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-Reductase Inhibitor, on Rat Epididymal Sperm Maturation and Fertility1
Departments of Pharmacology and Therapeutics3 and of
Obstetrics and Gynecology,4 McGill University, Montréal, Quebec, Canada H3G 1Y6
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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-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 5-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 5-reductase inhibitors in further studies of epididymal physiology and as a potential component of a male contraceptive.
dihydrotestosterone, epididymis, inhibitor, male reproductive tract, maturation, sperm, sperm maturation, sperm motility and transport, steroid hormones
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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The maintenance of epididymal structure and functions is known to be highly dependent on the presence of androgens [10, 11]. Furthermore, several in vivo and in vitro studies have confirmed that it is not testosterone, but rather dihydrotestosterone (DHT), that is the main androgen acting in this tissue [12–15]. In contrast, testicular spermatogenesis is a testosterone-dependent process [16, 17, reviewed in 18]. Therefore, the inhibition of DHT production is an interesting experimental approach for studying the regulation of sperm maturation in the epididymis while presumably having little or no effect on sperm production in the testis.
5-Reductase (EC 1.3.1.22) is the enzyme that catalyzes the conversion of testosterone into DHT. Two isoforms of 5-reductase have been identified and are termed type 1 and type 2 [19, 20]; the contributions of each isozyme to male reproductive biology have not been fully elucidated. It is known that both 5-reductase isozymes are present in the epididymis, where they are differentially distributed and regulated [21]. Though the importance of each isozyme in the epididymis remains unclear, a synergistic or additive contribution of both isozymes to epididymal functions is a plausible likelihood. Supporting this concept is the observation of a more pronounced virilization defect in dual 5-reductase knockout mice when compared with the single 5-reductase type 2 knockout animals [22] as well as greater decreases in serum and prostate DHT levels when both isozymes are inhibited simultaneously [23–25].
The utility of 5-reductase type 2 inhibitors (e.g., finasteride) for treatment of androgen-dependent disorders, such as benign prostatic hyperplasia and alopecia, has been demonstrated [reviewed in 24–26]. Recently, drug-development efforts to create even more therapeutically effective compounds have led to the production and testing of a novel class of 5-reductase inhibitors, termed dual inhibitors, that concurrently inhibit both 5-reductase type 1 and type 2. PNU157706 is one such inhibitor developed for the potential treatment of benign prostatic hyperplasia. PNU157706, in vitro, has been shown to be a specific competitive inhibitor of epididymal 5-reductase activity and it decreases DHT-dependent tissue weights (i.e., seminal vesicle and ventral prostate) in vivo [23, 27]. Furthermore, PNU157706 was shown to be more potent than finasteride in decreasing DHT levels [23]. In a previous study, we have shown that treatment of adult male rats with PNU157706 has a highly segment-specific effect on epididymal gene expression [27]. Importantly, some of the more dramatically affected genes are potentially involved in essential processes contributing to the formation of an optimal luminal microenvironment required for proper sperm maturation. For example, genes involved in fatty acid and lipid metabolism, regulation of ion and fluid transport, luminal acidification, oxidative defense, and protein processing and degradation were affected [27].
Studying the consequences of inhibiting DHT formation is likely to provide important information regarding the androgen regulation of epididymal functions; however, very few recent studies have examined the effects of 5-reductase inhibition, specifically in the epididymis. The present study was designed to evaluate the effects of an altered androgen environment on epididymal sperm maturation by taking advantage of a novel drug and sperm analysis technology to determine the effects of 5-reductase inhibition on sperm morphology, motility, and fertilizing ability.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Adult Sprague-Dawley rats were obtained from Charles River Canada (St. Constant, PQ, Canada), maintained under controlled light (14L:10D) and temperature (22°C), and provided with food and water ad libitum. Male rats (325–350 g) were randomly divided into two groups and gavaged with 0.5 ml/kg of 0 (control) or 10 mg/kg (treated) PNU157706 (Pharmacia and Upjohn, Italy) suspended in 0.5% methylcellulose solution (BDH, Montreal, QC) containing 0.4% Tween 80 (A&C American Chemicals Ltd., Montreal, PQ, Canada) for 28 consecutive days. The dose was selected based on reported effects on rat ventral prostate; the dosing regimen with 10 mg/kg/day PNU157706 has been shown to decrease prostatic dihydrotestosterone levels by >90% [23]. Virgin female rats (200–250 g) were used for the mating study. 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.
Sperm Motility
All chemicals were purchased from Sigma Chemical Co. (St. Louis, MO) unless otherwise noted. At the time of killing, animals were anesthetized and seminal vesicles; testes; ventral prostates; and one epididymis, sectioned into initial segment, caput, corpus, and cauda segments; were collected and immediately frozen in liquid nitrogen. Tissues were stored at –80°C for future use. Sperm samples were collected from the distal caput and distal cauda of the remaining epididymis and used for computer-assisted sperm analysis (CASA) as previously described [28, 29], with the exception that the medium used for motility analysis was as follows: Hanks balanced salts solution (Gibco Invitrogen Co., Grand Island, NY), buffered with 4.2 g/L HEPES and 0.35 g/L NaHCO3 and containing 2.0 g/L BSA, 0.9 g/L D-glucose, 0.1 g/L sodium pyruvate, and 0.025 g/L soybean trypsin inhibitor, pH 7.4, at 37°C [30]. Briefly, the epididymis was trimmed free of fat and clamped at the caput-corpus and corpus-cauda junctions, then severed on the corpus side of the clamp and blotted. While still clamped, the caput and cauda epididymides were rinsed in separate 35-mm Petri dishes containing 2 ml of motility media and then transferred to separate Petri dishes containing 3 ml of fresh media. A #11 scalpel blade was used to pierce several tubules of the distal caput and distal cauda regions, allowing sperm to diffuse into the medium. The tissue was removed and sperm were allowed to disperse for approximately 5 min. Sperm concentrations were optimized before aliquoting into an 80-µm-deep glass cannula for CASA on the HTM-IVOS (Hamilton-Thorne Research, Beverly, MA) using version 12 of the Toxicology software. Approximately 3500 caput epididymal sperm and 6000 cauda epididymal sperm were analyzed for each treatment group (n = 5–7). The percentages of motile and progressively motile sperm and the following kinematic parameters were determined by CASA: average-path velocity (VAP), curvilinear velocity (VCL), straight-line velocity (VSL), amplitude of lateral head displacement (ALH), beat cross frequency (BCF), linearity (LIN = VSL/VCL x 100) and straightness (STR = VSL/VAP x 100).
Sperm Morphology
The morphology of spermatozoa from the cauda epididymidis was evaluated by phase-contrast microscopy using the HTM-IVOS. Sperm from the original dilution for motility analysis were fixed with 10% neutral buffered formalin (1:3, Sigma-Aldrich, Oakville, ON, Canada). Samples were aliquoted onto regular slides and sperm were analyzed using the 10x phase-contrast objective. This system allowed clear visualization of cytoplasmic droplets [28]. Approximately 300–500 sperm per sample were analyzed and the percentage of sperm retaining their cytoplasmic droplet was calculated. Similarly, broken sperm (head only, tail only, other breakages) and angulated sperm (bent at midpiece) were also clearly detectable using this system, and the frequency of these abnormalities was calculated.
In addition to the HTM-IVOS analysis of sperm morphology, spermatozoa from the cauda epididymidis of four control and four treated rats were also processed for electron microscopic analysis. A #11 scalpel blade was used to pierce the cauda several times and sperm were collected by shaking in Hanks minimum essential medium (Invitrogen Canada Inc., Burlington, ON, Canada). Spermatozoa were washed in Hanks media, fixed in the same media containing 1% glutaraldehyde (Mecalab Ltd., Montréal, PQ, Canada), and embedded for electron microscope analysis. Briefly, the samples were washed three times in sodium cacodylate buffer (0.1 M) containing 3% sucrose, pH 7.4, postfixed in 1% osmium tetroxide and 1.5% potassium ferrocyanide, and embedded in epoxy resin. Sperm ultrastructure was then analyzed on the electron microscope (Philips 410 electron microscope, Eindhoven, The Netherlands). For analysis of cytoplasmic droplet retention, a minimum of 100 midpiece sperm tail (flagellum) cross-sections per animal were photographed and the number of sperm tails with cytoplamsic droplets recorded.
Mating Study
A mating study was designed to determine the effects of PNU157706 treatment on male fertility. Vaginal smears from female rats were analyzed daily for a minimum of 2 wk to confirm normal estrous cycles. On the last day of treatment, each male rat was paired overnight with two female rats in proestrus. Female rats were examined the next morning for the presence of sperm in vaginal smears; this was defined as Day 0 of gestation for sperm-positive animals. The male reproductive indices that were calculated are: copulation index (number of sperm-positive females/number of pairings), pregnancy index (number of pregnancies/number of sperm-positive females), and fertility index (number of pregnancies/number of pairings).
Pregnancy Outcome
To examine the effect of inhibitor treatment on pregnancy outcome, female rats underwent a cesarean section on Day 20 of gestation. The ovaries were removed and the number of corpora lutea was counted; uteri were opened, and the numbers of implantation sites were counted. The status of each implantation site was classified as resorption, dead or live fetus. For each treatment group, the overall implantation rate (number of implantations/number of corpora lutea), preimplantation loss per female ([number of corpora lutea – number of implantation sites]/number of corpora lutea), overall resorption rate (number of resorptions/number of total implantations), and postimplantation loss per female ([number of implantations – number of live fetuses]/number of implantations) were calculated. Fetuses were sexed, weighed, and examined for external malformations.
Statistical Analysis
Statistical analysis of frequency data (reproductive indices, fetal sex ratio) was done by the Fisher exact test. The implantation and resorption rates were analyzed by the test of proportions (Z-test). The remaining data assessing male fertility and pregnancy outcome, sperm motility, and sperm morphology were analyzed for differences between the control and treated groups by Student t-test or the Mann-Whitney U-test (in cases of failed normality or equal variance tests). Data are presented as mean ± SEM. For all analyses, the level of significance was set at P < 0.05.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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In the control group, a pattern of spermatozoal motility from circular, in the caput epididymidis, to progressively forward, in the cauda epididymidis, was observed. This progression was reflected in the sperm motion parameters measured by CASA where sperm from the cauda epididymides had higher VSL and STR (Table 1), as well as an increase in the percent that was progressively motile (Fig. 1). For sperm taken from the caput epididymidis, there was no effect of PNU157706 treatment on any of the motion parameters (Table 1) or on the percentage of motile and progressively motile sperm (Fig. 1). In contrast, treatment with the 5-reductase inhibitor caused a significant decrease in the VSL, STR, and LIN of sperm from the cauda epididymidis (Table 1). There was also a significant decrease in the percentage of motile and progressively motile sperm (approximately 12.5% and 20%, respectively) from the cauda epididymides of PNU157706-treated rats when compared with control rats (Fig. 1).
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Effects of PNU157706 Treatment on Sperm Morphology
A major morphological feature of epididymal sperm maturation is the displacement of the cytoplasmic droplet along the midpiece of sperm traveling from the caput to the cauda epididymal regions [7, 31]. Analysis of sperm from the cauda epididymidis under phase-contrast microscopy using the CASA system allowed clear visualization of the cytoplasmic droplet (Fig. 2A). As expected, a low percentage (approximately 12%) of sperm from this segment of the tissue of control males retained their cytoplasmic droplet. However, sperm samples from the cauda epididymides of males treated with PNU157706 had a significantly higher percentage of retained cytoplasmic droplets compared with controls, doubling to nearly 25% (Fig. 2B).
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Some sperm abnormalities, such as bent (angulated) sperm and sperm breakages (i.e., headless, tailless), were also easily visualized using the CASA system; examples of these defects are shown in Figure 3. Sperm samples from the cauda epididymides of control and treated males had similar percentages of bent sperm (Table 2). In contrast, there was a higher incidence of sperm breakage in sperm from the cauda epididymides of males treated with the inhibitor; the percentages of headless, tailless, and other breaks all increased significantly with treatment, as did the percentage of overall abnormalities observed using CASA (Table 2).
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Analysis of the ultrastructure of sperm from the cauda epididymidis revealed an apparent increase in the proportion of midpiece spermatozoa having cytoplasmic droplets; examples of sperm cross-sections with and without the droplet are shown in Figure 4. The droplet appears as a membrane-bound structure surrounding the sperm tail. There was a significant effect of treatment on the percentage of midpiece sperm cross-sections surrounded by cytoplasmic droplets (Fig. 4C). This is consistent with our CASA analysis of cytoplasmic droplet retention in sperm from the cauda epididymidis. Very few ultrastructural abnormalities were observed in sperm samples from either control or treated rats.
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Effects of PNU157706 Treatment on Male Fertility
In this study, each male (n = 8/treatment group) was paired with two virgin females in proestrus, for a total of 16 pairings for each treatment group. The effects of 5-reductase inhibitor treatment on indices of male reproduction are shown in Table 3. All of the control males successfully mated with at least one female, with the majority mating with both females. All of these successful matings resulted in pregnancies. In contrast, there was a decrease in the number of sperm-positive (successfully mated) females exposed to PNU157706-treated males. In some cases, approximately 50 or more sperm were present and easily detected in the vaginal smears of females exposed to treated males (comparable with controls), while in other cases, as few as 2 sperm were detected. Furthermore, not all of the sperm-positive females exposed to treated males became pregnant. Overall, the number of females with successful pregnancies that resulted from pairings with treated males was significantly reduced.
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Effects of PNU157706 Treatment on Pregnancy Outcome
As expected, females exposed to either control or PNU157706-treated males had similar numbers of corpora lutea (17.1 ± 0.7 and 17.2 ± 1.8, respectively). In contrast, the implantation rate, reflecting the number of released ova that were successfully fertilized and implanted in the uterus, was significantly decreased in females exposed to treated males (80.6%) compared with those exposed to control males (95%). Approximately half of the pregnant females that were mated with control males had some degree of preimplantation loss (7/13) compared with 83% of the pregnant females exposed to treated males (5/6). The mean preimplantation loss per litter increased from 5.3% in females exposed to control males to nearly 19% in females exposed to treated males (Fig. 5). Upon examination of the data for individual pregnant females, it is apparent that there is a heterogeneous effect of treatment on preimplantation loss. Of the females mated with PNU157706-treated males, three had low rates of preimplantation loss (<15%) that were comparable with females mated with control males. Of the three remaining females, one had a higher rate of preimplantation loss (>18%) while two females had exceedingly high preimplantation loss at approximately 37% and 41%. In addition, three females exposed to treated males showed evidence of successful copulation (sperm-positive vaginal smears) that failed to result in pregnancy. This occurrence can be considered as 100% preimplantation loss due to failure of sperm to fertilize the released ova. Analysis of preimplantation loss based on the number of sperm-positive females showed a significant increase in females exposed to males treated with PNU157706 (nearly 46%) when compared with control-mated females.
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There was no significant effect of inhibitor treatment on postimplantation loss (Fig. 5). There were no late fetal deaths (nonlive fetuses) observed in this study; therefore, the postimplantation loss observed was due solely to resorptions. Approximately half of the pregnant females exposed to either control or treated males had resorptions (control 6/13; treated 3/6), and the resorption rate was not significantly higher for those exposed to treated males.
The fetuses sired by control and treated males showed no gross external malformations. There was no difference in ano-genital distance between fetuses sired by PNU157706-treated males (male 3.69 ± 0.12 mm; female 1.97 ± 0.02 mm) and control males (male 3.92 ± 0.04 mm; female 1.95 ± 0.02 mm). The sex ratio (male/female) was also unaffected by treatment (control 96/107; treated 39/40). Fetuses sired by males treated with PNU157706 did not differ in weight (males 3.95 ± 0.36 g; females 3.81 ± 0.41 g) when compared with those sired by control males (males 4.23 ± 0.23 g; females 3.98 ± 0.23 g).
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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-reductase inhibitor, was first marketed for the treatment of benign prostatic hyperplasia in 1992 [32]. Consequently, the long-term efficacy and safety of finasteride have been extensively studied for this indication [reviewed in 24, 25]. Finasteride treatment decreases serum and prostate DHT levels by approximately 65–70% and 85–90%, respectively [25]. Efforts to discover more therapeutically effective compounds, i.e., that decrease DHT levels to a greater extent, have led to the development of a novel class of dual 5-reductase inhibitors. In early 2003, dutasteride (GI198745) became the first dual 5-reductase inhibitor available for the treatment of BPH [33]. In comparison with finasteride, dutasteride and other dual compounds achieve almost total suppression of DHT levels [23, 33, 34]. There are a limited number of studies that examine the effects of finasteride on other reproductive tissues and/or fertility [35–38] and fewer still that examine the extraprostatic effects of the novel class of dual inhibitors [33, 34]. In 1991, Cukierski et al. [35] reported a partial decrease in fertility following chronic finasteride treatment (80 mg/kg/ day for 24 wk) of adult male rats. Wise et al. [36] reported a similar effect in a related study. In these earlier studies, testis weight and histology were unaffected by treatment and the subfertility was attributed solely to a deficit in the formation of copulatory plugs due to the effects of treatment on seminal vesicle and prostate weights. However, any data pertaining to the epididymis were lacking; therefore, it is possible that the compromised fertility could have been due to an effect on this tissue as well.
In recent years, Mahendroo et al. [22] have developed and studied type-specific and dual 5-reductase null mutation mice. The dual knockout mice were fertile (pregnancy rates of 60% compared with 96% for wild-type animals). Unexpectedly, type 2 or dual knockout male mice showed only a mild virilization defect such that their internal and external genitalia were fully formed, but their prostates and seminal vesicles were smaller. These findings were markedly different from the phenotype of type 2 5-reductase deficiency in humans that results in a form of male pseudohermaphroditism [for review, see 39]. The authors concluded that, in mice, testosterone alone is required for differentiation of the male urogenital tract and that the synthesis of DHT serves largely as a signal amplification mechanism. In contrast, both testosterone and DHT are required for proper development of the reproductive system in men, and studies in male rats with pharmacological inhibitors also support a two-androgen model of sexual differentiation [40–42]. Thus, the rat appears to be a more suitable animal model for understanding the roles of DHT within the human male reproductive system and comparisons with the mouse should be made with caution.
With the recent novel use of finasteride for the treatment of androgenic alopecia in young men of reproductive age [26] and the emergence of dual inhibitors that decrease DHT levels to a greater extent [33, 34], a renewed focus on the effects of 5-reductase inhibition on the testis and fertility has emerged. To date, no adverse effects of 5-reductase inhibitor treatment on testis weight, histology, and/or sperm counts, and hence spermatogenesis, have been reported [24, 38, 43, 44], nor do 5-reductase knockout mice show any testicular phenotype [22]; however, epididymal studies still remain scarce. Given the important role of the epididymis in sperm maturation and the dependence of this tissue on DHT, a closer examination of the effects of 5-reductase inhibition on epididymal sperm was considered necessary.
In the current study, we show that PNU157706 treatment of adult male rats for 28 days resulted in decreased fertility. It is unlikely that the effects of PNU157706 treatment on male fertility were due to an effect on sexual behavior since 100% of the treated males showed evidence of successful copulation and 75% sired at least one litter. Current evidence does not suggest a role for DHT in regulating sexual behavior. While the physiological functions of brain 5-reductase activity have not been fully elucidated, evidence points to roles predominantly during brain ontogenesis and the sexual differentiation of specific brain regions as well as possible anesthetic/anxiolytic actions evoked when progesterone and/or corticoid levels are high, i.e., during stress, and neuroprotective roles mediated via the catabolism of potentially neurotoxic levels of steroids [reviewed in 45]. Additionally, the very low incidence of sexual side effects in men treated with finasteride or the dual inhibitor dutasteride provides support for a minimal or nonexistent role for DHT in regulating sexual behavior [32, 33].
Examination of pregnancy outcome demonstrated an effect of treatment on implantation rate and preimplantation loss per pregnant female. Preimplantation loss represents either failure of sperm to fertilize the released ova or the death of the fertilized embryo before implantation. The latter suggests involvement of genetic damage (chromosomal aberrations) leading to early death and is proposed as the reason for increased preimplantation loss and decreased fetal weight seen with increased paternal age and long-term paternal exposure to the alkylating agent cyclophosphamide [46, 47]. It is unlikely that the genetic integrity of spermatozoa in this study is compromised because the testis, the site of continuous sperm cell division, has not been shown to be affected by 5-reductase inhibitor treatment [24, 38, 43, 44]. Furthermore, treatment had no effect on any of the fetal parameters measured, including fetal weight, which might be indicative of genetic impairment, nor were there any abnormal or dead fetuses sired by treated males. It is more plausible that the sperm were unable to reach and fertilize the released ova due to compromised sperm motility and/or morphology. Supporting this is the observation of fewer sperm in the vaginal smears of some females mated with treated males and a lower pregnancy index.
We showed a decrease in both overall motility and progressive motility of sperm from the cauda epididymidis, accompanied by decreases in some motion parameters. The sperm tail encompasses the structural components directly involved in sperm motility [for review, see 48]; consequently, compromised tail ultrastructure could lead to impaired motility. In the present study, however, electron microscopic analysis of midpiece flagellar ultrastructure revealed no effects of treatment that could account for the reduced sperm motility. Again, these findings are consistent with a lack of effect of 5-reductase inhibitor treatment on the testis, where sperm ultrastructure is established. The acquisition of sperm motility is a key element of epididymal sperm maturation [3]. By the time sperm reach the distal regions of the epididymis, maximum progressive motility is achieved that enables sperm to reach and penetrate the egg [3, 49]. This transition to progressive motility requires interactions between sperm and the surrounding epididymal luminal environment [3, 4, 50]. In fact, in reproductive toxicology studies, altered sperm motility is a valuable indicator of toxicity arising from an exclusive effect on the epididymis or sperm within the epididymis [51]. In the current study, it is likely that the reduced sperm motility observed is due to exposure of sperm to a compromised epididymal luminal environment. Notably, in a previous study, we have shown that PNU157706 treatment affects the segment-specific expression of many genes in the epididymis potentially involved in the creation of the luminal environment that is critical for proper sperm maturation [27].
While the characteristic head, midpiece, and tail structures of spermatozoa are already present before sperm leave the testis, other morphological changes occur during sperm transit through the epididymis. One prominent change is the migration of the cytoplasmic droplet along the midpiece of the sperm and its eventual shedding in the distal regions of the epididymis [7, 31]. The percentage of sperm retaining their cytoplasmic droplet was increased in rats treated with PNU157706. Interestingly, while the reason for shedding of the droplet remains unclear, some evidence suggests that cytoplasmic droplet retention can be correlated with altered epididymal function and decreased fertility [52–55]. Shedding of the droplet is thought to be related to the well-characterized changes in sperm plasma membrane lipid composition and fluidity that occur during epididymal transit [5, 56]. It is tempting to speculate that the increased cytoplasmic droplet retention and possibly the increased sperm breakages observed in the current study are due to altered sperm membrane composition and dynamics arising from exposure to a suboptimal epididymal milieu. In fact, we have shown that PNU157706 treatment alters the epididymal expression of genes involved in oxidative defense that protect the highly susceptible sperm membrane lipids from oxidative damage [27]. Indeed, proper redox chemistry is essential within the epididymis as it regulates pathways involved in important sperm functions such as motility and sperm-egg interaction [57]; thus, any perturbation of the redox system likely has manifold consequences on epididymal sperm. Additionally, PNU157706 treatment also alters the expression of genes involved in fatty acid and lipid metabolism within the epididymis [27]. Of relevance to the current study is the reported reproductive impairment observed in mouse models of sphingolipid storage disorders (Niemann-Pick, Tay-Sachs, and Sandhoff diseases) that exhibit gross lipid accumulation within cells and tissues, including those of the male reproductive tract [58– 60]. Interestingly, there do not appear to be any changes in testis weight, morphology, or sperm counts in these animals; hence, the reproductive pathologies are believed to result from lipid accumulation in lysosomes of the epididymis that give rise to a dysfunctional sperm maturation environment. These models not only highlight the importance of lipid regulation within the epididymis but also clearly support an important role for the epididymis in sperm-related functions that are unique and separate from the testis.
In conclusion, while an effect on sperm fertilizing ability was reported in earlier 5-reductase inhibitor studies, to our knowledge, this is the first study that clearly demonstrates an effect of inhibitor treatment on sperm maturation in the epididymis. It is highly likely that the compromised sperm motility and morphology observed in the current study contribute to the subfertility observed in 5-reductase inhibitor-treated rats. Furthermore, our results support a role for both 5-reductase isozymes within the epididymis and indicate the potential of dual 5-reductase inhibition as an experimental model for more in-depth study of epididymal sperm-related functions separate from testicular sperm-related functions.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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2 Correspondence: B. Robaire, Department of Pharmacology and Therapeutics, McGill University, 3655 Promenade Sir-William-Osler, Montréal, QB, Canada H3G1Y6. FAX: 514 398 7120; bernard.robaire{at}mcgill.ca
Received: 23 June 2004.
First decision: 13 July 2004.
Accepted: 23 September 2004.
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REFERENCES |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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-androstan-3-one (dihydrotestosterone) by rat prostate and epididymis. J Clin Endocrinol Metab 1969 29:970-977[Medline]-reductase-2 deficiency. Mol Cell Endocrinol 2002 198:51-59[CrossRef][Medline]-reductase. Endocrinology 1985 16:807-812-reductase inhibitor on androgen physiology in the immature male rat. Endocrinology 1989 125:2434-2438-reductase inhibitor-induced abnormalities. Teratology 1990 42:483-496[CrossRef][Medline]This article has been cited by other articles:
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  T. Saether, T. N Tran, H. Rootwelt, H. J Grav, B. O Christophersen, and T. B Haugen Essential fatty acid deficiency induces fatty acid desaturase expression in rat epididymis, but not in testis Reproduction, February 1, 2007; 133(2): 467 - 477. [Abstract] [Full Text] [PDF]
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 N. A. Henderson, G. M Cooke, and B. Robaire Region-specific expression of androgen and growth factor pathway genes in the rat epididymis and the effects of dual 5{alpha}-reductase inhibition. J. Endocrinol., September 1, 2006; 190(3): 779 - 791. [Abstract] [Full Text] [PDF]
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