Ion Pair Strategy in Solid Lipid Nanoparticles: a Targeted Approach to Improve Epidermal Targeting with Controlled Adapalene Release, Resulting Reduced Skin Irritation.

PURPOSE: Adapalene (AD) is one of the main retinoids used in the topical therapy of acne, an extremely common skin disease usually associated with psychological morbidity. However, like other retinoids, AD is frequently associated with skin irritation. To overcome the skin irritation, we proposed the encapsulation of AD in solid lipid nanoparticles (SLNs) using the ion pair strategy. METHODS: The developed SLN-AD was characterized by high-performance liquid chromatography, differential scanning calorimetry, X-ray diffraction, synchrotron small-angle X-ray scattering, and transmission electron microscopy. In vitro permeation tests using porcine skin and in vivo mice skin irritation test were performed to evaluate, respectively, the drug's skin distribution and the skin irritation. RESULTS: The characterization studies were able to demonstrate that the proposed strategy effectively provided high AD encapsulation in SLNs and its incorporation into a hydrophilic gel. Sustained release, epidermal targeting, and less skin irritation were observed for SLN-AD gel in comparison to the marketed AD gel. CONCLUSIONS: The studies demonstrated that the encapsulation of AD in SLNs through the formation of an ion pair is a valuable alternative to diminish the adverse skin reactions caused by AD and can optimize patient adherence to treatment.

Seasonal variation of cell proliferation and apoptosis in the efferent ductules and epididymis of the Neotropical bat Artibeus lituratus (Chiroptera, Phyllostomidae).

The balance between cell proliferation and apoptosis is important for maintenance of male fertility, being influenced by a variety of stimuli including androgens and estrogens. However, studies concerning regulation of these processes along the male reproductive tract under physiological conditions are scarce. Therefore, in this study, we investigated the profile of cell proliferation and apoptosis in the efferent ductules and epididymis of the Neotropical bat Artibeus lituratus, a seasonal breeder that presents natural variation in components of the androgen and estrogen responsive systems along the circannual cycle. Low rates of cell proliferation and apoptosis were found in the efferent ductules and epididymis of A. lituratus during the reproductive period, as few epithelial cells were positive for MCM7 (proliferation marker) and cleaved caspase-3 or TUNEL (apoptosis markers). In contrast, during the regressive period, the rate of both proliferating and apoptotic cells was significantly higher in the epithelium lining the efferent ductules as well as throughout the epididymis. The increased proliferative activity at this phase was positively correlated with the expression of estrogen receptor alpha (ERα), whereas the variation in apoptosis appears to be unrelated to the local expression of androgen and estrogen receptors. Together, these data suggest that cell proliferation and apoptosis are differentially modulated in the efferent ductules and epididymis of A. lituratus during the annual reproductive cycle, and support the hypothesis that ERα may be important in preparing the male reproductive tract for sexual recrudescence.

Reduced vitamin D receptor (VDR) expression and plasma vitamin D levels are associated with aging-related prostate lesions.

BACKGROUND: Protective roles have been proposed for vitamin D in prostate cancer, which has the advanced age as the major risk factor. However, little is known about the expression of the vitamin D receptor (VDR) in the aging prostate and its association with the development of epithelial lesions that affect tissue homeostasis and may precede prostate tumors. METHODS: VDR expression in the prostatic complex of young adults to senile Wistar rats, a natural model to study age-related prostatic disorders, was evaluated by immunohistochemical, Western blotting, and image-assisted analyzes. Results were correlated with the plasma levels of vitamin D and testosterone, the occurrence of punctual histopathological changes in the aging prostate, and the expression of retinoid X receptors (RXR). RESULTS: VDR was widely distributed in the prostatic complex at all ages analyzed, with the highest immunoexpression found in basal epithelial cells. As the animals aged, VDR levels increased, except in punctual areas with intraepithelial proliferation, metaplasia, or proliferative inflammatory atrophy, which had reduced expression of this receptor concomitantly with increased cell proliferation. Interestingly, RXR expression in the aging prostate was similar to that found for its partner VDR, indicating that components of the VDR/RXR complex required for vitamin D signaling are affected in aging-related prostatic lesions. Moreover, plasma vitamin D levels declined at the same ages when prostatic alterations appeared. Although circulating levels of testosterone also decreased with aging, the changes observed in the components of the vitamin D system were not correlated with androgens. CONCLUSIONS: Our data indicate that the aging prostate suffers from an imbalance on the intricate mechanism of tissue regulation by the vitamin D responsive system. We argue that the status of VDR expression might be determinant for the development of histopathological alterations in the aging prostate, which include premalignant lesions.

Profile of cell proliferation and apoptosis activated by the intrinsic and extrinsic pathways in the prostate of aging rats.

BACKGROUND: Estrogens acting through the receptors ERα and ERß participate in prostate normal growth and cancer. ERß is highly expressed in the prostate epithelium, playing pro-apoptotic, anti-proliferative, and pro-differentiation roles. Apoptosis is activated by the intrinsic pathway after castration and by the extrinsic pathway after ERß agonist treatment. This differential activation of apoptotic pathways is important since a major problem in the treatment of prostate cancer is the recurrence of tumors after androgen withdrawal. However, a comprehensive study about the pattern of apoptosis in the aging prostate is lacking, a knowledge gap that we aimed to address herein. METHODS: Cellular age-related proliferative and apoptotic profiles of prostate tissue obtained from aging Wistar rats were evaluated. Cell death (caspase-3, -8, -9, TNFα) was assessed by immunohistochemistry, immunofluorescence, and TUNEL. Cell proliferation (MCM7) and cell survival factors (ERK1/2, p-ERK1/2, p-Akt, and NF-κB) were determined by immunohistochemistry. RESULTS: As the rats aged, the number of proliferating cells gradually reduced in the normal epithelium of all prostate lobes, while increasing in focal areas of intraepithelial proliferation. Interestingly, in areas of intraepithelial proliferation, we observed a reduction in the number of cells positive for caspase-3, -8, and -9. Regardless the animal's age, few prostate epithelial cells were positive for caspase-3, caspase-9, and TUNEL. In contrast, a progressive increase was seen in the positivity for caspase-8, especially in the atrophic epithelium of ventral prostate, which coincided with a reduction in TNFα immunoreaction. However, morphology of most caspase-8 positive cells suggests that they were not apoptotic. We also found reduced ERß expression in the same areas. Possibly, low levels of the pro-apoptotic inductors TNFα and ERß direct caspase-8 activity to an alternative pro-survival role in the atrophic epithelium. This hypothesis is supported by the increased expression of the key survival factors (ERK1/2, p-ERK1/2, p-Akt, and NF-κB) in these areas. CONCLUSIONS: Our findings reveal that, as the animals age, there is an increase of proliferation in restricted areas of the prostate epithelium, and a concomitant reduction of the apoptosis rate with an increase in cell survival induced by caspase-8, indicating a focused and spontaneous disruption of tissue homeostasis.

The insulin receptor translocates to the nucleus to regulate cell proliferation in liver.

UNLABELLED: Insulin's metabolic effects in the liver are widely appreciated, but insulin's ability to act as a hepatic mitogen is less well understood. Because the insulin receptor (IR) can traffic to the nucleus, and Ca(2+) signals within the nucleus regulate cell proliferation, we investigated whether insulin's mitogenic effects result from activation of Ca(2+)-signaling pathways by IRs within the nucleus. Insulin-induced increases in Ca(2+) and cell proliferation depended upon clathrin- and caveolin-dependent translocation of the IR to the nucleus, as well as upon formation of inositol 1,4,5,-trisphosphate (InsP3) in the nucleus, whereas insulin's metabolic effects did not depend on either of these events. Moreover, liver regeneration after partial hepatectomy also depended upon the formation of InsP3 in the nucleus, but not the cytosol, whereas hepatic glucose metabolism was not affected by buffering InsP3 in the nucleus. CONCLUSION: These findings provide evidence that insulin's mitogenic effects are mediated by a subpopulation of IRs that traffic to the nucleus to locally activate InsP3 -dependent Ca(2+)-signaling pathways. The steps along this signaling pathway reveal a number of potential targets for therapeutic modulation of liver growth in health and disease.

Subchronic intake of arsenic at environmentally relevant concentrations causes histological lesions and oxidative stress in the prostate of adult Wistar rats.

The prostate gland is one of the main sites of hyperplasia and cancer in elderly men. Numerous factors have been demonstrated to disrupt prostate homeostasis, including exposure to environmental pollutants. Arsenic is a metalloid found ubiquitously in soil, air, and water, which favors human poisoning through the involuntary intake of contaminated drinking water and food and has harmful effects by increasing the oxidative stress response. This study aimed to investigate the effects of prolonged exposure to arsenic at environmentally relevant concentrations on the prostate biology of adult Wistar rats. Thirty 80-day-old male rats were divided into three experimental groups. Rats from the control group received filtered water, whereas animals from the arsenic groups ingested 1 mg L-1 and 10 mg L-1 of arsenic, in the form of sodium arsenite, daily. The arsenic solutions were provided ad libitum in the drinking water for eight weeks. Our results showed that 1 mg L-1 and 10 mg L-1 of arsenic made the prostate susceptible to evolving benign and premalignant histopathological changes. While the ingestion of 1 mg L-1 of arsenic reduced SOD activity only, 10 mg L-1 diminished SOD and CAT activity in the prostate tissue, culminating in high MDA production. These doses, however, did not affect the intraprostatic levels of DHT and estradiol. In conclusion, exposure to arsenic at environmentally relevant concentrations through drinking water induces histological and oxidative stress-related changes in the prostate of adult rats, strengthening the between arsenic exposure and prostate disorders.

Differential expression and seasonal variation on aquaporins 1 and 9 in the male genital system of big fruit-eating bat Artibeus lituratus.

Efferent ductules and epididymis are involved in water and solute transport, which is indispensable for storage and maintenance of the sperm viability. The reabsorption process involves proteins such as aquaporins (AQP), which has been described in the male genital system of limited species, including primate, rodents, cats and dogs. To contribute with information about AQPs in the male system, here we investigated the distribution of AQP1 and AQP9 in the tropical bat Artibeus lituratus, along the annual reproductive cycle. A. lituratus is a seasonal breeder with natural variation in components of the androgen and estrogen responsive system, thus being a good model for exploring the AQPs modulation. AQP1 was found restricted to differentiating spermatids, efferent ductules epithelium and venular endothelia along the male tract. AQP9 was detected throughout the epididymis being more abundant in the cauda and ductus deferens, but was not found in testis, rete testis and efferent ductules. Contrasting with AQP1 which appear to be constitutively expressed, there was seasonal variation in AQP9 expression, which was reduced in regressed epididymis. The AQP9 does not appear to be modulated by estradiol or androgens, but possibly by other factor related to luminal sperm. The establishment of specific function for aquaporins in the male tract remains undetermined; however, the cellular distribution presently found are compatible with the main function of AQP1, as a selective water channel, and AQP9, which is a conduct for water and a plethora of neutral solutes present in the epididymis milieu such as glycerol and urea.

Dietary Vitamin D Mitigates Coronavirus-Induced Lung Inflammation and Damage in Mice.

The COVID-19 pandemic caused by the SARS-CoV-2 (ß-CoV) betacoronavirus has posed a significant threat to global health. Despite the availability of vaccines, the virus continues to spread, and there is a need for alternative strategies to alleviate its impact. Vitamin D, a secosteroid hormone best known for its role in bone health, exhibits immunomodulatory effects in certain viral infections. Here, we have shown that bioactive vitamin D (calcitriol) limits in vitro replication of SARS-CoV-2 and murine coronaviruses MHV-3 and MHV-A59. Comparative studies involving wild-type mice intranasally infected with MHV-3, a model for studying ß-CoV respiratory infections, confirmed the protective effect of vitamin D in vivo. Accordingly, mice fed a standard diet rapidly succumbed to MHV-3 infection, whereas those on a vitamin D-rich diet (10,000 IU of Vitamin D3/kg) displayed increased resistance to acute respiratory damage and systemic complications. Consistent with these findings, the vitamin D-supplemented group exhibited lower viral titers in their lungs and reduced levels of TNF, IL-6, IL-1ß, and IFN-γ, alongside an enhanced type I interferon response. Altogether, our findings suggest vitamin D supplementation ameliorates ß-CoV-triggered respiratory illness and systemic complications in mice, likely via modulation of the host's immune response to the virus.

Seasonal variation in estrogen receptor ERα, but not ERß, androgen receptor and aromatase, in the efferent ductules and epididymis of the big fruit-eating bat Artibeus lituratus.

The efferent ductules (ED) are a major target for estrogens, which act via the estrogen receptors ERα (ESR1) and ERß (ESR2). ERα has been found in the ED of all species studied so far. However, in the epididymis (EP), the expression of ERα is controversial, as is data about the occurrence of aromatase in the epithelium lining the excurrent ducts. Therefore, to further investigate this estrogen-responsive system, we used a seasonal breeder, the Neotropical bat, Artibeus lituratus, in which testicular expression of androgen (AR) and estrogen (ER) receptors vary with reproductive phase. The localization of aromatase, ERα, ERß and AR in the ED and EP of A. lituratus was investigated. The results showed that aromatase, AR and ERß were distributed throughout the excurrent ducts and did not vary during the annual reproductive cycle. Conversely, ERα was detected primarily in the ED epithelium, had marked seasonal variation and was increased during regression, especially in the EP epithelium. The results suggest that ERα may be involved in preparing the male genital tract for recrudescence. Together, the data obtained under natural conditions emphasize that specific segments of the excurrent ducts downstream of the testis are the primary targets for estrogen action via ERα, which is similar to previous findings in animals lacking functional ERα.

Roosters affected by epididymal lithiasis present local alteration in vitamin D3, testosterone and estradiol levels as well as estrogen receptor 2 (beta) expression.

Epididymal lithiasis is a reproductive dysfunction of roosters that is associated with loss of fertility and is characterized by the formation of calcium stones in the lumen of the efferent ductules of the epididymal region. The efferent ductules of birds are responsible for the reabsorption of the fluid coming from the testis as well as luminal calcium. It has been hypothesized that the epididymal stone formation may be related to the impairment of local fluid or calcium homeostasis, which depends on hormones such as estradiol (E(2)). Therefore, this study aimed to investigate possible alterations in the expression of ERα (ESR1) and ERß (ESR2) in the epididymal region of roosters affected by epididymal lithiasis. The study was performed by immunohistochemistry and western blotting assays. In addition, the concentrations of E(2), vitamin D3, and testosterone, which are also key hormones in maintenance of calcium homeostasis, were determined in the plasma and epididymal region, by ELISA. It was observed that ESR2 expression is increased in all segments of the epididymal region of affected roosters, whereas ESR1 levels are not altered. Moreover, the hormone concentration profiles were changed, as in the epididymal region of roosters with lithiasis the E(2) levels were increased and vitamin D3 as well as testosterone concentrations were significantly decreased. These results suggest that a hormonal imbalance may be involved with the origin and progression of the epididymal lithiasis, possibly by affecting the local fluid or calcium homeostasis.

Angiotensin-(1-7) induces ovulation and steroidogenesis in perfused rabbit ovaries.

A local renin-angiotensin system has been described in several organs, including the ovary; however, data indicating a role for angiotensin II in the induction of ovulation are controversial. We have previously shown the presence of a novel peptide, angiotensin-(1-7) [Ang-(1-7)], in the rat ovary and its effect on steroidogenesis. The objective of the present study was to determine whether Ang-(1-7) plays a role in ovulation. We first determined the presence and distribution of Ang-(1-7) and the receptor Mas in rabbit ovaries by immunohistochemistry. Angiotensin-(1-7) and Mas immunoreactivity were observed in interstitial cells and oocytes of immature ovaries. Immunoreactivity for Ang-(1-7) and Mas was also observed in theca and granulosa cells of preovulatory follicles in ovaries of gonadotrophin-stimulated rabbits. To verify the effect of Ang-(1-7) in ovulation and steroidogenesis, we used isolated ovaries from immature rabbits pretreated with equine chorionic gonadotrophin (50 i.u., 48 h before the experiment) and then perfused in vitro. The ovulatory efficiency was determined by the number of oocytes compared with the number of preovulatory follicles present in the ovary. Angiotensin-(1-7) stimulated oestradiol production and enhanced ovulatory efficiency, which was blocked by the specific Ang-(1-7) antagonist, A-779. Ovulation induced by human chorionic gonadotrophin was also antagonized by A-779. These results show, for the first time, the involvement of a novel regulatory peptide system, Ang-(1-7) and Mas, in the ovulatory process. More importantly, because A-779 antagonized hCG-induced ovulation, it may be inferred that Ang-(1-7) plays an important role in ovulation, possibly as a mediator of gonadotrophin action.

Occurrence and cellular distribution of estrogen receptors ERα and ERß in the testis and epididymal region of roosters.

Estrogen signaling is required for the maintenance of male reproductive function and is mediated by the estrogen receptors ERα and ERß. These receptors are widely distributed in mammalian reproductive tissues, but information is limited in non-mammalian species including birds. The aim of this study was to investigate the occurrence and cellular distribution of ERα and ERß in the testis and epididymal region of roosters. The results showed for the first time that ERß was the predominant receptor detected in the testis, being expressed in the somatic and some germ cells. Within the epididymal region, ERß was strongly expressed in all segments, whereas the most intense reaction for ERα was found in the distal efferent ductules. The differential expression of ERα and ERß within the rooster testis and epididymal region suggests that these organs may be a target for different actions of estrogen.

Tumor-Associated Macrophages (TAM) are recruited to the aging prostate epithelial lesions and become intermingled with basal cells.

BACKGROUND: Prostate cancer remains one of the most common cancers in men. Macrophages are thought to be important regulators in cancers, and their potential involvement in prostate cancer should not be overlooked. Therefore, the association between macrophages and the pre-tumorous changes in prostate epithelium during aging deserves further investigation. OBJECTIVES: We sought to investigate whether macrophages would be recruited into the prostate epithelium that display pathological lesions commonly found during aging. MATERIALS AND METHODS: Prostates of aging rats, with and without treatment with a combination of testosterone and estradiol, were examined for premalignant and malignant epithelial lesions. For comparison, prostates of castrated rats were also investigated. RESULTS: Intraepithelial macrophages were found restricted to areas of premalignant and malignant lesions. An unprecedented interaction between macrophages and basal cells was observed in the aging pathological lesions. The intraepithelial macrophages were associated with autophagy, in contrast to those found after castration. In prostate lesions, the intraepithelial macrophages had TAM phenotype (CD68+/iNOS+/CD206+/ARG+), denoting a possible involvement in cancer progression. However, M2 macrophages (CD68+/CD163+) were recruited into the epithelium after castration, possibly to phagocytize cells undergoing apoptosis. DISCUSSION AND CONCLUSION: In conclusion, macrophages were recruited into the prostate epithelium and presented diverse phenotypes and morphology, consistent with changes reflected in the hormonal environment. Macrophages with the TAM phenotype were found restricted to areas of premalignant and malignant lesions in aging prostates, denoting a possible involvement in cancer progression. In contrast, M2 macrophages were found in the regressed epithelium after castration.

Retinoic acid-loaded solid lipid nanoparticles surrounded by chitosan film support diabetic wound healing in in vivo study.

Repair of tissue damaged in diabetic wounds is essential to minimize the cases of amputation of the limbs in millions of diabetic people around the world. Although the all-trans retinoic acid (ATRA) is described as a potential wound healing agent, however its effects are controversial due to adverse reactions that may impair the wound healing during the treatment schedules. Our aim was to design and characterize an ATRA-loaded solid lipid nanoparticles surrounded by chitosan film to promote an ATRA controlled release and to evaluate its effectiveness in promoting wound healing in a diabetic mouse model. The SLN-ATRA were developed using biocompatible lipids without using organic solvent. The SLN-ATRA had high drug entrapment efficiency (98.0 %) and low polydispersity index (PDI) and average diameter, respectively, 0.24 ± 0.02 and 83.0 ± 6 nm. The transmission electron microscope (TEM) image presented that the SLN-ATRA were homogeneous in size and had spherical structures. The incorporation of SLN-ATRA in the chitosan films propitiated a homogeneous distribution of the drug and a controlled drug release. Furthermore, in vivo assay proved that chitosan films containing SLN-ATRA accelerated the closure of wounds of diabetic mice when compared to the control chitosan films without ATRA. SLN-ATRA chitosan films also reduced leukocyte infiltrate in the wound bed, improved collagen deposition, and reduced scar tissue. No sign of skin irritation was observed. These results indicated that SLN-ATRA surrounded in chitosan films are a promising candidate to treat diabetic wounds, improving tissue healing.

Targeting Wistar rat as a model for studying benign, premalignant and malignant lesions of the prostate.

AIMS: The purpose of this study was to describe a suitable experimental model for studying aging-related prostate disorders including cancer. MATERIALS AND METHODS: 12-month old Wistar rats were kept in control conditions (n = 12) or treated (n = 16) for 6 months with Silastic implants filled with testosterone (T) and estradiol (E2). After the experiment period (at 18 months of age), animals were euthanized and the prostate and other organs were harvested, dissected, weighed, and processed for morphological, ultrastructural and molecular analyses. KEY FINDINGS: We demonstrated that male rats of Wistar strain nicely recapitulate the carcinogenesis process taking place in the aging prostate through the arising of benign, precancerous and malignant lesions, and above all yields a modest incidence of spontaneous PCa (~36%). Moreover, our results highlight that 100% incidence of PCa and precancerous lesions such as prostatic intraepithelial neoplasia and proliferative inflammatory atrophy were achieved in this rat strain after T + E2 treatment, without changing the broad spectrum of changes that naturally emerge in the prostate at advanced ages. Such enhancement of precancerous lesions and tumors was linked to a decreased expression of E-cadherin and ß-catenin in parallel with an increase in Vimentin and N-cadherin, hallmark modifications of epithelial-mesenchymal transition. SIGNIFICANCE: Our findings provide solid evidence that aged Wistar rats may be an excellent model for studies regarding human prostate biology and related disorders including cancer.

Distribution of estrogen receptors (ERalpha and ERbeta) and androgen receptor in the testis of big fruit-eating bat Artibeus lituratus is cell- and stage-specific and increases during gonadal regression.

The testis is a classical target for androgens, especially testosterone, acting via androgen receptor (AR). Alternatively, androgens can be aromatized to produce estrogens which act via specific receptors ERalpha and ERbeta. Although estrogen action is essential for maintenance of male fertility, studies regarding the expression of ERalpha and ERbeta in testis are restricted to a few species of rodent and domestic animals, but rarely in wild species. To our knowledge, there are no studies in Chiroptera species. Chiroptera represent one of the largest and most diversified orders of mammals, which possess several interesting reproductive features, including higher affinity of SHBG for estrogens than androgens. Therefore, we thought that bats would constitute a good model for investigation of the role of estrogens in the male. In this study, the distribution of ERalpha, ERbeta and AR were evaluated in the testis of the big fruit-eating bat Artibeus lituratus and their levels were compared during reproductive and regressive periods. The results showed that ERalpha and AR were restricted to the somatic cells of the testis, whereas ERbeta was widely distributed in both somatic and spermatogenic cells in a cellular and stage-specific fashion. We demonstrated for the first time by immunohistochemistry, and confirmed by Western blotting, that ERbeta and AR increased during regression. The localization of ERalpha, ERbeta and AR in a seasonal, cell and stage-specific fashion in the testis of A. lituratus suggests that these receptors may play important roles in testis function during reproductive and non-reproductive periods.

Basal Cells Show Increased Expression of Aromatase and Estrogen Receptor α in Prostate Epithelial Lesions of Male Aging Rats.

Besides androgens, estrogen signaling plays a key role in normal development and pathologies of the prostate. Irreversible synthesis of estrogens from androgens is catalyzed by aromatase. Interestingly, animals lacking aromatase do not develop cancer or prostatitis, whereas those with overexpression of aromatase and, consequently, high estrogen levels develop prostatitis and squamous metaplasia via estrogen receptor 1 (ERα). Even with this evidence, the aromatase expression in the prostate is controversial. Moreover, little is known about the occurrence of age-dependent variation of aromatase and its association with histopathological changes commonly found in advanced age, a knowledge gap that is addressed herein. For this purpose, the immunoexpression of aromatase was evaluated in the prostatic complex of young adult to senile Wistar rats. ERα was also investigated, to extend our understanding of estrogen responsiveness in the prostate. Moderate cytoplasmic immunoreactivity for aromatase was detected in the glandular epithelium. Eventually, some basal cells showed intense staining for aromatase. The expression pattern for aromatase appeared similar in the normal epithelium when young and senile rats were compared; this result was corroborated by Western blotting. Conversely, in senile rats, there was an increase in the frequency of basal cells intensely stained for aromatase, which appeared concentrated in areas of intraepithelial proliferation and prostatitis. These punctual areas also presented increased ERα positivity. Together, these findings suggest a plausible source for hormonal imbalance favoring estrogen production, which, by acting through ERα, may favor the development of prostatic lesions commonly found in advanced age.

5alpha-Androstane-3beta,17beta-diol (3beta-diol), an estrogenic metabolite of 5alpha-dihydrotestosterone, is a potent modulator of estrogen receptor ERbeta expression in the ventral prostrate of adult rats.

Prostate is one of the major targets for dihydrotestosterone (DHT), however this gland is also recognized as a nonclassical target for estrogen as it expresses both types of estrogen receptors (ER), especially ERbeta. Nevertheless, the concentrations of aromatase and estradiol in the prostate are low, indicating that estradiol may not be the only estrogenic molecule to play a role in the prostate. It is known that DHT can be metabolized to 5alpha-androstane-3beta,17beta-diol (3beta-diol), a hormone that binds to ERbeta but not to AR. The concentration of 3beta-diol in prostate is much higher than that of estradiol. Based on the high concentration of 3beta-diol and since this metabolite is a physiological ERbeta ligand, we hypothesized that 3beta-diol would be involved in the regulation of ERbeta expression. To test this hypothesis, adult male rats were submitted to castration followed by estradiol, DHT or 3beta-diol replacement. ERbeta and AR protein levels in the prostate were investigated by immunohistochemistry and Western blotting assays. The results showed that after castration, the structure of the prostate was dramatically changed and ERbeta and AR protein levels were decreased. Estradiol had just minor effects on the parameters analyzed. DHT-induced partial recovery of ERbeta while it was the most effective inductor of AR expression. Replacement with 3beta-diol-induced the highest levels of ERbeta, but was comparatively less effective in recovering the AR expression and the gland structure. These results offer evidence that one functional role of 3beta-diol in the prostate may be autoregulation of its natural receptor, ERbeta.

Effects of the herbicide Roundup on the epididymal region of drakes Anas platyrhynchos.

Exposure to the Roundup has been shown to affect StAR protein and aromatase expression and activity, pointing out that this herbicide may cause adverse effects in animal reproduction by affecting androgen and estrogen synthesis. We tested this hypothesis by investigating the in vivo effects of the Roundup on the testis and epididymal region of drake Anas platyrhynchos. The exposure to the herbicide resulted in alterations in the structure of the testis and epididymal region as well as in the serum levels of testosterone and estradiol, with changes in the expression of androgen receptors restricted to the testis. The harmful effects were more conspicuous in the proximal efferent ductules and epididymal ducts, suggesting higher sensitivity of these segments among the male genital organs. The effects were mostly dose dependent, indicating that this herbicide may cause disorder in the morphophysiology of the male genital system of animals.

Effects of 3-beta-diol, an androgen metabolite with intrinsic estrogen-like effects, in modulating the aquaporin-9 expression in the rat efferent ductules.

BACKGROUND: Fluid homeostasis is critical for normal function of the male reproductive tract and aquaporins (AQP) play an important role in maintenance of this water and ion balance. Several AQPs have been identified in the male, but their regulation is not fully comprehended. Hormonal regulation of AQPs appears to be dependent on the steroid in the reproductive tract region. AQP9 displays unique hormonal regulation in the efferent ductules and epididymis, as it is regulated by both estrogen and dihydrotestosterone (DHT) in the efferent ductules, but only by DHT in the initial segment epididymis. Recent data have shown that a metabolite of DHT, 5-alpha-androstane-3-beta-17-beta-diol (3-beta-diol), once considered inactive, is also present in high concentrations in the male and indeed has biological activity. 3-beta-diol does not bind to the androgen receptor, but rather to estrogen receptors ER-alpha and ER-beta, with higher affinity for ER-beta. The existence of this estrogenic DHT metabolite has raised the possibility that estradiol may not be the only estrogen to play a major role in the male reproductive system. Considering that both ER-alpha and ER-beta are highly expressed in efferent ductules, we hypothesized that the DHT regulation of AQP9 could be due to the 3-beta-diol metabolite. METHODS: To test this hypothesis, adult male rats were submitted to surgical castration followed by estradiol, DHT or 3-beta-diol replacement. Changes in AQP9 expression in the efferent ductules were investigated by using immunohistochemistry and Western blotting assay. RESULTS: Data show that, after castration, AQP9 expression was significantly reduced in the efferent ductules. 3-beta-diol injections restored AQP9 expression, similar to DHT and estradiol. The results were confirmed by Western blotting assay. CONCLUSION: This is the first evidence that 3-beta-diol has biological activity in the male reproductive tract and that this androgen metabolite has estrogen-like activity in the efferent ductules, whose major function is the reabsorption of luminal fluid.