Cimetidine-induced androgenic failure causes cell death and changes in actin, EGF and V-ATPase immunoexpression in rat submandibular glands.

Submandibular gland (SMG) is responsive to androgens via androgen receptor (AR). We verified whether cimetidine induces androgenic dysfunction in SMG, and evaluated the structural integrity, cell death and immunoexpression of actin, EGF and V-ATPase in androgen-deficient SMG. Male rats received cimetidine (CMTG) and control animals (CG) received saline. Granular convoluted tubules (GCTs) diameter and number of acinar cell nuclei were evaluated. TUNEL and immunofluorescence reactions for detection of AR, testosterone, actin, EGF and V-ATPase were quantitatively analysed. In CG, testosterone immunolabelling was detected in acinar and ductal cells cytoplasm. AR-immunolabelled nuclei were observed in acinar cells whereas ductal cells showed AR-immunostained cytoplasm, indicating a non-genomic AR action. In CMTG, the weak testosterone and AR immunoexpression confirmed cimetidine-induced androgenic failure. A high cell death index was correlated with decreased number of acinar cells, GCTs diameter and EGF immunoexpression under androgenic dysfunction. Actin immunofluorescence decreased in the SMG cells, but an increased and diffuse cytoplasmic V-ATPase immunolabelling was observed in striated ducts, suggesting a disruption in the actin-dependent V-ATPase recycling due to androgenic failure. Our findings reinforce the androgenic role in the maintenance of SMG histophysiology, and point to a potential clinical use of cimetidine against androgen-dependent glandular tumour cells.

Venlafaxine-induced damage to seminiferous epithelium, spermiation, and sperm parameters in rats: A correlation with high estrogen levels.

BACKGROUND: Venlafaxine (selective serotonin and norepinephrine reuptake inhibitor) use has increased worldwide. However, the impact of venlafaxine on testes and sperm parameters has not been investigated. OBJECTIVES: We evaluated venlafaxine impact on testicular and sperm parameters and verified whether the changes are reversible. METHODS: Animals from venlafaxine-35 days and venlafaxine-65 days groups received 30 mg/kg of venlafaxine for 35 days. Control-35 days and control-65 days received distilled water. In control-65 days and venlafaxine-65 days, the treatment was interrupted for 30 days. Sperm concentration, morphology, motility, and mitochondrial activity were analyzed. Number of step 19 spermatids (NLS), frequency of tubules with spermiation failure, Sertoli cells number, and TUNEL-positive germ cells were quantified. Testicular aromatase, connexin 43 (Cx43) immunoexpression, Cx43 protein levels, and Cx43 expression were evaluated. Either intratesticular testosterone or estrogen levels were measured. RESULTS: Venlafaxine impaired sperm morphology, reduced sperm concentration, mitochondrial activity, and sperm motility. The frequency of tubules with spermiation failure and NLS increased in parallel to increased Cx43 immunoexpression; mRNA and protein levels; and aromatase, testosterone, and estrogen levels. An increase in germ cell death and decreased Sertoli cells number were observed. In venlafaxine-65 days, except for sperm motility, mitochondrial activity, Sertoli cells number, and germ cell death, all other parameters were partially or totally recovered. CONCLUSION: Venlafaxine increases testosterone aromatization and Cx43. This drug, via high estrogen levels, disturbs Sertoli cells, induces germ cell death, and impairs spermiation and sperm parameters. The restoration of spermiation associated with the decreased Cx43 and hormonal levels in venlafaxine-65 days reinforces that high estrogen levels are related to venlafaxine-induced changes. The presence of damaged Sertoli cells, germ cell death, and low sperm motility in venlafaxine-65 days indicates that interruption of treatment for 30 days was insufficient for testicular recovery and points to a long-term estrogen impact on the seminiferous epithelium.

The antineoplastic busulphan impairs peritubular and Leydig cells, and vitamin B12 stimulates spermatogonia proliferation and prevents busulphan-induced germ cell death.

Busulphan (Bu), an alkylating agent used for bone marrow and spermatogonial stem cell transplantation (SSCT), impairs Sertoli (SC) cells, which are necessary for the spermatogonial stem cell (SSC) homing during transplantation. As Leydig (LC) and peritubular myoid (PMC) cells are essential for SC support and maintenance of spermatogonial niche, we evaluated the impact of Bu on the LC and PMC structural integrity. Vitamin B12 (B12) has demonstrated beneficial effects against drug-induced testicular changes; thus, we also examined whether this vitamin is able to stimulate spermatogonia mitotic activity and prevent Bu-induced germ cell death. Rats received 10mg/kg of Bu in the 1st and 4th days, and daily B12 supplementation during Bu treatment and for 6days after the last injection of Bu (Bu-6d), totaling 10days of treatment. Other animals received the same treatment as Bu-6d, and B12 supplementation (Bu+7dB12) or saline (Bu+7dS) for 7 more days, totaling 17days of treatment. Serum testosterone levels were measured. In the historesin-embedded testis sections, the seminiferous tubule and epithelial areas were measured, and the number of spermatogonia and PMC was quantified. Actin and 17ß-HSD6 immunofluorescence was detected, and the number of TUNEL-positive LC and germ cells was computed. In Bu-6d, PMC number reduced, and a weak actin immunoexpression and death in these cells was observed. The testosterone levels reduced, and the interstitial tissue showed a weak 17ß-HSD6 immunoexpression and increased number of TUNEL-positive LC. In Bu+7dB12, the number of spermatogonia was higher than in Bu-6d and Bu+7dS, and the number of TUNEL-positive germ cells was significantly lower than in Bu+7dS. Bu exerts a harmful impact on PMC and LC, reducing the testosterone levels. Vitamin B12 prevents significantly Bu-induced germ cell death and stimulates spermatogonia proliferation, being a useful strategy for the enrichment of SSC in vitro and an adjuvant therapy for spermatogenesis recovery in oncologic patients.

Vitamin B12 supplement exerts a beneficial effect on the seminiferous epithelium of cimetidine-treated rats.

Treatment of gastric ulcer with cimetidine reduces acid secretion and interferes in the vitamin B(12) absorption. Regarding the harmful effect of cimetidine on the seminiferous tubules, the aim of the present study was to verify if prolonged treatment with cimetidine causes vitamin B(12) deficiency and whether the testicular damages are attenuated by vitamin B(12) supplementation. Adult male rats received, for 50 days, cimetidine (CMTG), cimetidine and vitamin B(12) (CMT/B(12)G), vitamin B(12) (B(12)G) and saline solution (CG). Vitamin B(12) and homocysteine plasma levels were evaluated and the testes were embedded in glycol methacrylate for the morphometric analyses of total, epithelial and luminal areas of the seminiferous tubules, number of Sertoli cells and frequencies of tubules according to stages and containing Sertoli and germ cells in the lumen. Terminal deoxynucleotidyl-transferase mediated dUTP nick end labeling (TUNEL) method and proliferating cell nuclear antigen (PCNA) immunohistochemistry were carried out. CMTG showed TUNEL-positive Sertoli cells and significant reductions in the epithelial and total tubular areas, number of Sertoli cells and frequency of tubules VII-VIII. In the CMT/B(12)G, the number of Sertoli cells and the epithelial and total tubular areas were similar to CG. The number of Sertoli cells (in B(12)G) and the frequency of tubules at stages VII-VIII (in B(12)G and CMT/B(12)G) increased significantly; PCNA-positive Sertoli cells were found in these groups. Although cimetidine was not able to induce vitamin B(12) deficiency, this drug causes tubular atrophy due to Sertoli cell damage and loss of germ cells. However, vitamin B(12) supplement is able to stimulate spermatogenesis and restore the number of Sertoli cells, softening the harmful effect of cimetidine on spermatogenesis.