Varicocele-Mediated Male Infertility: From the Perspective of Testicular Immunity and Inflammation.

Background: Varicocele (VC) is present in 35 - 40% of men with infertility. However, current surgical and antioxidant treatments are not completely effective. In addition to oxidative stress, it is likely that other factors such as testicular immune microenvironment disorder contribute to irreversible testicular. Evidence suggests that VC is associated with anti-sperm antibodies (ASAs), spermatogenesis and testosterone secretion abnormalities, and testicular cytokine production. Moreover, inhibition of inflammation can alleviate VC-mediated pathogenesis. The normal function of the testis depends on its immune tolerance mechanism. Testicular immune regulation is complex, and many infectious or non-infectious diseases may damage this precision system. Results: The testicular immune microenvironment is composed of common immune cells and other cells involved in testicular immunity. The former includes testicular macrophages, T cells, dendritic cells (DCs), and mast cells, whereas the latter include Leydig cells and Sertoli cells (SCs). In animal models and in patients with VC, most studies have revealed an abnormal increase in the levels of ASAs and pro-inflammatory cytokines such as interleukin (IL)-1 and tumor necrosis factor (TNF)-alpha in the seminal plasma, testicular tissue, and even peripheral blood. It is also involved in the activation of potential inflammatory pathways, such as the nucleotide-binding oligomerization domain-like receptor family pyrin domain containing (NLRP)-3 pathway. Finally, the development of VC-mediated infertility (VMI) may be facilitated by abnormal permeability of proteins, such as claudin-11, that constitute the blood-testis barrier (BTB). Conclusions: The testicular immune response, including the production of ASAs and inflammatory factors, activation of inflammatory pathways, and destruction of the BTB may be involved in the pathogenesis of VMI it is necessary to further explore how patient outcomes can be improved through immunotherapy.

Bone Status in a Mouse Model of Experimental Autoimmune-Orchitis.

Investigations in male patients with fertility disorders revealed a greater risk of osteoporosis. The rodent model of experimental autoimmune-orchitis (EAO) was established to analyze the underlying mechanisms of male infertility and causes of reduced testosterone concentration. Hence, we investigated the impact of testicular dysfunction in EAO on bone status. Male mice were immunized with testicular homogenate in adjuvant to induce EAO (n = 5). Age-matched mice were treated with adjuvant alone (adjuvant, n = 6) or remained untreated (control, n = 7). Fifty days after the first immunization specimens were harvested. Real-time reverse transcription-PCR indicated decreased bone metabolism by alkaline phosphatase and Cathepsin K as well as remodeling of cell-contacts by Connexin-43. Micro computed tomography demonstrated a loss of bone mass and mineralization. These findings were supported by histomorphometric results. Additionally, biomechanical properties of femora in a three-point bending test were significantly altered. In summary, the present study illustrates the induction of osteoporosis in the investigated mouse model. However, results suggest that the major effects on bone status were mainly caused by the complete Freund's adjuvant rather than the autoimmune-orchitis itself. Therefore, the benefit of the EAO model to transfer laboratory findings regarding bone metabolism in context with orchitis into a clinical application is limited.

Prior and novel coronaviruses, Coronavirus Disease 2019 (COVID-19), and human reproduction: what is known?

OBJECTIVE: To summarize current understanding of the effects of novel and prior coronaviruses on human reproduction, specifically male and female gametes, and in pregnancy. DESIGN: Review of English publications in PubMed and Embase to April 6, 2020. METHOD(S): Articles were screened for reports including coronavirus, reproduction, pathophysiology, and pregnancy. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Reproductive outcomes, effects on gametes, pregnancy outcomes, and neonatal complications. RESULT(S): Seventy-nine reports formed the basis of the review. Coronavirus binding to cells involves the S1 domain of the spike protein to receptors present in reproductive tissues, including angiotensin-converting enzyme-2 (ACE2), CD26, Ezrin, and cyclophilins. Severe Acute Respiratory Syndrome Coronavirus 1 (SARS-CoV-1) may cause severe orchitis leading to germ cell destruction in males. Reports indicate decreased sperm concentration and motility for 72-90 days following Coronavirus Disease 2019 (COVID-19) infection. Gonadotropin-dependent expression of ACE2 was found in human ovaries, but it is unclear whether SARS-Coronavirus 2 (CoV-2) adversely affects female gametogenesis. Evidence suggests that COVID-19 infection has a lower maternal case fatality rate than SARS or Middle East respiratory syndrome (MERS), but anecdotal reports suggest that infected, asymptomatic women may develop respiratory symptoms postpartum. Coronavirus Disease 2019 infections in pregnancy are associated with preterm delivery. Postpartum neonatal transmission from mother to child has been reported. CONCLUSION(S): Coronavirus Disease 2019 infection may affect adversely some pregnant women and their offspring. Additional studies are needed to assess effects of SARS-CoV-2 infection on male and female fertility.

Exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin promotes inflammation in mouse testes: The critical role of Klotho in Sertoli cells.

Increasing evidence shows that 2,3,7,8-tetrachlorodibenzo-p-dioxin(TCDD) enhances inflammation, and inflammation has a significant negative impact on fertility. Therefore, the aim of this study was to investigate the effects of TCDD on testis inflammation. Pregnant mice and primary Sertoli cells were treated with TCDD, and male offspring and Sertoli cells were treated with lipopolysaccharides(LPS). We then measured testis apoptotic cells, proinflammatory cytokines, and observed the Klotho/PDLIM2/p65 pathway. In vivo results revealed that TCDD further enhanced LPS-increased testis apoptotic cells and concentrations of testicular proinflammatory cytokines (IL1ß, IL18, and IL12) (p < 0.05). An in vitro investigation showed the levels of proinflammatory cytokines were increased in TCDD + LPS-treated cells compared with LPS-treated cells (p < 0.05). Compared with the LPS-treated cells, expression of Klotho and PDLIM2 was significantly decreased in TCDD + LPS-treated cells (p < 0.05), while expression of p65 and NLRP3 were significantly increased in the cotreatment cells (p < 0.05). However, the addition of Klotho to the TCDD + LPS-cotreated cells significantly increased PDLIM2 and decreased p65 activation and NLRP3 (p < 0.05). Meanwhile, mRNA levels and the secretion of proinflammatory cytokines were both suppressed by exogenous Klotho (p < 0.05). Administration of Klotho decreased TCDD + LPS-induced cytokines and apoptosis in mice (p < 0.05). Taken together, TCDD may increase testicular inflammation by affecting the secretion of proinflammatory cytokines in Sertoli cells via the Klotho/PDLIM2/p65 pathway, which influences the testicular microenvironment and induces germ cell apoptosis.

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.

ASSOCIATION OF MEAN PLATELET VOLUME AND THE MONOCYTE/LYMPHOCYTE RATIO WITH BRUCELLA-CAUSED EPIDIDYMO-ORCHITIS.

We evaluated the association between the mean platelet volume (MPV) and monocyte/lymphocyte ratio (MLR) with brucella-caused epididymo-orchitis to determine if they could be used to differentiate between brucella and non-brucella epididymo-orchitis. The charts of 88 patients with non-brucella and 14 patients with brucella epididymo-orchitis were retrospectively reviewed. Brucellosis was diagnosed by isolating Brucella spp from a blood culture or from a serum agglutination titer ≥ 1:160 along with accompanying clinical findings. The patients with brucella epididymo-orchitis were significantly more likely to have a lower MPV and a higher MLR than those with non-brucella epididymo-orchitis. Using a MPV cut-off level of less than 9.25 fl to differentiate brucella from non-brucella epididymo-orchitis gives a sensitivity of 78.6%, a specifity of 78.4%, a positive predictive value of 36.7% and a negative predictive value of 95.8%. Using a MLR cut-off level of greater than 0.265 to differentiate brucella from non-brucella epididymo-orchitis gives a sensitivity of 71.4%, a specifity of 65.9%, a positive predictive value of 25% and a negative predictive value of 93.5.%. MPV and MLR values may assist in differentiating between brucella and non-brucella epididymo-orchitis.

Thymoquinone ameliorates testicular tissue inflammation induced by chronic administration of oral sodium nitrite.

Although sodium nitrite has been widely used as food preservative, building bases of scientific evidence about nitrite continues to oppose the general safety in human health. Moreover, thymoquinone (TQ) has therapeutic potential as antioxidant, anti-inflammatory, antibacterial and anticancer. Therefore, we investigated the effects of both sodium nitrite and TQ on testicular tissues of rats. Forty adult male Sprague Dawley rats were used. They received either 80 mg kg(-1) sodium nitrite or 50 mg kg(-1) TQ daily for twelve weeks. Serum testosterone was measured. Testis were weighed and the testicular tissue homogenates were used for measurements of tumour necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-4, IL-6, IL10, caspase-3, caspase-8 and caspase-9. Sodium nitrite resulted in significant reduction in serum testosterone concentration and elevation in testis weight and Gonado-Somatic Index. We found significant reduction in testicular tissues levels of IL-4 and IL-10 associated with elevated levels of TNF-α, IL-1ß, IL-6, caspase-3, caspase-8 and caspase-9. In conclusion, chronic oral sodium nitrite induced changes in the weight of rat testis accompanied by elevation in the testicular tissue level of oxidative stress markers and inflammatory cytokines. TQ attenuated sodium nitrite-induced testicular tissue damage through blocking oxidative stress, restoration of normal inflammatory cytokines balance and blocking of apoptosis.

Roles of Toll-like receptors 2 and 4 in mediating experimental autoimmune orchitis induction in mice.

The mammalian testis is an immunoprivileged site where male germ cell antigens are immunologically tolerated under physiological conditions. However, some pathological conditions can disrupt the immunoprivileged status and induce autoimmune orchitis, an etiological factor of male infertility. Mechanisms underlying autoimmune orchitis induction are largely unknown. The present study investigated the roles of Toll-like receptor 2 (TLR2) and TLR4 in mediating the induction of experimental autoimmune orchitis (EAO) in mice after immunization with male germ cell antigens emulsified with complete Freund adjuvant. Wild-type mice developed severe EAO after three immunizations, which was characterized by leukocyte infiltration, autoantibody production, and impaired spermatogenesis. Tlr2 or Tlr4 deficient mice showed relatively low susceptibility to EAO induction compared with wild-type mice. Notably, Tlr2 and Tlr4 double knockout mice were almost completely protected from EAO induction. Moreover, we demonstrated that TLR2 was crucial in mediating autoantibody production in response to immunization. The results imply that TLR2 and TLR4 cooperatively mediate EAO induction.

[Clinical characteristics of mumps infection in children and adult patients].

The aim of the article was to study clinical manifestations of mumps infection (infectious parotitis) - a viral illness that affects glands that produce saliva, pancreas, and nervous system in children and adult patients. 219 patients (42 children and 177 adults) with mumps infection were studied. The investigation showed that parotid salivary gland disorder was the most common in adults; sublinguitis - inflammation of the sublingual gland was the most common in children. Serous meningitis occurred exclusively in preschool and early school age. Pancreatitis was less common in children than in adults. Infectious parotitis involving the parotid salivary gland was taking its normal course with positive outcome. Pancreatitis and serous meningitis occurred at the 3-5 day of illness with infectious parotitis. Pancreatitis was with positive outcome, with the exceptions of adult patients with pain syndrome (repair process delayed to 1-1.5 months). Mean duration of hospitalization for children with infectious parotitis was 7 days, for adults - 10-14 days. Mean duration of hospitalization for patients with serous meningitis was 14 days. Study showed that in 20,1% of 16-27 years old males developed orchitis.

Effects of moderate exercise over different phases on age-related physiological dysfunction in testes of SAMP8 mice.

Oxidative stress and chronic inflammation have been implicated in the testicular aging process. Different types and moderate-intensity of regular exercise may reduce age-related physiological dysfunction associated with inflammation and oxidative stress, but such effects of moderate-intensity of exercise over different phases of life in testes have not been reported. In this study, male SAMP8 mice, a senescence-accelerated strain, were maintained as sedentary (sed) or subjected to daily 15-min periods of swimming exercise between ages of 2-7 months (lifelong), 2-4 months (earlier) or 5-7 months (late). Age-related changes, including serum testosterone levels and biomarkers of inflammation and oxidative stress were analyzed at the end of the experiment. All exercise groups showed significantly greater serum testosterone levels and decreased age-related inflammation and oxidative stress compared with the sedentary group. Exercise also increased expression and activity of the nuclear factor erythroid 2-related factor (Nrf2), a transcriptional regulator of the cellular anti-oxidant system, and decreased expression and activity of nuclear factor kappa beta (NF-κB), a mediator of inflammatory molecules, in the nucleus of testicular cells. However, lifelong and earlier groups generally showed significantly better protective effects than the late group against age-related physiological dysfunction in testes. Thus, lifelong exercise and earlier phase exercise were most effective in counteracting oxidative stress and inflammation and in preserving testes function through regulation of Nrf2 and NF-κB. These results advocate the benefits of lifelong exercise and emphasize a greater protection against male aging by instituting exercise earlier rather than late in life.

Electrophysiologic evaluation of cremasteric reflex in experimental orchitis.

AIM: Absent cremasteric reflex (CR) is a well known but not reliable sign of testicular torsion. We hypothesized that CR can also be altered in other causes of acute scrotum in children. An experimental study was performed to evaluate the clinical and electrophysiological features of CR in orchitis. METHOD: Eighteen Wistar albino rats were allocated into three groups: control (CG), sham (SG) and orchitis (OG). In CG, after anesthetization with ketamine hydrochloride, the medial site of the anterior superior iliac spine was stimulated to obtain CR electrophysiologically, and latency and duration were recorded with a needle electrode placed in the cremasteric muscle. Electrophysiologic evaluations were performed 24 h after injection of 0.1 ml of 10(6) cfu/ml Escherichia coli (0:6 strain) in 1 ml of physiologic saline into the right testicle in OG, and 1 ml of saline only in SG. All testicles were sampled to check for orchitis after the electrophysiologic evaluations. RESULTS: CR was obtained in all rats in CG and in 83.3% and 66.6% in SG and OG respectively (p < 0.05). The latency of CR was significantly higher in OG (15.1 ± 0.9 ms) and SG (15.5 ± 1.2 ms) than CG (10.5 ± 0.7 ms) (p < 0.017). The duration of CR was 15.1 ± 3.2 ms in CG, 16.2 ± 4.9 ms in SG and 18.5 ± 3 ms in OG (p > 0.05). Histopathologic confirmation of orchitis was obtained in all testicle samples in OG, and number of neutrophils and total orchitis score was significantly higher in OG than the other groups (p < 0.05). CONCLUSION: Electrophysiologic parameters of CR may be altered in orchitis. Prolonged latency of CR in orchitis may be due to inflammation of the genitofemoral nerve or cremasteric muscle.

Early regression of spermatogenesis in boars of an inbred Duroc strain caused by incident orchitis/epididymo-orchitis.

In the process of establishment of an inbred Duroc pig strain, males with size asymmetry of the testes were frequently observed. To clarify the possible causes of this asymmetry, we examined the testes and epididymides of 67 males of the F4-F7 generations at 35-100 weeks of age. Testicular weights showed a wide variation (120-610 g). When the weights of the testes were compared bilaterally, 35 of the 67 males showed more than a 10% difference. Histological examination of testes from this asymmetry group revealed a range of seminiferous tubule disruption including disappearance of all germ cells, but not Sertoli cells, in the epithelium. Focal lesions associated with the degenerated tubules were observed. Trends of incident fibrosis or hyalinization of these lesions were seen in aged males of the asymmetry group. Besides this abnormality of spermatogenesis, infiltration of mononuclear inflammatory cells around the tubule was frequently observed in the asymmetry group (32.9%, compared with 1.6% in males showing testis symmetry). In severe cases, the inflammatory cells were concentrated in the intertubular region instead of Leydig cells. Cellular infiltration was also observed around the epididymal duct and blood vessels, but its incidence did not differ between the symmetry and asymmetry groups. Testicular testosterone levels were significantly increased in the asymmetry group, but those of E2 and inhibin did not differ between the two groups. These histopathological features indicate that disruption of spermatogenesis after orchitis/epididymo-orchitis could induce testicular atrophy. Genetic predispositions for this trait may cause prevalent retrograde infections, resulting in orchitis/epididymo-orchitis.

Lymphangiogenesis in chronic inflammation in the testis.

Lymphangiogenesis occurs in various organs under inflammatory conditions. Recently, it was demonstrated that activated macrophages play an important role in the process of lymphangiogenesis. However, lymphangiogenesis during testicular inflammation has not yet been studied. Here, we investigated lymphangiogenesis in experimental autoimmune orchitis, a immunologic male infertility model, in mice. Histological changes were observed using immunohistochemical staining with the monoclonal antibodies against F4/80 (mature macrophage marker), lymph vessel endothelium HA-receptor 1 (LYVE-1) (lymphatic endothelial cells marker) and CD31 (endothelial cells marker). The expression of angiogenesis and lymphangiogenesis factors, such as vascular endothelial growth factor (VEGF)-A, VEGF-C, VEGF-D and TNF-α, which are secreted by activated macrophages, were examined using real-time RT-PCR. The results showed that lymphangiogenesis occurred along the undersurface of the tunica albuginea but not into the interstitium proper between the seminiferous tubules (STs) during the orchitis. It was noted that some F4/80-positive macrophages expressed LYVE-1 at the undersurface of the tunica albuginea and also in the testicular interstitium proper. RT-PCR analysis revealed that the expressions of VEGF-A, VEGF-D and TNF-α were significantly increased but that of VEGF-C remained unchanged in the inflammatory testes. This study suggests that testicular macrophages are involved in the specific lymphangiogenesis in the chronic inflammation.

First report of orchitis in man caused by Brucella abortus biovar 1 in Ecuador.

We present a 44-year-old man from a rural community in northern Ecuador who worked on a cattle farm where he was involved with primary veterinary care, including assistance during births (or calving) and placenta retention and artificial insemination, with minimal precautions. In September of 2009, quite abruptly, he developed asthenia and hypersomnia without any apparent cause or symptoms like fever, chills, or night sweats. On November 14, 2009, he suffered from pain and edema in the right testicle that coincided with pain in the abdomen. Clinical, serological, and bacteriological investigations confirmed the first case of unilateral orchitis in man in Ecuador caused by Brucella abortus biovar 1. Because brucellosis is a neglected disease, special attention should be given to it in the training of medical and veterinary students.

Postinflammation stage of autoimmune orchitis induced by immunization with syngeneic testicular germ cells alone in mice.

We previously established an immunological infertility model, experimental autoimmune orchitis (EAO), which can be induced by two subcutaneous injections of viable syngeneic testicular germ cells on days 0 and 14 in mice without using any adjuvant. In this EAO model, CD4+ T-cell-dependent lymphocytic infiltration and immune deposits were found with spermatogenic disturbance on day 120. However, the late stage of EAO (= postactive inflammation stage on day 365) has not yet been investigated. Therefore, we investigated the histopathological characteristics of the late stage. The results revealed that the lymphocytic infiltration finally resolved; however, the seminiferous epithelium persistently showed maturation arrest and the Sertoli cell-only feature. In the seminiferous tubules showing maturation arrest, both proliferation and apoptosis of germ cells had occurred simultaneously. It was also noted that there were deposits of immunoglobulin G and the third component of complement on the thickened basement membrane of seminiferous tubules in the late stage of EAO. These results indicate that histopathology after active inflammation in EAO comprises persistent damage to the seminiferous epithelium and may resemble the histopathology of "idiopathic disturbance of spermatogenesis" in man.

[Effectiveness of cycloferon in complex treatment of brucellosis patients with lesions of the scrotum].

The article presents the results of urological examination (spermograms and data of ultrasound examination) of 22 patients with chronic brucellosis and diseases of the scrotum (6 patients with orchitis, 16 with orchiepididymitis) before and after conventional therapy (10 patients) and combined treatment with the inclusion of cycloferon (2 courses of 5 intramuscular injection [0.25 g] with an interval of 10 days)--12 patients. It is shown that the administration of cycloferon leads to more effective relief of intoxication symptoms and inflammation in the testes and appendages (reduction of scrotal wall thickness, size of testes and/or adjuncts, and the incidence and severity of hydrocele), and has a positive effect on spermatogenesis (reduction of semen viscosity, the number of white blood cells in semen, sperm agglutination associated with the formation of sperm antibodies in most patients after treatment), as well as reduces the number of exacerbations of chronic orchitis/orchiepididymitis by 2.4 times.

Immunoprivileged sites: the testis.

The testis is an immunological privileged tissue as evidenced by its ability to support grafts with minimal rejection. Immune privilege is essential for the tolerance of neo-antigens from developing germ cells that appear after the constitution of self-tolerance, but imposes the paradoxical task of also providing efficient protection against pathogens and tumor cells. It is becoming increasingly clear that immune privilege cannot be attributed to a single factor such as the sequestration of neo-antigens from the immune system behind the blood-testis barrier, but is based on a complex multifaceted interplay between cells and factors that are essential for the reproductive function of the testis and the testicular immune system. This review summarizes the evidence that has accumulated regarding the role of Sertoli cells, androgens, and selected population of leukocytes in the maintenance of immune privilege and its perturbation in testicular inflammatory sub- and infertility.

[Orchitis and male infertility]. / Orchitis und Infertilität.

Infections and inflammations of the genital tract are considered the most frequent causes of reduced male fertility, but conclusive epidemiological data are not available. In view of the exposure of germ cells to pathogenic components as well as the cells and mediators involved in the inflammatory processes, irreversible damage to spermatogenesis and corresponding decline of ejaculate quality are to be expected, particularly in cases of chronic orchitis. While the consequences of orchitis and epididymo-orchitis that exhibit clinical symptoms due to systemic or local infections are well known, including testicular atrophy and complete loss of fertility, those cases of inflammatory reactions of the testicles that manifest an asymptomatic or subclinical course, or are not even due to an infection, have received little attention until now. However, systematic histopathological analyses have shown a high prevalence of asymptomatic inflammatory reactions in testicular biopsies from infertile men. The mostly focal lymphocytic infiltrates correlate with the degree of damage to spermatogenesis and corresponding clinical and endocrinological parameters of testicular function. Noninvasive diagnostic techniques are not yet available so that chronic asymptomatic inflammations of the testicles as the primary cause or cofactor of male fertility disorders are underestimated. Except for administration of pathogen-specific antibiotics, treatment recommendations are to a large extent still lacking.