Chlamydia Muridarum Genital Infection Research Articles

Characterization of CD4+ t cell subsets during Chlamydia muridarum genital infection

Characterization of CD4+ T cell subsets in a stressed mouse model during Chlamydia muridarum genital infection. Katrina Davis, Tesfaye Belay. Department of Applied Sciences and Mathematics. Chlamydia genital infection caused by Chlamydia trachomatis is highly prevalent in low socioeconomic status populations. Whether stress is a risk factor for the high incidence of chlamydia remains to be investigated. CD4+ T cells are the major immune cells that fight chlamydia genital infection; however, the dynamics of CD4+ T cells during stressful conditions and chlamydia infection are unclear. The profiles of CD4+ T cell signature cytokines: T helper 1 (IL-12, Interferon-gamma (IFN-γ), Thelper2 (IL-4, IL-5, IL-13), Thelper17(IL-17), and Treg (IL-10). Mice were stressed for 5 minutes daily for 21 days and infected with 1x 105 IFU/ml of Chlamydia muridarum intravaginally. Non-stressed mice were infected as controls. Mice were sacrificed 7 days post-infection, the spleen and lymph nodes were harvested, and T cells were purified and proliferated for 72 h. Determination of the levels of cytokines in culture supernatants of T cells from stressed and non-stressed mice are underway for poster presentation. Supported by the NIH grant # 1R15AI124156-01 awarded to BSU

Active Hexose Correlated Compound Feeding to Stress Mice Induces Differential Cytokine Production during Chlamydia muridarum Genital Infection

Oral supplementation of Active Hexose Correlated Compound (AHCC) mushroom extract is reported to enhance resistance to infection. However, AHCC feeding to stressed mice during Chlamydia muridarum. genital infection and host immune response remain unexplored. This study aimed to explore cytokine production profiles of immune cells of AHCC-fed stressed mice. Wildtype C57BL/6J mice that were stressed and infected with C. muridarum intravaginally. After seven days, splenic T cells and bone marrow-derived dendritic cells (DCs), and macrophages (MØs) were proliferated, and culture supernatants were collected to determine cytokine production by ELISA. Phenotyping of surface markers of immune cells using flow cytometry is underway. ELISA results show that AHCC increases the production of Th1 (IL-12 and IFN-g) and Th2 (IL-10, IL-13, IL.23) cytokines compared to PBS-fed stressed mice. Increased production of TNF-α in LPS-treated DCs and MØs of WT was observed. Overall, AHCC-feeding increases the production of protective and suppressive cytokines.

Cytokine production profiles of immune cells of stressed beta2-adrenergic receptor knockout mouse model during Chlamydia muridarumgenital infection

Abstract Chlamydia genital infection by Chlamydia trachomatisis the most common bacterial sexually transmitted disease worldwide. The relationships between stress and chlamydia genital infection remain unknown. Beta2-adrenergic receptor(β2-AR), the major receptor of the stress hormone norepinephrine (NE), is known to impair the function of immune cells. Stress was induced by immersing mice in cold water for five minutes daily for 21 days before infection. This study determined profiles of immune cells of stressed β2-AR KO and stressed WT C57BL/6J after 48 h of Chlamydia muridarumgenital infection. Splenic T cells and differentiated bone marrow-derived macrophages and dendric cells (DCs) were purified by negative selection and stimulated by seeding T cells on anti-CD3/CD28 antibodies coated wells and by treating macrophages and DCs with LPS. After 48 h proliferation, cytokine levels in culture supernatants of the cells were determined using ELISA. Results show increased IFN-γ and IL-1β secretions but deceased IL-4 secretion by CD4+ T cells of β2-AR KO compared to WT (p<0.05). The secretion of IL-13, IL-5, and IL-10 but not IL-23 in CD4 T cells of stressed β2-AR KO mice was two-fold lower than that of stressed WT. TNF-α and IL-1β secretions in macrophages and DCs or IL-12 secretion in DCs was high in stressed β2-AR KO compared to WT (p<0.05). Overall, this study provides insights into the β2-AR signaling pathway’s role in suppressing the secretion of protective cytokines suggesting that inhibiting β2-AR signaling pathway may restore the function of immune cells during chlamydia genital infection. However, further study is warranted. This work was funded by NIH grant # 1R15AI124156-01 awarded to Bluefield State University

Roles of dendritic cells and macrophages during Chlamydia muridarum genital infection in stressed mice

It is well known that stress has a significant impact on public health and more evidence shows that stressors generally are greater in populations of lower socioeconomic status. Cold-induced stress inhibits immune cell activity and has been shown to cause different expressions of cytokine profiles during Chlamydia muridarum genital infection. Dendritic cells (DCs) and macrophages(MØ) are immune cells that express beta2-adrenergic receptor (β2-AR) and with key roles during chlamydia genital infection. This study is aimed to investigate the role of β2-AR by using a β2-AR knockout mouse. Wildtype (WT) and β2-AR knockout(KO) mice were stressed for five minutes daily and infected with C. muridarum intravaginally. Non-stressed infected mice of the groups were used as controls. Bone-marrow-derived DCs and MØ were tested isolated, counted differentiated, and proliferated for cytokine production. The proliferation of DCs and macrophages in the presence/absence of β2-AR agonists and antagonists was tested. Data show that non-stressed mice had a higher production of cytokines than stressed mice. Increased production of TNF-α in LPS-treated DCs and MØ of WT and β2-AR KO was observed. The effect of fenoterol and ICI118,55 antagonist showed no significant difference in cytokine production. The data indicate that β2-AR KO and WT had a similar pattern of cytokine production suggesting that deficiency in β2-AR restores the function of immune cells during genital infection. Experiments are undergoing to fully understand the mechanisms involved in modulating the function of DC and MØ during chlamydia genital infection.

Active Hexose Correlated Compound as a Countermeasure against Cold-induced Stress during Chlamydia muridarum Genital Infection in Mice.

Chronic stress is known to suppress the immune system, but the influence of stress on the pathogenesis of Chlamydia trachomatis genital infection and host immune response remains unexplored A stress mouse model has been developed in our lab and we have demonstrated that cold-induced stress (CIS) suppresses the immune system and subsequently leads to increased intensity of Chlamydia muridaum in mice. We have started feeding mice with Active hexose correlated compound (AHCC) that may restore the function of the immune system and result in reduced C. muridarum shedding from the genital tract. During the feeding of stressed mice with AHCC, dendritic cells (DCs) and macrophages bring the AHCC to T cells in the lymph node; however, the specific functions of DCs subsets in our stress model are not defined. We determined the production of cytokines by macrophages, CDs, and CD4+T in the AHCC-fed stress mice during C. muridarum genital infection. CD4+ T cells isolated from the spleen or lymph node of stressed-AHCC-fed, PBS-stressed, or non-stressed mice proliferated, and cytokine secretion was measured by ELISA. AHCC-feeding led to lower production of IL-5, IL-13, IL-10 and higher IL-12 (P<0.01) and IFN-

Production of CD4+ T Cell Subsets in a Beta2-Adrenergic Receptor Knockout Stress Mouse Model during Chlamydia muridarum Genital Infection

Kaitlyn Cook*, Tesfaye Belay, Ph.D*., *Department of Applied Science and Mathematics, Bluefield State College, Bluefield WV 24701 Production of CD4+ T Cell Subsets in a Beta2-Adrenergic Receptor Knockout Stress Mouse Model during Chlamydia muridarum Genital Infection
 Chlamydia is the most widely spread sexually transmitted disease, with more than one hundred million new cases occurring every year worldwide. Studies show the effect of stress promoting infection, but little is known about Chlamydia and the impact of stress. Cold-induced stress was used to simulate stress in mice to understand the relationship between the stress hormone norepinephrine (NE) and the immune system. This study aims to determine the role of beta2-adrenergic receptor (β2-AR) under stressful conditions during infection. We hypothesize the β2-AR suppresses T helper cell 1 (Th1) and promotes the production of Th2 cytokines. Following stressing, infection, sacrificing, and dissection of mice, the spleen, lymph node, and genital tract cells were proliferated for 72 hours. ELISA data for IL-13 concentration shows stressed knockout (KO) mice had 273.6pg/mL compared to stressed wildtype (WT) mice with 120.6pg/mL. Various treatment of CD4+ T cells resulted in production of TNF- α, ranging from 595pg/mL to 398.7pg/mL. Viability of CD4+ T cells treated with β2-AR agonist Fenoterol, and antagonist ICI 118,551 were analyzed. The viability of CD4+ T cells treated with NE was 70.7% in KO, compared to 69.1% and 70.2% for the agonist and antagonist, respectively. NE, Fenoterol, and ICI 118,551 treatment resulted in 72.2%, 83.6% and 84.4% respectively for WT. No significant difference is found between the agonist and antagonist. Chlamydia muridarum isolation from the genital tract of treatment groups is underway. The ELISA data implies that β2-AR mice restored increased production of protective cytokines more than their WT counterparts.

Profile of CD8+ T-cells in a Stressed beta2- Adrenergic Receptor Knockout Mouse Model during Chlamydia muridarum Genital Infection.


 Chlamydia trachomatis is the most common bacterial sexually transmitted disease. A negative correlation between the immune system and stress has been reported. We have demonstrated that cold-induced stress (CIS) increases the intensity of Chlamydia muridarum genital infection in a mouse model, but the status of CD8+ T cells is still unknown. This study profiles CD8+ T cell’s interaction with stress hormone; norepinephrine (NE) and the beta2-adrenergic receptor (β2-AR) in the presence of agonists and antagonists; and examines cytokines produced in the β2-AR knockout (KO) stress mouse model during C. muridarum genital infection. We hypothesize that CD8+ T cells produce high-level IFN-g and low-level TNF-a in stressed β2-AR KO mice than C57BL/6J wildtype (WT). Two groups of mice were cold-water stressed for 21 days, infected with C. muridarum, and then sacrificed for CD8+ T cell isolation. Viability was tested. The proliferation of CD8+ T cells was carried out for 72 h and culture supernatant was collected for cytokine production using ELISA. To confirm genital tract infection, Chlamydia muridarum isolation is underway. Our data show that the viability of CD8+ T cells treated with NE was significantly reduced compared to cells treated with LPS or Con A, indicating the suppressive effect of NE on immune cells. Stressed β2-AR KO mice displayed increased production of protective cytokines compared to stressed WT mice, indicating the absence of β2-AR leads to the functional restoration of protective immune cells.

Active Hexose-Correlated Compound Restores Gene Expression and Protein Secretion of Protective Cytokines of Immune Cells in a Murine Stress Model during Chlamydia muridarum Genital Infection.

Chlamydia trachomatis genital infection is the most common bacterial sexually transmitted disease worldwide. Previously, we reported that cold-induced stress results in immune suppression of mice that subsequently leads to increased intensity of Chlamydia muridarum genital infection. Furthermore, we demonstrated that stressed mice orally fed with active hexose-correlated compound (AHCC) have reduced shedding of C. muridarum from the genital tract. However, the mechanism of AHCC in reducing the organ load and changing the immune response in the stress model is not well known. This study evaluated infection and changes in immunological parameters of stressed AHCC-fed mice with or without C. muridarum genital infection. We hypothesized that AHCC feeding to stressed mice restores protective immune function and reduces susceptibility to C. muridarum genital infection. The results show that oral feeding of stressed mice with AHCC resulted in decreased shedding of C. muridarum from the genital tract, reduced production of plasma catecholamines, increased expression of T-bet and reduced GATA-3 in CD4+ T cells, increased production of interleukin-12 (IL-12) and interferon gamma (IFN-γ) and reduced production of IL-4 in CD4+ T cells, and enhanced expression of surface markers and costimulatory molecules of CD4+ T cells, bone marrow-derived dendritic cells (BMDCs), and natural killer cells. Coculturing of mature BMDCs with splenic CD4+ T cells led to the increased and decreased production of T helper 1 and T helper 2 cytokines, respectively. Overall, our results show that AHCC fosters the restoration of Th1 cytokine production while reducing Th2 cytokine production, which would promote C. muridarum clearance in the murine stress model.

Decreased Chlamydia muridarum genital infection and improved immune response in a β2-Adrenergic Receptor Knockout Mouse Stress Model

Chlamydia genital infection caused by Chlamydia trachomatis shows a high prevalence in low socioeconomic populations; however, whether stress is associated with the high prevalence of infection is not known. During chronic stress conditions, the interaction of beta2-adrenergic receptor (β2-AR) with norepinephrine (NE) is known to suppress the immune system in humans and animal models. However, the mechanism of suppression that leads to chlamydia infection is not well known. This study aims to measure the susceptibility of mice and cytokine production of a β2-AR knockout (KO) during Chlamydia muridarum genital infection. Stressed and non-stressed C57BL/6J wildtype (WT) and β2-AR gene KO mice will be infected with C. muridarum intravaginally post 21-day stressing period. Swabbing of mice at the 3-day interval for 42 days will be performed to determine C. muridarum counts and isolation. Moreover, splenic CD4+ T cells, bone marrow-derived dendritic cells at day 7 post-infection will be performed for immune response analysis by ELISA. We hypothesize that the deficiency of β2-AR leads to significantly less C. muridarum shedding from the genital tract of stressed KO and leads to an increase and decrease in the production of protective and suppressive cytokines, respectively. We anticipate results obtained will align with our central hypothesis that stressed WT mice have increased C. muridarum shedding of genital tracts and increased production of protective cytokines in stressed β2-AR KO, suggesting the importance of stress in β2-AR stimulation.

Modulation of T helper 1 and T helper 2 immune balance in a murine stress model during Chlamydia muridarum genital infection.

A murine model to study the effect of cold-induced stress (CIS) on Chlamydia muridarum genital infection and immune response has been developed in our laboratory. Previous results in the lab show that CIS increases the intensity of chlamydia genital infection, but little is known about the effects and mechanisms of CIS on the differentiation and activities of CD4+ T cell subpopulations and bone marrow-derived dendritic cells (BMDCs). The factors that regulate the production of T helper 1 (Th1) or T helper 2 (Th2) cytokines are not well defined. In this study, we examined whether CIS modulates the expressions of beta-adrenergic receptor (β-AR), transcription factors, hallmark cytokines of Th1 and Th2, and differentiation of BMDCs during C. muridarum genital infection in the murine model. Our results show that the mRNA level of the beta2-adrenergic receptor (β2-AR) compared to β1-AR and β3-AR was high in the mixed populations of CD4+ T cells and BMDCs. Furthermore, we observed decreased expression of T-bet, low level of Interferon-gamma (IFN-γ) production, increased expression of GATA-3, and Interleukin-4 (IL-4) production in CD4+ T cells of stressed mice. Exposure of BMDCs to Fenoterol, β2-AR agonist, or ICI118,551, β2-AR antagonist, revealed significant β2-AR stimulation or inhibition, respectively, in stressed mice. Moreover, co-culturing of mature BMDCs and naïve CD4+ T cells increased the production of IL-4, IL-10, L-17, and IL-23 cytokines, suggesting that stimulation of β2-AR leads to the increased production of Th2 cytokines. Overall, our results show for the first time that CIS promotes the switching from a Th1 to Th2 cytokine environment. This was evidenced in the murine stress model by the overexpression of GATA-3 concurrent with elevated IL-4 production, reduced T-bet expression, and IFN-γ secretion.

Outcome of Chlamydia muridarum genital infection and immune response in a beta2-adrenergic receptor knockout stress mouse model

Abstract The beta-2-adrenergic receptor (β2-AR) is one of the β-AR subsets found in most immune cells and is stimulated by norepinephrine (NE) binding. We showed previously that exposure of a mouse model to cold water stress leads to high level production of NE that may result in increased intensity of Chlamydia muridarum genital infection but the mechanism is not well defined. The purpose of this study was to develop and characterize a β2-AR knockout (KO) stress mouse model to investigate its susceptibility to C. muridarum genital infection. We hypothesized that lacking the β2-AR leads to a decreased C. muridarum shedding from the genital tract of stressed mice compared to that of stressed wild type (WT) mice. Stressed and non-stressed C57BL/6J WT and β2-AR KO mice were infected intravaginally with C. muridarum. Swabbing at 3-day intervals during primary infection and C. muridarum isolation was performed by culturing in McCoy tissue culture following standard methods. β2-AR KO stressed mice had a mean of 8540 inclusion forming units/ml (IFU/ml) compared to a mean of 18000 IFU/ml of stressed wild type at day 9 post infection. The overall course of primary infection in the β2-AR KO mouse model showed slightly reduced Chlamydia shedding. In addition, transfer of CD4+ T cells from black J6 WT to β2-AR KO resulted in a significant increase of shedding compared to that of non-stressed WT transfer (6000 versus 1000 IFU/ml). Secretion of interferon gamma was restored in β2-AR KO mice compared to WT. Overall, these results imply that deficiency in β2-AR leads to decreased C. muridarum shedding compared to the WT. However, other members of the alpha- and beta-adrenergic receptor family may compensate for the absence in β2-AR KO mice thus more investigation is needed.

Gene Expression of Transcription factors and Secretion of Cytokines in Co-culturing of Dendritic Cells and Naïve T cells in Stressed AHCC-fed Mice Courtney Rolen1, Elisha Martin1, Rajnish Sahu2Tesfaye Belay1, 1Bluefield State College, Bluefield, WV; 2Alabama State University, Montgomery, AL.

Cold-induced stress is known to inhibit the activity of immune cells and has been shown to cause different expression of cytokine profiles during Chlamydia muridarum genital infection. Active Hexose Correlated Compound (AHCC) is a mushroom extract known to modulate the immune system, the mechanism is not well known. The purpose of this study was to investigate if AHCC feeding to stressed mice would affect the pattern of surface marker expression of transcription factors and CD4+ T cell cytokines. We hypothesized that AHCC feeding to stressed mice decreases the expression and secretion of GATA-3 and IL-4 in stressed mice compared to PBS-fed stressed mice. We used magnet enrichment protocols to isolate DCs from bone marrow and naïve T cells from the spleen. Naive T cells and matured DCs were co-cultured for 24 and 48 hours. A quantitative RT-PCR and ELISA were used to determine gene expression and secretion of cytokines, respectively. Our data show downregulation of transcription factor GATA-3 in AHCC-fed mice (-0.45 ) and upregulation of GATA-3 in PBS-fed stressed mice (+1.5). In contrast, transcription factors T-bet was upregulated in stress AHCC-fed (0.9) and non-stressed (+2.0) during infection. Moreover, non-stressed (+2.9) and AHCC-fed (+1.85) mice showed a significant upregulation of T-bet during Chlamydia muridarum genital infection. Increased secretion of IL-12 and IFN-g and decreased secretion of IL-4 and IL-23 was observed in AHCC-fed mice compared to PBS-fed mice. Overall, our results show that AHCC-feeding leads to the restoration of the function of the protective immune system against chlamydia genital infection. (Supported by NIH Grant P20GM103434 to the West Virginia IDeA Network for Biomedical Research Excellence and NIH Grant P20GM103434 awarded to Bluefield State College).

Gene expression profiles T helper cell chemokines, cytokines and transcription factors in the genital tract of a stress mouse model during Chlamydia muridarum infection

A stress mouse model established in our laboratory showed that prolonged exposure of mice to cold water-induced stress results in elevated plasma norepinephrine levels, increased intensity of Chlamydia muridarum genital infection, and decreased functions of the immune system. The current study was designed to test whether Active Hexose Correlated Compound (AHCC) modulates the gene expression profiles of chemokines associated with either Thelper 1 (Th1) or Th2 in the stress model. We hypothesized that oral administration of AHCC feeding up-regulates the gene expression profiles of Th1 chemokines and Th1 transcription factors and increases the secretion of cytokines in Th cells and lysates of the genital tract. During the stressing period of 21 days, we mice either 300 mg/kg of AHCC or PBS per day using a gavage feeding technique. We used real PCR to determine gene expression of chemokines favoring Th1 or Th2 chemokines and cytokines. We used ELISA to assess cytokine production in the genital tract lysates. Oral administration of AHCC to stressed mice resulted in the up-regulation of Th1 chemokines. The level of mRNA fold-changes of CXCL10 (+2.2 fold), CXCL9 (+1.8 fold), and CCL5 (+2.fold) was observed. Moreover, AHCC feeding resulted in a significant increase in productions of interleukin-1b, and interleukin 10(IL-10) in the genital tract of stressed mice. However, the production of TNF-a and IL-6 were undetectable by ELISA. Although further study is needed, preliminary data suggest that AHCC feeding favors the production of protective cytokines in our stress mouse model. (Supported by NIH Grant P20GM103434 to the West Virginia IDeA Network for Biomedical Research Excellence and NIH Grant P20GM103434 awarded to Bluefield State College).

Chlamydia-infected macrophages are resistant to azithromycin treatment and are associated with chronic oviduct inflammation and hydrosalpinx development.

Chlamydia infection remains the leading sexually-transmitted bacterial infection worldwide, causing damaging sequelae such as tubal scarring, infertility and ectopic pregnancy. As infection is often asymptomatic, prevention via vaccination is the optimal strategy for disease control. Vaccination strategies aimed at preventing bacterial infection have shown some promise, although these strategies often fail to prevent damaging inflammatory pathology when Chlamydia is encountered. Using a murine model of Chlamydia muridarum genital infection, we employed two established independent models to compare immune responses underpinning pathologic development of genital Chlamydia infection. Model one uses antibiotic treatment during infection, with only early treatment preventing pathology. Model two uses a plasmid-cured variant strain of C. muridarum that does not cause pathologic outcomes like the plasmid-containing wild-type counterpart. Using these infection models, contrasted by the development of pathology, we identified an unexpected role for macrophages. We observed that mice showing signs of pathology had greater numbers of activated macrophages present in the oviducts. This may have been due to early differences in macrophage activation and proinflammatory signaling leading to persistent or enhanced infection. These results provide valuable insight into the cellular mechanisms driving pathology in Chlamydia infection and contribute to the design and development of more effective vaccine strategies for protection against the deleterious sequelae of Chlamydia infection of the female reproductive tract.

Cold-induced stress alters the dynamics of T helper 1 and T helper 2 cytokine production in a mouse model during Chlamydia muridarum genital infection.

Chlamydia caused by Chlamydia trachomatis is the leading cause of bacterial STDs worldwide and the USA. Although there are more clinical and epidemiological studies of chlamydia genital infection, the effect of stress on chlamydia disease is not well explored. Previous results from our lab show that cold-induced stress results in increased intensity of chlamydia genital infection in mice; however, the mechanisms are unknown. Thelper 1(Th1) is known to be protective against chlamydia, whereas Th2 leads to infection. However, the differentiation of Th1 and Th2 during exposure to cold stress is not well understood. Several studies show GATA-3 and T-bet are associated with Th1 and Th2 cytokine production. The purpose of this study was to evaluate the effect of cold water on the activity off Th1 and Th2 during Chlamydia muridarum genital infection. We hypothesized that cold stress alters the differentiation and variation in gene expression profile of transcription factors and cytokine productions in T cells. T cells isolated from the genital tract were proliferated for 48 h incubated at 37OC and 5% CO2. The mRNA levels and detection of cytokines in culture supernatants was determined by quantitative real time PCR and ELISA, respectively. Our results showed increased IL-4 production, which was associated with upregulated expression of transcription factor GATA-3. In contrast, decreased gamma interferon production was associated with decreased T-bet gene expression in T cells. Based on this data, cold stress modulates the dynamics of signature Th1 and Th2 cytokine productions that are regulated by expression of GATA-3 and T-bet. This work was supported by NIH Grant P20GM103434 to the West Virginia IDeA Network for Biomedical Research Excellence and NIH Grant P20GM103434 awarded to Bluefield State College.

. Deficiency in beta2 adrenergic receptor in mice leads to decreased Chlamydia muridarum shedding in genital tract.

Chlamydia caused by Chlamydia trachomatis is the most common bacterial STD that causes pelvic inflammatory disease and infertility, especially in childbearing women. We have shown that stressing mice in cold water for 5 minutes daily for 21 days leads to increased production of stress hormones. It is known that beta2 adrenergic receptor (b2-AR) as a ligand to norepinephrine hormone is shown to regulate immune function in mice. However, the effect and mechanism of b2-AR on chlamydia infection is not well known. The purpose of the study was to examine the susceptibility of a b2-AR knock out model to Chlamydia muridarum genital infection. We hypothesized that deficiency in b2-AR leads i) a decreased C. muridarum shedding from the genital tract of stressed mice; ii) increased gene expression of protective cytokines in the genital tract during infection. Mouse cervico-vaginal swabbing at 3-day intervals for 28 days was performed. C. muridarum was isolated from genital swabs in McCoy tissue culture according to standard methods, and inclusion bodies were visualized and counted after staining with fluorescein isothiocyanate-labeled, anti-chlamydial antibody. Quantitative real time PCR and ELISA methods, respectively, were used to measure cytokine mRNA expressions and secretions from genital tract lysates. Results showed no significance differences between stressed and non-stressed of IFU/ml. Moreover, no significant gene expression of b-AR subsets in stressed and non-stressed mice was observed. Interestingly, up-regulation of interferon gamma was observed in stressed compared to non-stressed suggesting that mice lacking b2 adrenergic receptor have reverted to protective cytokine production to decrease infection. This work was supported by NIH Grant P20GM103434 to the West Virginia IDeA Network for Biomedical Research Excellence and NIH Grant P20GM103434 awarded to Bluefield State College.

Cold-induced stress modulates the maturation of immune cells of mice during Chlamydia muridarum genital infection

Abstract Chlamydia genital infection caused by Chlamydia trachomatis is the most common bacterial sexually transmitted disease (STD) in the United States. Results in a cold-induced stress mouse model established in our laboratory show that cold-induced stress increases the intensity of chlamydia genital infection. This study was to survey the production of T helper cell type 1 (Th1) and (Th2) cytokines in the stress mouse model. Cells harvested from bone marrow or T cells from spleens of stressed and non-stressed mice during Chlamydia muridarum genital infection were allowed to proliferate in vitro. Cytokine production in culture supernatants was tested by ELISA. Gene expression of GATA 3, T-bet and INF-γ in splenic T cells were determined by QPCR. Production of IL-12 by dendritic cells of stressed mice was 0μg compared to 41.06 μg in non stressed mice. However, C. muridarum infection induced increased IL-12 production (97.27 μg and 123 μg) in stressed and non-stressed mice, respectively. Stress induced a 2-fold decrease in T-bet gene expression in T-cells compared to non-stressed group. In contrast, gene expression of GATA 3 in T cells of stressed mice was significantly increased compared to all other groups tested. INF-γ gene expression in T cells of stressed mice showed a 4-fold decrease compared to T cells of non stressed mice. Stress resulted in decreased IL-12 production in dendritic cells and macrophages, INF-γ production and gene expression of T-bet by Th1 cells. Furthermore, the increased gene expression of GATA 3 only in stressed chlamydia infected T cell suggests that cold-induced stress leads to a skewed immune response toward marked increase in production of Th2 cytokines that favor immunosuppression during chlamydia genital infection.

Effect of cold water-induced stress on immune response, pathology and fertility in mice during Chlamydia muridarum genital infection.

Genital infection by Chlamydia trachomatis is the most common bacterial sexually transmitted disease worldwide. It causes serious reproductive health complications, including pelvic inflammatory disease and infertility. Stress is implicated as a risk factor for various infections; however, its effect on chlamydia genital infection is unknown. We previously showed that repeated exposure of mice to cold water results in increased severity of chlamydia genital infection. In this study, cold water-induced stress resulted in (i) elevated levels of norepinephrine (NE) and epinephrine in the spleen and genital tract of stressed mice; (ii) elevated IL-1β, TNF-α, IL-6 and nitric oxide production in macrophage-rich peritoneal cells of mice; (iii) supplement of NE in vitro exerts an immunosuppressive effect on splenic T-cell production of cytokines; (iv) decreased C. muridarum shedding in the genital tract of β1Adr/β2Adr receptor KO mice; and (v) a higher rate of infertility in infected mice. These results suggest that cold water stress induces the production of catecholamines, which may play a critical role in the modulation of the immune system leading to increased intensity of C. muridarum genital infection.

IFNγ is Required for Optimal Antibody-Mediated Immunity against Genital Chlamydia Infection.

Defining the mechanisms of immunity conferred by the combination of antibody and CD4+ T cells is fundamental to designing an efficacious chlamydial vaccine. Using the Chlamydia muridarum genital infection model of mice, which replicates many features of human C. trachomatis infection and avoids the characteristic low virulence of C. trachomatis in the mouse, we previously demonstrated a significant role for antibody in immunity to chlamydial infection. We found that antibody alone was not protective. Instead, protection appeared to be conferred through an undefined antibody-cell interaction. Using gene knockout mice and in vivo cellular depletion methods, our data suggest that antibody-mediated protection is dependent on the activation of an effector cell population in genital tract tissues by CD4+ T cells. Furthermore, the CD4+ T cell-secreted cytokine gamma interferon (IFN-γ) was found to be a key component of the protective antibody response. The protective function of IFN-γ was not related to the immunoglobulin class or to the magnitude of the Chlamydia-specific antibody response or to recruitment of an effector cell population to genital tract tissue. Rather, IFN-γ appears to be necessary for activation of the effector cell population that functions in antibody-mediated chlamydial immunity. Our results confirm the central role of antibody in immunity to chlamydia reinfection and demonstrate a key function for IFN-γ in antibody-mediated protection.

Interferon-gamma and Effector Cell Activation are Required for Antibody-Mediated Immunity against Genital Chlamydia Infection

Abstract Defining the mechanisms of immunity conferred by the combination of antibody and CD4+ T cells is fundamental to designing an efficacious chlamydial vaccine. Using the Chlamydia muridarum genital infection model of mice, which replicates many features of human C. trachomatis infection and avoids the characteristic low virulence of C. trachomatis in the mouse, we previously demonstrated a central role for antibody in immunity to chlamydial infection. We found that antibody alone was not protective. Instead, protection appeared to be conferred through an undefined antibody-cellular interaction. Using gene knockout mice and in vivo cellular depletion methods, antibody-mediated protection was shown to be solely dependent on the activation of an effector cell population in genital tract tissues by CD4+ T cells. Furthermore, the CD4+ T cell-secreted cytokine interferon-gamma (IFNγ) was found to be a key component of the protective antibody response. The function of IFNγ was not related to the immunoglobulin class or magnitude of the Chlamydia-specific antibody response or to recruitment of an effector cell population to genital tract tissue. Rather, IFNγ appears to be necessary for activation of the effector cell population that functions in antibody-mediated chlamydial immunity. Ongoing studies aim to identify the effector cell population(s) involved in the antibody-mediated protective response. Our results confirm the central role of antibody and CD4+ T cells in immunity to chlamydia reinfection, and demonstrate a key function for IFNγ in antibody-mediated protection.