Mechanisms underlying impaired spermatogenic function in orchitis induced by busulfan.

Busulfan is an alkylating agent commonly used in cancer chemotherapy. It is also an ideal agent for preparing transplant recipients of spermatogonial stem cells because of its high efficiency in destroying endogenous germ cells in the testis. However, its toxicity mechanism remains unclear, affecting its clinical use and applications. Based on reports of busulfan causing orchitis and a previous study by our team, this article summarizes the relationship between busulfan and orchitis, cytokines, the blood-testis barrier, and the cytoskeleton, unravels the regulatory pathways and mechanism behind busulfan-induced orchitis, and reveals the molecular mechanism underlying impaired spermatogenic function in orchitis, providing new ideas for the clinical application of busulfan while reducing its testicular toxicity.

Sodium-glucose cotransporter 2 inhibitor-associated severe epididymo-orchitis.

A man in his late 50s, with uncontrolled type 2 diabetes mellitus (T2DM) and morbid obesity, presented to the hospital with complicated epididymo-orchitis. The onset of symptoms (scrotal pain, erythema and swelling) occurred after the use of empagliflozin, a sodium-glucose cotransporter 2 (SGLT2) inhibitor, for 2 months. His baseline antidiabetic medications were insulin, glipizide and metformin. Initially, he had failed treatment of epididymo-orchitis with oral levofloxacin for 3 weeks, followed by 2 weeks of doxycycline therapy. At the presentation to the hospital, an ultrasound of the scrotum revealed scrotal and right testicular abscess. The patient underwent right inguinal orchiectomy. Postoperatively, pus culture was positive for Enterococcus faecalis and Candida glabrata, and hence, he was treated with oral antibiotics including high-dose antifungal medications. Adequate wound care and regular follow-up demonstrated resolution of infection. This case highlights the risk of severe urogenital infection associated with the use of SGLT2 inhibitors in the setting of uncontrolled T2DM.

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.

The ameliorative effect of ellagic acid on di-(2-ethylhexyl) phthalate-induced testicular structural alterations, oxidative stress, inflammation and sperm damages in adult mice.

BACKGROUND: Phthalates such as di (2-ethylhexyl) phthalate (DEHP) are well known exogenous substances, disrupting reproductive system function and structure. The current research demonstrated the effect of ellagic acid (EA) on DEHP-induced testicular injury in mice. METHODS: Thirty-five healthy adult male mice were randomly divided to five groups; normal saline receiving group, DEHP (2 g/kg/day, dissolved in corn oil, p.o.) receiving group, DEHP (2 g/kg/day, dissolved in corn oil, p.o.) and EA receiving groups (25, 50 and 100 mg/kg/day, p.o.). Treatment duration of animals was 14 days. Body and testes weights and sperm characteristics and histological changes of testes were evaluated. Serum testosterone, luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels were analyzed. In the testicular tissue, oxidative/nitrosative stress markers and inflammatory cytokine levels were measured. RESULTS: Ellagic acid significantly reduced DEHP-induced reduction of body and testes weights. The DEHP-induced reduction of spermatogonia, primary spermatocyte and sertoli cells numbers as well as reduction of sperm vitality and progressive motility were reversed by EA. Furthermore, EA inhibited DEHP-induced alterations in serum hormone levels. These effects were associated with the reduction of DEHP-induced increased level of oxidative stress and inflammatory responses. CONCLUSIONS: Ellagic acid considerably inhibits testicular toxicity of DEHP through reducing oxidative/nitrosative stress and inflammatory responses. Our data suggest that EA may be considered as a promising agent to inhibit male reproductive toxicity induced by endocrine disrupting chemicals such as DEHP.

Dysfunction in Sertoli cells participates in glucocorticoid-induced impairment of spermatogenesis.

The effect of stress on male fertility is a widespread public health issue, but less is known about the related signaling pathway. To investigate this, we established a hypercortisolism mouse model by supplementing the drinking water with corticosterone for four weeks. In the hypercortisolism mice, the serum corticosterone was much higher than in the control, and serum testosterone was significantly decreased. Moreover, corticosterone treatment induced decrease of sperm counts and increase of teratozoospermia. Increased numbers of multinucleated giant cells and apoptotic germ cells as well as downregulated meiotic markers suggested that corticosterone induced impaired spermatogenesis. Further, upregulation of macrophage-specific marker antigen F4/80 as well as inflammation-related genes suggested that corticosterone induced inflammation in the testis. Lactate content was found to be decreased in the testis and Sertoli cells after corticosterone treatment, and lactate metabolism-related genes were downregulated. In vitro phagocytosis assays showed that the phagocytic activity in corticosterone-treated Sertoli cells was downregulated and accompanied by decreased mitochondrial membrane potential, while pyruvate dehydrogenase kinase-4 inhibitor supplementation restored this process. Taken together, our results demonstrated that dysfunctional phagocytosis capacity and lactate metabolism in Sertoli cells participates in corticosterone-induced impairment of spermatogenesis.

Methane Ameliorates Lipopolysaccharide-Induced Acute Orchitis by Anti-inflammatory, Antioxidative, and Antiapoptotic Effects via Regulation of the PK2/PKR1 Pathway.

OBJECTIVE: The present study is aimed at investigating the anti-inflammatory, antioxidative, and antiapoptotic effects of methane on lipopolysaccharide- (LPS-) induced acute orchitis and its potential mechanisms. METHODS: Adult male rats were intraperitoneally (i.p.) injected with methane-rich saline (MS, 20 mL/kg) following LPS (5 mg/kg, i.p.). The survival rate was assessed every 12 h until 72 h after LPS induction, and surviving rats were sacrificed for further detection. The wet/dry (W/D) ratio was determined, and testicular damage was histologically assessed. Inflammatory cytokines in the testes and serum, including interleukin-1ß (IL-1ß), IL-6, IL-10, and tumor necrosis factor-α (TNF-α), were measured using ELISA and RT-qPCR. Oxidative stress was evaluated by the level of superoxide dismutase (SOD) and malondialdehyde (MDA). Testicular apoptosis was detected via TUNEL staining. The expression of prokineticin 2 (PK2)/prokineticin receptor 1 (PKR1) was also analyzed using RT-qPCR, western blotting, and immunohistochemistry. RESULTS: It was found that methane significantly prolonged rat survival, decreased the W/D ratio, alleviated LPS-induced histological changes, and reduced apoptotic cells in the testes. Additionally, methane suppressed and promoted the production of pro- and anti-inflammatory cytokines, respectively. Furthermore, methane significantly increased SOD levels, decreased MDA levels, and decreased testicular expression of PK2 and PKR1. Therefore, methane exerts therapeutic effects on acute orchitis and might be a new and convenient strategy for the treatment of inflammation-related testicular diseases.

Protective effect of Alpha-Linolenic acid on Lipopolysaccharide-Induced Orchitis in mice.

Previous studies have demonstrated that polyunsaturated fatty acids (PUFAs) have anti-inflammatory effects. One specific PUFA, alpha-linolenic acid (ALA), shows both anti-inflammatory and antioxidant properties. In the testes, inflammatory mediators are known to increase when orchitis is induced by bacterial lipopolysaccharide (LPS). This study aimed to determine whether the anti-inflammatory properties of ALA could have a protective effect against LPS-induced orchitis in mice. The mice were divided into untreated control, orchitis and ALA-treated orchitis groups. Orchitis was induced by intraperitoneal injection of LPS. The ALA-treated group was administered ALA by gavage three days before intraperitoneal LPS injection. Cyclooxygenase-2 (COX-2), cytosolic phospholipase A2 (cPLA2), inducible nitric oxide synthase (iNOS) enzymes and nuclear factor kappa-B (NF-κB) in the testes, as well as serum interleukin 6 (IL-6) and tumour necrosis factor-alpha (TNF-α), were analysed using enzyme-linked immunosorbent assay (ELISA) tests. LPS administration increased the expression of several inflammatory mediators, including IL-6, TNF-α and NF-κB, as well as the COX-2, cPLA2 and iNOS enzymes. ALA administration significantly prevented the LPS-induced increases in these inflammatory mediators and enzymes (p < .05). The anti-inflammatory and antioxidant effects of ALA may make it a useful candidate for the treatment of orchitis caused by bacterial LPS.

Evaluating the effects of acacetin versus a low dose of cisplatin drug on male reproductive system and kidney in mice: With emphasis on inflammation process.

This study aimed to compare acacetin adverse effect besides a cisplatin low dose on male reproductive and urinary systems on mice. In this study, 36 male Balb/c mice were received Dimethyl sulfoxide, cisplatin (1 mg/kg) or acacetin (10, 25, 50 mg/kg) for 3 days, while the sixth group, treated with acacetin (50 mg/kg) for 10 days. All treatments were done consequence daily and intraperitoneally. Histological and biochemical factors to male reproductive and urinary systems were assayed. Only in cisplatin exposed group, significant differences were seen with the others. So that, some reproductive criteria were significantly decreased; serum levels of follicle-stimulating hormone, luteinizing hormone, testosterone, sperm parameters, the diameters of seminiferous tubules, Johnsen's score and from the urinary system; renal corpuscles' space, Mg2+ concentration (p < .01). Moreover, significant increases were seen in serum levels of tumour necrosis factor-alpha, Blood urea nitrogen, creatinine, testis myeloperoxidase activity and tumour necrosis factor-alpha expression, kidney histological damage and renal corpuscle diameter (p < .01). Cisplatin exposure disrupts histological and functional characteristics of either male reproductive or urinary systems, suggesting by inflammation-promoting. Acacetin does not induce any harmful impact on these two systems and could be considered as a safe flavonoid by high dose and prolonged usage.

Fluoride Induces Autoimmune Orchitis Involved with Enhanced IL-17A Secretion in Mice Testis.

Fluoride (F) widely exists in the water and food. Recent studies reported that F induced testicular toxicity via inflammation reaction. This study was aimed to explore the mechanism of F-induced inflammation in testis. 100 healthy male mice (BALB/cJ strain) were randomly divided into five groups including: control, experimental autoimmune orchitis (EAO), and three F groups (25, 50, and 100 mg/L sodium fluoride (NaF)). After 150 d, the results showed a significant increase in testicular cytokines levels including of IL-17A, IL-6, IFN-γ, and TNF-α in NaF and EAO groups compared with control group. Interestingly, the presence of specific antisperm autoantibodies in antitesticular autoantibodies and the notable recruitment of immunocyte (T cells and dendritic cells) were also observed in NaF and EAO groups. In addition, findings showed that in NaF and EAO groups macrophages and T cells both significantly secreted IL-17A, and the protein and mRNA levels of cytokines (IL-6 and TGF-ß) were significantly increased. From these results, it can be concluded that autoimmune orchitis and IL-17A are implicated in F-induced testicular inflammation.

Severe Epididymo-Orchitis and Encephalitis Complicating Anti-PD-1 Therapy.

BACKGROUND: Immune checkpoint inhibitors such as pembrolizumab and nivolumab have emerged as active treatment options for patients with many cancers, including metastatic melanoma, but can also cause symptomatic or life-threatening immune-related adverse events, including encephalitis. Epididymitis and orchitis are rare complications of these therapies. CASE PRESENTATION: We describe herein a patient with metastatic melanoma who developed epididymo-orchitis followed by encephalitis while receiving pembrolizumab. The patient developed testicular pain and fever after his third dose of pembrolizumab; ultrasound evaluation demonstrated bilateral epididymo-orchitis. He then developed headaches, fever, and altered mental status over the next week and was admitted to the hospital. Lumbar puncture revealed inflammatory changes consistent with meningoencephalitis; he did not improve with broad-spectrum antibiotics, and an extensive workup for infectious etiologies, including cerebrospinal fluid testing using a clinical metagenomic next-generation sequencing assay, was negative. He received high-dose steroids for suspected autoimmune encephalitis, and both his orchitis and meningoencephalitis improved rapidly after one dose. He fully recovered after a 5-week taper of oral steroids. DISCUSSION: Here, we report a case of epididymo-orchitis complicating immune checkpoint inhibitor therapy. This patient subsequently developed severe encephalitis but rapidly improved with steroids. Clinicians should be aware of rare complications of these agents. KEY POINTS: Epididymo-orchitis is a rare and potentially life-threatening complication of anti-programmed death protein 1 (anti-PD-1) therapy.For patients on anti-PD-1 therapy who develop either epididymo-orchitis or epididymitis without clear infectious cause, immune-related adverse events should be considered in the differential diagnosis.If severe, epididymo-orchitis related to anti-PD-1 therapy may be treated with high-dose corticosteroids.

Systemic LPS induces toll-like receptor 3 (TLR3) expression and apoptosis in testicular mouse tissue.

It is well known that sepsis and inflammation reduce male fertility. Within the testis, toll-like receptor 3 (TLR3) is constitutively expressed and recognizes double-stranded RNA (dsRNA) from viruses, degraded bacteria, damaged tissues and necrotic cells. To characterize the potential role of TLR3 in response to testicular infections, its expression and downstream signaling were investigated upon challenge with lipopolysaccharides (LPS) in two mouse strains that differ in their immuno-competence regarding T cell-regulated immunity. Thereto, Balb/c and Foxn1nu mice were randomized into six interventional groups treated with either i.v. application of saline or LPS followed by 20 min, 5 h 30 min and 18 h of observation and two sham-treated control groups. LPS administration induced a significant stress response; the amplification was manifested for TLR3 and interleukin 6 (IL6) mRNA in the impaired testis 5 h 30 min after LPS injection. TLR3 immunostaining revealed that TLR3 was primarily localized in spermatocytes. The TLR3 expression displayed different temporal dynamics between both mouse strains. However, immunofluorescence staining indicated only punctual interferon regulatory factor 3 (IRF3) expression upon LPS treatment along with minor alterations in interferon ß (IFNß) mRNA expression. Induction of acute inflammation was closely followed by a significant shift of the Bax/Bcl2 ratio to pro-apoptotic signaling accompanied by augmented TUNEL-positive cells 18 h after LPS injection with again differing patterns in both mouse strains. In conclusion, this study shows the involvement of TLR3 in response to LPS-induced testicular inflammation in immuno-competent and -incompetent mice, yet lacking transmission into its signaling pathway.

[Testicular injection of lipopolysaccharide: An effective method for establishing the mouse model of orchitis].

OBJECTIVE: To search for a method of establishing a reliable mouse model of orchitis and investigate the association of orchitis with the activation of the inflammasome. METHODS: We equally randomized 40 adult male KM mice into groups A (sham operation), B (intraperitoneal injection of lipopolysaccharide ï¼»LPSï¼½), C (unilateral testicular injection of glacial acetic acid ï¼»GAAï¼½), and D (unilateral testicular injection of LPS). At 3 weeks after modeling, we measured the sperm concentration and percentage of progressively motile sperm (PMS) in the epididymis by computer-assisted semen analysis, observed the pathological changes in the testis tissue by HE staining, and determined the expressions of the Caspase-1 and interleukin (IL)-1ß proteins by Western blot. RESULTS: The sperm concentration in the epididymis was significantly decreased in groups B (ï¼»25.74 ± 3.19ï¼½ ×106/ml), C (ï¼»17.16 ± 4.41ï¼½ ×106/ml) and D (ï¼»16.92 ± 7.13ï¼½ ×106/ml) as compared with that in group A (ï¼»28.20 ± 1.63ï¼½ ×106/ml) (all P < 0.05), even more significantly in B than in C and D (P < 0.01), and so was PMS in groups B (ï¼»29.57 ± 2.16ï¼½%), C (ï¼»18.10 ± 2.38ï¼½%) and D (ï¼»7.34 ± 1.63ï¼½%) in comparison with group A (ï¼»59.34 ± 1.10ï¼½%) (P < 0.01), even more significantly in B and C than in D (P < 0.01). Light microscopy revealed different degrees of pathological changes in the testis tissue, most significant in group D, followed by C and B. Both the expressions of Caspase-1 and IL-1ß were remarkably up-regulated in groups B, C and D compared with those in group A (P < 0.01), even more markedly in D than in B and C (P < 0.05). CONCLUSIONS: Unilateral testicular injection of LPS is a more efficient method than either unilateral testicular injection of GAA or intraperitoneal injection of LPS for establishing the mouse model of orchitis. Orchitis may be pathologically associated with the activation of the NLRP3 inflammasome.

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.

Microcystin-leucine arginine exhibits immunomodulatory roles in testicular cells resulting in orchitis.

Microcystin-leucine arginine (MC-LR) causes testicular inflammation and hinders spermatogenesis. However, the molecular mechanisms underlying the immune responses to MC-LR in the testis have not been elucidated in detail. In this study, we show that MC-LR induced immune responses in Sertoli cells (SC), germ cells (GC), and Leydig cells (LC) via activating phosphatidylinositol 3-kinase (PI3K)/AKT/nuclear factor kappa B (NF-κB), resulting in the production of pro-inflammatory cytokines and chemokines including tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1), and chemokine (C-X-C motif) ligand 10 (CXCL10). The observed effects were attributed to reduced activity of protein phosphatases 2A (PP2A) as a result of binding of MC-LR to the catalytic subunit of PP2A in SC and GC. By contrast, innate immune responses were triggered by Toll-like receptor 2 (TLR2) in LC because MC-LR could not enter into the LC and subsequently inhibit the PP2A activity. PI3K/AKT/NF-κB were also activated in SC, GC, and LC in vivo, with the enrichment of TNF-α, IL-6, MCP-1, and CXCL10 in the testis. Following chronic exposure, MC-LR-treated mice exhibited decreased sperm counts and abnormal sperm morphology. Our data demonstrate that MC-LR can activate innate immune responses in testicular cells, which provides novel insights to explore the mechanism associated with MC-LR-induced 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.

Low-dose exposure to di-(2-ethylhexyl) phthalate (DEHP) increases susceptibility to testicular autoimmunity in mice.

Exposure to di-(2-ethylhexyl) phthalate (DEHP) induces spermatogenic disturbance (SD) through oxidative stress, and affects the immune system by acting as an adjuvant. Recently, we reported that in mice, a low dose of DEHP, which did not affect spermatogenesis, was able to alter the testicular immune microenvironment. Experimental autoimmune orchitis (EAO) can be induced by repeated immunization with testicular antigens, and its pathology is characterized by production of autoantibodies and SD. In the present study, we investigated the effect of a low-dose DEHP on the susceptibility of mice to EAO. The exposure to DEHP-containing feed (0.01%) caused a modest functional damage to the blood-testis barrier (BTB) with an increase in testicular number of interferon gamma (IFN-γ)-positive cells and resulted in the production of autoantibodies targeting haploid cells, but did not affect spermatogenesis. While only single immunization with testicular antigens caused very mild EAO, the concurrent DEHP exposure induced severe EAO with significant increases in number of interferon gamma-positive cells and macrophages, as well as lymphocytic infiltration and serum autoantibody titer accompanied by severe SD. To summarize, the exposure of mice to the low-dose DEHP does not induce significant SD, but it may cause an increase in IFN-γ positive cells and modest functional damage to the BTB in the testis. These changes lead to an autoimmune response against haploid cell autoantigens, resulting in increased susceptibility to EAO.

Inhibition of NOS-NO System Prevents Autoimmune Orchitis Development in Rats: Relevance of NO Released by Testicular Macrophages in Germ Cell Apoptosis and Testosterone Secretion.

BACKGROUND: Although the testis is considered an immunoprivileged organ it can orchestrate immune responses against pathological insults such as infection and trauma. Experimental autoimmune orchitis (EAO) is a model of chronic inflammation whose main histopathological features it shares with human orchitis. In EAO an increased number of macrophages infiltrate the interstitium concomitantly with progressive germ cell degeneration and impaired steroidogenesis. Up-regulation of nitric oxide (NO)-NO synthase (NOS) system occurs, macrophages being the main producers of NO. OBJECTIVE: The aim of our study was to evaluate the role of NO-NOS system in orchitis development and determine the involvement of NO released by testicular macrophages on germ cell apoptosis and testosterone secretion. METHOD AND RESULTS: EAO was induced in rats by immunization with testicular homogenate and adjuvants (E group) and a group of untreated normal rats (N) was also studied. Blockage of NOS by i.p. injection of E rats with a competitive inhibitor of NOS, L-NAME (8mg/kg), significantly reduced the incidence and severity of orchitis and lowered testicular nitrite content. L-NAME reduced germ cell apoptosis and restored intratesticular testosterone levels, without variations in serum LH. Co-culture of N testicular fragments with testicular macrophages obtained from EAO rats significantly increased germ cell apoptosis and testosterone secretion, whereas addition of L-NAME lowered both effects and reduced nitrite content. Incubation of testicular fragments from N rats with a NO donor DETA-NOnoate (DETA-NO) induced germ cell apoptosis through external and internal apoptotic pathways, an effect prevented by N-acetyl-L-cysteine (NAC). DETA-NO inhibited testosterone released from Leydig cells, whereas NAC (from 2.5 to 15 mM) did not prevent this effect. CONCLUSIONS: We demonstrated that NO-NOS system is involved in the impairment of testicular function in orchitis. NO secreted mainly by testicular macrophages could promote oxidative stress inducing ST damage and interfering in Leydig cell function.

Endogenous interleukin 18 regulates testicular germ cell apoptosis during endotoxemia.

Orchitis (testicular swelling) often occurs during systemic inflammatory conditions, such as sepsis. Interleukin 18 (IL18) is a proinflammatory cytokine and is an apoptotic mediator during endotoxemia, but the role of IL18 in response to inflammation in the testes was unclear. WT and IL18 knockout (KO) mice were injected lipopolysaccharide (LPS) to induce endotoxemia and examined 12 and 48  h after LPS administration to model the acute and recovery phases of endotoxemia. Caspase activation was assessed using immunohistochemistry. Protein and mRNA expression were examined by western blot and quantitative real-time RT-PCR respectively. During the acute phase of endotoxemia, apoptosis (as indicated by caspase-3 cleavage) was increased in WT mice but not in IL18 KO mice. The death receptor-mediated and mitochondrial-mediated apoptotic pathways were both activated in the WT mice but not in the KO mice. During the recovery phase of endotoxemia, apoptosis was observed in the IL18 KO mice but not in the WT mice. Activation of the death-receptor mediated apoptotic pathway could be seen in the IL18 KO mice but not the WT mice. These results suggested that endogenous IL18 induces germ cell apoptosis via death receptor mediated- and mitochondrial-mediated pathways during the acute phase of endotoxemia and suppresses germ cell apoptosis via death-receptor mediated pathways during recovery from endotoxemia. Taken together, IL18 could be a new therapeutic target to prevent orchitis during endotoxemia.

Lack of TCRalphabeta+ CD8+ and TCRgammadelta+ lymphocytes ameliorates LPS induced orchitis in mice--preliminary histological observations.

The inflammation of the reproductive system can affect reproduction causing partial or complete infertility. It is well known that lipopolysaccharide (LPS) triggers an inflammatory response in the whole organism, including immunologically privileged organs, e.g. the testicles. Adult male TCRalpha-/-, TCRdelta-/-, CD1d-/- and beta2m-/- on B10.PL (H-2(u)) and B10.PL control mice were intraperitonealy (i.p.) injected with lipopolysaccharide (LPS). The animals were killed 24h and 10 days post LPS treatment and their gonads were prepared for microscopic examination. Histological changes in the testes after LPS injection were found only in control B10PL and CD1d-/- mice. The experiments revealed disturbances in Leydig's glands structure, blood vessel dilatation in the interstitial tissue as well as degeneration of seminal tubule epithelium, disruption ofspermatogenesis and subsequent decrease of sperm cell number in the tubule lumen. These changes were noticed mainly 10 days after LPS treatment. Lack of either TCRalphabeta+ CD8+ or TCRgammadelta+ lymphocytes diminishes the response of testicular macrophages to LPS whereas the absence of CD1d-dependent NKT cells does not affect macrophage reactivity.