Chronic Testicular Inflammation Research Articles

Monocytes expressing activin A and CCR2 exacerbate chronic testicular inflammation by promoting immune cell infiltration.

Monocytes expressing activin A and CCR2 exacerbate chronic testicular inflammation by promoting immune cell infiltration.

Alterations of small non-coding RNA in the spermatozoa of mice with paternal experimental autoimmune epididymo-orchitis are associated with metabolic dysfunction in offspring.

Infection and inflammation of the genital tract are major potentially treatable factors contributing to male infertility. The profile of small non-coding RNA (sncRNAs) in spermatozoa can be altered by environmental exposures and inflammatory conditions. Experimental autoimmune epididymo-orchitis (EAEO) is a well-established model of autoimmune-induced chronic testicular and epididymal inflammation. This model investigates the effect of chronic inflammation on sperm sncRNA profiles and offspring phenotypes. Regarding the EAEO model, mice were immunized with testis homogenates thrice. Subsequently, flow cytometry and histological analyses were conducted on EAEO mice. Next-generation sequencing was used to profile small RNA of spermatozoa from the caput, corpus, and cauda epididymis. We performed a comprehensive integrative analysis of sperm sncRNAs and chronic epididymitis and identified their molecular signatures. The metabolic functions of the first-generation (F1) offspring were evaluated using a glucose tolerance test (GTT). Body weight and metabolic function were significantly altered in F1 offspring from EAEO sperm donors. The analysis of cauda sperm sncRNA profiles revealed that the proportions of miRNAs and tsRNAs increased and decreased, respectively, after autoimmunization. Three differentially expressed miRNAs and seven differentially expressed tsRNAs were significantly correlated with F1 metabolic dysfunction. The expression patterns of miRNAs and tsRNAs in mice partially overlapped with those observed in the spermatozoa from human patients with chronic epididymitis. We revealed that autoimmune epididymo-orchitis alters sncRNA profiles in mouse spermatozoa. Offspring from mice with autoimmune orchitis develop metabolic disorders. A comprehensive analysis of human and mouse inflammation data revealed an association between alterations in the miRNA and tsRNA profiles of epididymal spermatozoa and offspring phenotypes.

Activin A and CCR2 regulate macrophage function in testicular fibrosis caused by experimental autoimmune orchitis

Experimental autoimmune-orchitis (EAO), a rodent model of chronic testicular inflammation and fibrosis, replicates pathogenic changes seen in some cases of human spermatogenic disturbances. During EAO, increased levels of pro-inflammatory and pro-fibrotic mediators such as TNF, CCL2, and activin A are accompanied by infiltration of leukocytes into the testicular parenchyma. Activin A levels correlate with EAO severity, while elevated CCL2 acting through its receptor CCR2 mediates leukocyte trafficking and recruits macrophages. CCR2 + CXCR4 + macrophages producing extracellular matrix proteins contribute widely to fibrogenesis. Furthermore, testicular macrophages (TMs) play a critical role in organ homeostasis. Therefore, we aimed to investigate the role of the activin A/CCL2-CCR2/macrophage axis in the development of testicular fibrosis. Following EAO induction, we observed lower levels of organ damage, collagen deposition, and leukocyte infiltration (including fibronectin+, collagen I+ and CXCR4+ TMs) in Ccr2−/− mice than in WT mice. Furthermore, levels of Il-10, Ccl2, and the activin A subunit Inhba mRNAs were lower in Ccr2−/− EAO testes. Notably, fibronectin+ TMs were also present in biopsies from patients with impaired spermatogenesis and fibrotic alterations. Overexpression of the activin A antagonist follistatin reduced tissue damage and collagen I+ TM accumulation in WT EAO testes, while treating macrophages with activin A in vitro increased the expression of Ccr2, Fn1, Cxcr4, and Mmp2 and enhanced migration along a CCL2 gradient; these effects were abolished by follistatin. Taken together, our data indicate that CCR2 and activin A promote fibrosis during testicular inflammation by regulating macrophage function. Inhibition of CCR2 or activin A protects against damage progression, offering a promising avenue for therapeutic intervention.

Aspirin Alleviates Particulate Matter Induced Asymptomatic Orchitis of Mice via Suppression of cGAS-STING Signaling.

As an important source of air pollutant, airborne particulate matter (PM) has become a major threat to public health. Orchitis is characterized by acute or chronic testicular inflammation and is a primary cause of male infertility. Although accumulating evidence indicates that PM exposure is associated with increased male infertility rates, the mechanism by which PM is involved is not well understood. Here, we found that short-term PM exposure activated NF-κB signaling in mouse Leydig cells and testes and leading to asymptomatic orchitis. Analyzing the mitochondrial abundance and cGAMP levels in PM exposed mouse Leydig cells, we found that PM exposure induced mitochondrial injury and mtDNA release, leading to inflammation via the cGAS-STING axis. We also found that aspirin-induced acetylation of cGAS inhibited the inflammation in mice after PM exposure, especially in the testes. Moreover, aspirin pretreatment rescued offspring growth in PM-exposed mice. In summary, our study not only provides evidence that PM-induced asymptomatic orchitis in mice may be amenable to aspirin pre-treatment by acetylating cGAS, but also provides a potential explanation for male infertility caused by air pollutants.

Experimental Cryptorchidism Causes Chronic Inflammation and a Progressive Decline in Sertoli Cell and Leydig Cell Function in the Adult Rat Testis.

Cryptorchidism causes spermatogenic failure and reduced serum androgen levels, as well as testicular oedema and fibrosis, which are hallmarks of inflammation. However, the role of inflammation and the effects of cryptorchidism on Sertoli cell and Leydig cell function at the molecular level remain ill-defined. Bilateral cryptorchidism was surgically induced in adult rats for 7 and 14 weeks. Testis weights decreased to 40% of normal within 7 weeks, due to loss of all developing spermatogenic cells except spermatogonia, but did not decrease further at 14 weeks. Serum FSH and LH were increased at both time points, consistent with a loss of feedback by inhibin and testosterone. This damage was accompanied by progressive accumulation of interstitial fluid and peritubular fibrosis, and a progressive decline of several critical Sertoli cell genes (Sox9, Inha (inhbin α-subunit), Cldn11 (claudin 11), Gja1 (connexin 43), and Il1a (interleukin-1α)) and the Leydig cell steroidogenic enzymes, Cyp11a1, Hsd3b1, and Hs17b3. Activin B and the activin-binding protein, follistatin, also declined, but the intratesticular concentration of activin A, which is a regulator of inflammatory responses, was largely unaffected at either time point. Expression of genes involved in inflammation (Tnf, Il10, Il1b, Mcp1) and fibrosis (Acta2, Col1a1) were considerably elevated at both time points. These data indicate that induction of experimental cryptorchidism, which causes complete failure of spermatogenesis in the adult rat, also induces chronic testicular inflammation, manifesting in oedema and fibrosis, and a progressive decline of Sertoli and Leydig cell gene expression and function.

Relevance of angiogenesis in autoimmune testis inflammation.

Experimental autoimmune orchitis (EAO) is a useful model to study organ-specific autoimmunity and chronic testicular inflammation. This model reflects testicular pathological changes reported in immunological infertility in men. Progression of EAO in rodents is associated with a significantly increased percentage of testicular endothelial cells and interstitial testicular blood vessels, indicating an ongoing angiogenic process. Vascular endothelial growth factor A (VEGFA), the main regulator of physiological and pathological angiogenesis, can stimulate endothelial cell proliferation, chemotaxis and vascular permeability. The aim of this study was to explore the role of VEGFA in the pathogenesis of testicular inflammation. Our results found VEGFA expression in Leydig cells, endothelial cells and macrophages in testis of rats with autoimmune orchitis. VEGFA level was significantly higher in testicular fluid and serum of rats at the end of the immunization period, preceding testicular damage. VEGF receptor (VEGFR) 1 is expressed mainly in testicular endothelial cells, whereas VEGFR2 was detected in germ cells and vascular smooth muscle cells. Both receptors were expressed in testicular interstitial cells. VEGFR2 increased after the immunization period in the testicular interstitium and VEGFR1 was downregulated in EAO testis. In-vivo-specific VEGFA inhibition by Bevacizumab prevented the increase in blood vessel number and reduced EAO incidence and severity. Our results unveil relevance of VEGFA-VEGFR axis during orchitis development, suggesting that VEGFA might be an early marker of testicular inflammation and Bevacizumab a therapeutic tool for treatment of testicular inflammation associated with subfertility and infertility.

Pathomechanisms of Autoimmune Based Testicular Inflammation.

Infection and inflammation of the male reproductive tract are relevant causes of infertility. Inflammatory damage occurs in the special immunosuppressive microenvironment of the testis, a hallmark termed testicular immune privilege, which allows tolerance to neo-antigens from developing germ cells appearing at puberty, long after the establishment of systemic immune tolerance. Experimental autoimmune orchitis (EAO) is a well-established rodent model of chronic testicular inflammation and organ specific autoimmunity that offers a valuable in vivo tool to investigate the pathological and molecular mechanisms leading to the breakdown of the testicular immune privilege. The disease is characterized by the infiltration of the interstitium by immune cells (mainly macrophages, dendritic cells, and T cells), formation of autoantibodies against testicular antigens, production of pro-inflammatory mediators such as NO, MCP1, TNFα, IL6, or activins and dysregulation of steroidogenesis with reduced levels of serum testosterone. EAO leads to sloughing of germ cells, atrophic seminiferous tubules and fibrotic remodeling, parameters all found similarly to changes in human biopsies from infertile patients with inflammatory infiltrates. Interestingly, testosterone supplementation during the course of EAO leads to expansion of the regulatory T cell population and inhibition of disease development. Knowledge of EAO pathogenesis aims to contribute to a better understanding of human testicular autoimmune disease as an essential prerequisite for improved diagnosis and treatment.

Induction of experimental autoimmune orchitis in mice: responses to elevated circulating levels of the activin-binding protein, follistatin

Experimental autoimmune orchitis (EAO) is a rodent model of chronic testicular inflammation that mimics the pathology observed in some types of human infertility. In a previous study, testicular expression of the inflammatory/immunoregulatory cytokine, activin A, was elevated in adult mice during the onset of EAO, indicating a potential role in the regulation of the disease. Consequently, we examined the development of EAO in mice with elevated levels of follistatin, an endogenous activin antagonist, as a potential therapeutic approach to testicular inflammation. Prior to EAO induction, mice received a single intramuscular injection of a non-replicative recombinant adeno-associated viral vector carrying a gene cassette of the circulating form of follistatin, FST315 (FST group). Serum follistatin levels were increased 5-fold in the FST group compared with the control empty vector (EV) group at 30 and 50 days of EAO, but intra-testicular levels of follistatin or activin A were not significantly altered. Induction of EAO was reduced, but not prevented, with mild-to-severe damage in 75% of the EV group and 40% of the FST group, at 50 days following immunisation with testicular homogenate. However, the EAO damage score (based on disruption of the blood-testis barrier, apoptosis, testicular damage and fibrosis) and extent of intratesticular inflammation (expression of inflammatory mediators) were directly proportional to the levels of activin A measured in the testis at 50 days. These data implicate activin A in the progression of EAO, thereby providing a potential therapeutic target; however, elevating circulating follistatin levels were not sufficient to prevent EAO development.

Predomination of IL-17-producing tryptase-positive/chymase-positive mast cells in azoospermic chronic testicular inflammation.

Chronic testicular inflammation and infection have been regarded as important factors in the pathogenesis of azoospermia. As key effector cells in innate and adaptive immune system, mast cells (MCs) were observed in inflammation and autoimmune disease. Furthermore, increased expression of tryptase-positive MCs has been reported in testicular disorders associated with male infertility/subfertility. However, little is known about the potential relationship between MCs and chronic testicular inflammation in azoospermic patients. Moreover, the preferential expression of MCs' subtypes in testis of these patients is still far from being understood. Thus, this study aimed to investigate characteristics of testicular MCs as well as their subtypes in azoospermic men with chronic testicular inflammation (AZI, n=5) by immunohistochemical techniques. Our results showed significant increase of MCs in AZI, and more importantly, considerable numbers of tryptase-positive/chymase-positive MCs could also be demonstrated in AZI, when compared to control groups representing azoospermia without chronic testicular inflammation (AZW, n=5) and normal spermatogenesis (NT, n=5) respectively. Most interestingly, immunofluorescence staining revealed autoimmune-associated interleukin (IL)-17-producing MCs in AZI, whereas co-expression of MC markers with tumour necrosis factor (TNF)-α, IL-10 and IL-1β could not be detected. In conclusion, AZI is associated with significant increase of tryptase-positive/chymase-positive MCs expressing IL-17, and these MCs might contribute to the pathogenesis of AZI.

Autoantibodies against protein disulfide isomerase ER-60 are a diagnostic marker for low-grade testicular inflammation.

Is there a non-invasive biomarker for the diagnosis of testicular inflammatory lesions? In sera from infertile azoospermic patients with histologically confirmed low-grade testicular inflammation, significantly elevated titers of autoantibodies against disulfide isomerase family A, member 3 (ER-60) were found. Infection and inflammation of the genital tract are supposed to be responsible for up to 15% of cases among infertile males. However, specific seminal or serological markers are not available to assess subacute or chronic inflammatory conditions in the testis. This study consisted of the identification of autoantibodies for testicular antigens in sera of patients with low-grade testicular inflammation, validation of candidates, development of an ELISA for the most promising target antigen and measurement of autoantibodies titers in healthy normozoospermic men (n = 20); male blood donors (n = 14); men with impaired semen quality without (n = 14) or with (n = 26) symptoms of genital tract infection/inflammation; azoospermic men with histologically confirmed testicular inflammatory lesions (n = 16); men after pharmacotherapy of genital tract infection/inflammation (n = 15) and men with acute epididymo-orchitis (n = 30). Proteins in lysates of normal testicular tissue were separated by high-resolution 2D gel electrophoresis and probed with sera of 13 patients with histologically confirmed chronic testicular inflammation. There were 14 proteins that immunoreacted with a majority of these sera and could be identified by mass spectrometry. Of these 14 proteins, disulfide isomerase family A, member 3 (ER-60), transferrin and chaperonin containing TCP1 complex, subunit 5 (epsilon) (CCT5) were considered as specific. Since ER-60 reacted with 92% of patient sera, an ER-60-autoantibody ELISA was developed. The newly established ELISA detected significantly elevated titers of autoantibodies against ER-60 in the sera from infertile men with histologically confirmed chronic testicular inflammation (median 8.6; P < 0.01) compared with the control groups. Moreover, elevated levels of anti-ER-60 titers were detected in patients suffering from acute epididymo-orchitis (median 3.3; P < 0.05) as compared with healthy normozoospermic men (median 2.13; P < 0.001), male blood donors with unknown fertility status (median 2.72; P < 0.01), patients with impaired semen quality but no infection/inflammation (median 2.59; P < 0.001) and patients with symptoms of genital tract infections and/or inflammation (median 2.18; P < 0.001). Significantly lower levels of anti-ER-60 antibodies were measured in sera from patients after application of anti-inflammatory pharmacotherapy (median 1.9; P < 0.01) compared with those with histologically confirmed chronic testicular inflammation. The cut-off value of the assay was set to 6.6 U/ml based on a calculated sensitivity of 100% and a specificity of 81.2%. The results obtained in this study showed statistically significant elevated titers of ER-60 antibodies in sera from patients with histologically confirmed testicular inflammatory lesions and from a few patients with acute epididymo-orchitis. However, the number of serum samples tested was limited. Severe testicular damage seen in azoospermic patients could represent a bias towards ER-60 reactivity, while the assay does not allow for different etiologies of the lesions to be distinguished. Due to ethical reasons, the prevalence of testicular inflammatory lesions among controls and non-azoospermic men cannot be studied at the histological level. Measurement of ER-60 autoantibody titers in serum could be a novel non-invasive marker for the diagnosis of asymptomatic testicular inflammation causing male fertility disturbances. This study was supported by a grant of the Deutsche Forschungsgemeinschaft (ME 1323/4-4) and the Translational Science Fund (Wirtschafts-und Strukturbank Hessen-WI Bank). M.F., A.P., W.W., H.-C.S. and A.M. are supported by the LOEWE focus group 'MIBIE' (Male infertility during infection and inflammation). The ER-60 ELISA is protected by a patent to the Justus-Liebig-University of Giessen with A.M. and M.F. as inventors (patent no. DE 10 2008 053 503). T.Z. as employee of the DRG Company was responsible for the ELISA development.

7: Cell-specific action of extracellular HMGB1 in sterile inflammation of rat testis

High mobility group box protein 1 (HMGB1), the non-histone chromosomal protein, plays an important role in onset and chronification of autoimmune diseases once released from the nuclei. In this study, we analyzed how HMGB1 can regulate inflammatory reactions in vivo in a rat model of experimental autoimmune orchitis (EAO) and in vitro in primary testicular cells. HMGB1 was translocated from the nuclei in EAO testis and in the testis of infertile men with leukocytic infiltrates. HMGB1 levels in EAO testis were elevated at late chronic phase of disease as compared to early proinflammatory cytokines such as IL-6 and TNF- α . We found that testicular somatic cells show a cell-specific expression profile of HMGB1 receptors TLR4 and receptor for advanced glycation end products (RAGE). The highly sensitive and specific proximity ligation assay was used to analyze HMGB1 receptor binding in testicular cells. HMGB1-TLR4 binding was dominant in testicular macrophages. However, Sertoli and peritubular cells showed higher levels of HMGB1-RAGE interaction. In support, HMGB1 triggered RAGE-dependent ERK1/2 MAPK and CREB activation in Sertoli and peritubular cells, whilst in testicular macrophages HMGB1 induced TLR4-signaling as evidenced by p38 MAPK and p65 NF-?B phosphorylation which stimulated an increase in mRNA levels of TNF- α and IL-6. Recent data shows that RAGE induced ERK activation leads to enhanced autophagy levels. In line with these data, extracellular HMGB1 triggered formation of autophagosomes in Sertoli cells. Increased autophagy levels in Sertoli cells might explain how these cells survive the inflammatory environment in EAO testis. Considering HMGB1’s late phase of action, inhibition of HMGB1 may be a putative target for therapeutic intervention in treatment of chronic testicular inflammation.

P086 HMGB1 modulates immune responses in a cell-specific manner in the rat autoimmune orchitis

Introduction High mobility group box protein 1 (HMGB1) is a non-histone nucleosomal protein. Under inflammatory conditions HMGB1 is secreted from activated immune cells or released from necrotic cells and acts as an endogenous danger signal. Despite its immune privileged status, the testis is prone to inflammatory infertility. In this study we analyzed the role of HMGB1 in mediating autoimmune responses in experimental autoimmune orchitis (EAO) – a rodent model of chronic testicular inflammation. Methods EAO was induced in WKY rats by immunization with testicular homogenate in complete Freund’s adjuvant. HMGB1 localization was analyzed in EAO and adjuvant control rat testes by immunofluorescence staining. Levels of HMGB1, IL-6 and TNF- α were measured in testes and serum using ELISA. Isolated testicular macrophages (TM), Sertoli cells (SC) and peritubular cells (PTC) were stimulated with recombinant HMGB1. Cytokine mRNA levels were measured by qRT-PCR. Testosterone production was evaluated using RIA assay in HMGB1 treated Leydig cells (LC). Expression of RAGE as well as activation of NF-?B (p65), MAPK (p38, ERK1/2) and Akt pathway were investigated in TM, SC and pPTC by Western blot. Possible binding of HMGB1 to TLR4 was investigated in isolated testicular cells using Duolink PLA assay. Results In control testes, HMGB1 was expressed mainly in the nucleus of testicular somatic cells. In contrast, EAO testes showed translocation of HMGB1 from the nucleus into the cytoplasm. Concentrations of IL-6 and TNF- α were increased 50 days after first immunization in EAO testes, whilst HMGB1 levels increased 80 days post-immunization. RAGE was highly expressed in SC as well as PTC and to a lower level in TM. In contrast, TLR4 levels were higher in TM. Our preliminary results show that HMGB1 binding to TLR4 is significantly higher in TM than in PTC. In vitro stimulation of isolated TM using recombinant HMGB1 activates phosphorylation of p38 and p65. However, HMGB1 treated PTC and SC activated ERK1/2. HMGB1 induced a significant increase in mRNA expression of TNF- α and IL-6 in TM. In HMGB1 treated SC the levels of IL-6 and TNF- α mRNA did not show a statistically significant difference. HMGB1 induced a significant increase in testosterone production by LC. Conclusion In control testis, HMGB1 was expressed mainly in the nucleus of testicular somatic cells. In contrast, EAO testes showed translocation of HMGB1 from the nucleus into the cytoplasm. Concentrations of IL-6 and TNF- α were increased 50 days after first immunization in EAO testis, whilst HMGB1 levels increased 80 days post-immunization. RAGE was highly expressed in SC as well as PTC and to a lower level in TM. In contrast, TLR4 levels were higher in TM. Our preliminary results show that HMGB1 binding to TLR4 is significantly higher in TM than in PTC. In vitro stimulation of isolated TM using recombinant HMGB1 activates phosphorylation of p38 and p65. In contrast, HMGB1 treated PTC and SC activated ERK1/2. HMGB1 induced a significant increase in mRNA expression of TNF- α and IL-6 in TM. In HMGB1 treated SC the levels of IL-6 and TNF- α mRNA did not show a statistically significant difference. HMGB1 induced a significant increase in testosterone production by LC.

Isolated granulomatous orchitis: MR imaging findings

Isolated granulomatous orchitis is a rare chronic testicular inflammation, which should be differentiated from testicular malignancies on scrotal imaging. We present a case of a 33-year-old man, with a diffuse pattern of granulomatous orchitis, confirmed by means of testicular biopsy. MR imaging of the scrotum revealed the presence of two small-sized intratesticular lesions, hypointense on T2-weighted images, not enhancing after gadolinium administration suggesting the diagnosis of benignity.

CD4+ and CD8+ T cells producing Th1 and Th17 cytokines are involved in the pathogenesis of autoimmune orchitis

Experimental autoimmune orchitis (EAO) is a useful model to study chronic testicular inflammation and infertility. EAO is characterized by severe damage of seminiferous tubules with germ cells that undergo apoptosis and sloughing. We previously reported an increase in CD4+ and CD8+ effector T cells in the testes of rats with EAO. Since cytokine patterns determine T cell effector functions, in the present work we analyzed the cytokines expressed by these cells during disease development. By flow cytometry, we detected an increase in the number of tumor necrosis factor-α (TNF) and interferon -γ (IFNG)-producing CD4+ T cells in the testis at EAO onset. As the severity of the disease progressed, these cells declined while CD8+ T cells producing TNF and IFNG increased, with the predominance of IFNG expression. As a novel finding, we identified by immunofluorescence CD4+ interleukin 17 (IL17)+ and CD8+ IL17+ cells in the testes of EAO rats, with CD4+ and CD8+ T cells predominating at the onset and in the chronic phase of EAO respectively. Moreover, IL17 (western blot) and IL23 content (ELISA) increased in EAO, with maximum levels in the chronic phase. These results suggest the involvement of CD4+ T helper (Th) 1 and Th17 subsets as co-effector cells governing EAO onset, as well as the central contribution of CD8+ T cells producing Th1 and Th17 cytokines in the maintenance of chronic inflammation. The expression of T-bet and RORγt (western blot) in the testis over the course of disease also supports the presence of Th1 and Th17 cells in the testes of EAO rats.

Differential changes in CD4 + and CD8 + effector and regulatory T lymphocyte subsets in the testis of rats undergoing autoimmune orchitis

Experimental autoimmune orchitis (EAO) is a useful model to study organ-specific autoimmunity and chronic testicular inflammation. EAO is characterized by an interstitial lymphomononuclear cell infiltration and damage of the seminiferous tubules showing germ cell sloughing and apoptosis. Using flow cytometry, we analysed the phenotype and number of T lymphocytes present in the testicular interstitium of rats during EAO development. A large increase in the number of testicular CD3 + T lymphocytes was detected. The number of CD4 + and CD8 + effector T lymphocytes (T effector cells) dramatically increased in the testis at EAO onset, with the CD4 + cell subset predominating. As the severity of the disease progressed, CD4 + T effector cells declined in number while the CD8 + T effector cell subset remained unchanged, suggesting their involvement in maintenance of the chronic phase of EAO. As a novel finding, we detected by immunohistochemistry and flow cytometry Foxp3 expressing CD4 + and CD8 + regulatory T lymphocytes (T regs) in chronically inflamed testis of EAO rats. The numbers of both T reg cell subsets increased in the testis of rats with orchitis, mainly at the onset of EAO; CD4 +Foxp3 + T reg cells were more abundant than CD8 +Foxp3 + T reg cells. Unexpectedly, CD25 − T lymphocytes were more abundant than CD25 + cells within CD4 +Foxp3 + and CD8 +Foxp3 + T reg cell populations. Although T reg subsets are actively accumulated into the testis in EAO rats, these cells are outnumbered by an even more vigorously expanding T effector subset. Further, it is possible that factors present in the inflamed testis might limit the ability of T regs to abrogate tissue damage.

Cytokines and chemokines in testicular inflammation: A brief review

A wide spectrum of data in the literature shows the relevance of cytokines as paracrine regulators of spermatogenesis and steroidogenesis in the normal testis. In this brief review, we highlight the relevance of cytokines in the testis during inflammation. This phenomenon involves complex and multiple interactions among immune and germ cells generally resulting in the alteration of spermatogenesis. The complexity of these cell interactions is multiplied because Sertoli and Leydig cells are also producers of pro- and anti-inflammatory cytokines and chemokines. Also, cytokines are pleiotropic and they exert opposite and/or redundant effects in different conditions. However, in spite of this bidirectional immunoregulatory function of cytokines, the mass of the data, reported from experiments of acute testicular inflammation, shows upregulation of interleukin (IL)-1beta, IL-1alpha, IL-6, and tumor necrosis factor alpha (TNF-alpha), which induce adverse effects on germ cells. In autoimmune orchitis, a chronic testicular inflammation, chemokines such as CCL2, CCL3, and CCL4 induce attraction and extravasation of immune cells within the testicular interstitium. These cells alter the normal immunosuppressor microenvironment principally through the secretion of proinflammatory cytokines, interferon-gamma initially, and IL-6 and TNF-alpha thereafter. Germ cells expressing TNFR1, IL-6R, and Fas increase in number and undergo apoptosis, through the TNF-alpha/TNFR1, IL-6/IL-6R, and Fas/Fas L systems. The knowledge of immune-germ and somatic testicular cell interactions will contribute to the understanding of the mechanisms by which chronic inflammatory conditions of the testis can disrupt the process of spermatogenesis.

Dendritic Cell Based Intervention Effectively Prevents the Development of Experimental Autoimmune Orchitis in Rats.

In recent years, dendritic cells (DC) have received increasing attention as key players in both, the initiation and maintenance of autoimmune disease and the induction of peripheral tolerance. The maturation and/or activation state of DC is regarded as a control point for the induction of either peripheral tolerance or autoimmunity. Especially immature or semi-mature DC have a potent ability to tolerize T cells and prevent undesired immune reactions. Immature DC have the highest capacity to internalize antigens, but low T cell stimulatory activity, whereas mature DC downregulate their endocytic activity and are excellent T lymphocyte stimulators. Infection and inflammation of the male genital tract are amongst the most important identifiable etiologies of infertility and experimental autoimmune orchitis (EAO) serves as a model to study chronic testicular inflammation and organ specific autoimmunity. The histopathology of EAO is characterized by a substantial increase of interstitial lymphocytes and macrophages, different degrees of germ cell loss caused by apoptosis resulting in aspermatogenesis and atrophy of the seminiferous tubules. Our recent characterization of the testicular autoantigens heat shock protein 70 (hsp70) and heteronuclear ribonucleoprotein H1 (hn RNP H1) in EAO animals as well as the isolation and characterization of rat testicular DC prompted us to explore the potential of DC in inducing tolerance to EAO development. DC precursor cells were isolated from the bone marrow of inbred Wistar rats and differentiated in vitro to immature DC by addition of granulocyte-monocyte-colony stimulating factor (GM-CSF) and interleukin (IL)-4 to the culture medium. After 7 days in culture DC were pulsed either with the recombinant autoantigens hsp70, hn RNP H1, whole testicular homogenate or left unpulsed. DC were injected s.c. 4 weeks prior induction of EAO using testicular homogenate in adjuvant. Disease progression was intermittantly monitored using flat-panel volumetric computerized tomography (fpvCT) and by histological examination 80 days after injection of DC (50 days after EAO induction). In vivo injection of unpulsed DC showed a strong inhibitory effect on EAO development with only 33% of rats revealing signs of disease in comparison to control (100% EAO). DC pulsed with hsp70, hn RNP H1 and testicular homogenate proved to be less effective than unpulsed immature DC (66% of diseased animals). Significantly elevated levels of the anti-inflammatory cytokine IL-10 in the supernatants of isolated T cells from the spleen of rats injected with unpulsed DC point to a Th2 response which could be an explanation for the effective induction of tolerance by this cell based immunization.