Involvement of tumor necrosis factor-α in the pathogenesis of autoimmune orchitis in rats

We studied the testicular macrophages of rats with experimental autoimmune orchitis (EAO) and analyzed whether the tumor necrosis factor-α (TNFα) is involved in germ cell apoptosis and in Leydig cell steroidogenesis. The EAO was induced in adult male Sprague-Dawley rats by active immunization with testicular homogenate and adjuvants. In the experimental group, a severe orchitis was observed 80 days after the first immunization. ED1- and ED2-positive macrophages were quantified by immunohistochemistry. The TNFα concentration of conditioned media from testicular macrophages (TMCM) was determined by ELISA. The number of apoptotic TNF receptor 1 (TNFR1)-positive germ cells was identified by combining in situ end labeling of apoptotic DNA and immunohistochemical techniques. The effect of TNFα on Leydig cell testosterone production was determined by RIA. In rats with EAO, we observed a significant increase in the number of TNFα-positive testicular macrophages, the TNFα concentration in TMCM, and the number of TNFR1-positive germ cells. Sixty percent of TNFR1-positive germ cells were apoptotic. These results suggest that TNFα could be involved in the pathogenesis of EAO. Acting together with other local factors such as Fas-FasL, TNFα could trigger germ cell apoptosis. We also demonstrated that TNFα inhibited in vitro testosterone production in basal and hCG-stimulated Leydig cells from rats with orchitis.Facultad de Ciencias Exacta

Correlation between inhibin secretion and damage of seminiferous tubules in a model of experimental autoimmune orchitis

The aim of the present study was to evaluate inhibin secretion in rats with autoimmune orchitis. As we have previously described, experimental autoimmune orchitis (EAO) induced in rats by active immunization with testis homogenate and adjuvants is characterized by an interstitial mononuclear cell infiltrate and sloughing of the germinal epithelium. At 120 days after the first immunization 60% of the rats exhibited a severe orchitis with large areas of aspermatogenic seminiferous tubules in which only spermatogonia and Sertoli cells with cytoplasmic vacuolization remained attached to the tubular wall. None of the untreated (N) or control (C) rats revealed pathological alterations. Sixty percent decrease in testis weight was observed in rats with EAO compared with N or C groups. A 3-fold increase in serum FSH levels was observed in rats with EAO compared with N or C groups (19·8 ± 3·7 vs 5·6 ± 0·3 and 5·9 ± 0·1 ng/ml respectively). A significant decrease in inhibin B levels was observed in rats with EAO when compared with N or C groups (40 ± 4·6 vs 207 ± 38·8 and 221·4 ± 28·6 pg/ml respectively). An inverse correlation between inhibin B and FSH serum levels and a direct correlation between inhibin B and testis weight were found. Strong expression of the inhibin α-subunit in Sertoli cells of untreated and control rats was observed; this subunit was undetectable or poorly detectable in rats with orchitis. Positive staining for the inhibin α-subunit was also observed in Leydig cells of all groups studied. In conclusion, using a model of autoimmune orchitis our results show that circulating inhibin B levels and inhibin α-subunit expression in Sertoli cell cytoplasm closely correlate with the degree of damage of the germinal epithelium.Instituto Multidisciplinario de Biología CelularFacultad de Ciencias Exacta

Cyclooxygenase-2 and prostaglandin F2α in syrian hamster leydig cells : Inhibitory role on luteinizing hormone/human chorionic gonadotropin-stimulated testosterone production

We have previously found that cyclooxygenase-2 (COX-2), a key enzyme in the biosynthesis of prostaglandins (PGs), is present in the testicular interstitial cells of infertile men, whereas it is absent in human testes with no evident morphological changes or abnormalities. To find an animal model for further investigating COX-2 and its role in testicular steroidogenesis, we screened testes from adult species ranging from mice to monkeys. By using immunohistochemical assays, we found COX-2 expression only in Leydig cells of the reproductively active (peripubertal, pubertal, and adult) seasonal breeder Syrian hamster. COX-2 expression in hamster Leydig cells was confirmed by RT-PCR. In contrast, COX-1 expression was not detected in hamster testes. Because COX-2 expression implies PG synthesis, we investigated the effect of various PGs on testosterone production and found that PGF2α stood out because it significantly reduced human chorionic gonadotropin-stimulated testosterone release from isolated hamster Leydig cells in a dose-dependent manner. This mechanism involves a decreased expression of testicular steroidogenic acute regulatory protein and 17β-hydroxysteroid dehydrogenase. Testicular concentration and content of PGF2α in reproductively active hamsters as well as production of PGF2α from isolated hamster Leydig cells were also determined. Moreover, PGF2α receptors were localized in Leydig cells of hamsters and testicular biopsies from patients with Sertoli cell only and germ arrest syndromes. Thus, in this study, we described a COX-2-initiated pathway that via PGF2α production, PGF2α receptors, steroidogenic acute regulatory protein, and 17β-hydroxysteroid dehydrogenase represents a physiological local inhibitory system of human chorionic gonadotropin-stimulated testosterone production in the Syrian hamster testes.Instituto Multidisciplinario de Biología Celula

Augmented serum levels of the IGF-I/IGF-binding protein-3 ratio in pre-menopausal patients with type I breast cysts

Objective: Gross cystic disease (GCD) is the most common benign breast pathology. Although breast cysts are not considered pre-malignant lesions, an increased risk of breast cancer has been reported for patients with type I cysts (Na+/K+ < 3). Furthermore, an augmented IGF-I/IGF-binding protein-3 (IGFBP-3) ratio has been described in breast cancer patients. The objective was to evaluate serum IGF-I and binding protein concentrations of type I and type II cyst patients as compared with healthy women. Methods: Twenty-four patients with type I cysts, 17 with type II cysts and 25 healthy women were evaluated. Serum IGF-I, IGFBP-3 and IGFBP-1 concentrations were measured by IRMA. Results: IGF-I concentrations were significantly higher in sera from patients with type I cysts than in patients with type II cysts. A highly significant decrease of IGFBP-3, the major IGFBP, was found in patients with type I cysts with respect to healthy women, whereas no significant difference was evident between the different cyst types. The IGF-I/IGFBP-3 ratio, an estimate of biologically active IGF-I, was very significantly higher in patients with type I cysts than in both type II patients and healthy women. IGFBP-1 levels were significantly lower in patients with type I than in controls and type II cysts. The IGF-I/IGFBP-1 ratio was significantly higher in patients with type I cysts than in type II bearers and healthy women. Estrogen levels correlated with IGF-I in patients and controls. Conclusions: The enhanced levels of IGF-I/IGFBP-3 found in patients with type I cysts could eventually be associated with the increased risk of breast cancer described for this group.Facultad de Ciencias Exacta

Augmented serum levels of the IGF-I/IGF-binding protein-3 ratio in pre-menopausal patients with type I breast cysts

Objective: Gross cystic disease (GCD) is the most common benign breast pathology. Although breast cysts are not considered pre-malignant lesions, an increased risk of breast cancer has been reported for patients with type I cysts (Na+/K+ < 3). Furthermore, an augmented IGF-I/IGF-binding protein-3 (IGFBP-3) ratio has been described in breast cancer patients. The objective was to evaluate serum IGF-I and binding protein concentrations of type I and type II cyst patients as compared with healthy women. Methods: Twenty-four patients with type I cysts, 17 with type II cysts and 25 healthy women were evaluated. Serum IGF-I, IGFBP-3 and IGFBP-1 concentrations were measured by IRMA. Results: IGF-I concentrations were significantly higher in sera from patients with type I cysts than in patients with type II cysts. A highly significant decrease of IGFBP-3, the major IGFBP, was found in patients with type I cysts with respect to healthy women, whereas no significant difference was evident between the different cyst types. The IGF-I/IGFBP-3 ratio, an estimate of biologically active IGF-I, was very significantly higher in patients with type I cysts than in both type II patients and healthy women. IGFBP-1 levels were significantly lower in patients with type I than in controls and type II cysts. The IGF-I/IGFBP-1 ratio was significantly higher in patients with type I cysts than in type II bearers and healthy women. Estrogen levels correlated with IGF-I in patients and controls. Conclusions: The enhanced levels of IGF-I/IGFBP-3 found in patients with type I cysts could eventually be associated with the increased risk of breast cancer described for this group.Facultad de Ciencias Exacta

Correlation between inhibin secretion and damage of seminiferous tubules in a model of experimental autoimmune orchitis

The aim of the present study was to evaluate inhibin secretion in rats with autoimmune orchitis. As we have previously described, experimental autoimmune orchitis (EAO) induced in rats by active immunization with testis homogenate and adjuvants is characterized by an interstitial mononuclear cell infiltrate and sloughing of the germinal epithelium. At 120 days after the first immunization 60% of the rats exhibited a severe orchitis with large areas of aspermatogenic seminiferous tubules in which only spermatogonia and Sertoli cells with cytoplasmic vacuolization remained attached to the tubular wall. None of the untreated (N) or control (C) rats revealed pathological alterations. Sixty percent decrease in testis weight was observed in rats with EAO compared with N or C groups. A 3-fold increase in serum FSH levels was observed in rats with EAO compared with N or C groups (19·8 ± 3·7 vs 5·6 ± 0·3 and 5·9 ± 0·1 ng/ml respectively). A significant decrease in inhibin B levels was observed in rats with EAO when compared with N or C groups (40 ± 4·6 vs 207 ± 38·8 and 221·4 ± 28·6 pg/ml respectively). An inverse correlation between inhibin B and FSH serum levels and a direct correlation between inhibin B and testis weight were found. Strong expression of the inhibin α-subunit in Sertoli cells of untreated and control rats was observed; this subunit was undetectable or poorly detectable in rats with orchitis. Positive staining for the inhibin α-subunit was also observed in Leydig cells of all groups studied. In conclusion, using a model of autoimmune orchitis our results show that circulating inhibin B levels and inhibin α-subunit expression in Sertoli cell cytoplasm closely correlate with the degree of damage of the germinal epithelium.Instituto Multidisciplinario de Biología CelularFacultad de Ciencias Exacta