5α-reductase Isoforms Research Articles

Insight on the intracrinology of menopause: androgen production within the human vagina

ABSTRACT Introduction The human vagina is extremely sensitive to hormonal changes occurring during the woman's lifetime, in particular the decline in estrogens that characterizes the menopause is known to induce a range of genital and sexual symptoms (dryness, burning, irritation, lack of lubrication, discomfort, pain), known as “genitourinary syndrome of menopause” (GSM). However, this reduction in estrogen levels is also accompanied by a progressive ageing-dependent decline in androgens for which the role of androgens in the development and treatment of GSM has received more attention. It is known that peripheral tissues compensate for circulating hormonal imbalances through the in-situ production of small quantities of sex hormones deriving from inactive precursors metabolized by specific cellular enzymes. This non-classical production and local activity of hormones is defined as “intracrinology”. Objective The aim of the study was to investigate whether the human vagina possesses the specific enzymatic machinery to convert weak androgen precursors, such as DHEA, into active androgens [Δ4-androstenedione (A), testosterone (T) and dihydrotestosterone (DHT)] as already described in other species. Methods In this study, a well established model of human smooth muscle cells (hSMCs), isolated from vagina tissues of women undergoing surgery for benign gynecological diseases, was used. Semi-quantitative RT-PCR was employed in order to evaluate the mRNA expression of the steroidogenic markers in hSMCs and corrispective tissue. The production of androgens was quantified in hSMCs medium by liquid chromatography tandem-mass spectrometry (LC-MS/MS). Results In vaginal tissues, AR mRNA was significantly less expressed than estrogen receptor α (ERα), whereas in hvSMCs, its mRNA expression was higher than progestin and both estrogen receptors. In hvSMCs and in vaginal tissue, when compared to ovaries, the mRNA expression of pro-androgenic steroidogenic enzymes (HSD3β1/β2, HSD17β3/β5), along with 5α-reductase isoforms and sulfo-transferase, resulted as being more abundant. In addition, enzymes involved in androgen inactivation were less expressed than in the ovaries. The LC-MS/MS analysis revealed that, in hvSMCs, short term DHEA supplementation increased Δ4-androstenedione levels in spent medium, while increasing testosterone (T) and dihydrotestosterone (DHT) secretion after longer incubation. Finally, androgenic signaling activation was evaluated through AR-dependent marker mRNA expression, after DHEA and T stimulation. Conclusions This study provides evidence for the presence of steroidogenic machinery in the human vagina, supporting, for the first time, the construct of intracrinology in this organ, emphasizing that the vagina is an androgen-dependent organ. These observations suggest the potential mechanism and resulting action of intravaginally administered DHEA, recently indicated for the treatment of GSM and open new perspectives for the treatment of this common and bothersome condition. Disclosure Work supported by industry: no.

Dihydrotestosterone through blockade of TGF-β/Smad signaling mediates the anti-fibrosis effect under hypoxia in canine Sertoli cells

The hypoxic microenvironment of cryptorchidism is an important factor to induce the impairment of the structure and function of Sertoli cells and thus lead to spermatogenesis loss or tumorigenesis. Dihydrotestosterone (DHT), as a potent nonaromatizable 5α-reduced androgen, has both positive and negative effect on pathological fibrosis process. However, it is still unknown whether DHT can regulate hypoxia-induced fibrosis of Sertoli cells. Herein, in this study, we evaluate the DHT level, two 5α-reductase isoforms, 5α-red1 and 5α-red2, as well as HIF-1α expression pattern in canine cryptorchidism and contralateral normal testis. Results showed that the abdominal testes presented low DHT levels and 5α-red1 and 5α-red2 expression, while significantly higher HIF-1α expression and ECM production compared with the scrotum. Moreover, we established a hypoxia-induced fibrosis model in canine Sertoli cells induced by cobalt chloride (CoCl2), and found that DHT inhibited the fibrosis of Sertoli cells in a dose-dependent manner. Meanwhile, DHT interfered with the TGF-β signaling by reducing the expression of TGF-βRI and TGF-βRII and inhibiting the expression and phosphorylation of Smad2 and Smad3, while flutamide (androgen receptor inhibitor) inhibited these effects of DHT. Furthermore, use of LY2109761 (TGF-β receptor type I/II inhibitor) to interfere with the TGF-β/Smad pathway showed a similar effect with DHT suppression of the fibrosis in Sertoli cells. Our research data demonstrated that cryptorchidism is located in a hypoxic and DHT deficiency microenvironment. Moreover, supplementing DHT can alleviate the fibrosis process of Sertoli cells caused by hypoxia, which is associated with AR regulating the inhibition of TGF-β/Smad signaling.

LC-MS/MS determination of dutasteride and its major metabolites in human plasma

Dutasteride is a specific and selective inhibitor of both 5α-reductase isoforms used mainly in benign prostatic hyperplasia and lower urinary tract symptoms. Although the drug is extensively metabolized in humans, data on the concentrations of its main metabolites are lacking. There is also a lack of data on dutasteride stability in frozen plasma samples. Our method was used to determine dutasteride and its active metabolites: 4′-hydroxydutasteride, 6β-hydroxydutasteride, and 1,2-dihydrodutasteride in plasma after a single administration of 0.5 mg of dutasteride. We also assessed the long-term stability (two years in the freezer) of dutasteride in clinical samples. The developed method covered the range of 0.1–3.5 ng/mL for dutasteride and 0.08–1.2 ng/mL for 1,2-dihydrodutasteride, 4′-hydroxydutasteride, 6β-hydroxydutasteride. It was proved to be reliable as it met all validation criteria required by the European Medicine Agency for bioanalytical methods. 4′-hydroxydutasteride and 1,2-dihydrodutasteride concentrations in plasma were higher than 6β-hydroxydutasteride. Dutasteride was stable in the freezer for up to 2 years in clinical samples. Thus within 1014 days of storage (below − 65 °C), samples can be reanalyzed without the risk of unreliable results.

Insight on the Intracrinology of Menopause: Androgen Production within the Human Vagina.

In this study, we investigated steroidogenic gene mRNA expression in human vaginas and verified the ability of human vagina smooth muscle cells (hvSMCs) to synthesize androgens from upstream precursor dehydroepiandrosterone (DHEA). As a readout for androgen receptor (AR) activation, we evaluated the mRNA expression of various androgen-dependent markers. hvSMCs were isolated from vagina tissues of women undergoing surgery for benign gynecological diseases. In these cells, we evaluated mRNA expression of several steroidogenic enzymes and sex steroid receptors using real time reverse transcription-polymerase chain reaction. Androgen production was quantified with liquid chromatography tandem-mass spectrometry (LC-MS/MS). In vaginal tissues, AR mRNA was significantly less expressed than estrogen receptor α, whereas in hvSMCs, its mRNA expression was higher than progestin and both estrogen receptors. In hvSMCs and in vaginal tissue, when compared to ovaries, the mRNA expression of proandrogenic steroidogenic enzymes (HSD3β1/β2, HSD17β3/β5), along with 5α-reductase isoforms and sulfotransferase, resulted as being more abundant. In addition, enzymes involved in androgen inactivation were less expressed than in the ovaries. The LC-MS/MS analysis revealed that, in hvSMCs, short-term DHEA supplementation increased Δ4-androstenedione levels in spent medium, while increasing testosterone and DHT secretion after longer incubation. Finally, androgenic signaling activation was evaluated through AR-dependent marker mRNA expression, after DHEA and T stimulation. This study confirmed that the human vagina is an androgen-target organ with the ability to synthesize androgens, thus providing support for the use of androgens for local symptoms of genitourinary syndrome in menopause.

The Cardiovascular Safety of Dutasteride

Randomized controlled trials suggest an increased risk of heart failure with dutasteride, which inhibits both the type 1 and type 2 isoforms of 5α-reductase. In contrast, no such association has been suggested for finasteride, which selectively inhibits the type 2 isoform. We investigated the risk of cardiovascular events among patients receiving dutasteride relative to finasteride. We performed a population based cohort study of Ontario men 66 years old or older who commenced treatment with dutasteride or finasteride between October 1, 2005 and March 31, 2015. For each individual treated with dutasteride, we identified 1 treated with finasteride, matching on a propensity score and calendar quarter of treatment initiation to account for temporal changes in prescribing. The primary outcome was hospitalization for heart failure. Secondary analyses were done to examine acute myocardial infarction and stroke. Cox proportional hazards regression was used to adjust for differences between groups. We studied 36,311 men who commenced dutasteride and 36,311 treated with finasteride. In the primary analysis, we found no difference in the risk of heart failure among patients receiving dutasteride relative to those receiving finasteride (adjusted HR 0.98, 95% CI 0.88-1.08). Similarly, we found no difference in the risk of acute myocardial infarction (HR 0.94, 95% CI 0.82-1.08) or stroke (HR 1.03, 95% CI 0.88-1.20). In this population based cohort study of more than 72,000 older men, dutasteride was not associated with an increased risk of cardiovascular events relative to finasteride.

Expression of 5α- and 5β-reductase in spinal cord and muscle of birds with different courtship repertoires.

BackgroundThrough the actions of one or more isoforms of the enzyme 5α-reductase in many male reproductive tissues, circulating testosterone (T) undergoes metabolic conversion into 5α-dihydrotestosterone (DHT), which binds to and activates androgen receptors (AR) with greater potency than T. In birds, T is also subject to local inactivation into 5β-DHT by the enzyme 5β-reductase. Male golden-collared manakins perform an androgen-dependent and physically elaborate courtship display, and these birds express androgen receptors in skeletal muscles and spinal cord at levels far greater than those expressed in species with more limited courtship routines, including male zebra finches. To determine if local T metabolism facilitates or impedes activation of male manakin courtship, we examined expression of two isoforms of 5α-reductase, as well as 5β-reductase, in forelimb muscles and spinal cords of males and females of the two aforementioned species.ResultsWe found that all enzymes were expressed in all tissues, with patterns that partially predict a functional role for 5α-reductase in these birds, especially in both muscle and spinal cord of male manakins. Moreover, we found that 5β-reductase was markedly different between species, with far lower levels in golden-collared manakins, compared to zebra finches. Thus, modification to neuromuscular deactivation of T may also play a functional role in adaptive behavioral modulation.ConclusionsGiven that such a role for 5α-reductase in androgen-sensitive mammalian skeletal muscle is in dispute, our data suggest that, in birds, local metabolism may play a key role in providing active androgenic substrates to peripheral neuromuscular systems. Similarly, we provide the first evidence that 5β-reductase is expressed broadly through an organism and may be an important factor that regulates androgenic modulation of neuromuscular functioning.

Herbal Extracts Induce Dermal Papilla Cell Proliferation of Human Hair Follicles.

BackgroundThe number of people suffering from balding or hair thinning is increasing, despite the advances in various medical therapies. Therefore, it is highly important to develop new therapies to inhibit balding and increase hair proliferation.ObjectiveWe investigated the effects of herbal extracts commonly used for improving balding in traditional medicine to identify potential agents for hair proliferation.MethodsThe expression levels of 5α-reductase isoforms (type I and II) were analyzed using quantitative real-time reverse transcription polymerase chain reaction in the human follicular dermal papilla cells (DPCs). The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenylteterazolium bromide and bromodeoxyuridine tests were used to evaluate the cell proliferation effect of herbal extracts in DPCs. The expression levels of extracellular signal-regulated kinase (ERK), Akt, cyclin D1, cyclin-dependent kinase 4 (Cdk4), B-cell lymphoma (Bcl-2) and Bcl-2-associated X protein (Bax) were measured using western blot analysis.ResultsThe 5α-reductase isoform mRNAs and proteins were detected in the cultured DPCs, and the expression level of 5α-R2 in DPCs in the presence of the herbal extracts was gradually decreased. Herbal extracts were found to significantly increase the proliferation of human DPCs at concentrations ranging from 1.5% to 4.5%. These results show that the herbal extracts tested affected the protein expressions of ERK, Akt, cyclin D1, Cdk4, Bcl-2, and Bax in DPCs.ConclusionThese results suggest that herbal extracts exert positive effects on hair proliferation via ERK, Akt, cyclin D1, and Cdk4 signaling in DPCs; they also suggest that herbal extracts could be a great alternative therapy for increasing hair proliferation.

Cellular localization of 5α-reductase in the rat cerebellum

The enzyme 5α-reductase catalyzes the transformation of progesterone, testosterone, and deoxycorticosterone into 5α-reduced metabolites, which are recognized as neurosteroids in the brain with variable potential neuroactivity. Two isoforms of 5α-reductase were identified in rodents, and, of these, 5α-reductase type 1 (5α-R1) is abundantly expressed in the brain. To understand the multiple influences of neurosteroids in the central nervous system, we need to know their region-specific synthesis. The present study reports the detailed localization of 5α-R1 in the adult rat cerebellum. The occurrence of 5α-R1 was detected by reverse transcription-polymerase chain reaction. The enzyme activity was also detected by thin layer chromatography. Immunocytochemistry showed 5α-R1 immunoreactive cells in all cerebellar layers. Multiple immunolabeling revealed that 5α-R1 was mainly localized in glia, such as astrocytes and oligodendrocytes. The most intense immunoreactivity for 5α-R1 was found in Bergmann glia, and the processes of these glia were associated with dendrites of both Purkinje cells and interneurons in the molecular layer. The 5α-R1 in the cerebellum was expressed consistently throughout different ages and sexes, in both gonadectomized and hypophysectomized rats. Thus, 5α-R1 may contribute to the formation and maintenance of the cerebellar neurons through 5α-reduced metabolites, which are synthesized through a complex interaction between neurons and glia.

Mode of action of human pharmaceuticals in fish: The effects of the 5-alpha-reductase inhibitor, dutasteride, on reproduction as a case study

In recent years, a growing number of human pharmaceuticals have been detected in the aquatic environment, generally at low concentrations (sub-ng/L–low μg/L). In most cases, these compounds are characterised by highly specific modes of action, and the evolutionary conservation of drug targets in wildlife species suggests the possibility that pharmaceuticals present in the environment may cause toxicological effects by acting through the same targets as they do in humans. Our research addressed the question of whether or not dutasteride, a pharmaceutical used to treat benign prostatic hyperplasia, may cause adverse effects in a teleost fish, the fathead minnow (Pimephales promelas), by inhibiting the activity of both isoforms of 5α-reductase (5αR), the enzyme that converts testosterone into dihydrotestosterone (DHT). Mammalian pharmacological and toxicological information were used to guide the experimental design and the selection of relevant endpoints, according to the so-called “read-across approach”, suggesting that dutasteride may affect male fertility and steroid hormone dynamics. Therefore, a 21-day reproduction study was conducted to determine the effects of dutasteride (10, 32 and 100μg/L) on fish reproduction. Exposure to dutasteride significantly reduced fecundity of fish and affected several aspects of reproductive endocrine functions in both males and females. However, none of the observed adverse effects occurred at concentrations of exposure lower than 32μg/L; this, together with the low volume of drug prescribed every year (10.34kg in the UK in 2011), and the extremely low predicted environmental concentration (0.03ng/L), suggest that, at present, the potential presence of dutasteride in the environment does not represent a threat to wild fish populations.

Association of genetic variants in the two isoforms of 5α-reductase, SRD5A1 and SRD5A2, in lean patients with polycystic ovary syndrome

ObjectiveGiven its role in converting testosterone to dihydrotestosterone and cortisol to dihydrocortisol, 5α-reductase may be important in the pathophysiology of the polycystic ovary syndrome (PCOS). Increased activity of this enzyme has already been demonstrated in ovaries of affected women, and might be caused by genetic alterations. The aim of this study was to analyze representative genetic variants of both isoforms of 5α-reductase with regard to PCOS parameters in lean and obese women. Study designWe analyzed one single nucleotide polymorphism (SNP) (rs523349) of the isoform 2 (SRD5A2) and one haplotype of the isoform 1 (SRD5A1), consisting of the two SNPs rs39848 and rs3797179, in 249 women with PCOS and 226 healthy women using a 5′-exonuclease-assay. The genotypes were associated with anthropometric, metabolic and hormonal as well as functional tests in these women. ResultsIn the investigated haplotype of SRD5A1, the TA variant was associated with an increased frequency of PCOS (P=0.022) and an increased Ferriman–Gallwey Score (hirsutism) (P=0.016) in women with normal weight. The G allele at the examined position of the SRD5A2 showed a decreased frequency of PCOS (P=0.03) in women with normal weight. ConclusionOne of the keys in the development of the PCOS is hyperandrogenism, which might be caused by an increased 5α-reductase activity, as it is often seen in obesity. This mechanism might therefore be of importance in lean PCOS patients and contribute to the clinical findings.

Expression of 5α-reductase type 1 in breast cancer and adjacent non-malignant tissue: an immunohistochemical study.

Intratumoral biosynthesis of sex steroids is thought to play a role in the pathogenesis and development of human breast cancer. There is evidence that androgens can inhibit the development and progression of breast cancer. Among the enzymes involved in the biosynthesis of androgens, 5α-reductase plays a key role by reducing testosterone to dihydrotestosterone, the most potent androgen. Two isoforms of 5α-reductase have been characterized and 5α-reductase type 1 is predominant in breast cancer tissue. We developed specific antibodies to 5α-reductase type 1 and studied the expression of the enzyme in 84 specimens of breast carcinoma and adjacent non-malignant tissues by immunohistochemistry. The results were correlated with the expression of androgen receptor, estrogen receptor α, progesterone receptor and CDC47, a cell division marker as well as the tumor stage, tumor size, nodal status and menopausal status. The expression of 5α-reductase type 1 in 61% of breast cancer specimens appeared significantly lower than that observed in normal adjacent tissues (87% of cases being positive). There was no significant correlation between 5α-reductase type 1 expression and the clinicopathological parameters studied. The decrease in 5α-reductase type 1 expression in breast cancer as compared to that observed in the adjacent normal tissues could play a role in the development and/or progression of the cancer by modifying the intratumoral levels of androgens.

Genetic Variation of Genes Involved in Dihydrotestosterone Metabolism and the Risk of Prostate Cancer

Dihydrotestosterone (DHT) is an important factor in prostate cancer (PCA) genesis and disease progression. Given PCA's strong genetic component, we evaluated the possibility that variation in genes involved in DHT metabolism influence PCA risk. We investigated copy number variants (CNV) and single nucleotide polymorphisms (SNP). We explored associations between CNV of uridine diphospho-glucuronosyltransferase (UGT) genes from the 2B subclass, given their prostate specificity and/or involvement in steroid metabolism and PCA risk. We also investigated associations between SNPs in genes (HSD3B1, SRD5A1/2, and AKR1C2) involved in the conversion of testosterone to DHT, and in DHT metabolism and PCA risk. The population consisted of 426 men (205 controls and 221 cases) who underwent prostate-specific antigen screening as part of a PCA early detection program in Tyrol, Austria. No association between CNV in UGT2B17 and UGT2B28 and PCA risk was identified. Men carrying the AA genotype at SNP rs6428830 (HSD3B1) had an odds ratio (OR) of 2.0 [95% confidence intervals (95% CI), 1.1-4.1] compared with men with GG, and men with AG or GG versus AA in rs1691053 (SRD5A1) had an OR of 1.8 (95% CI, 1.04-3.13). Individuals carrying both risk alleles had an OR of 3.1 (95% CI, 1.4-6.7) when compared with men carrying neither (P = 0.005). Controls with the AA genotype on rs7594951 (SRD5A2) tended toward higher serum DHT levels (P = 0.03). This is the first study to implicate the 5alpha-reductase isoform 1 (SRD5A1) and PCA risk, supporting the rationale of blocking enzymatic activity of both isoforms of 5alpha-reductase for PCA chemoprevention.

Comparison of the Potency of Different Brands of Serenoa Repens Extract on 5α-Reductase Types I and II in Prostatic Co-Cultured Epithelial and Fibroblast Cells

Background:Serenoa repens extract is the phytotherapeutic agent most frequently used for the treatment of the urological symptoms caused by benign prostatic hyperplasia. There are many extracts in the market and each manufacturer uses different extraction processes; for this reason, it’s possible that one product is not equivalent to another. The aim of this study was to compare the activity of different extracts of Serenoa repens marketed in Italy. Methods: The following extracts were tested on 10 day co-cultured epithelial and fibroblast cells by a 5α-reductase activity assay: Permixon®, Saba®, Serpens®, Idiprost®, Prostamev®, Profluss® and Prostil®. In order to assess the variability in Serenoa repens products, 2 different batches for each brand were evaluated. Results and Conclusions: All extracts tested, albeit variably, are able to inhibit both isoforms of 5α-reductase. However, the potency of the extracts appears to be very different, as well as the potencies of 2 different batches of the same extract. This is probably due to qualitative and quantitative differences in the active ingredients. So, the product of each company must be tested to evaluate the clinical efficacy and bioactivity.

The promoter of the rat 5α-reductase type 1 gene is bidirectional and Sp1-dependent

In many androgen target tissues, testosterone is reduced to the more potent androgen, dihydrotestosterone, by steroid 5α-reductase. Two isoforms of 5α-reductase, type 1 and type 2, have been cloned. They are differentially expressed and regulated. To determine the mechanisms of regulation of 5α-reductase type 1 expression, we have cloned its 5′upstream region and defined its promoter. The proximal 5′upstream region of 5α-reductase type 1 displays all the features of a CpG island and has numerous Sp1 binding sites. By transient transfection assays, we have identified a bidirectional promoter activity in this region; this activity was highest in the negative orientation, in the direction of the methyltransferase Nsun2 (predicted) gene. Promoter activity, in either orientation, was lost in Sp1 deficient cells but was rescued following co-transfection with a Sp1 expression vector. Thus, the 5′upstream region of rat 5α-reductase type 1 contains a bidirectional promoter with an activity that is Sp1-dependent.

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

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 5alpha-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 DHT-dependence using two different dual 5alpha-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 5alpha-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 5alpha-reductase inhibition; the two inhibitors had parallel effects. Specifically, in proximal regions, 5alpha-reductase 1, androgen receptor, and TGF-beta1 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.

Variants in the 5α-Reductase Type 1 and Type 2 Genes Are Associated with Polycystic Ovary Syndrome and the Severity of Hirsutism in Affected Women

Despite the importance of dihydrotestosterone in androgen action, polymorphisms in the genes for the two isoforms of 5alpha-reductase (SRD5A1 and SRD5A2) have not been evaluated as risk factors for polycystic ovary syndrome (PCOS). The objective of the study was to test the hypothesis that haplotypes in the SRD5A1 and SRD5A2 genes are risk factors for PCOS and the severity of hirsutism in affected women. PCOS and control subjects were genotyped for seven single-nucleotide polymorphisms in SRD5A1 and eight single-nucleotide polymorphisms in SRD5A2. Haplotypes were determined and tested for association with PCOS diagnosis and component phenotypes. Subjects were recruited from the reproductive endocrinology clinic at the University of Alabama at Birmingham; control subjects were recruited from the general surrounding community. Genotyping took place at Cedars-Sinai Medical Center in Los Angeles. A total of 287 White women with PCOS and 187 controls participated. SRD5A1 and SRD5A2 genotype, quantitative hirsutism score, and hormonal and metabolic phenotypes were assessed. Haplotypes within both genes were associated with PCOS risk. The Leu allele of the Val89Leu variant in SRD5A2 was associated with protection against PCOS; this allele is known to modestly reduce 5alpha-reductase activity. Haplotypes in SRD5A1 but not SRD5A2 were also associated with the degree of hirsutism in affected women. This study presents genetic evidence suggesting an important role of both isoforms of 5alpha-reductase in the pathogenesis of PCOS. That only SRD5A1 haplotypes were associated with hirsutism suggests that only this isoform is important in the hair follicle.

Regulation of 5alpha-reductase isoforms by oxytocin in the rat ventral prostate.

Oxytocin (OT) is present in the male reproductive tract, where it is known to modulate contractility, cell growth, and steroidogenesis. Little is known about how OT regulates these processes. This study describes the localization of OT receptor in the rat ventral prostate and investigates if OT regulates gene expression and/or activity of 5alpha-reductase isoforms I and II. The ventral prostates of adult male Wistar rats were collected following daily sc administration of saline (control), OT, a specific OT antagonist or both OT plus antagonist for 3 d. Expression of the OT receptor was identified in the ventral prostate by RT-PCR and Western blot, and confirmed to be a single active binding site by radioreceptor assay. Immunohistochemistry localized the receptor to the epithelium of prostatic acini and to the stromal tissue. Real-time RT-PCR determined that OT treatment significantly reduced expression of 5alpha-reductase I but significantly increased 5alpha-reductase II expression in the ventral prostate. Activity of both isoforms of 5alpha-reductase was significantly increased by OT, resulting in increased concentration of prostatic dihydrotestosterone. In conclusion, OT is involved in regulating conversion of testosterone to the biologically active dihydrotestosterone in the rat ventral prostate. It does so by differential regulation of 5alpha-reductase isoforms I and II.

The mechanism on which the lipido-sterolic extract of serenoa repens (permixon®) inhibits both isoforms of 5αreductase without interfering with PSA production in the prostate

The mechanism on which the lipido-sterolic extract of serenoa repens (permixon®) inhibits both isoforms of 5αreductase without interfering with PSA production in the prostate

Comparative study of human steroid 5α-reductase isoforms in prostate and female breast skin tissues: sensitivity to inhibition by finasteride and epristeride

Steroid 5α-reductase (5-AR) catalyses the reduction of testosterone (T) to dihydrotestosterone (DHT). The 5α-reductase found in human benign prostatic hyperplasia (BPH) has been compared with that found in human breast skin tissue in respect of sensitivity to inhibition by Finasteride and Epristeride. Kinetic studies showed the presence of two isoforms of 5α-reductase in benign prostatic hyperplasia indicated by low and high Km isoforms for testosterone, while female breast skin tissue contained only one isoform. The isoforms differ in their affinity for the inhibitors Finasteride and Epristeride, both compounds being more effective for the low Km 5α-reductase isoform than the high Km 5α-reductase of prostatic tissue, with Finasteride displaying competitive inhibition and Epristeride uncompetitive. Finasteride and Epristeride are also inhibitors of skin 5α-reductase, which possesses a comparable Ki for Finasteride to that of the low Km prostatic enzyme, but Epristeride was a less potent inhibitor of the skin enzyme relative to the prostate isoform. These results suggest that the inhibitors have therapeutic potential, other than for treatment of benign prostatic hyperplasia, for treating skin disorders influenced by the action of dihydrotestosterone and warrant further investigation.

5α‐reductase isozymes and aromatase are differentially expressed and active in the androgen‐independent human prostate cancer cell lines DU145 and PC3

BACKGROUND The presence and possible role of androgen-metabolizing enzymes in androgen-independent prostate carcinoma (CaP) are still unclear. The aim of the present study was: 1) to evaluate the pattern of androgen metabolism (relative production of 5α-reduced vs. 17-keto androgens); and 2) to analyze whether one or both the two known 5α-reductase isoforms (5α-R1 and 5α-R2) and the aromatase (Aro) are expressed and active in this pathology. METHODS Two different cell lines (DU145 and PC3) were used as a model of androgen-independent human CaP. In these cells, the expression of the two 5α-Rs and of Aro were evaluated by reverse transcription-polymerase chain reaction (RT-PCR) and Southern blot, using specific sets of oligoprimers and of [32P]-labeled oligoprobes; the enzymatic activities of 5α-R and of Aro were evaluated by radioenzymatic methods. The pH optimum for the activity of the two 5α-Rs was assessed in cell homogenates at different pH (from 3.5–8), using substrate concentrations similar either to 5α-R1 or to 5α-R2 Kms. RESULTS The two CaP cell lines DU145 and PC3, although unresponsive to androgens, possess the enzymatic machinery involved in the metabolism of this class of hormonal steroids: 5α-Rs, which allow their transformation into 5α-reduced steroids (5α-dihydrotestosterone, DHT, and 5α-androstandione, 5α-A), and 17β-hydroxysteroid-oxidoreductase (17β-HSD), which interconverts testosterone (T) and androstenedione (ADIONE); however, the two cell lines show differences in the rate of formation of these metabolites. Furthermore, two cell lines expressed the type 1 isoform of 5α-R, but only DU145 cells also possess 5α-R2. Aro is expressed and active in DU145 as well as in PC3 cells. CONCLUSIONS The present findings suggest that T might still be indirectly active in androgen-unresponsive CaP through its local conversion into estrogens by the action of Aro; the biological role played by the two 5α-Rs in androgen-independent CaP deserves further investigation. Prostate 41:224–232, 1999. © 1999 Wiley-Liss, Inc.