Reproductive morphophysiology of an iteroparous batch spawner fish: from early follicle development to ovulation.

Batch spawner fishes develop successive clutches of oocytes which allows them to participate in many reproductive cycles during their adult life (iteroparous) and spawn in multiple events within each breeding cycle. Here, ovarian follicular development was morpho-functionally analyzed in females of the iteroparous batch spawner fish Gymnocorymbus ternetzi. To obtain better insights into the reproductive morpho-physiology in batch spawners, the objective of this research was to analyze the dynamics of the follicular development, with its hormonal regulation between two active reproduction events. We found that over 16 days, follicles progressed asynchronously to chromatin nucleolus, Primary and Secondary growth stages of oogenesis with progressive secretion of 17ß-estradiol (E2). During the end of secondary growth, the increase in 17α,20ß-dihydroxy-4-pregnen-3-one (17,20ß-p) was measured relative to the maturation process of the ovarian follicles (e.g., nuclear migration and its rupture during the resumption of meiosis). Interestingly, an additional increase in E2 was observed after fish reproduction, probably related to the recruitment of new batch follicles for secondary growth. We also measured the high values of multiple condition factor post-reproduction measurements, reflecting more energy invested during the pre-reproductive process. We also quantified high concentrations of 17,20ß-p, probably related to the recruitment of a new batch of oogonia to meiosis, presumably secreted by post-ovulatory follicles, after fish reproduction. We finally found that fish without exposure to reproductive stimulus developed a regression phase at day 24, characterized by massive follicle atresia, that allow them to recycle energy and constitutive materials of the follicles invested during oogenesis for another reproductive cycle.

Activation of G Protein-Coupled Estrogen Receptor (GPER) Negatively Modulates Cardiac Excitation-Contraction Coupling (ECC) through the PI3K/NOS/NO Pathway.

The G-protein-coupled estrogen receptor (GPER) has been described to exert several cardioprotective effects. However, the exact mechanism involved in cardiac protection remains unclear. The aim of this study is to investigate the role of GPER activation on excitation-contraction coupling (ECC) and the possibility that such effect participates in cardioprotection. The cardiac myocytes of male Wistar rats were isolated with a digestive buffer and loaded with Fura-2-AM for the measurement of intracellular calcium transient (CaT). Sarcomere shortening (SS) and L-type calcium current (ICaL) were also registered. The confocal technique was used to measure nitric oxide (NO) production in cells loaded with DAF-FM-diacetate. Cardiac myocytes exposed to 17-ß-estradiol (E2, 10 nM) or G-1 (1 µM) for fifteen minutes decreased CaT, SS, and ICaL. These effects were prevented using G-36 (antagonist of GPER, 1 µM), L-Name (NO synthase -NOS- inhibitor, 100 nM), or wortmannin (phosphoinositide-3-kinase -PI3K- inhibitor, 100 nM). Moreover, G1 increased NO production, and this effect was abolished in the presence of wortmannin. We concluded that the selective activation of GPER with E2 or G1 in the isolated cardiac myocytes of male rats induced a negative inotropic effect due to the reduction in ICaL and the decrease in CaT. Finally, the pathway that we proposed to be implicated in these effects is PI3K-NOS-NO.

Sexual hormones changes in burning mouth syndrome: A systematic review.

BACKGROUND: Burning mouth syndrome (BMS) is a chronic pain condition affecting the oral cavity. This condition mostly affects peri- or postmenopausal women; for this reason, sexual hormonal changes have been implicated in BMS pathogenesis. METHODS: A systematic review was performed in MEDLINE/PubMed, Scopus, Web of Science, Cochrane Library and EMBASE without restriction for language or year. Eligibility criteria were controlled studies addressing the PICO question: (P) patients with BMS; (I) detection of the sex hormones; (C) patients without BMS; (O) changes on sexual hormones as a risk factor for BMS severity. Risk of bias was performed with Newcastle-Ottawa Quality Assessment Scale. RESULTS: Four studies were included. Salivary levels were evaluated in three studies and serum blood was used in one. Three studies analysed oestradiol and/or dehydroepiandrosterone (DHEA), two assessed progesterone and one evaluated follicle-stimulating hormone (FSH). Oestradiol results were contradictory, with two studies reporting lower levels in BMS patients compared to controls and one finding the opposite. DHEA was statistically lower in the BMS group in one study. Progesterone showed opposite results in two studies, although none with statistical significance. FSH was statistically higher in the BMS group compared to controls. Correlation of hormones with quality of life was performed in three studies and there was no significant correlation with self-perceived symptoms severity. CONCLUSION: Sexual hormones can be altered in BMS, especially oestradiol. Despite these changes, we did not find correlation between hormone fluctuation and BMS symptoms intensity affecting quality of life. These findings suggested the need for further investigation on hormonal alterations, which may be a promising target on BMS management.

The modulatory role of serotonin-1A receptors of the basolateral amygdala and dorsal periaqueductal gray on the impact of hormonal variation on the conditioned fear response.

Despite significant advances in the study of fear and fear memory formation, little is known about fear learning and expression in females. This omission has been proven surprising, as normal and pathological behaviors are highly influenced by ovarian hormones, particularly estradiol and progesterone. In the current study, we investigated the joint influence of serotonin (5-HT) neurotransmission and estrous cycle phases (low or high levels of estradiol and progesterone) on the expression of conditioned fear in a group of female rats that were previously divided according to their response to stressful stimuli into low or high anxiety-like subjects. The baseline amplitude of the unconditioned acoustic startle responses was high in high-anxiety female rats, with no effect on the estrous cycle observed. Data collected during the proestrus-estrus phase revealed that low-anxiety rats had startle amplitudes similar to those of high-anxiety rats. It is supposed that high-anxiety female rats benefit from increased estradiol and progesterone levels to achieve comparable potentiated startle amplitudes. In contrast, female rats experienced a significant decrease in hormone levels during the Diestrus phase. This decrease is believed to play a role in preventing them from displaying a heightened startle response when faced with strongly aversive stimuli. Data collected after 5-HT and 8-OH-DPAT were administered into the basolateral nuclei and dorsal periaqueductal gray suggest that 5-HT neurotransmission works with progesterone and estrogen to reduce startle potentiation, most likely by activating the serotonin-1A receptor subtype.

Proliferation and apoptosis dynamics of the normal canine mammary gland during the oestrous cycle evaluated by Ki-67 and Caspase-3 indexes.

Understanding the normal physiology of the canine mammary gland (CMG) is crucial, as it provides a foundational reference for understanding canine mammary neoplasms. The relation between the Proliferation Index (PI) indicated by Ki-67 expression, along with the Apoptotic Index (AI) determined through Caspase-3 expression during the oestrous cycle, is inadequately documented in existing literature. This study seeks to offer insights into the interplay between PI and AI in the CMG across oestrous cycle phases. An extensive investigation was conducted on a diverse case series of bitches (n = 18). Oestrous cycle stages were determined through vaginal cytology, histological examination of the reproductive tract and serum progesterone and oestradiol concentrations. The entire mammary chain was histologically examined, and proliferation and apoptosis were assessed via double immunohistochemistry employing anti-Ki-67 and Caspase-3 antibodies. PI and AI were evaluated through a systematic random sampling approach, counting a minimum of 200 cells for each cell type. There was a significantly higher PI during early dioestrus in all mammary gland components, with a greater proportion of positive cells observed in epithelial cells compared to stromal cells. The highest PI was detected in epithelial cells within the end buds. Significant differences were found in Ki-67 labelling across the cranial mammary glands. A positive and strong correlation was noted between progesterone concentration and PI in epithelial cells. The AI remained consistently low throughout the oestrous cycle, with few differences observed across histological components. Caspase-3 labelling displayed the highest positivity in caudal mammary pairs. A negative and moderate correlation was identified between progesterone concentration and AI in interlobular mesenchymal cells. This study highlights the influence of endocrine regulation on cell proliferation indices in mammary tissue, emphasizing the need to consider these hormonal variations in toxicopathological studies involving canine mammary gland.

Tauroursodeoxycholate prevents estradiol 17ß-d-glucuronide-induced cholestasis and endocytosis of canalicular transporters by switching off pro-cholestatic signaling pathways.

AIMS: Estradiol 17ß-d-glucuronide (E217G) induces cholestasis by triggering endocytosis and further intracellular retention of the canalicular transporters Bsep and Mrp2, in a cPKC- and PI3K-dependent manner, respectively. Pregnancy-induced cholestasis has been associated with E217G cholestatic effect, and is routinely treated with ursodeoxycholic acid (UDCA). Since protective mechanisms of UDCA in E217G-induced cholestasis are still unknown, we ascertained here whether its main metabolite, tauroursodeoxycholate (TUDC), can prevent endocytosis of canalicular transporters by counteracting cPKC and PI3K/Akt activation. MAIN METHODS: Activation of cPKC and PI3K/Akt was evaluated in isolated rat hepatocytes by immunoblotting (assessment of membrane-bound and phosphorylated forms, respectively). Bsep/Mrp2 function was quantified in isolated rat hepatocyte couplets (IRHCs) by assessing the apical accumulation of their fluorescent substrates, CLF and GS-MF, respectively. We also studied, in isolated, perfused rat livers (IPRLs), the status of Bsep and Mrp2 transport function, assessed by the biliary excretion of TC and DNP-SG, respectively, and Bsep/Mrp2 localization by immunofluorescence. KEY FINDINGS: E217G activated both cPKC- and PI3K/Akt-dependent signaling, and pretreatment with TUDC significantly attenuated these activations. In IRHCs, TUDC prevented the E217G-induced decrease in apical accumulation of CLF and GS-MF, and inhibitors of protein phosphatases failed to counteract this protection. In IPRLs, E217G induced an acute decrease in bile flow and in the biliary excretion of TC and DNP-SG, and this was prevented by TUDC. Immunofluorescence studies revealed that TUDC prevented E217G-induced Bsep/Mrp2 endocytosis. SIGNIFICANCE: TUDC restores function and localization of Bsep/Mrp2 impaired by E217G, by preventing both cPKC and PI3K/Akt activation in a protein-phosphatase-independent manner.

NADPH oxidase-generated reactive oxygen species are involved in estradiol 17ß-d-glucuronide-induced cholestasis.

The endogenous metabolite of estradiol, estradiol 17ß-D-glucuronide (E17G), is considered the main responsible of the intrahepatic cholestasis of pregnancy. E17G alters the activity of canalicular transporters through a signaling pathway-dependent cellular internalization, phenomenon that was attributed to oxidative stress in different cholestatic conditions. However, there are no reports involving oxidative stress in E17G-induced cholestasis, representing this the aim of our work. Using polarized hepatocyte cultures, we showed that antioxidant compounds prevented E17G-induced Mrp2 activity alteration, being this alteration equally prevented by the NADPH oxidase (NOX) inhibitor apocynin. The model antioxidant N-acetyl-cysteine prevented, in isolated and perfused rat livers, E17G-induced impairment of bile flow and Mrp2 activity, thus confirming the participation of reactive oxygen species (ROS) in this cholestasis. In primary cultured hepatocytes, pretreatment with specific inhibitors of ERK1/2 and p38MAPK impeded E17G-induced ROS production; contrarily, NOX inhibition did not affect ERK1/2 and p38MAPK phosphorylation. Both, knockdown of p47phox by siRNA and preincubation with apocynin in sandwich-cultured rat hepatocytes significantly prevented E17G-induced internalization of Mrp2, suggesting a crucial role for NOX in this phenomenon. Concluding, E17G-induced cholestasis is partially mediated by NOX-generated ROS through internalization of canalicular transporters like Mrp2, being ERK1/2 and p38MAPK necessary for NOX activation.

Biochemical profiling of the follicular environment to predict oocyte competence in cattle.

To identify markers of oocyte competence, we compared the biochemical characteristics of fluid and cells from follicles containing oocytes with different capacities to form an embryo. Follicles (5-6 mm) were dissected, and follicular fluid (FF), granulosa cells (GC), cumulus cells (CC) from immature and mature cumulus-oocyte-complexes (COC) were individually collected. The oocytes were matured, fertilized, and cultured individually until day 8 (D8) of development. On D8, the samples were grouped according to embryo production into those that gave rise to blastocysts (EMB) and those that did not reach the blastocyst stage (NEMB). In CCs from immature and mature COCs and GCs, expression of CASP3, SERPINE2, VCAN, LUM, FSHR, EGFR, PGR, and GHR genes was quantified. Cell-free DNA (cfDNA), progesterone, and estradiol concentrations in the FF were determined. Data were analyzed by Mann-Whitney U test (GraphPad Prism 9). GHR was highly expressed in immature CCs from the EMB group, whereas CASP3 was highly expressed in mature CCs from the NEMB group (P<0.05). During maturation, the expression of CASP3 and GHR genes increased only in the NEMB group. ART2 cfDNA was highly detected in FF of the NEMB compared to the EMB group. Progesterone concentration was similar between the groups, whereas estradiol concentration was higher (P<0.05) in the EMB than in the NEMB group. It was concluded that a higher level of GHR transcripts in immature CCs, lower CASP3 expression in CCs from matured COCs, lower levels of ART2, and higher estradiol concentrations in FF may indicate oocytes with greater potential for development.

Branching morphogenesis of the mouse mammary gland after exposure to benzophenone-3.

Pubertal mammary branching morphogenesis is a hormone-regulated process susceptible to exposure to chemicals with endocrine disruptive capacity, such as the UV-filter benzophenone-3 (BP3). Our aim was to assess whether intrauterine or in vitro exposure to BP3 modified the branching morphogenesis of the female mouse mammary gland. For this, pregnant mice were dermally exposed to BP3 (0.15 or 50 mg/kg/day) from gestation day (GD) 8.5 to GD18.5. Sesame oil treatment served as control. Changes of the mammary glands of the offspring were studied on postnatal day 45. Further, mammary organoids from untreated mice were cultured under branching induction conditions and exposed for 9 days to BP3 (1 × 10-6 M, 1 × 10-9 M, or 1 × 10-12 M with 0.01% ethanol as control) to evaluate the branching progression. Mice that were exposed to BP3 in utero showed decreased mRNA levels of progesterone receptor (PR) and WNT4. However, estradiol and progesterone serum levels, mammary histomorphology, proliferation, and protein expression of estrogen receptor alpha (ESR1) and PR were not significantly altered. Interestingly, direct exposure to BP3 in vitro also decreased the mRNA levels of PR, RANKL, and amphiregulin without affecting the branching progression. Most effects were found after exposure to 50 mg/kg/day or 1 × 10-6 M of BP3, both related to sunscreen application in humans. In conclusion, exposure to BP3 does not impair mammary branching morphogenesis in our models. However, BP3 affects PR transcriptional expression and its downstream mediators, suggesting that exposure to BP3 might affect other developmental stages of the mammary gland.

The differential sensitivity of the hypothalamic-hypophysial-ovarian axis to 5-hydroxytryptophan alters the secretion of estradiol.

Serotonin [5-hydroxytryptamine (5-HT)] modulates ovarian function. The precursor of 5-HT, 5-hydroxytryptophan (5-HTP), has been used to treat depression. However, the effects of 5-HTP on ovarian and reproductive physiology remain unknown. In this research, we analysed the impact of 5-HTP on the monoaminergic system and its interactions with the reproductive axis and ovarian estradiol secretion when administered by distinct routes. Female rats 30 days of age were injected with 5-HTP i.p. (100 mg/kg), into the ovarian bursa (1.5 µg/40 µL) or into the median raphe nucleus (20 µg/2.5 µL) and were killed 60 or 120 min after injection. As controls, we used rats of the same age injected with vehicle (0.9% NaCl). Monoamine, gonadotrophin and steroid ovarian hormone concentrations were measured. The injection of 5-HTP either i.p. or directly into the ovarian bursa increased the concentrations of 5-HT and the metabolite 5-hydroxyindole-3-acetic acid in the ovary. For both routes of administration, the serum concentration of estradiol increased. After i.p. injection of 5-HTP, the concentrations of luteinizing hormone were decreased and follicle-stimulating hormone increased after 120 min. Micro-injection of 5-HTP into the median raphe nucleus increased the concentrations of 5-HT in the anterior hypothalamus and dopamine in the medial hypothalamus after 120 min. Our results suggest that the administration of 5-HTP either i.p. or directly into the ovarian bursa enhances ovarian estradiol secretion.

Incubation as a driver of maternal effects: Temperature influences levels of yolk maternally derived 5α-dihydrotestosterone.

In birds, maternal hormones deposited into eggs in response to environmental stimuli can impact offspring phenotype. Although less studied, environmental conditions can also influence females' incubation behavior, which might play a role in regulating embryo exposure to maternal hormones through changes in incubation temperature that affect the activity of the enzymes responsible for converting testosterone (T) to 5α-dihydrotestosterone (DHT) or estradiol. Here, we tested the hypothesis that the initial T content of the yolk and incubation temperature determine exposure to T metabolites during early embryo development. In the Japanese quail (Coturnix japonica), we experimentally manipulated yolk T and incubation temperature (38° C versus 36° C) and analyzed DHT and estradiol titers on day four of incubation. We found that eggs with experimentally increased T and those incubated at 36° C showed higher DHT concentration in egg yolk (with no synergistic effect of the two treatments). Estradiol titers were not affected by T manipulation or incubation temperature. Our study suggests that incubation temperature influences DHT titers and may act as an understudied source of maternal influence on offspring phenotype.

Pyometra alters the redox status and expression of estrogen and progesterone receptors in the uterus of domestic cats.

OBJECTIVES: The aim of this study was to evaluate the expression profile of sex steroid receptors and redox mediators in the uterus of domestic cats with pyometra. METHODS: Twelve cats were used and divided into groups: (1) non-gestational healthy diestrus (n = 7) and (2) pyometra (n = 5). The plasma profiles of estradiol and progesterone (P4) as well as uterine expression levels of estradiol alpha (ERα), progesterone (PR) and androgen (AR) receptors, of the antioxidant enzymes superoxide dismutase 1 (SOD1), catalase and glutathione peroxidase 1 (GPX1), and of the oxidative damage marker 8-hydroxy-2'-deoxyguanosine (8-OHdG) were evaluated. RESULTS: Cats with pyometra showed higher plasma P4 levels and increased uterine messenger RNA (mRNA) and protein expression of ERα and PR, mainly in the glandular epithelium for ERα and in stromal and myometrial cells for PR. In addition, there was an increase in 8-OHdG immunostaining and GPX1 mRNA and protein expression in cats with pyometra compared with those in non-gestational diestrus, while catalase showed a reduction in endometrial immunostaining in cats with pyometra. There were no differences in uterine AR and SOD1 expression between the groups. CONCLUSION AND RELEVANCE: The findings of this study showed that pyometra is associated with oxidative stress in the uterus of domestic cats and alterations of the profile of sex steroid receptors, especially ERα and PR, and of antioxidant enzymes, suggesting that changes in these mediators may play a role with the etiopathogenesis of this disease.

Increased proliferation and neuronal fate in prairie vole brain progenitor cells cultured in vitro: effects by social exposure and sexual dimorphism.

BACKGROUND: The prairie vole (Microtus ochrogaster) is a socially monogamous rodent that establishes an enduring pair bond after cohabitation, with (6 h) or without (24 h) mating. Previously, we reported that social interaction and mating increased cell proliferation and differentiation to neuronal fate in neurogenic niches in male voles. We hypothesized that neurogenesis may be a neural plasticity mechanism involved in mating-induced pair bond formation. Here, we evaluated the differentiation potential of neural progenitor cells (NPCs) isolated from the subventricular zone (SVZ) of both female and male adult voles as a function of sociosexual experience. Animals were assigned to one of the following groups: (1) control (Co), sexually naive female and male voles that had no contact with another vole of the opposite sex; (2) social exposure (SE), males and females exposed to olfactory, auditory, and visual stimuli from a vole of the opposite sex, but without physical contact; and (3) social cohabitation with mating (SCM), male and female voles copulating to induce pair bonding formation. Subsequently, the NPCs were isolated from the SVZ, maintained, and supplemented with growth factors to form neurospheres in vitro. RESULTS: Notably, we detected in SE and SCM voles, a higher proliferation of neurosphere-derived Nestin + cells, as well as an increase in mature neurons (MAP2 +) and a decrease in glial (GFAP +) differentiated cells with some sex differences. These data suggest that when voles are exposed to sociosexual experiences that induce pair bonding, undifferentiated cells of the SVZ acquire a commitment to a neuronal lineage, and the determined potential of the neurosphere is conserved despite adaptations under in vitro conditions. Finally, we repeated the culture to obtain neurospheres under treatments with different hormones and factors (brain-derived neurotrophic factor, estradiol, prolactin, oxytocin, and progesterone); the ability of SVZ-isolated cells to generate neurospheres and differentiate in vitro into neurons or glial lineages in response to hormones or factors is also dependent on sex and sociosexual context. CONCLUSION: Social interactions that promote pair bonding in voles change the properties of cells isolated from the SVZ. Thus, SE or SCM induces a bias in the differentiation potential in both sexes, while SE is sufficient to promote proliferation in SVZ-isolated cells from male brains. In females, proliferation increases when mating is performed. The next question is whether the rise in proliferation and neurogenesis of cells from the SVZ are plastic processes essential for establishing, enhancing, maintaining, or accelerating pair bond formation. Highlights 1. Sociosexual experiences that promote pair bonding (social exposure and social cohabitation with mating) induce changes in the properties of neural stem/progenitor cells isolated from the SVZ in adult prairie voles. 2. Social interactions lead to increased proliferation and induce a bias in the differentiation potential of SVZ-isolated cells in both male and female voles. 3. The differentiation potential of SVZ-isolated cells is conserved under in vitro conditions, suggesting a commitment to a neuronal lineage under a sociosexual context. 4. Hormonal and growth factors treatments (brain-derived neurotrophic factor, estradiol, prolactin, oxytocin, and progesterone) affect the generation and differentiation of neurospheres, with dependencies on sex and sociosexual context. 5. Proliferation and neurogenesis in the SVZ may play a crucial role in establishing, enhancing, maintaining, or accelerating pair bond formation.

In this study, researchers evaluated whether social interactions and copulation induce changes in the proliferation and differentiation of neural progenitor cells in adult male and female voles using an in vitro neurosphere formation assay. The following groups were assigned: control animals without any exposure to another vole outside their litter, another group with social exposure consisting of sensory exposure to a vole of the opposite sex and a third group with social cohabitation and copulation. Forty eight hours after social interactions, cells were isolated from the neurogenic niche subventricular zone (SVZ) and cultured to assess their self-renewal and proliferation abilities to form neurospheres. The results showed in the social interaction groups, a greater number and growth of neurospheres in both males and females. Differentiation capacity was assessed by immunodetection of MAP2 and GFAP to identify neurons or glia, respectively, arise from neurospheres, with an increase in neuronal fate in groups with social interaction. In the second part of the study, the researchers analyzed the effect of different hormone and growth factor treatments and found that the response in both proliferation and differentiation potential may vary depending on the sociosexual context or sex. This study suggests that social interactions leading to pair bond formation alter the properties of SVZ cells, whereby proliferation and neurogenesis may have an impact on the establishment and maintenance of pair bonding.

Quantitative Analysis of Cellular Morphology During In Vitro Decidualization.

Decidualization is a differentiation process involving shape reorganization from a fibroblast to an epithelioid-like appearance characteristic of endometrial stromal cells. For the study of in vitro decidualization, one needs to check that the cells have undergone this process effectively. Verification is usually done by analyzing the expression of decidual markers, but changes in morphology are a more comprehensive feature. However, morphological specificities (i.e., flatness) of endometrial cells prevent the use of existing automated tools. A simple and accurate methodology was developed to quantify the phenotypic changes that occur in an in vitro decidualization system. This approach analyzes cell circularity directly from light microscopy images to follow the effects of progesterone or progestin R5020 in combination with estradiol (E2) and cAMP in inducing the decidualization of human endometrial cells. A statistical model to detect the differences in the kinetics of decidualization of the two hormonal stimuli before all the cell population acquire the decidual phenotype was implemented. It was found that statistical differences in morphology between decidualized and control cells could be detected 2 days after the treatments. Here we detail the model applied, scripts, and input files in order to provide a useful, practical, and low-cost tool to evaluate morphological aspects of endometrial stromal differentiation. This method allows the verification of the effectiveness of the decidualization process of the stromal endometrial cells without having to use cell replicates, as other methods such as immunofluorescence and RT-qPCR assays require. Consequently, this approach can follow the kinetics of a living single replicate throughout the experiment. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Cell circularity quantification of human stromal endometrial cells using ImageJ Basic Protocol 2: Statistical analysis of cell circularity of human stromal endometrial cells.

Evaluation of the therapeutic effects of oestradiol on the systemic inflammatory response and on lung injury caused by the occlusion of the proximal descending aorta in male rats.

OBJECTIVES: Ischaemia and reperfusion-induced microvascular dysfunction is a serious problem encountered during a variety surgical procedures, leading to systemic inflammation and affecting remote organs, specially the lungs. 17ß-Oestradiol reduces pulmonary repercussions from various acute lung injury forms. Here, we focused on the 17ß-oestradiol therapeutic effects after aortic ischaemia and reperfusion (I/R) by evaluating lung inflammation. METHODS: Twenty-four Wistar rats were submitted to I/R by insufflation of a 2-F catheter in thoracic aorta for 20 min. Reperfusion took 4 h and 17ß-oestradiol (280 µg/kg, i.v.) was administered after 1 h of reperfusion. Sham-operated rats were controls. Bronchoalveolar lavage was performed and lung samples were prepared for histopathological analysis and tissue culture (explant). Interleukin (IL)-1ß, IL-10 and tumour necrosis factor-α were quantified. RESULTS: After I/R, higher number of leukocytes in bronchoalveolar lavage were reduced by 17ß-oestradiol. The treatment also decreased leukocytes in lung tissue. I/R increased lung myeloperoxidase expression, with reduction by 17ß-oestradiol. Serum cytokine-induced neutrophil chemoattractant 1 and IL-1ß increased after I/R and 17ß-oestradiol decreased cytokine-induced neutrophil chemoattractant 1. I/R increased IL-1ß and IL-10 in lung explants, reduced by 17ß-oestradiol. CONCLUSIONS: Our results showed that 17ß-oestradiol treatment performed in the period of reperfusion, modulated the systemic response and the lung repercussions of I/R by thoracic aortic occlusion. Thus, we can suggest that 17ß-oestradiol might be a supplementary approach leading the lung deterioration after aortic clamping in surgical procedures.

Distribution of estrogen receptors alpha and beta in the brain of male rats with same-sex preference.

Two distinct estrogen receptors (ERs) exist, ERα and ERß. Both receptors participate in sexual differentiation of the rat brain and likely participate in the regulation of adult sexual orientation (i.e. partner preference). This last idea was investigated herein by examining males treated with the aromatase inhibitor, letrozole, administered prenatally (0.56 µg/kg G10-22). This treatment usually provokes same-sex preference in 1-2 males per litter. Vehicle-treated males (with female preference) and females in spontaneous proestrus (with male preference) were included as controls. ERα and ERß expression was analyzed by immunohistochemistry in brain areas known to control masculine sexual behavior and partner preference, like the medial preoptic area (MPOA), bed nucleus of the stria terminalis (BNST), medial amygdala (MeA) and ventromedial hypothalamic nucleus (VMH), as well as other brain regions suspected to participate in these processes. In addition, serum levels of estradiol were determined in all male groups. Letrozole-treated male rats that preferred sexually experienced males (LPM) showed over-expressed ERα in the hippocampal cornu Ammonis (CA 1, 3, 4) and dentate gyrus. The LPM group showed up-regulated ERß expression in the CA2 and reticular thalamic nucleus. The levels of estradiol did not differ between the groups. Higher expression of ERs in these males was different than their expression in females, with male sex-preference. This suggests that males with same-sex preference showed a unique brain, this sui generis steroid receptor expression probably participates in the biological underpinnings of sexual preference.

Deletion of GABAB receptors from Kiss1 cells affects glucose homeostasis without altering reproduction in male mice.

Kisspeptin and γ-amino butyric acid (GABA), synthesized in the central nervous system, are critical for reproduction. Both are also expressed in peripheral organs/tissues critical to metabolic control (liver/pancreas/adipose). Many kisspeptin neurons coexpress GABAB receptors (GABABR) and GABA controls kisspeptin expression and secretion. We developed a unique mouse lacking GABABR exclusively from kisspeptin cells/neurons (Kiss1-GABAB1KO) to evaluate the impact on metabolism/reproduction. We confirmed selective deletion of GABABR from Kiss1 cells in the anteroventral periventricular nucleus/periventricular nucleus continuum (AVPV/PeN; immunofluorescence and PCR) and arcuate nucleus (ARC), medial amygdala (MeA), pituitary, liver, and testes (PCR). Young Kiss1-GABAB1KO males were fertile, with normal LH and testosterone. Kiss1 expression was similar between genotypes in AVPV/PeN, ARC, MeA, bed nucleus of the stria terminalis (BNST), and peripheral organs (testis, liver, pituitary). Kiss1-GABAB1KO males presented higher fasted glycemia and insulin levels, an impaired response to a glucose overload, reduced insulin sensitivity, and marked insulin resistance. Interestingly, when Kiss1-GABAB1KO males got older (9 mo old) their body weight (BW) increased, in part due to an increase in white adipose tissue (WAT). Old Kiss1-GABAB1KO males showed higher fasted insulin, increased pancreatic insulin content, insulin resistance, and significantly decreased pancreatic kisspeptin levels. In sum, lack of GABABR specifically in Kiss1 cells severely impacts glucose homeostasis in male mice, reinforcing kisspeptin involvement in metabolic regulation. These alterations in glucose homeostasis worsened with aging. We highlight the impact of GABA through GABABR in the regulation of the pancreas kisspeptin system in contrast to liver kisspeptin that was not affected.NEW & NOTEWORTHY We developed a unique mouse lacking GABAB receptors specifically in Kiss1 cells to evaluate the impact on reproduction and metabolism. Knockout males showed a severe impact on glucose homeostasis, which worsened with aging. These results reinforce the proposed kisspeptin involvement in metabolic regulation and highlight the impact of GABA through GABABR in the regulation of the peripheral pancreas kisspeptin system.

Exposure to acetaminophen impairs gametogenesis and fertility in zebrafish (Danio rerio).

Acetaminophen (ACE; paracetamol) is one of the most widely used nonsteroidal anti-inflammatory drugs worldwide and is often found in aquatic systems, where it can act on nontarget species and impair fish reproduction. This study aimed to investigate the effects of chronic exposure to environmentally relevant ACE concentrations (0.5, 5 and 50 µg/L) on multiple reproductive parameters in zebrafish (Danio rerio). Gametogenesis was analyzed using histology, morphometry, cell proliferation, and apoptosis. This study also evaluated sex steroids, and prostaglandin E2 (PGE2) levels, gene expression for sex steroids and PGE2 receptors, fertilization rate, and semen quality. In females, exposure to 5 and 50 µg/L ACE induced larger and more abundant vitellogenic follicles and increased follicular atresia. In these treatments, males showed a lower proportion and proliferation of undifferentiated spermatogonia and a higher proportion of TUNEL-positive differentiated spermatogonia, spermatids, and spermatozoa, resulting in lower sperm production. ACE increased 17ß-estradiol (E2) and reduced 11-ketotestosterone levels in the testis, whereas only E2 increased in the ovaries. In both sexes, gonadal PGE2 levels were reduced. ACE at 50 µg/L induced an increase in the gene expression of androgen, estrogen, and PGE2 receptors in the ovaries, and reduced expression in the testes. Results also showed lower egg production and fertilization rate from 28 days of exposure with reduced sperm quality. These results demonstrated that ACE impairs the reproductive performance of zebrafish, affecting multiple reproductive parameters, which may be caused by the synergistic action of the imbalance of sex steroids, with a reduction of PGE2 and its receptors.

Biotransformation of 17ß-Estradiol through a Denitrifying Sludge.

17ß-estradiol (E2) is the natural estrogen with the most significant potential for endocrine disruption in the biota of aquatic ecosystems at trace concentrations. It is, therefore, essential to study treatments for water polluted with E2 that would guarantee its complete elimination and mineralization. Denitrification is a biological process shown to have the capacity to completely biodegrade drugs, such as ampicillin. This work is aimed to evaluate the biotransformation of 17ß-estradiol by employing a denitrifying sludge. The assays performed were: (I) abiotic with 3.5 mg E2-C L-1 and (II) denitrifying with 10 mg CH3COO--C L-1 as the reference, 10 mg E2-C L-1 as the sole electron donor, and a mixture of (mg L-1) 10 E2-C with 10 CH3COO--C at C N-1 of 1.1. The E2-C and NO3--N consumption efficiencies were greater than 99%, and HCO3--C and N2 production yields were close to 1 in all assays. The denitrifying sludge could biodegrade up to 10 mg E2-C L-1 as the sole electron donor and when mixed with 10 mg CH3COO--C L-1. No intermediate metabolites were generated from the process.

Ageing decreases the healing of wounds in the skin of alcohol-preferring rats.

OBJECTIVE: Alcohol consumption combined with ageing alters the healing process of the skin. We evaluated whether ageing decreases the healing of incisional wounds in the skin of Wistar rats of Universidade de Chile of variety B (UChB). METHOD: A total of 20 adult rats and 20 older UChB rats, divided into two groups which underwent surgical aggression in the anterior region of the abdomen, were used: G1, adult rats (100 days old, control) with water and 10% ethanol; G2, aged rats (540 days old, experimental) with water and 10% ethanol; evaluated at 4, 7, 14 and 21 days after surgery. RESULTS: Ageing did not alter the rupture force and collagen elasticity and resistance. There were increases in telomerase with the implementation of cellular senescence, in interleukin 1-alpha (IL-1α) at 14 days of healing, in epidermal growth factor (EGF) at 14 and 21 days of healing with delayed growth and development of keratinocytes, also an increase of IL-ß at 4 days, and decrease in tumour necrosis factor (TNFα) at 7 days, associated with chronic scarring. There was an increase in vascular endothelial growth factor (VEGF) at 4 and 7 days, responsible for the early vessels re-establishment. There was a decrease in transforming growth factor 2-beta (TGFß2) and ß3 at 4 and 7 days of healing respectively, and estradiol at 4 days. CONCLUSION: Ageing decreases the skin healing in incisional wounds in alcohol-preferring rats.