RESUMO
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.
Assuntos
Glucocorticoides/toxicidade , Células de Sertoli/fisiologia , Espermatogênese/efeitos dos fármacos , Animais , Proteínas de Ligação ao Cálcio/análise , Corticosterona/toxicidade , Síndrome de Cushing/sangue , Síndrome de Cushing/induzido quimicamente , Síndrome de Cushing/fisiopatologia , Ácido Dicloroacético/farmacologia , Hormônio Foliculoestimulante/sangue , Ácido Láctico/metabolismo , Hormônio Luteinizante/sangue , Masculino , Meiose/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Orquite/induzido quimicamente , Orquite/metabolismo , Fagocitose/efeitos dos fármacos , Piruvato Desidrogenase Quinase de Transferência de Acetil/antagonistas & inibidores , Receptores Acoplados a Proteínas G/análise , Células de Sertoli/metabolismo , Contagem de Espermatozoides , Espermatozoides/patologia , Testículo/metabolismo , Testosterona/sangueRESUMO
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.
Assuntos
Doenças Autoimunes/prevenção & controle , Inibidores Enzimáticos/farmacologia , Células Intersticiais do Testículo/metabolismo , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico/antagonistas & inibidores , Orquite/prevenção & controle , Espermatozoides/metabolismo , Acetilcisteína/farmacologia , Adjuvantes Imunológicos , Animais , Apoptose/efeitos dos fármacos , Doenças Autoimunes/induzido quimicamente , Doenças Autoimunes/imunologia , Doenças Autoimunes/patologia , Técnicas de Cocultura , Misturas Complexas , Regulação da Expressão Gênica , Humanos , Células Intersticiais do Testículo/efeitos dos fármacos , Células Intersticiais do Testículo/imunologia , Células Intersticiais do Testículo/patologia , Hormônio Luteinizante/sangue , Hormônio Luteinizante/genética , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/patologia , Masculino , Óxido Nítrico/biossíntese , Doadores de Óxido Nítrico/farmacologia , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Orquite/induzido quimicamente , Orquite/imunologia , Orquite/patologia , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Espermatozoides/efeitos dos fármacos , Espermatozoides/imunologia , Espermatozoides/patologia , Testosterona/biossíntese , Testosterona/metabolismo , Triazenos/farmacologiaRESUMO
BACKGROUND: Male reproductive tract infection and inflammation are important aetiological factors of infertility. Experimental Autoimmune Orchitis (EAO) is a model of chronic inflammation useful to study mechanisms of inflammatory reactions leading to testicular impairment. EAO is characterised by interstitial cell infiltrate of lymphomonocytes, producers of pro-inflammatory cytokines involved in germ cell apoptosis. Nitric oxide (NO), a free radical promoting immune cell activation and apoptosis, is synthesised by conversion of l-arginine to l-citrulline catalysed by NO synthase (NOS). The NOS isoforms are: constitutively endothelial (e) and neuronal (n) NOS and inducible (i) NOS. OBJECTIVES: Although the NO-NOS system was found to be up-regulated by pro-inflammatory mediators in immune and non immune testicular cells, data on its regulation in chronic inflammatory states is lacking. METHODS AND RESULTS: EAO was induced in rats by active immunisation with spermatic antigens and adjuvants; control (C) rats were injected with adjuvants. Untreated normal (N) rats were also studied. We demonstrated that iNOS, eNOS and nNOS was mainly expressed by interstitial cells in N and C rats and that in EAO NOS was up-regulated and also expressed by tubular cells. Constitutive and inducible NOS content (Western blot) as well as NO production and activity increased in the testis of rats with EAO. iNOS content and activity were selectively up-regulated in the testis of rats with orchitis. Flow cytometric analysis of NOS isoforms in testicular macrophages (M) showed that the percentage of ED1(+)ED2(-) and ED1(+)ED2(+) M subsets, expressing constitutive and iNOS isoforms was significantly higher in EAO, but no change in the percentage of ED1(-)ED2(+) resident M was observed compared to C rats. M from EAO rats also released more NO than C and N rats. CONCLUSIONS: In testis of rats with EAO, NO-NOS system was up-regulated and both testicular M and cells from seminiferous tubules contributed to NO increase. NO over production in orchitis was generated mainly by increased iNOS content and activity.
Assuntos
Doenças Autoimunes/metabolismo , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico/metabolismo , Orquite/metabolismo , Testículo/metabolismo , Adjuvantes Imunológicos , Animais , Doenças Autoimunes/induzido quimicamente , Doenças Autoimunes/enzimologia , Western Blotting , Modelos Animais de Doenças , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Macrófagos/enzimologia , Macrófagos/metabolismo , Masculino , Óxido Nítrico Sintase Tipo I/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Orquite/induzido quimicamente , Orquite/enzimologia , Ratos , Ratos Sprague-Dawley , Testículo/enzimologia , Testículo/patologia , Regulação para CimaRESUMO
The influence of acute and chronic uranium poisoning on the induction and course of experimental autoimmune thyroiditis and orchitis was studied. It has been demonstrated that the testes and thyroid gland are involved in the general pathological process in uranium poisoning. Autoantibodies to the testes and to the thyroid gland were found to circulate in the blood of the poisoned animals. Uranium poisoning caused destructive changes in the testes and in the thyroid, similar to the picture of an autoimmune process. No summation of the effect was observed in the synthesis of autoantibodies during the induction of experimental autoimmune thyroiditis and orchitis on the background of chronic uranium poisoning although the most conspicuous histological changes were found in these groups and were characteristic of a pronounced autoimmune process.