C-type natriuretic peptide: a link between hyperandrogenism and anovulation in a mouse model of polycystic ovary syndrome.

The polycystic ovary (PCO) syndrome (PCOS) is the most common cause of anovulatory infertility in women and is associated with several clinical disorders. Despite the great amount of research in the area, mechanisms involved in the genesis of this syndrome remain poorly understood. In a recent issue of Clinical Science (vol. 132, issue 7, 759-776), Wang and colleagues, highlight the important role of overactivated C-type natriuretic peptide (CNP) and natriuretic peptide receptor 2 (CNP/NPR2) system in preventing oocyte maturation and ovulation in PCOS mice model induced by androgen. Dehydroepiandrosterone (DHEA) treatment caused anovulation, high levels of androgen and estrogen receptors (AR and ER) in the ovary, high expression of CNP and natriuretic peptide receptor 2 (NPR2) in granulosa cells (GC), and an increase in testosterone and estradiol (E2) levels in sera. The high level of CNP/NPR2 was associated with oocyte meiotic arrest and very low ovulation rate. Treatment with human chorionic gonadotropin (hCG) or inhibitors of AR or ER reduced the level of CNP/NPR2, which resulted in meiotic resumption and ovulation. The article provided important information for understanding the effect of ovarian steroids on control of oocyte maturation and fertility and highlighted CNP/NPR2 as a specific pathway that is potentially involved in the ovulatory disruption in PCOS.

Morphological study of apoptosis in granulosa cells and ovulation in a model of atresia in rat preovulatory follicles.

SummaryPrevious studies have established a model of atresia in preovulatory follicles after stimulation of immature rats with equine chorionic gonadotropin (eCG). This gonadotropin recruits a follicular pool and the deprivation of preovulatory luteinizing hormone (LH) surge induces the atresia in preovulatory follicles. The present study investigated the occurrence of ovulation and provided some morphological features of granulosa cell (GC) apoptosis of atretic follicles at 0, 48, 72 and 120 h after eCG stimulation. Histological sections of ovaries from untreated animals (0 h) showed primordial, primary, secondary and early antral follicles. After 48 h ovaries showed large antral follicles. Preovulatory follicles were observed at 72 h, and two out of five rats displayed cumulus-oocyte complexes (COCs) in the oviducts. All animals exhibited corpora lutea after 120 h. We observed increased estradiol (E2) levels 48 h after eCG treatment that might trigger an endogenous preovulatory gonadotropin surge. Higher progesterone (P4) level, which is the hallmark of a functional corpus luteum, was observed at 120 h. Atresia in secondary and antral follicles was observed by pyknotic granulosa cell nuclei in histology and positive immunolabelling for cleaved caspase 3. We also observed macrophages in secondary and antral follicles in atresia. Transmission electron microscopy revealed GCs with compacted chromatin against the nuclear envelope, nuclear fragmentation, cell shrinkage and fragmentation. No preovulatory follicles showed apoptosis of GCs. In conclusion, our results suggested the occurrence of an endogenous gonadotropin surge, promoting ovulation and preventing atresia of preovulatory follicles.

Effect of Polyhexamethylene Biguanide Solution on Bacterial Load and Biofilm in Venous Leg Ulcers: A Randomized Controlled Trial.

PURPOSE: The purpose of this study was to investigate the effect of polyhexamethylene biguanide (PHMB) solution as a wound cleanser on bacterial load and bacterial biofilm in venous leg ulcers. DESIGN: Randomized controlled trial. SUBJECTS AND SETTING: The target population was adults attending the dermatology outpatient clinic of the Clinical Hospital of the Federal University of Minas Gerais, Brazil. The sample comprised 44 patients with venous leg ulcers recruited over a 6-month period. METHODS: Participants were divided into 2 groups: the intervention group had their wounds cleansed with PHMB, and the control group had their wound cleansed with a 0.9% saline solution. Tissue fragments of the wounds were collected for bacteriological analysis; transmission electron microscopy was used to identify the presence of biofilm. RESULTS: The bacterial load was reduced in both groups compared to baseline values; no significant difference was found when groups were compared. Correlation analysis between wound duration (months), wound area (cm²), necrosis (%), variables, and bacterial count (colony forming units [CFUs]/g) after cleansing showed a significant relationship between area of the wound and CFUs/g (P = .0070, r = 0.51). Transmission electron microscopy revealed the presence of bacterial biofilm in the wounds after cleansing with both solutions. CONCLUSIONS: The results of this study indicate that both PHMB and saline solution are effective in reducing the bacterial load in venous leg ulcers. However, bacterial biofilm was present after cleansing with both solutions. These findings provide important evidence regarding effectiveness of 2 common wound cleansers on bacterial presence in wounds.

Evidence that angiotensin-(1-7) is an intermediate of gonadotrophin-induced oocyte maturation in the rat preovulatory follicle.

Several studies have shown the presence of components of the renin-angiotensin system in mammalian ovaries and their involvement in ovarian physiology. We have previously shown the presence of angiotensin-(1-7) [Ang-(1-7)], an important biologically active component of the renin-angiotensin system, and its receptor, Mas, in rat, rabbit and human ovaries. We have also shown the involvement of Ang-(1-7) in the rabbit ovulatory process in vitro. In the present study, we observed that Ang-(1-7) stimulated the resumption of meiosis in oocytes of rat preovulatory follicles, reaching more than 30% of oocytes with germinal vesicle breakdown. The specific antagonist of the Mas receptor, A-779, inhibited the germinal vesicle breakdown induced by Ang-(1-7) and reduced the oocyte maturation stimulated by luteinizing hormone (LH). Immunohistochemistry showed that LH increased both Ang-(1-7) and angiotensin-converting enzyme 2 (ACE2) staining in preovulatory follicles. The effect of gonadotrophins on mRNA expression of Mas and ACE2 in ovaries of immature equine chorionic gonadotrophin-primed rats was analysed by real-time PCR after 6 h of human chorionic gonadotrophin (hCG) injection, which exhibits LH-like effects. After hCG treatment, ACE2 mRNA expression was higher in the ovaries of treated rats than in the ovaries of control rats, whereas Mas mRNA levels were unchanged. A-779 changed the steroidogenesis stimulated by LH. An increased testosterone concentration and decreased progesterone levels were measured in the follicle medium. In conclusion, our results suggest that LH upregulates the ACE2-Ang-(1-7)-Mas axis and that Ang-(1-7) promotes meiotic resumption, possibly as a gonadotrophin intermediate.

Disruption of polycystin-1 cleavage leads to cardiac metabolic rewiring in mice.

Cardiovascular manifestations account for marked morbi-mortality in autosomal dominant polycystic kidney disease (ADPKD). Pkd1- and Pkd2-deficient mice develop cardiac dysfunction, however the underlying mechanisms remain largely unclear. It is unknown whether impairment of polycystin-1 cleavage at the G-protein-coupled receptor proteolysis site, a significant ADPKD mutational mechanism, is involved in this process. We analyzed the impact of polycystin-1 cleavage on heart metabolism using Pkd1V/V mice, a model unable to cleave this protein and with early cardiac dysfunction. Pkd1V/V hearts showed lower levels of glucose and amino acids and higher lipid levels than wild-types, as well as downregulation of p-AMPK, p-ACCß, CPT1B-Cpt1b, Ppara, Nppa and Acta1. These findings suggested decreased fatty acid ß-oxidation, which was confirmed by lower oxygen consumption by Pkd1V/V isolated mitochondria using palmitoyl-CoA. Pkd1V/V hearts also presented increased oxygen consumption in response to glucose, suggesting that alternative substrates may be used to generate energy. Pkd1V/V hearts displayed a higher density of decreased-size mitochondria, a finding associated with lower MFN1, Parkin and BNIP3 expression. These derangements were correlated with increased apoptosis and inflammation but not hypertrophy. Notably, Pkd1V/V neonate cardiomyocytes also displayed shifts in oxygen consumption and p-AMPK downregulation, suggesting that, at least partially, the metabolic alterations are not induced by kidney dysfunction. Our findings reveal that disruption of polycystin-1 cleavage leads to cardiac metabolic rewiring in mice, expanding the understanding of heart dysfunction associated with Pkd1 deficiency and likely with human ADPKD.

The superior beneficial effects of exercise training versus hormone replacement therapy on skeletal muscle of ovariectomized rats.

Previous studies have highlighted the positive effects of Estradiol (E2) replacement therapy and physical exercise on skeletal muscle during menopause. However, the comparison effects of exercise training (ET) and estradiol replacement therapy during menopause on skeletal muscle have not been investigated to date. This study aimed to compare the effects of endurance exercise training versus E2 replacement therapy on mitochondrial density, redox status, and inflammatory biomarkers in the skeletal muscle of ovariectomized rats. Thirty female Wistar rats (12-week-old) were randomly assigned into three groups: Untrained ovariectomized rats (UN-OVX, n = 10); untrained ovariectomized rats treated with estradiol replacement therapy (E2-OVX); and, trained ovariectomized rats (TR-OVX). After ovariectomy, the E2-OVX rats were treated subcutaneously with E2 (implanted Silastic® capsule containing 360 µg of 17ß-estradiol/mL) while the TR-OVX group performed an exercise training protocol (50-70% of maximal running speed on a treadmill, 60 min/day, 5 days/week for 8 weeks). After euthanasia, the soleus muscle was processed for histological and biochemical evaluations. Only exercise prevented the reduction of maximal oxygen consumption and increased mechanical efficiency (ME). While mitochondrial muscle density, total antioxidant capacity (FRAP), catalase (CAT) activity, and interleukin 10 levels were higher in TR-OVX, only OVX-E2 presented higher CAT activity and lower interleukin 6 levels. Endurance exercise training compared with E2 replacement therapy maintains the aerobic capacity improving the ME of OVX rats. In addition, only endurance exercise training raises the skeletal muscle mitochondrial content and tends to balance the redox and inflammatory status in the skeletal muscle of OVX rats.

Superior cardiometabolic and cellular adaptive responses to multiple versus single daily sessions of high-intensity interval training in Wistar rats.

This study aimed to compare in rats the cardiometabolic and cellular adaptative responses to 8 weeks of high-intensity interval training (HIIT) performed in a single (1xHIIT) or three shorter daily sessions (3xHIIT). Male Wistar rats were assigned to untrained (n = 10), 1xHIIT (n = 10), and 3xHIIT (n = 10) groups. Both HIIT groups performed 15 min of a treadmill run five times per week for 8 weeks. The 1xHIIT performed single daily sessions of 15 min, and the 3xHIIT performed three daily sessions of 5 min with an interval of 4 h between sessions. Resting VO2 and VO2max were measured using a metabolic chamber; blood pressure and heart rate were measured by plethysmography; body composition was estimated by DEXA; Glucose and insulin tolerance tests were performed; after euthanasia, hearts, gastrocnemius, and visceral fat were harvested for analysis of cardiac function, histology, and morphology. Mitochondrial densities of the gastrocnemius and left ventricle muscles were determined by electron microscopy. 3xHIIT induced similar positive adaptative responses to 1xHIIT on resting VO2 and VO2max, cardiac function, and mitochondria density. 3xHIIT was superior to 1xHIIT in reducing visceral fat weight and adipocyte size and improving insulin tolerance. Multiple short daily bouts of HIIT may be superior to single HIIT daily sessions in improving cardiometabolic and cellular adaptations in rats.

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.

Cardioprotective effects of severe calorie restriction from birth in adult ovariectomized rats.

AIMS: Menopause is a female condition induced by a reduction of ovarian hormone and is related to an increase in cardiovascular diseases in women. We have shown that severe calorie restriction (SCR) from birth reduces the cardiometabolic risk in adult male Wistar rats. In this study, we investigated the effects of SCR from birth to adulthood on cardiovascular function of ovariectomized rats. MAIN METHODS: From birth to adulthood, rats were daily fed ad libitum (control group - C) or with 50% of the amount consumed by the control group (calorie-restricted group - R). At 90 days, half of the rats in each group underwent bilateral ovariectomy (OVX), totaling 4 groups: C-Sham, C-OVX, R-Sham, R-OVX. Systolic blood pressure (SBP), heart rate (HR) and, double product (DP) index were recorded by tail-cuff plethysmography. Cardiac function was analyzed by the Langendorff technique and cardiomyocyte diameter was accessed by histologic analysis. Additionally, cardiac SERCA2 content and redox status were evaluated. KEY FINDINGS: C-OVX rats exhibited reduced cardiac function and cardiac non-enzymatic total antioxidant capacity (TAC). R-Sham animals showed reduced SBP, DP, HR, improved cardiac function, reduced cardiac protein carbonyl derivatives and increased TAC, catalase, and superoxide dismutase activities. R-OVX rats maintained reduced SBP, DP, HR, and increased contractility and relaxation indexes. R-Sham and R-OVX rats exhibited preserved heart mass and reduced cardiomyocyte diameter. Cardiac SERCA2 content did not differ between the groups. SIGNIFICANCE: Taken together, our findings show cardioprotective effects of SCR from birth in adult ovariectomized rats.

Genetic deletion of the Angiotensin-(1-7) receptor Mas leads to a reduced ovulatory rate.

Angiotensin-(1-7) [Ang-(1-7)] is a component of Renin-Angiotensin System (RAS) that acts through activation of the G-protein-coupled receptor Mas. Recent studies highlight Ang-(1-7) as an intermediate of gonadotropin in ovarian physiology. Genetically Mas-deficient mice allow the investigation of Ang-(1-7) in the ovulatory process. Therefore, the present study aimed to analyze the effects of Mas gene deletion on ovulation to confirm our hypothesis that Mas Knockout (Mas-KO) mice exhibit impairment in the ovulatory outcome. First, we evaluated the breeding data from our animal facilities and from a breeding experiment. The ovulation was observed directly from oviducts after a superovulation protocol and in the estrus morning. We also checked the follicular pool and mRNA expression of Insulin-like growth factor-1 (IGF-1) in ovaries to investigate a possible reason underlying the reduced ovulation. Mas-KO mice showed a reduced litter size and decreased spontaneous ovulatory rate. Ovarian stimulation by gonadotropins reversed ovulation outcome in Mas-KO mice. Mas deficiency also promoted a reduced ovarian follicular pool and lower IGF-1 mRNA levels, suggesting that Mas receptor plays a role in the survival of ovarian follicle. The reduction of ovulatory rate highlights the relevance of Ang-(1-7)/Mas axis in female reproduction, probably through a reduction of IGF-1 mRNA levels.

Physical activity prevents alterations in mitochondrial ultrastructure and glucometabolic parameters in a high-sugar diet model.

Endurance exercise is a remarkable intervention for the treatment of many diseases. Mitochondrial changes on skeletal muscle are likely important for many of the benefits provided by exercise. In this study, we aimed to evaluate the effects that a regular physical activity (swimming without workload) has on mitochondrial morphological alterations and glucometabolic parameters induced by a high-sugar diet (HSD). Weaned male Wistar rats fed with a standard diet or a HSD (68% carbohydrate) were subjected to 60 minutes of regular physical activity by swimming (without workload) for four- (20 sessions) or eight-week (40 sessions) periods. After training, animals were euthanized and the sera, adipose tissues, and skeletal muscles were collected for further analysis. The HSD increased body weight after an 8-week period; it also increased the fat pads and the adipose index, resulting in glucose intolerance and insulin resistance (IR). Transmission electron microscopy showed an increase in alterations of mitochondrial ultrastructure in the gastrocnemius muscle, as well as a decrease in superoxide dismutase (SOD) activity, and an increase in protein carbonylation. Regular physical activity partially reverted these alterations in rats fed a HSD, preventing mitochondrial morphological alterations and IR. Moreover, we observed a decrease in Pgc1α expression (qPCR analysis) in STD-EXE group and a less pronounced reduction in HSD-EXE group after an 8-week period. Thus, regular physical activity (swimming without workload) in rats fed a HSD can prevent mitochondrial dysfunction and IR, highlighting the crucial role for physical activity on metabolic homeostasis.

Comparative morphology of the oocyte surface and early development in four characiformes from the São Francisco River, Brazil.

Early development from the egg fertilization to complete resorption of the yolk-sac is a critical period in the life cycle of teleost fish. Knowledge of this process provides essential parameters for aquaculture and identification of spawning sites in the wild. In the present study, a comparative morphological analysis of the oocyte surface as well as early development was performed in four commercially valuable species from the São Francisco River: Brycon orthotaenia, Leporinus obtusidens, Prochilodus argenteus, and Salminus franciscanus. Stripped oocytes, embryo, and yolk-sac larvae were analyzed by scanning electron microscopy (SEM) and histology. A set of 10 lectins was used for investigation of lectin-binding pattern in oocytes. In the four species, the outer layer of the zona radiata reacted to most lectins, indicating complex polysaccharides at the oocyte surface while no reactivity was detected in the inner zona radiata and yolk globules. Typical structural arrangements were recognized at the micropylar region by SEM. The four species showed nonadhesive eggs, short embryonic period (18-20 h at 24 ± 1°C), and poorly developed larvae at hatching. At 24 h posthatching (hph), larvae of the four species had neuromasts on the body surface. Rudimentary cement glands for larval attachment were identified on the cephalic region at 24 and 48 hph in B. orthotaenia and S. franciscanus, and following they were in regression. The time for whole yolk resorption varied among species from 48 to 120 hph, occurring earlier in S. franciscanus, followed by B. orthotaenia, P. argenteus, and L. obtusidens. The formation of the digestive tract and the mouth opening indicated initiation of exogenous feeding 24 h before complete resorption of the yolk. Together, our data indicate similarities in the early development among species that may be related to the life cycle strategies and phylogeny.

The effect of angiotensin-converting enzyme inhibition throughout a superovulation protocol in ewes.

Many studies identified new components of the renin­angiotensin system (RAS), such as Angiotensin-(1-7) [Ang-(1­7)] and Angiotensin-converting enzyme type 2 (ACE2), in mammalian ovaries.We previously showed Angiotensin-Converting Enzyme (ACE) inhibition, which increases the level of Ang-(1­7), stimulated ovarian estradiol output in ewe after estrous synchronization. Considering that Ang-(1­7) stimulates ovarian function and elevated estradiol before ovulation is associated with increased chance of achieving pregnancy, the present study investigated whether ACE inhibition throughout a superovulation protocol in ewe might improve ovulation outcome. At first, immunohistochemistry in ovaries of nonpregnant ewes revealed localization of Angiotensin II (Ang II), Ang-(1­7) and ACE2 in theca cells of antral follicles and in corpus luteum. Ang II and Ang-(1­7)were also detected in follicular fluid (FF) by Radioimmunoassay (RIA). Enalapril treatment throughout the superovulation protocol decreased 17ß-estradiol (E2) output and raised progesterone:estradiol (P4:E2) ratio without a direct influence on ovulation and quality of embryos.

Downregulation of natriuretic peptide system and increased steroidogenesis in rat polycystic ovary.

Atrial natriuretic peptide (ANP) is known to regulate ovarian functions, such as follicular growth and steroid hormone production. The aim of the present study was to investigate the natriuretic peptide system in a rat model of chronic anovulation, the rat polycystic ovary. Adult female Wistar rats received a single subcutaneous injection of 2mg estradiol valerate to induce polycystic ovaries, while the control group received vehicle injection. Two months later, their ovaries were quickly removed and analyzed. Polycystic ovaries exhibited marked elevation of testosterone and estradiol levels compared to control ovaries. The levels of ANP and the expression of ANP mRNA were highly reduced in the polycystic ovaries compared to controls. By immunohistochemistry, polycystic ovaries showed weaker ANP staining in stroma, theca cells and oocytes compared to controls. Polycystic ovaries also had increased activity of neutral endopeptidase, the main proteolytic enzyme that degrades natriuretic peptides. ANP receptor C mRNA was reduced and ANP binding to this receptor was absent in polycystic ovaries. Collectively, these results indicate a downregulation of the natriuretic peptide system in rat polycystic ovary, an established experimental model of anovulation with high ovarian testosterone and estradiol levels. Together with previous evidence demonstrating that ANP inhibits ovarian steroidogenesis, these findings suggest that low ovarian ANP levels may contribute to the abnormal steroid hormone balance in polycystic ovaries.

Preparation, physicochemical characterization, and cell viability evaluation of long-circulating and pH-sensitive liposomes containing ursolic acid.

Cancer is one of the leading causes of death worldwide. Although several drugs are used clinically, some tumors either do not respond or are resistant to the existing pharmacotherapy, thus justifying the search for new drugs. Ursolic acid (UA) is a triterpene found in different plant species that has been shown to possess significant antitumor activity. However, UA presents a low solubility in aqueous medium, which presents a barrier to its biological applications. In this context, the use of liposomes presents a promising strategy to deliver UA and allow for its intravenous administration. In this work, long-circulating and pH-sensitive liposomes containing UA (SpHL-UA) were developed, and their chemical and physicochemical properties were evaluated. SpHL-UA presented adequate properties, including a mean diameter of 191.1 ± 6.4 nm, a zeta potential of 1.2 ± 1.4 mV, and a UA entrapment of 0.77 ± 0.01 mg/mL. Moreover, this formulation showed a good stability after having been stored for 2 months at 4 °C. The viability studies on breast (MDA-MB-231) and prostate (LNCaP) cancer cell lines demonstrated that SpHL-UA treatment significantly inhibited cancer cell proliferation. Therefore, the results of the present work suggest the applicability of SpHL-UA as a new and promising anticancer formulation.

Tissue specific localization of angiotensin-(1-7) and its receptor Mas in the uterus of ovariectomized rats.

The vasoactive peptide angiotensin (Ang)-(1-7) has vasodilator, antifibrotic and antihypertrophic properties, but little is known about its regulation in the uterus. The aim of this study was to evaluate Ang-(1-7) and its receptor Mas expression throughout rat uterine tissues, in ovariectomized animals treated with estrogen alone or combined with progestin. Adult Wistar rats (n = 19) were ovariectomized and randomly assigned into three different groups 1 week later. One group received a single dose of estradiol benzoate (1.5 mg/kg, i.m. injection, n = 6). Another group received estradiol associated with depot medroxyprogesterone acetate (3 mg/kg, i.m. injection, n = 6). Control group (n = 7) received oil injection. One week later, the rats were euthanized and their uteri were fixed and stained by immunohistochemistry, using a polyclonal antibody specific to Ang-(1-7) and its receptor Mas. Ang-(1-7) was detected in all uterine tissues, but it was weak or absent in the circular myometrium of treated animals. The intensity of the immunostaining decreased in the glandular epithelium of hormonally treated animals when compared to controls. In estrogen treated rats, Ang-(1-7) labeling was scattered and sometimes included the nuclei of glandular cells. We also detected Ang-(1-7) expression in longitudinal myometrium and uterine serosa. Mas receptor was present in all tissues with similar intensity among the tissue types in the control and estrogen plus progestin groups. In the estrogen group, Mas staining was stronger in the luminal and glandular epithelium when compared with stroma or circular myometrium. In conclusion, ovarian steroids are not required to allow endometrial expression of Ang-(1-7) and its receptor Mas in rats, as it remains abundant in ovariectomized animals. However, estrogen and progestin may modulate the distribution pattern of this peptide in the endometrium, especially in the glandular compartment.

Gonadotropin stimulation increases the expression of angiotensin-(1--7) and MAS receptor in the rat ovary.

We have previously shown the presence of immunoreactive angiotensin-(1-7) [Ang-(1-7)] in rat ovary homogenate and its stimulatory effect on estradiol and progesterone production in vitro. In the current study, we investigated the presence and cellular distribution of Ang-(1-7) and the Mas receptor, the expression of Mas and angiotensin-converting enzyme 2 (ACE2) messenger RNA (mRNA), and the enzymatic activity in the rat ovary following gonadotropin stimulation in vivo. Immature female Wistar rats (25 days old) were injected subcutaneously (SC) with equine chorionic gonadotropin (eCG, 20 IU in 0.2 mL) or vehicle 48 hours before euthanasia. Tissue distributions of Ang-(1-7), Mas receptor, and ACE2 were evaluated by immunohistochemistry, along with angiotensin II (Ang II) localization, while the mRNA expression levels of Mas receptor and ACE2 were evaluated by real-time polymerase chain reaction (PCR). In addition, we determined the activity of neutral endopeptidase (NEP), prolyl endopeptidase (PEP), and ACE by fluorometric assays. After eCG treatment, we found strong immunoreactivity for Ang-(1-7) and Mas primarily in the theca-interstitial cells, while Ang II appeared in the granulosa but not in the thecal layer. Equine chorionic gonadotropin treatment increased Mas and ACE2 mRNA expression compared with control animals (3.3- and 2.1-fold increase, respectively; P < .05). Angiotensin-converting enzyme and NEP activities were lower, while PEP activity was higher in the eCG-treated rats (P < .05). These data show gonadotropin-induced changes in the ovarian expression of Ang-(1-7), Mas receptor, and ACE2. These findings suggest that the renin-angiotensin system (RAS) branch formed by ACE2/Ang-(1-7)/Mas, fully expressed in the rat ovary and regulated by gonadotropic hormones, could play a role in the ovarian physiology.