithelial monolayer responds with active expulsion of the bacteria into the lumen and with compensatory cellular proliferation-demonstrating a role of epithelial homeostasis in the defense against this pathogen. In addition, Ctr infection activates LIF signaling, which we find to be an essential regulator of stemness in the organoids. Infected organoids exhibit a less differentiated phenotype with higher stemness potential, as confirmed by increased organoid forming efficiency. Moreover, Ctr increases hypermethylation of DNA, which is an indicator of accelerated molecular aging. Thus, the chronic organoid infection model suggests that Ctr has a long-term impact on the epithelium. These heritable changes might be a contributing factor in the development of tubal pathologies, including the initiation of high grade serous ovarian cancer.

Correlation of clinical data with fallopian tube specimen immune cells and tissue culture capacity.

Human fallopian tube fimbria secretory epithelial cells (hFTSECs) are considered an origin of ovarian cancer and methods for their culture from fallopian tube specimens have been reported. Our objective was to determine whether characteristics of the donors or surgeries were associated with the capacities of fimbria specimens to generate hFTSEC cultures or their immune profiles. There were no surgical complications attributable to fallopian tube removal. Attempts to establish primary hFTSEC cultures were successful in 37 of 55 specimens (67%). Success rates did not differ significantly between specimens grouped by patient or surgery characteristics. Established cultures could be revived after cryopreservation and none became contaminated with microorganisms. Two cultures evaluated for long term growth senesced between passages 10 and 15. M1 macrophages were the predominant cell type, while all other immune cells were present at much lower percentages. IL-10 and TGF-ß exhibited opposing trends with M1 and M2 macrophages. Plasma IL-10 levels exhibited significant positive correlation with patient age. In conclusion, fallopian tube fimbria specimens exhibit a pro-inflammatory phenotype and can be used to provide a source of hFTSECs that can be cultured for a limited time regardless of the donor patient age or race, or the type of surgery performed.

The prevalence of Chlamydia trachomatis and Mycoplasma genitalium tubal infections and their effects on the expression of IL-6 and leukaemia inhibitory factor in Fallopian tubes with and without an ectopic pregnancy.

This was a prospective case-control study that measured the prevalence of Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG) and Mycoplasma genitalium (MG) by an IVD CE multiplex PCR kit in fresh Fallopian tubes (FT) obtained from 96 ectopic pregnancies (EP) and 61 controls in the midluteal phase of the cycle. We later measured the expression profile of IL-6, leukaemia inhibitory factor (LIF) and their signalling molecules, in respect to the type and number of infections, by immunohistochemistry, ELISA and quantitative RT-PCR. The frequencies of CT, and MG mono- and co-infections were significantly higher in EP. IL-6, LIF, their receptors and intracellular mediators were significantly up-regulated at the gene and protein levels in positive compared with negative FTs within each group (P < 0.05). EP tubal samples with co-infections showed the highest significant expression of the candidate cytokines by all techniques (P < 0.05). CT and MG are frequent in EP and up-regulate the tubal expression of IL-6, LIF and their signalling molecules. Both cytokines could be involved in the tubal immune response against bacterial infections, as well as the pathogenesis of EP. Further studies are needed to explore the roles of IL-6 family in infection-induced tubal inflammation and EP.

Prognostic impact of tumour-associated B cells and plasma cells in epithelial ovarian cancer.

BACKGROUND: The critical role of the immune system in controlling cancer progression has become evident and immune modulatory therapy is now approved for clinical use. However, while the majority of studies on the inflammatory tumour microenvironment have focused on the cellular immune response, in particular the prognostic and predictive role of various T cell infiltrates, the role of the humoral immune response in this context has long been overlooked. This study aimed to investigate the clinicopathological correlates and prognostic impact of B cell and plasma cell infiltration in epithelial ovarian cancer (EOC). METHODS: Immunohistochemical expression of immunoglobulin kappa C (IGKC), CD20 and CD138 was analysed in tissue microarrays with tumours from 154 incident cases of EOC from two pooled prospective population-based cohorts. Subsets of corresponding benign-appearing fallopian tubes (n = 38) and omental metastases (n = 33) were also analysed. Kaplan-Meier analysis and Cox regression analysis were used to determine the impact of immune-cell specific IGKC, CD20 and CD138 expression on overall survival and ovarian cancer-specific survival. RESULTS: High IGKC expression correlated significantly with expression of CD20 (p = 0.001) and CD138 (p = 0.035). Expression of IGKC as well as CD138 was significantly higher in primary tumours than in fallopian tubes (p = 0.004 and p = 0.001, respectively). High CD20 and CD138 expression correlated significantly with high tumour grade (p = 0.032 and p = 0.030, respectively). CD20 and IGKC expression was not prognostic but univariable Cox regression analysis revealed high CD138 expression to correlate with a significantly reduced overall survival (HR = 2.20; 95 % CI 1.34-3.55; p-0.001) as well as ovarian cancer-specific survival (HR = 1.95; 95 % CI 1.28-2.98; p = 0.002). The prognostic impact was independent of established clinical parameters (age, grade, clinical stage) as shown in multivariable analysis (HR = 2.28; 95 % CI 1.39-3.75; p = 0.001). CONCLUSIONS: In conclusion, our results demonstrate that plasma cell infiltration in epithelial ovarian cancer has a significant impact on tumour progression and prognosis. The important role of the humoral immune system merits further study and may be harnessed as immune modulatory strategies in cancer therapy.

Secrets of Endometrial Receptivity: Some Are Hidden in Uterine Secretome.

PROBLEM: Endometrium, the innermost mucosal layer of the uterus, serves as a lodge for the embryo in eutherian mammals. The endometrium is constituted of various cell types, and each cell type executes specific functions to facilitate embryo implantation and development. It is well established that the endometrium, despite being non-permissive to the embryo for the major period of a menstrual cycle, is irreplaceable in the scheme of events essential for procreation. However, the embryo, before initiating physical contact with the endometrium, encounters the uterine cavity that remains bathed in uterine fluid. Uterine fluid is an admixture of endometrial secretions, plasma transudates, and oviductal fluid. Uterine fluid components are believed to play important roles in immunosuppression and embryo development during peri-implantation period. Uterine fluid is also involved in defense against pathogens, sperm migration, and lubrication of endometrium. The advent of high-throughput functional genomics tools has created enormous opportunities to investigate the uterine fluid for its protein repertoire and modulation during the receptive phase of an endometrial cycle in animals and humans. Towards this, few investigations have been conducted in recent years. The data obtained using non-targetted functional genomics approaches need to be assimilated with the existing information on specific components of uterine fluid. METHOD: This review compiles existing information on the composition of uterine fluid and its significance in endometrial functions and dysfunctions. RESULT: Collectively, investigations based on targetted and non-targetted approaches have revealed the presence of several cytokines, growth factors, ions, carbohydrates, and steroids, in human uterine fluid. CONCLUSION: Detailed investigations of human uterine fluid, especially directed towards the elucidation of functional relevance of different proteins in uterine fluid, will help identify novel markers of endometrial receptivity and also gain significant insights into the mechanisms underlying unexplained infertility, recurrent pregnancy losses, and other endometrial pathologies.

Role of STAT1 in Chlamydia-Induced Type-1 Interferon Production in Oviduct Epithelial Cells.

We previously reported that Chlamydia muridarum-infected murine oviduct epithelial cells (OE cells) secrete interferon ß (IFN-ß) in a mostly TLR3-dependent manner. However, C. muridarum-infected TLR3-deficient OE cells were still able to secrete detectable levels of IFN-ß into the supernatants, suggesting that other signaling pathways contribute to Chlamydia-induced IFN-ß synthesis in these cells. We investigated the role of STAT1 as a possible contributor in the Chlamydia-induced type-1 IFN production in wild-type (WT) and TLR3-deficient OE cells to ascertain its putative role at early- and late-times during Chlamydia infection. Our data show that C. muridarum infection significantly increased STAT1 gene expression and protein activation in WT OE cells; however, TLR3-deficient OE cells showed diminished STAT1 protein activation and gene expression. There was significantly less IFN-ß detected in the supernatants of C. muridarum-infected OE cells derived from mice deficient in STAT1 when compared with WT OE cells, which suggest that STAT1 is required for the optimal synthesis of IFN-ß during infection. Real-time quantitative polymerase chain reaction analyses of signaling components of the type-1 IFN signaling pathway demonstrated equal upregulation in the expression of STAT2 and IRF7 genes in the WT and TLR3-deficient OE cells, but no upregulation in these genes in the STAT1-deficient OE cells. Finally, experiments in which INFAR1 was blocked with neutralizing antibody revealed that IFNAR1-mediated signaling was critical to the Chlamydia-induced upregulation in IFN-α gene transcription, but had no role in the Chlamydia-induced upregulation in IFN-ß gene transcription.

Characterization of tubal occlusion after transcervical polidocanol foam (PF) infusion in baboons.

OBJECTIVE: Our long-term goal is to develop a nonsurgical method of fallopian tubal occlusion for the purpose of permanent contraception. We have previously demonstrated that transcervical administration of 5% polidocanol foam (PF) can create tubal occlusion in macaques but that multiple treatments are required. In this study, we assessed the efficacy of various regimens of PF with and without depomedroxyprogesterone acetate (DMPA) (to control ovarian cycle phase) in the baboon. STUDY DESIGN: Adult cycling female baboons were evaluated for tubal patency by hysterosalpingography and then received a transcervical infusion of PF with (+) or without (-) an intramuscular injection of DMPA (3.5 mg/kg). Two concentrations of PF were compared: 1% [(+) DMPA, n=5; (-) DMPA, n=3] and 5% [(+) DMPA, n=4; (-) DMPA, n=3]. Controls received (+) DMPA (n=2) or (-) DMPA, (n=3) only. The reproductive tracts were removed 1-3 months after treatment for examination. RESULTS: No fallopian tubal occlusion was observed in negative controls (±DMPA). Histologic complete tubal occlusion was observed in 3/8 of females treated with 1% PF and in 6/7 treated with 5% PF. Histologic evaluation suggested that 1% PF is associated with prolonged chronic inflammation (more than 2-3 months), while 5% treatment eliminates the epithelial lining, at least focally, and resolves into complete occlusion within 1-2 months. This pattern of complete occlusion was seen in all 4 females that received 5% PF (+DMPA) and in 2/3 that received 5% PF (-DMPA). CONCLUSION: In a baboon model of transcervical permanent contraception, a single treatment with 5% PF resulted in complete tubal occlusion more reliably (85%) than 1% PF (38%). Cotreatment with DMPA may improve treatment results with 5% PF but requires additional study. IMPLICATIONS: A finding that a single transcervical treatment with 5% PF can occlude the fallopian tubes of baboon supports further study of this approach as a novel strategy for permanent contraception for women.

Expression pattern of inflammatory response genes and their regulatory micrornas in bovine oviductal cells in response to lipopolysaccharide: implication for early embryonic development.

In the present study, we used an in vitro model to investigate the response of the oviduct with respect to inflammatory mediators and their regulatory microRNAs in case of bacterial infection and subsequent association with embryo survival. For this, we conducted two experiments. In the first experiment, cultured primary bovine oviductal cells (BOEC) were challenged with lipopolysaccharide (LPS) for 24h and the temporal expression pattern of inflammatory mediators and their regulatory microRNAs were measured at 0, 3, 6, 12, 24 and 48h after LPS treatment. Intriguingly, the temporal patterns of all miRNAs except miR-21 were significantly up-regulated at 6h after LPS treatment. Whereas, we observed significant overexpression of pro-inflammatory mediators as tumor necrosis factor alpha (TNFα) and interleukin-1 beta (IL1ß) after LPS challenge for 24h. On the other hand, the expression level of essential elements like oviductal glycoprotein 1 (OVGP1) and insulin-like growth factor 2 (IGF2) was significantly decreased in challenged groups compared with control. Moreover, miR-155, miR-146a, miR-223, miR-21, miR-16 and miR-215 have shown a clear suppression in challenged group after LPS treatment. In the 2nd experiment there were four groups of blastocysts produced, namely embryo+LPS free media, embryo+LPS, BOEC+embryo and BOEC+embryo+LPS. The suboptimal oviduct environment due to LPS challenge is found to have a significant influence on the expression of inflammatory response genes (TNFα and CSF1), stress response genes (SOD and CAT), mitochondrial activity, reactive oxygen species (ROS) accumulation and apoptotic level either in cultured or co-cultured blastocysts. Collectively, LPS challenge led to aberrant changes in oviductal transcriptome profile, which could lead to a suboptimal environment for embryo development.

Pathogenesis of fallopian tube damage caused by Chlamydia trachomatis infections.

Chlamydia trachomatis is the leading cause of bacterial sexually transmitted disease worldwide resulting in 4-5 million new cases of Chlamydia annually and an estimated 100 million cases per annum. Infections of the lower female genital tract (FGT) frequently are asymptomatic; thus, they often remain undiagnosed or untreated. If infections are either not resolved or left untreated, chlamydia can ascend to the upper FGT and infect the fallopian tubes (FTs) causing salpingitis that may lead to functional damage of the FTs and tubal factor infertility (TFI). Clinical observations and experimental data have indicated a role for antibodies against C. trachomatis proteins such as the 60-kDa heat shock protein 60 (cHSP60) in the immunopathogenesis of TFI. When released from infected cells, cHSP60 can induce proinflammatory immune responses that may functionally impair the FTs leading to fibrosis and luminal occlusion. Chlamydial pathogenesis of irreversible and permanent tubal damage is a consequence of innate and adaptive host immune responses to ongoing or repeated infections. The extracellular matrix that is regulated by metalloproteinases may also be modified by chlamydial infections of the FGT. This review will highlight protective and pathogenic immune responses to ongoing and repeated chlamydial infections of the FGT. It will also present two recent hypotheses to explain mechanisms that may contribute to FT damage during a C. trachomatis infection. If Chlamydia immunopathology can be controlled, it might yield a method of inducing fibrosis and thus provide a means of nonsurgical permanent contraception for women.

Evaluation of immunological interaction between spermatozoa and fallopian tube epithelial cells.

Toll-like receptors (TLR) are one of the major compartments of innate immune system. It was revealed that the TLR have relevance in ovulation, sperm capacitation and fertilisation. So, in this study, the expression of TLR, their adaptor molecules and cytokines in human fallopian tube cell line under the effect of human normal spermatozoa was evaluated. TLR mRNA and protein were evaluated in OE-E6/E7 cell line. Semen samples from 10 donors were collected and co-incubated with OE-E6/E7 cell line and used as sperm group, and cell line without spermatozoa was used as control group. Afterwards, the level of TLR, their adaptor molecule and cytokine mRNA expression was compared using qPCR in sperm and control groups, and supernatant was used for ELISA. To determine whether elevated cytokine reaction to spermatozoa in OE-E6/E7 cell line is mediated via TLR, TLR3 function-blocking antibody was used. OE-E6/E7 cell line expressed TLR1-6 genes and proteins. TLR expressions, especially TLR3 and TLR5, in OE-E6/E7 cell line under the effect of spermatozoa were significantly higher. Also, levels of adaptor molecules and cytokine production were increased in sperm group than in control group (P < 0.05). So, it may be hypothesised that TLR are essential for spermatozoa and fallopian tube immunological interaction and for preparing safe environment for important events in fallopian tube.

Characterization of the immune cell repertoire in the normal fallopian tube.

Recent studies implicating the fallopian tube as the site of putative precursors of ovarian serous carcinoma, and the hypothesis that injury, inflammation, and repair of the ovarian surface epithelium at the time of ovulation, may be contributing factors to ovarian carcinogenesis, prompted us to undertake a comprehensive analysis of the immune cells in the normal fallopian tube. Accordingly, the aim of this study was to provide a baseline for future studies exploring the relationship of inflammation with the early events of ovarian carcinogenesis by characterizing the immune cell repertoire in 13 normal human fallopian tubes, combining digital microscopy of immunostained slides and flow cytometry of fresh single-cell suspensions, with a panel of markers that identify the most important adaptive and innate immune cells. We found that CD45(+) leukocytes are regularly observed in the fallopian tube and are mainly composed of CD163(+) macrophages, CD11c dendritic cells, and CD8(+) T cells. In addition, there are minor populations of CD56(+) NK cells, CD4(+) T cells, CD20(+) B cells, TCRγδ(+) T cells, and, among dendritic cells, CD207(Langerin)(+) Langerhans cells. The cellular mapping that we performed indicates that the local immune system in the human fallopian tube is composed of a mixture of innate and adaptive immune cells, many of which are recognized as playing a role in cancer immune surveillance. This local immune system could provide a first line of defense against early precancerous lesions and could potentially be exploited for immune-based therapies.

Different subsets of Langerhans cells in human uterine tubes and uterus.

AIM: Langerhans cells (LC) are antigen-presenting cells present in tissues with high antigenic exposure. Their role in the upper female reproductive tract is not fully understood. This study aims to determine the distribution and morphology of LC in the normal and post-partum human uterine tubes and uterus by staining with the specific LC markers, CD1a and zinc iodide-osmium (ZIO), and to determine their association with helper and cytotoxic T cells. MATERIAL AND METHODS: Normal and post-partum uterine tube and uterine specimens were stained with CD1a and ZIO and their morphology and distribution noted. Double immune staining with CD1a-CD4 and CD1a-CD8 in post-partum uterine tube were also done. RESULTS: It was noted that CD1a-positive cells were significantly fewer and smaller in diameter than ZIO-positive cells in the uterine tube and both types of cells were significantly more prevalent in post-partum tubes. Perivascular clusters of ZIO-positive cells were seen in the post-partum tubes. Close association of CD1a-positive cells with CD4- and CD8-positive T cells was noted in the post-partum uterine tube. In the uterus, scanty CD1a-positive cells were present in the surface and glandular epithelium and endometrial stroma. ZIO-positive cells were absent. CONCLUSION: This study suggests that CD1a-positive and ZIO-positive cells may be different subsets of LC that are needed for presentation of antigen to immunocompetent cells. Their respective functions are yet to be determined.