Expression of HPV-16 E6 and E7 oncoproteins alters Chlamydia trachomatis developmental cycle and induces increased levels of immune regulatory molecules.

Introduction: Infection with Human Papillomavirus (HPV) is a recognized risk factor for Chlamydia trachomatis (CT) infection and vice versa. Coinfection of HPV and CT in women is a very common and usually asymptomatic finding that has been linked to increased risk of cervical cancer. It has been demonstrated that CT facilitates the entry of multiple high risk HPV genotypes, leading to damage of the mucosal barrier and interfering with immune responses and viral clearance, which ultimately favours viral persistence and malignant transformation. Although the facilitating effects elicited by CT infection on viral persistence have been reported, little is known about the consequences of HPV infection on CT development. Methods: Herein, we took advantage of a genetically modified human cervical cell line co-expressing HPV-16 major oncogenic proteins E6 and E7, as an experimental model allowing to investigate the possible effects that HPV infection would have on CT development. Results and discussion: Our results show that CT infection of HPV-16 E6E7 expressing cells induced an upregulation of the expression of E6E7 oncoproteins and host cell inhibitory molecules PD-L1, HVEM and CD160. Additionally, smaller chlamydial inclusions and reduced infectious progeny generation was observed in E6E7 cells. Ultrastructural analysis showed that expression of E6 and E7 did not alter total bacterial counts within inclusions but resulted in increased numbers of reticulate bodies (RB) and decreased production of infectious elementary bodies (EB). Our results indicate that during CT and HPV coinfection, E6 and E7 oncoproteins impair RB to EB transition and infectious progeny generation. On the other hand, higher expression of immune inhibitory molecules and HPV-16 E6E7 are cooperatively enhanced in CT-infected cells, which would favour both oncogenesis and immunosuppression. Our findings pose important implications for clinical management of patients with HPV and CT coinfection, suggesting that screening for the mutual infection could represent an opportunity to intervene and prevent severe reproductive health outcomes, such as cervical cancer and infertility.

Ureaplasma urealyticum and Mycoplasma hominis urogenital infections associate with semen inflammation and decreased sperm quality.

Ureaplasma urealyticum and Mycoplasma hominis are among the most prevalent sexually transmitted infections proposed to induce urogenital inflammation and impair sperm quality. However, the topic remains controversial since contradictory findings have been reported. Herein, we performed a comprehensive analysis of U. urealyticum and M. hominis urogenital infections and their association with urogenital inflammation (i.e., leukocyte subsets and inflammatory cytokines in semen,) and sperm quality parameters in a cohort of men with couple's primary infertility undergoing initial infertility evaluation or with lower urinary tract symptoms and no infertility-related complaints. Overall, U. urealyticum and M. hominis infection was detected in 17.0% and 23.6% of patients, respectively, whereas the coinfection was detected in 3.8% of patients only. Remarkably, similar infection frequencies were found in the different patient subpopulations analyzed. Moreover, infections were associated with elevated semen levels of TNF, IL-1ß, and IL-6 and/or increased counts of total leukocytes and their subsets, including CD4 and CD8 T lymphocytes and neutrophils. In addition, M. hominis infection and the coinfection with U. urealyticum were associated with impairments in sperm quality variables. Our results indicate that U. urealyticum and M. hominis male urogenital infections induce urogenital inflammation and decrease sperm quality, thus impairing male fertility potential. Screening for U. urealyticum and M. hominis infections and performing a comprehensive analysis of different leukocyte subsets and inflammatory cytokines in semen may be clinically helpful in the diagnosis and follow-up of male urogenital infection.

Immune Alterations in a Patient With Hyperornithinemia-Hyperammonemia-Homocitrullinuria Syndrome: A Case Report.

The hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome is a rare autosomal recessive inborn error of the urea cycle caused by mutations in the SLC25A15 gene. Besides the well-known metabolic complications, patients often present intercurrent infections associated with acute hyperammonemia and metabolic decompensation. However, it is currently unknown whether intercurrent infections are associated with immunological alterations besides the known metabolic imbalances. Herein, we describe the case of a 3-years-old girl affected by the HHH syndrome caused by two novel SLC25A15 gene mutations associated with immune phenotypic and functional alterations. She was admitted to the hospital with an episode of recurrent otitis, somnolence, confusion, and lethargy. Laboratory tests revealed severe hyperammonemia, elevated serum levels of liver transaminases, hemostasis alterations, hyperglutaminemia and strikingly increased orotic aciduria. Noteworthy, serum protein electrophoresis showed a reduction in the gamma globulin fraction. Direct sequencing of the SLC25A15 gene revealed two heterozygous non-conservative substitutions in the exon 5: c.649G>A (p.Gly217Arg) and c.706A>G (p.Arg236Gly). In silico analysis indicated that both mutations significantly impair protein structure and function and are consistent with the patient clinical status confirming the diagnosis of HHH syndrome. In addition, the immune analysis revealed reduced levels of serum IgG and striking phenotypic and functional alterations in the T and B cell immune compartments. Our study has identified two non-previously described mutations in the SLC25A15 gene underlying the HHH syndrome. Moreover, we are reporting for the first time functional and phenotypic immunologic alterations in this rare inborn error of metabolism that would render the patient immunocompromised and might be related to the high frequency of intercurrent infections observed in patients bearing urea cycle disorders. Our results point out the importance of a comprehensive analysis to gain further insights into the underlying pathophysiology of the disease that would allow better patient care and quality of life.

Association between Human Papillomavirus and Chlamydia trachomatis genital infections in male partners of infertile couples.

The prevalence of HPV infection and its relationship with other sexually transmitted infections was analyzed in a cohort of 117 male partners of infertile couples from Cordoba, Argentina. Semen samples and urethral swabs were obtained and the infection with HPV, Chlamydia trachomatis, HSV1, HSV2, Mycoplasma hominis and Ureaplasma urealyticum was analyzed. A prevalence of HPV infection of 27.4% was found. Interestingly, infections by exclusively low risk HPV genotypes or high/intermediate risk HPV genotypes were present in 64.5% and 22.6% of cases, respectively. Low risk-HPV6 was the most frequently detected genotype. Remarkably, HPV and C. trachomatis infections were significantly associated to each other (OR: 11.55, 95% CI 1.14-117.06). No significant differences in sperm quality were found between HPV-positive and HPV-negative patients indicating that HPV male urogenital infection does not impair sperm quality. Our results show a high prevalence of HPV urogenital infection among male partners of infertile couples, and that HPV and C. trachomatis infections are reciprocal risk factors of their co-infection. Moreover, our results suggest that men constitute a reservoir for continued transmission of C. trachomatis and HPV to women highlighting the need for routine screening for these two pathogens in male partners of infertile couples.

Patients with chronic prostatitis/chronic pelvic pain syndrome show T helper type 1 (Th1) and Th17 self-reactive immune responses specific to prostate and seminal antigens and diminished semen quality.

OBJECTIVES: To assess the presence of self-reactive immune responses to seminal and prostate antigens (PAg), biomarkers of inflammation of the male genital tract, and semen quality parameters in patients with chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS). PATIENTS, SUBJECTS AND METHODS: Peripheral blood and semen samples were collected from patients with CP/CPPS and age-matched healthy control volunteers. We analysed the lymphoproliferative responses of peripheral blood mononuclear cells (PBMC) to different seminal plasma (SP)-derived and purified PAg, serum autoantibodies specific to PAg, leucocyte subpopulations, and inflammatory cytokines in semen, sperm apoptosis/necrosis, and semen quality parameters. RESULTS: Significantly greater PBMC proliferative responses specific to PAg, with elevated secretion of interferon (IFN)γ and interleukin (IL)-17, were detected in the patients with CP/CPPS vs the controls. Moreover, the patients with CP/CPPS had significantly greater serum immunoglobulin G immune reactivity to SP proteins, such as prostate-specific antigen and prostatic acid phosphatase, than the controls. Inflammation of the male genital tract was exemplified by high levels of IFNγ, IL-17, IL-1ß and IL-8, as well as higher counts of leukocytes, mainly CD4 T lymphocytes and macrophages, in the semen. In addition, this local inflammation was associated with an overall diminished semen quality, i.e., reduced sperm concentration, motility and viability; and higher levels of sperm apoptosis/necrosis in patients with CP/CPPS vs controls. CONCLUSION: Patients with CP/CPPS show T helper type 1 (Th1) and Th17 immune responses specific to PAg associated with chronic inflammation of the male genital tract and reduced semen quality. These immune responses may underlie the induction and development of chronic pelvic pain and inflammation of the male genital tract, which in turn could alter normal prostate functioning and impair semen quality.

Differences in T regulatory cells between mouse strains frequently used in immunological research: Treg cell quantities and subpopulations in NOD, B6 and BALB/c mice.

Foxp3+ Regulatory T cells (Tregs) are essential for the maintenance of tolerance to self. Therefore, it is expected that lower numbers and/or less than optimal function could impact on the functioning of the immune system, and thereby contributing to the development of autoimmune diseases. In the present report, by comparing Tregs from most frequently used mouse strains in immunological research (C57BL/6 (B6), BALB/c and NOD), we provide evidence showing that the NOD mouse strain, highly predisposed to develop autoimmune responses, exhibit a generalized decreased in Tregs counts with enhanced proportions of CD44hiCD62Llow Tregs when compared with BALB/c mice. No major differences were observed in Helios+ or Helios- Tregs between strains. The expression of CXCR3, CCR5 and CCR6 on Tregs from all strains showed minor proportions of CXCR3+ and CCR5+ cells in NOD Tregs. Naïve CD4+CD25- T cells from NOD mice also showed decreased capacity to induce in vitro iTregs when compared with B6 and BALB/c mice. Lower expression of molecules involved in Treg suppressor mechanisms such as CD25, LAP-1, CD39 and PD-1 was observed both in NOD iTregs and Tregs from lymph nodes of NOD mice. Moreover, in vitro assays showed that Tregs from NOD mice exhibited reduced ability to suppress proliferation of CD4+CD25- responder T cells when compared with B6 and BALB/c mice. Major differences were consistently observed between NOD and BALB/c mice, whereas no major differences were found for many of the analyzed parameters between the NOD and B6 mice, suggesting that highly and mildly autoimmune prone mouse strains may share some Tregs features. On the contrary, BALB/c Tregs were in major quantities, expressed higher levels of Foxp3 and exhibited more potent ability to inhibit effector T cell proliferation, data that could be related to its natural resistance to the induction of different experimental autoimmune conditions. Altogether our results demonstrate a generalized Treg cell dysfunction in NOD mice, a strain characterized by its high predisposition to develop spontaneous and induced autoimmune diseases.

T Regulatory Cells From Non-obese Diabetic Mice Show Low Responsiveness to IL-2 Stimulation and Exhibit Differential Expression of Anergy-Related and Ubiquitination Factors.

Foxp3+ Regulatory T cells (Tregs) are pivotal for the maintenance of tolerance. Alterations in their number and/or function have been proposed to occur in the autoimmune-prone non-obese diabetic (NOD) mouse. Comparing the frequencies and absolute numbers of CD4+Foxp3+CD25+ Tregs among 4 to 6-week old NOD, B6, and BALB/c mice, we observed differences in counts and Foxp3 expression in Tregs from secondary lymphoid organs, but not in the thymus. Upon TCR and IL-2 stimulation, NOD Tregs showed lower responses than Tregs from B6 and BALB/c mice. Indeed, NOD Tregs responded with less proliferation and with smaller increments in the expression of CD25, LAP-1, CD39, PD-1, PD-L1, and LAG-3, when in vitro cultured for 3 days with anti-CD3/CD28 in the absence or presence of IL-2, Tregs from NOD mice showed to be highly dependent on IL-2 to maintain Foxp3 expression. Moreover, NOD Tregs become producers of IL-17 and INF-gamma more easily than Tregs from the other strains. In addition, NOD Tregs showed lower responsiveness to IL-2, with significantly reduced levels of pSTAT5, even at high IL-2 doses, with respect to B6 and BALB/c Tregs. Interestingly, NOD Tregs exhibit differences in the expression of SOCS3, GRAIL, and OTUB1 when compared with Tregs from B6 and BALB/c mice. Both, at steady state conditions and also after activation, Tregs from NOD mice showed increased levels of OTUB1 and low levels of GRAIL. In addition, NOD Tregs had differences in the expression of ubiquitin related molecules that play a role in the maintenance of Foxp3 cellular pools. Indeed, significantly higher STUB1/USP7 ratios were detected in NOD Tregs, both at basal conditions and after stimulation, compared to in B6 and BALB/c Tregs. Moreover, the addition of a proteasome inhibitor to cell cultures, conferred NOD Tregs the ability to retain Foxp3 expression. Herein, we provide evidence indicating a differential expression of SOCS3, GRAIL, and STUB1/USP7 in Tregs from NOD mice, factors known to be involved in IL-2R signaling and to affect Foxp3 stability. These findings add to the current knowledge of the immunobiology of Tregs and may be related to the known insufficiency of Tregs from NOD mice to maintain self-tolerance.

A Plasmonic Approach to Study Protein Interaction Kinetics through the Dimerization of Functionalized Ag Nanoparticles.

Understanding the kinetics of protein interactions plays a key role in biology with significant implications for the design of analytical methods for disease monitoring and diagnosis in medical care, research and industrial applications. Herein, we introduce a novel plasmonic approach to study the binding kinetics of protein-ligand interactions following the formation of silver nanoparticles (Ag NPs) dimers by UV-Vis spectroscopy that can be used as probes for antigen detection and quantification. To illustrate and test the method, the kinetics of the prototype biotin-streptavidin (Biot-STV) pair interaction was studied. Controlled aggregates (dimers) of STV functionalized Ag NPs were produced by adding stoichiometric quantities of gliadin-specific biotinylated antibodies (IgG-Biot). The dimerization kinetics was studied in a systematic way as a function of Ag NPs size and at different concentrations of IgG-Biot. The kinetics data have shown to be consistent with a complex reaction mechanism in which only the Ag NPs attached to the IgG-Biot located in a specific STV site are able to form dimers. These results help in elucidating a complex reaction mechanism involved in the dimerization kinetics of functionalized Ag NPs, which can serve as probes in surface plasmon resonance-based bioassays for the detection and quantification of different biomarkers or analytes of interest.

IL-10 Producing B Cells Dampen Protective T Cell Response and Allow Chlamydia muridarum Infection of the Male Genital Tract.

A significant proportion of individuals develop chronic, persistent and recurrent genital tract infections with Chlamydia trachomatis, which has been attributed to the numerous strategies that the bacterium uses to subvert host immune responses. Animal chlamydia models have demonstrated that protective immune response is mediated by CD4+ Th1 cytokine responses. Herein, we demonstrate that early after infecting the male genital tract, C. muridarum triggers the production of IL-10 by splenic and lymph node cells. In addition, C. muridarum triggers IL-6 and TNFα secretion. Data obtained from in vitro and in vivo experiments revealed B cells as the major IL-10 contributors. Indeed, purified B cells produced high amounts of IL-10 and also exhibited enhanced expression of inhibitory molecules such as CD39, PD-L1 and PD1 after C. muridarum stimulation. In vitro experiments performed with sorted cell subsets revealed that Marginal Zone B cells were the main IL-10 producers. In vitro and in vivo studies using TLR-deficient mice indicated that TLR4 signaling pathway was essential for IL-10 production. In addition, in vivo treatments to neutralize IL-10 or deplete B cells indicated that IL-10 and B cells played a significant role in delaying bacterial clearance ability. Moreover, the latter was confirmed by adoptive cell transfer experiments in which the absence of IL-10-producing B cells conferred the host a greater capability to induce Th1 responses and clear the infection. Interestingly, NOD mice, which were the least efficient in clearing the infection, presented much more Marginal Zone B counts and also enhanced TLR4 expression on Marginal Zone B cells when compared to B6 and BALB/c mice. Besides, treatment with antibodies that selectively deplete Marginal Zone B cells rendered mice more capable of inducing enhanced IFNγ responses and clearing the infection. Our findings suggest that B cells play a detrimental role in C. muridarum infection and that activation by innate receptors like TLR4 and IL-10 production by these cells could be used by Chlamydia spp. as a strategy to modulate the immune response establishing chronic infections in susceptible hosts.

Implications of prostate inflammation on male fertility.

The prostate is the seat of three major causes of morbidity: benign prostatic hyperplasia, prostate cancer and prostatitis, three conditions in which inflammation has been implicated. A state of inflammation of the prostate gland, originally incited by an infection, an autoimmune response, a neurogenic stimulus or another trigger may have consequences on prostate functionality. In fact, male fertility depends intrinsically on the content of prostatic fluid factors secreted by the prostatic epithelium. Taking into account that the prostate gland is the major male accessory gland that exerts essential functions for male fertility, a state of local inflammation can alter male fertility by either directly impairing sperm quality or, indirectly, by causing prostate dysfunction. In the present review, we summarise the current knowledge regarding prostatitis due to well-known infections such as Escherichia coli, Chlamydia trachomatis and other commonly identified microorganisms focusing on inflammatory markers detected during these infections and seminal quality and male fertility alterations reported. We also focused on type III prostatitis or chronic nonbacterial prostatitis/chronic pelvic pain syndrome, of unknown aetiology, in which inflammation of an autoimmune origin, neurogenic stimuli or another trigger have been proposed and fertility alterations reported.

Enzyme-Free Immunoassay Using Silver Nanoparticles for Detection of Gliadin at Ultralow Concentrations.

Determination of biomarkers in clinical or food samples is of crucial importance for monitoring, prevention, and care of public health. The standard procedure used for this purpose is the enzyme-linked immunosorbent assay (ELISA), which makes use of the specific antibody-antigen biorecognition and the catalytic effect of the enzymes. One of the main shortcomings of this technique is the use of enzymes that often present low chemical and thermal stabilities compared to other chemicals. Other drawbacks include the nonspecific binding process that could lead to false-positive results, the use of relatively large amounts of the sample, and the number of time-consuming steps involved. Recently, an enzyme-free and ultrasensitive analytical method for antigen detection denoted as intensity depletion immunolinked assay (IDILA) has been proposed by our laboratory. The assay is based on the inhibition to form Ag nanosphere dimers linked by a specific antibody in the presence of the corresponding antigen. In this work, we go a step further demonstrating how the performance of this method could be improved by using silver nanoparticles (Ag NPs) of different diameters (58 and 78 nm). The experiments are performed for detecting gliadin, an antigen of utmost importance in celiac disease, and the results are compared with ELISA, the standard technique homologated by the Food Codex Alimentarius. It is found that the IDILA assay could be around 1000 or 10 000 times more sensitive than ELISA, also having lower limits of detection, depending on the conditions explored (fraction of dimers and Ag NP diameter). Using the appropriate conditions, the IDILA assay is shown to be able to detect femtomolar concentrations of the antigen, besides being robust, reliable, cheap, rapid (around 2 h), and of easy implementation using the standard equipment and biomolecular reagents used for the ELISA assay.

Erratum: Chlamydia trachomatis neither exerts deleterious effects on spermatozoa nor impairs male fertility.

A correction to this article has been published and is linked from the HTML version of this paper. The error has not been fixed in the paper.

Immunological Mechanisms Underlying Chronic Pelvic Pain and Prostate Inflammation in Chronic Pelvic Pain Syndrome.

Chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) is the most common urologic morbidity in men younger than 50 years and is characterized by a diverse range of pain and inflammatory symptoms, both in type and severity, that involve the region of the pelvis, perineum, scrotum, rectum, testes, penis, and lower back. In most patients, pain is accompanied by inflammation in the absence of an invading infectious agent. Since CP/CPPS etiology is still not well established, available therapeutic options for patients are far from satisfactory for either physicians or patients. During the past two decades, chronic inflammation has been deeply explored as the cause of CP/CPPS. In this review article, we summarize the current knowledge regarding immunological mechanisms underlying chronic pelvic pain and prostate inflammation in CP/CPPS. Cumulative evidence obtained from both human disease and animal models indicate that several factors may trigger chronic inflammation in the form of autoimmunity against prostate, fostering chronic prostate recruitment of Th1 cells, and different other leukocytes, including mast cells, which might be the main actors in the consequent development of chronic pelvic pain. Thus, the local inflammatory milieu and the secretion of inflammatory mediators may induce neural sensitization leading to chronic pelvic pain development. Although scientific advances are encouraging, additional studies are urgently needed to establish the relationship between prostatitis development, mast cell recruitment to the prostate, and the precise mechanisms by which they would induce pelvic pain.

Chlamydia trachomatis neither exerts deleterious effects on spermatozoa nor impairs male fertility.

Chlamydia trachomatis is the most prevalent sexually transmitted bacterial infection. However, whether Chlamydia trachomatis has a negative impact on sperm quality and male fertility is still controversial. Herein, we report the effects on sperm quality of the in vitro exposure of spermatozoa to Chlamydia trachomatis, and also the effects of male genital infection on male fertility using an animal model. Human and mouse sperm were obtained from healthy donors and cauda epididimys from C57BL/6 mice, respectively. Highly motile human or mouse spermatozoa were in vitro exposed to C. trachomatis (serovar E or LGV) or C. muridarum, respectively. Then, sperm quality parameters were analyzed. Moreover, male fertility of Chlamydia muridarum infected male C57BL/6 mice was assessed. Human or murine sperm in vitro exposed to increasing bacterial concentrations or soluble factors from C. trachomatis or C. muridarum, respectively, did not show differences in sperm motility and viability, apoptosis, mitochondrial membrane potential, DNA fragmentation, ROS production and lipid peroxidation levels, when compared with control sperm (p > 0.05). Moreover, no differences in fertility parameters (potency, fecundity, fertility index, pre- and post-implantation loss) were observed between control and infected males. In conclusion, our results indicate that Chlamydia spp. neither directly exerts deleterious effects on spermatozoa nor impairs male fertility.

Chronic Infection of the Prostate by Chlamydia muridarum Is Accompanied by Local Inflammation and Pelvic Pain Development.

BACKGROUND: Chlamydia trachomatis urogenital infections are the leading cause of sexually transmitted bacterial infections. Although the prevalence of chlamydial infection is similar in men and women, current research is mainly focused on women, neglecting the study of male genital tract infections. We, therefore, investigated Chlamydia infection in the rodent male genital tract. MATERIALS AND METHODS: Male NOD and C57BL/6 mice were inoculated in the meatus urethra with C. muridarum. Bacterial DNA, leukocyte infiltration of male genital tract tissues, pelvic pain, and Chlamydia-specific immune responses were analyzed at different time points. RESULTS AND CONCLUSIONS: The inoculation of C. muridarum in the meatus urethra of male mice resulted in an ascending and widely disseminated infection of the male genital tract. C. muridarum remained longer and with the highest bacterial burdens in the prostate, thus showing a special tropism for this organ. Infection caused leukocyte infiltration, mainly composed by neutrophils, and also induced early pelvic pain development that rapidly dropped and resolved as the infection became chronic. Bacterial load and leukocyte infiltration was observed in all prostate lobes, although dorsolateral prostate was the most affected lobe. Interestingly, immune responses induced by both mice strains were characterized by the production of high levels of IL-10 during early stages of the infection, with highest and sustained levels observed in NOD mice, which showed to be less efficient in clearing the infection. Chronic infection of the prostate accompanied by local inflammation and pelvic pain development described herein have important implications for the improvement of the diagnosis and for the design of new efficient therapies. Prostate 77:517-529, 2017. © 2017 Wiley Periodicals, Inc.

Chronic Pelvic Pain Development and Prostate Inflammation in Strains of Mice With Different Susceptibility to Experimental Autoimmune Prostatitis.

BACKGROUND: Experimental autoimmune prostatitis (EAP) is an autoimmune inflammatory disease of the prostate characterized by peripheral prostate-specific autoimmune responses associated with prostate inflammation. EAP is induced in rodents upon immunization with prostate antigens (PAg) plus adjuvants and shares important clinical and immunological features with the human disease chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS). METHODS: EAP was induced in young NOD, C57BL/6, and BALB/c male mice by immunization with PAg plus complete Freund́s adjuvant. Tactile allodynia was assessed using Von Frey fibers as a measure of pelvic pain at baseline and at different time points after immunization. Using conventional histology, immunohistochemistry, FACS analysis, and protein arrays, an interstrain comparative study of prostate cell infiltration and inflammation was performed. RESULTS: Chronic pelvic pain development was similar between immunized NOD and C57BL/6 mice, although the severity of leukocyte infiltration was greater in the first case. Coversely, minimal prostate cell infiltration was observed in immunized BALB/c mice, who showed no pelvic pain development. Increased numbers of mast cells, mostly degranulated, were detected in prostate samples from NOD and C57BL/6 mice, while lower total counts and resting were observed in BALB/c mice. Prostate tissue from NOD mice revealed markedly increased expression levels of inflammatory cytokines, chemokines, adhesion molecules, vascular endothelial growth factor, and metalloproteinases. Similar results, but to a lesser extent, were observed when analyzing prostate tissue from C57BL/6 mice. On the contrary, the expression of the above mediators was very low in prostate tissue from immunized BALB/c mice, showing significantly slight increments only for CXCL1 and IL4. CONCLUSIONS: Our results provide new evidence indicating that NOD, C57BL/6, and BALB/c mice develop different degrees of chronic pelvic pain, type, and amount of prostate cell infiltration and secretion of inflammatory mediators. Our results corroborate and support the notion that mice with different genetic background have different susceptibility to EAP induction. Prostate 77:94-104, 2017. © 2016 Wiley Periodicals, Inc.

Regulatory T cells control strain specific resistance to Experimental Autoimmune Prostatitis.

Susceptibility to autoimmune diseases results from the encounter of a complex and long evolved genetic context with a no less complex and changing environment. Major actors in maintaining health are regulatory T cells (Treg) that primarily dampen a large subset of autoreactive lymphocytes escaping thymic negative selection. Here, we directly asked whether Treg participate in defining susceptibility and resistance to Experimental Autoimmune Prostatitis (EAP). We analyzed three common laboratory strains of mice presenting with different susceptibility to autoimmune prostatitis upon immunization with prostate proteins. The NOD, the C57BL/6 and the BALB/c mice that can be classified along a disease score ranging from severe, mild and to undetectable, respectively. Upon mild and transient depletion of Treg at the induction phase of EAP, each model showed an increment along this score, most remarkably with the BALB/c mice switching from a resistant to a susceptible phenotype. We further show that disease associates with the upregulation of CXCR3 expression on effector T cells, a process requiring IFNγ. Together with recent advances on environmental factors affecting Treg, these findings provide a likely cellular and molecular explanation to the recent rise in autoimmune diseases incidence.

Design of a novel plasmonic nanoconjugated analytical tool for ultrasensitive antigen quantification.

To date, while various diagnostic approaches for antigen detection have been proposed, most are too expensive, lengthy and limited in sensitivity for clinical use. Nanoparticle systems with unique material properties, however, circumvent these problems and offer improved accuracy and sensitivity over current methods like the enzyme-linked immunosorbent assay (ELISA). Herein, we present a novel functionalization strategy of plasmonic nanoparticle probes capable of specific quantification of antigens directly in clinical samples. A nanoconjugation strategy that allows one to perform an intensity depletion immuno-linked assay (IDILA), involving specific antibodies that target the antigen of interest was designed to obtain a calibration curve and achieve the quantification of the antigen in clinical samples in the same experiment using a microplate reader (i.e., an UV-vis spectrophotometer). Finally, the IDILA methodology allowed specific detection of various clinically relevant antigens, with significantly improved sensitivity over the ELISA. Furthermore, the assay was shown to be robust, reliable, cheap and rapid, diagnosing antigens in clinical serum samples within 2 hours.

IL-17 is not essential for inflammation and chronic pelvic pain development in an experimental model of chronic prostatitis/chronic pelvic pain syndrome.

Pain and inflammation in the absence of infection are hallmarks in chronic prostatitis and chronic pelvic pain syndrome (CP/CPPS) patients. The etiology of CP/CPPS is unclear, and autoimmunity has been proposed as a cause. Experimental autoimmune prostatitis (EAP) models have long been used for studying CP/CPPS. Herein, we studied prostate inflammation induction and chronic pelvic pain development in EAP using IL-12p40-KO, IL-4-KO, IL-17-KO, and wild-type (C57BL/6) mice. Prostate antigen (PAg) immunization in C57BL/6 mice induced specific Th1 and Th17 immune responses and severe prostate inflammation and cell infiltration, mainly composed of CD4 T cells and macrophages. Moreover, chronic pelvic pain was evidenced by increased allodynia responses. In immunized IL-17-KO mice, the presence of a prominent PAg-specific Th1 immune response caused similar prostate inflammation and chronic pelvic pain. Furthermore, markedly high PAg-specific Th1 immune responses, exacerbated prostate inflammation, and chronic pelvic pain were detected in immunized IL-4-KO mice. Conversely, immunized IL-12p40-KO mice developed PAg-specific Th2 immune responses, characterized by high IL-4 secretion and neither infiltration nor damage in the prostate. As observed in wild-type control animals, IL12p40-KO mice did not evidence tactile allodynia responses. Our results suggest that, as in patients, chronic pelvic pain is a consequence of prostate inflammation. After PAg immunization, a Th1-associated immune response develops and induces prostate inflammation and chronic pelvic pain. The absence of Th1 or Th2 cytokines, respectively, diminishes or enhances EAP susceptibility. In addition, IL-17 showed not to be essential for pathology induction and chronic pelvic pain development.

Agarofuran sesquiterpenes from Schaefferia argentinensis.

Sixteen dihydro-ß-agarofuran sesquiterpenes were isolated from the aerial parts of Schaefferia argentinensis Speg. Their structures were determined by a combination of 1D and 2D NMR and MS techniques. The in vitro antiproliferative activity of the major sesquiterpenes was examined in T47D, MCF7, and MDA-MB231 human cancer cell lines, but was found to be marginal.