Effects of PNU157706, a dual 5alpha-reductase inhibitor, on gene expression in the rat epididymis

Abstract

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.

Region-specific expression of androgen and growth factor pathway genes in the rat epididymis and the effects of dual 5α-reductase inhibition

“…The tissues were stored at K80 8C until used for RNA extraction. The changes in reproductive tissue weights with treatment were consistent with what has been reported previously for both compounds (Hirosumi et al 1995a,b, di Salle et al 1998, Henderson et al 2004. All animal studies were conducted in accordance with the principles and procedures outlined in the Guide to the Care and Use of Experimental Animals prepared by the Canadian Council on Animal Care (McGill Animal care Committee protocol no.…”

“…Frozen caput-corpus epididymides (only control, untreated tissues were used) were thawed, homogenized (Polytron) in DMG buffer (50 mM DMG, 5% glycerol, NaOH, and 0$5 mM NADPH) pH 6$5, and the nuclear and microsomal fractions were assayed for 5a-reductase activity as previously described for PNU157706 (Henderson et al 2004). Briefly, kinetic studies of 5a-reductase activity were done at pH 6$5 with testosterone (nine concentrations ranging from 5 to 375 nM).…”

“…Previously, using gene expression profiling, we have shown that the treatment of adult male rats with the dual 5a-reductase inhibitor, PNU157706, has a highly regionspecific effect on epididymal gene expression (Henderson et al 2004). The affected genes are involved in fatty acid and lipid metabolism, regulation of ion and fluid transport, luminal acidification, oxidative defense, and protein processing and degradation; these are essential processes that contribute to the formation of the optimal luminal microenvironment (Henderson et al 2004).…”

“…The affected genes are involved in fatty acid and lipid metabolism, regulation of ion and fluid transport, luminal acidification, oxidative defense, and protein processing and degradation; these are essential processes that contribute to the formation of the optimal luminal microenvironment (Henderson et al 2004).…”

“…The cumulative array data from this study and our previous PNU157706 study (Henderson et al 2004), as well as the present literature, were used as a guide in the selection of genes potentially involved in epididymal regulatory mechanisms for further characterization in the epididymis. The genes chosen include, both the isozymes of 5a-reductase and the androgen receptor, which are directly involved in mediating androgen action in the epididymis, as well as those involved in different growth factor signaling systems.…”