No abstract

Dihydrotestosterone (DHT) is the primary androgen acting in the epididymis, the site of sperm maturation. Previously, we showed that the treatment of male rats with PNU157706, an inhibitor that acts on both isoforms of 5a-reductase to prevent DHT formation, has effects on the expression of genes implicated in processes that create the optimal luminal microenvironment required for sperm maturation, and on sperm maturation itself. However, signaling pathways involved in regulating or mediating DHT actions in the epididymis remain largely unknown. The goals of this study were to determine the expression profiles of potential signaling systems in the epididymis and assess their DHTdependence using two different dual 5a-reductase inhibitors. Rats were untreated or gavaged with vehicle, 10 mg/kg per day PNU157706 or 32 mg/kg per day FK143 for 28 days and epididymal gene expression was analyzed. Gene array analysis revealed analogous effects of FK143 on overall epididymal gene expression when compared with previous PNU157706 studies. Quantitative RT-PCR analysis of the expression of the 5a-reductase isozymes, androgen receptor, and members of the IGF, FGF, TGF, and VEGF families revealed novel region-specific expression profiles in the epididymis that were differentially affected by 5a-reductase inhibition; the two inhibitors had parallel effects. Specifically, in proximal regions, 5a-reductase 1, androgen receptor, and TGF-b1 expression increased after treatment, while in distal regions expression of IGF-I, IGFBP-5, IGFBP-6, and FGF-10 decreased. These results provide insight into epididymal signaling mechanisms and indicate potential candidates acting either upstream or downstream of DHT to regulate and/or mediate its actions in the epididymis.

Sperm entering the epididymis gain progressive motility and fertilizing ability in a process termed maturation. The functional dependence of the epididymis on dihydrotestosterone (DHT) is well established, yet few studies have examined the consequences on the epididymis of inhibiting DHT formation. We have shown that inhibition of both isoforms of 5alpha-reductase (types 1 and 2), the enzyme that converts testosterone to DHT, has pronounced effects on epididymal gene expression. In the present study, we investigate whether inhibiting 5alpha-reductase has consequences on epididymal sperm maturation. Rats were treated with vehicle or 10 mg/kg/day PNU157706, a dual-type inhibitor, for 28 days. Fertility and several key facets of sperm maturation were analyzed. Changes in sperm motility were assessed by computer-assisted sperm analysis (CASA). Changes in sperm morphology were assessed by CASA and electron microscopy. The motility of spermatozoa from the cauda epididymidis of treated animals showed a significant decrease in both the percentage of motile and progressively motile sperm as well as altered motion parameters. The morphology of cauda epididymal spermatozoa was also adversely affected by the treatment; the most prominent effect was a markedly elevated proportion of sperm that retained their cytoplasmic droplet. Matings with treated males resulted in fewer successful pregnancies and a higher rate of preimplantation loss. Progeny outcome was unaffected. The compromised sperm motility and morphology likely contribute to the subfertility of inhibitor-treated rats. Our results indicate a role for dual 5alpha-reductase inhibitors in further studies of epididymal physiology and as a potential component of a male contraceptive.

The epididymis is the site of sperm maturation and storage. 5 -Reductases (types 1 and 2) are key enzymes in this tissue because they convert testosterone to dihydrotestosterone (DHT), the main androgen regulating epididymal functions. Examining the consequences of inhibiting DHT formation is likely to provide important information regarding the regulation of epididymal functions, yet few inhibitor studies have focused on this tissue. To understand better DHT-mediated regulation of epididymal gene expression, we employed a dual 5 -reductase inhibitor and cDNA microarrays to examine the effects of 5 -reductase inhibition on gene expression in the initial segment, caput, corpus, and cauda epididymidis. Inhibition of epididymal 5 -reductase activity by PNU157706 was confirmed by in vitro enzyme assays. Rats were treated with 0, 0·1, 1·0 or 10 mg/kg per day PNU157706 for 28 days. The weights of DHT-dependent tissues, including the epididymis, were decreased following treatment. The effect of treatment on gene expression was dose-dependent and highly segment-specific. The initial segment responded uniquely in that a similar number of genes increased and decreased in expression compared with the other segments where the majority of affected genes decreased in expression. Some of the more dramatically affected genes were involved in signal transduction as well as fatty acid and lipid metabolism, regulation of ion and fluid transport, luminal acidification, oxidative defense and protein processing and degradation. These are essential processes contributing to the formation of an optimal luminal microenvironment required for proper sperm maturation. These results provide a novel insight into the DHT-dependent mechanisms that control epididymal functions.