ProblemChlamydia trachomatis infections are often associated with acute syndromes including cervicitis, urethritis, and endometritis, which can lead to chronic sequelae such as pelvic inflammatory disease (PID), chronic pelvic pain, ectopic pregnancy, and tubal infertility. As epithelial cells are the primary cell type productively infected during genital tract Chlamydia infections, we investigated whether Chlamydia has any impact on the integrity of the host epithelial barrier as a possible mechanism to facilitate the dissemination of infection, and examined whether TLR3 function modulates its impact.Method of studyWe used wild-type and TLR3-deficient murine oviduct epithelial (OE) cells to ascertain whether C. muridarum infection had any effect on the epithelial barrier integrity of these cells as measured by transepithelial resistance (TER) and cell permeability assays. We next assessed whether infection impacted the transcription and protein function of the cellular tight-junction (TJ) genes for claudins1-4, ZO-1, JAM1 and occludin via quantitative real-time PCR (qPCR) and western blot.ResultsqPCR, immunoblotting, transwell permeability assays, and TER studies show that Chlamydia compromises cellular TJ function throughout infection in murine OE cells and that TLR3 deficiency significantly exacerbates this effect.ConclusionOur data show that TLR3 plays a role in modulating epithelial barrier function during Chlamydia infection of epithelial cells lining the genital tract. These findings propose a role for TLR3 signaling in maintaining the integrity of epithelial barrier function during genital tract Chlamydia infection, a function that we hypothesize is important in helping limit the chlamydial spread and subsequent genital tract pathology.

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-β), interleukin-6 (IL-6), interleukin-6 receptor alpha (IL-6Rα), soluble interleukin-6 receptor beta (sIL-6Rβ, 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.

Hormonally upregulated neu-associated kinase (HUNK) is a serine/threonine (S/T) protein kinase related to the adenosine monophosphate-activated protein kinase (AMPK) family of kinases. HUNK was originally discovered using a screen to identify kinases expressed in the mouse mammary gland. Therefore, the majority of studies to date have been carried out in models specific to this tissue, and the kinase was named to reflect its mammary gland-specific physiology and pathology. Prior studies show a clear pathogenic role for HUNK in breast cancer. HUNK is upregulated in response to oncogenic HER2/neu and Akt, and there is strong evidence that HUNK is critical for the survival of breast cancer cells. Further evidence shows that inhibiting HUNK using a variety of breast cancer models, including those that are resistant, inhibits tumorigenesis and metastasis. However, HUNK alterations are infrequent. Here, the incidence and consequence of HUNK alterations in breast cancer is reviewed using data mined from the online database cBioPortal and considered in relation to prior research studies.

21Problem: Chlamydia trachomatis infections are often associated with acute syndromes 22 including cervicitis, urethritis, and endometritis, which can lead to chronic sequalae such 23 as pelvic inflammatory disease (PID), chronic pelvic pain, ectopic pregnancy, and tubal 24 infertility. As epithelial cells are the major cell type productively infected during genital 25 tract Chlamydia infections, we investigated whether Chlamydia has any impact on the 26 integrity of the host epithelial barrier as a possible mechanism to facilitate the 27 dissemination of infection, and examined whether TLR3 function modulates its impact. 28 Method of Study:We used wild-type and TLR3-deficient murine oviduct epithelial (OE) 29 cells to ascertain whether C. muridarum infection had any effect on the epithelial barrier 30 integrity of these cells as measured by transepithelial resistance (TER) and cell 31 permeability assays. We next assessed whether infection impacted the transcription and 32 protein function of the cellular tight-junction (TJ) genes for claudins1-4, ZO-1, JAM1 and 33 occludin via quantitative real-time PCR (qPCR) and western blot. 34Results: qPCR, immunoblotting, transwell permeability assays, and TER studies show 35 that Chlamydia compromises cellular TJ function throughout the course of infection in 36 murine OE cells, and that TLR3 deficiency significantly exacerbates this effect. 37 Conclusion: Our data show that TLR3 plays a role in modulating epithelial barrier 38 function during Chlamydia infection of epithelial cells lining the genital tract. This 39 proposes a role for TLR3 signaling in maintaining the integrity of epithelial barrier function 40 during genital tract Chlamydia infection, a function that we hypothesize is important in 41 helping limit the chlamydial spread and subsequent genital tract pathology. 3 42 Introduction 43 Chlamydia trachomatis is a gram-negative intracellular bacterium and the cause of the 44 disease chlamydia, which is the most common sexually transmitted infection in the United 45 States, with over 1.7 million cases reported in the US in 2017 alone[1]. Genital tract 46 infections with C. trachomatis are associated with many acute syndromes including 47 cervicitis, urethritis, and endometritis [2]. Complications from chronic infections include 48 pelvic inflammatory disease (PID) and its sequelae of chronic pelvic pain, ectopic 49 pregnancy, and tubal infertility [3]. Although Chlamydia is treatable with antibiotics, 50 infected individuals are often asymptomatic; which facilitates the spread of the bacterium 51 through further sexual contact. As a result, Chlamydia infections have continued to rise 52 despite the implementation of screening and early intervention strategies [4]. The 53 ultimate goal in developing more effective therapeutic measures against Chlamydia 54infection is to identify aspects of host immunity that will augment clearance, while 55 minimizing immune responses that lead to genital tract pathology. 56As an obligate intracellular pathogen, Chlamy...