Is there potential for estradiol receptor signaling in lophotrochozoans?

Estrogen receptors (ERs) are thought to be the ancestor of all steroid receptors and are present in most lophotrochozoans studied to date, including molluscs, annelids, and rotifers. A number of studies have investigated the functional role of estrogen receptors in invertebrate species, although most are in molluscs, where the receptor is constitutively active. In vitro experiments provided evidence for ligand-activated estrogen receptors in annelids, raising important questions about the role of estrogen signalling in lophotrochozoan lineages. Here, we review the concordant and discordant evidence of estradiol receptor signalling in lophotrochozoans, with a focus on annelids and rotifers. We explore the de novo synthesis of estrogens, the evolution and expression of estrogen receptors, and physiological responses to activation of estrogen receptors in the lophotrochozoan phyla Annelida and Rotifera. Key data are missing to determine if de novo biosynthesis of estradiol in non-molluscan lophotrochozoans is likely. For example, an ortholog for the CYP11 gene is present, but confirmation of substrate conversion and measured tissue products is lacking. Orthologs CYP17 and CYP19 are lacking, yet intermediates or products (e.g. estradiol) in tissues have been measured. Estrogen receptors are present in multiple species, and for a limited number, in vitro data show agonist binding of estradiol and/or transcriptional activation. The expression patterns of the lophotrochozoan ERs suggest developmental, reproductive, and digestive roles but are highly species dependent. E2 exposures suggest that lophotrochozoan ERs may play a role in reproduction, but no strong dose-response relationship has been established. Therefore, we expect most lophotrochozoan species, outside of perhaps platyhelminths, to have an ER but their physiological role remains elusive. Mining genomes for orthologs gene families responsible for steroidogenesis, coupled with in vitro and in vivo studies of the steroid pathway are needed to better assess whether lophotrochozoans are capable of estradiol biosynthesis. One major challenge is that much of the data are divided across a diversity of species. We propose that the polychaetes Capitella teleta or Platyneris dumerilii, and rotifer Brachionus manjavacas may be strong species choices for studies of estrogen receptor signalling, because of available genomic data, established laboratory culture techniques, and gene knockout potential.

Effects of insulin-like growth factor-1 on the mRNA expression of estradiol receptors, steroidogenic enzymes, and steroid production in bovine follicles.

Insulin-like growth factor-1 (IGF-1) plays a crucial role in follicular growth and stimulates steroid hormone production in bovine follicles. Steroid hormones are synthesized through the actions of steroidogenic enzymes, specifically STAR, CYP11A1, HSD3B, and CYP19A1 in both theca cells (TCs) and granulosa cells (GCs), under the influence of gonadotropins. Particularly, estradiol 17ß (E2) assumes a central role in follicular development and selection by activating estrogen receptors ß (ESR2) in GCs. We assessed ESR2 mRNA expression in GCs of developing follicles and investigated the impact of IGF-1 on the mRNA expression of ESR2, CYP19A1, FSHR, and LHCGR, STAR, CYP11A1, and HSD17B in cultured GCs and TCs, respectively. Additionally, we assessed the influence of IGF-1 on androstenedione (A4), progesterone (P4), and testosterone (T) production in TCs. Small-sized follicles (< 6 mm) exhibited the highest levels of ESR2 mRNA expression, whereas medium-sized follicles (7-8 mm) displayed higher levels than large-sized follicles (≥ 9 mm) (P < 0.05). IGF-1 increased the mRNA expression of ESR2, CYP19A1, and FSHR in GCs of follicles of both sizes, except for FSHR mRNA in medium-sized follicles (P < 0.05). IGF-1 significantly elevated mRNA expression of LHCGR, STAR, CYP11A1, and CYP17B in TCs of small- and medium-sized follicles (P < 0.05). Moreover, IGF-1 augmented the production of A4 and P4 but had no impact on T production in TCs of small- and medium-sized follicles. Taken together, our findings indicate that IGF-1 upregulates steroidogenic enzymes and steroid hormone production, underscoring the crucial role of IGF-1 in follicle development and selection.

Activation of the G Protein-Coupled Estrogen Receptor (GPER) Increases Neurogenesis and Ameliorates Neuroinflammation in the Hippocampus of Male Spontaneously Hypertensive Rats.

It is known that spontaneously hypertensive rats (SHR) present a marked encephalopathy, targeting vulnerable regions such as the hippocampus. Abnormalities of the hippocampus of SHR include decreased neurogenesis in the dentate gyrus (DG), partial loss of neurons in the hilus of the DG, micro and astrogliosis and inflammation. It is also known that 17ß-estradiol (E2) exert neuroprotective effects and prevent hippocampal abnormalities of SHR. The effects of E2 may involve a variety of mechanisms, including intracellular receptors of the ERα and ERß subtypes or membrane-located receptors, such as the G protein-coupled estradiol receptor (GPER). We have now investigated the protective role of GPER in SHR employing its synthetic agonist G1. To accomplish this objective, 5 month-old male SHR received 150 µg/day of G1 during 2 weeks. At the end of this period, we analyzed neuronal progenitors by staining for doublecortin (DCX), and counted the number of glial fibrillary acidic protein (GFAP)-labeled astrocytes and Iba1-stained microglial cells by computerized image analysis. We found that G1 activation of GPER increased DCX+ cells in the DG and reduced GFAP+ astrogliosis and Iba1+ microgliosis in the CA1 region of hippocampus. We also found that the high expression of proinflammatory makers IL1ß and cyclooxygenase 2 (COX2) of SHR was decreased after G1 treatment, which correlated with a change of microglia phenotype from the activated to a resting morphology. Additionally, G1 treatment increased the anti-inflammatory factor TGFß in SHR hippocampus. Altogether, our results suggest that activation of GPER plays a neuroprotective role on the encephalopathy of SHR, an outcome resembling E2 effects but avoiding secondary effects of the natural hormone.

Anorexigenic effects of estradiol in the medial preoptic area occur through membrane-associated estrogen receptors and metabotropic glutamate receptors.

Activation of membrane-associated estrogen receptors (mER) decreases food and water intake in female rats. Additional studies suggest these effects are mediated, at least in part, by membrane-associated estrogen receptor alpha (ERα). Nevertheless, the critical site of action and the intracellular signaling required for the ingestive effects of ERα remain unclear. Estradiol given to the medial preoptic area (mPOA) decreases ingestive behaviors, and membrane-associated ERα has been shown to affect intracellular signaling through interactions with metabotropic glutamate receptor (mGluR) subtypes, but an involvement of this signaling pathway, in the mPOA, in ingestive behavior remains untested. To address these open questions, we first showed that activation of mER in the mPOA decreased both overnight food and water intake, and did so in a time course consistent with a genomic mechanism of action. Next, we tested the requirement of mGluR1a signaling in the mPOA for the anorexigenic and anti-dipsogenic effects of estradiol. As expected, estradiol in the mPOA decreased food intake, but only in the absence of an mGluR1a antagonist. The same was not true for estradiol effects on water intake, which were unaffected by an mGluR1a antagonist. These results suggest that estrogens require mGluR activation for at least some of their effects on ingestive behaviors, and indicate that the mPOA is a critical site of action. The results also reveal an interesting divergence in the estrogenic control of ingestive behavior by which mGluR signaling in the mPOA plays a role in the control of food intake, but not water intake.

Prostanoid receptors EP2, EP4, and FP are regulated by estradiol in bovine oviductal smooth muscle.

Gamete and embryo transport is an important function of the oviduct. This transport involves both smooth muscle contraction and epithelial cell secretions, the former of which is mediated by prostaglandins (PGs) and their receptors. Our aim was to study the regulation of prostaglandin E2 and prostaglandin F2α receptors (EP2, EP4, and FP receptor) by estradiol in bovine oviduct smooth muscle. EP2, EP4, and FP receptor mRNA and protein expression was investigated using real-time RT-PCR and Western blot analyses, respectively. To evaluate the contraction or relaxation of cultured bovine oviductal smooth muscle tissue, peristalsis was used to assess contractile activity. EP2, EP4, and FP receptor mRNA and protein expression was increased in oviductal smooth muscle tissue after treatment with different concentrations of estradiol for various durations. The expression of all receptors peaked at an estradiol concentration of 10(-11)mol/L after 8h of treatment, whereas no increase in expression was observed after fulvestrant (a selective antagonist of E2 receptor) treatment, indicating that E2 interacts with specific E2 nuclear receptors to regulate EP2, EP4, and FP receptor expression. Although PGF2α and PGE2 induced both contraction and relaxation, no significant differences were found in contractility between the estradiol-treated and control groups, with both groups of cultured smooth muscle strips showing similar vitality. In conclusion, estradiol increases EP2, EP4, and FP receptor mRNA and protein expression in bovine oviductal smooth muscle when added for different periods of time and at different concentrations. Additionally, E2 is transported intracellularly and interacts with specific E2 nuclear receptors to regulate their expression.

1,25-Dihydroxyvitamin D3 increases testosterone-induced 17beta-estradiol secretion and reverses testosterone-reduced connexin 43 in rat granulosa cells.

BACKGROUND: Aromatase converts testosterone into 17beta-estradiol in granulosa cells, and the converted 17beta-estradiol contributes to follicular maturation. Additionally, excessive testosterone inhibits aromatase activity, which can lead to concerns regarding polycystic ovary syndrome (PCOS). Generally, 1,25-dihydroxyvitamin D3 (1,25D3) supplements help to improve the symptoms of PCOS patients who exhibit low blood levels of 1,25D3. Therefore, this study investigated the interaction effects of 1,25D3 and testosterone on estrogenesis and intercellular connections in rat granulosa cells. METHODS: Primary cultures of granulosa cells were treated with testosterone or testosterone plus 1,25D3, or pre-treated with a calcium channel blocker or calcium chelator. Cell lysates were subjected to western blot analysis to determine protein and phosphorylation levels, and 17beta-estradiol secretion was examined using a radioimmunoassay technique. Cell viability was evaluated by MTT reduction assay. Connexin 43 (Cx43) mRNA and protein expression levels were assessed by qRT-PCR, western blot, and immunocytochemistry. RESULTS: Testosterone treatment (0.1 and 1 microg/mL) increased aromatase expression and 17beta-estradiol secretion, and the addition of 1,25D3 attenuated testosterone (1 microg/mL)-induced aromatase expression but improved testosterone-induced 17beta-estradiol secretion. Furthermore, testosterone-induced aromatase phosphotyrosine levels increased at 10 min, 30 min and 1 h, whereas 1,25D3 increased the longevity of the testosterone effect to 6 h and 24 h. Within 18-24 h of treatment, 1,25D3 markedly enhanced testosterone-induced 17beta-estradiol secretion. Additionally, pre-treatment with a calcium channel blocker nifedipine or an intracellular calcium chelator BAPTA-AM reduced 1,25D3 and testosterone-induced 17beta-estradiol secretion. Groups that underwent testosterone treatment exhibited significantly increased estradiol receptor beta expression levels, which were not affected by 1,25D3. Neither testosterone nor 1,25D3 altered 1,25D3 receptor expression. Finally, at high doses of testosterone, Cx43 protein expression was decreased in granulosa cells, and this effect was reversed by co-treatment with 1,25D3. CONCLUSIONS: These data suggest that 1,25D3 potentially increases testosterone-induced 17beta-estradiol secretion by regulating aromatase phosphotyrosine levels, and calcium increase is involved in both 1,25D3 and testosterone-induced 17beta-estradiol secretion. 1,25D3 reverses the inhibitory effect of testosterone on Cx43 expression in granulosa cells.

17ß-Estradiol induces supernumerary primordial germ cells in embryos of the polychaete Platynereis dumerilii.

In the polychaete Platynereis dumerilii exactly four primordial germ cells (PGCs) arise in early development and are subject to a transient mitotic arrest until the animals enter gametogenesis. In order to unravel the mechanisms controlling the number of PGCs in Platynereis, we tested whether the steroid 17ß-estradiol (E2) is able to induce PGC proliferation, as it had been described in other species. Our data provide strong support for such a mechanism, showing that E2 significantly increases the occurrence of larvae with supernumerary PGCs in Platynereis in a dose dependent manner. E2 responsiveness is restricted to early developmental stages, when the PGCs are specified. During these stages, embryos exhibit high expression levels of the estradiol receptor (ER). The ER transcript localizes to the yolk-free cytoplasm of unfertilized eggs and segregates into the micromeres during cleavage stages. Nuclear ER protein is found asymmetrically distributed between daughter cells. Neither transcript nor protein is detectable in PGCs at larval stages. Addition of the specific estradiol receptor inhibitor ICI-182,780 (ICI) abolishes the proliferative effect of E2, suggesting that it is mediated by ER signaling. Our study reports for the first time an ER mediated proliferative effect of E2 on PGCs in an invertebrate organism.

Effects of bisphenol A and triclocarban on brain-specific expression of aromatase in early zebrafish embryos.

Estrogen regulates numerous developmental and physiological processes. Most effects are mediated by estrogen receptors (ERs), which function as ligand-regulated transcription factors. Estrogen also regulates the activity of GPR30, a membrane-associated G protein-coupled receptor. Many different types of environmental contaminants can activate ERs; some can bind GPR30 as well. There is growing concern that exposure to some of these compounds, termed xenoestrogens, is interfering with the behavior and reproductive potential of numerous wildlife species, as well as affecting human health. Here, we investigated how two common, environmentally pervasive chemicals affect the in vivo expression of a known estrogen target gene in the brain of developing zebrafish embryos, aromatase AroB, which converts androgens to estrogens. We confirm that, like estrogen, the well-studied xenoestrogen bisphenol A (BPA, a plastics monomer), induces strong brain-specific overexpression of aromatase. Experiments using ER- and GPR30-selective modulators argue that this induction is largely through nuclear ERs. BPA induces dramatic overexpression of AroB RNA in the same subregions of the developing brain as estrogen. The antibacterial triclocarban (TCC) by itself stimulates AroB expression only slightly, but TCC strongly enhances the overexpression of AroB that is induced by exogenous estrogen. Thus, both BPA and TCC have the potential to elevate levels of aromatase and, thereby, levels of endogenous estrogens in the developing brain. In contrast to estrogen, BPA-induced AroB overexpression was suppressed by TCC. These results indicate that exposures to combinations of certain hormonally active pollutants can have outcomes that are not easily predicted from their individual effects.

Estradiol receptors agonists induced effects in rat intestinal microcirculation during sepsis.

The steroid hormone estradiol is suggested to play a protective role in intestinal injury during systemic inflammation (sepsis). Our aim was to determine the effects of specific estradiol receptor (ER-α and ER-ß) agonists on the intestinal microcirculation during experimental sepsis. Male and sham ovariectomized female rats were subjected to sham colon ascendens stent peritonitis (CASP), and they were compared to male and ovariectomized female rats underwent CASP and either estradiol receptor α (ER-α) agonist propyl pyrazole triol (PPT), estradiol receptor ß (ER-ß) agonist diarylpropiolnitrile (DPN), or vehicle treatment. Intravital microscopy was performed, which is sufficiently sensitive to measure changes in the functional capillary density (FCD) as well as the major steps in leukocyte recruitment (rolling and adhesion). The leukocyte extravasations were also quantified by using histological paraffin sections of formalin fixed intestine. We found that either DPN (ER-ß) or PPT (ER-α) significantly reduced (P<0.05) sepsis-induced leukocyte-endothelial interaction (rolling, adherent leukocytes and neutrophil extravasations) and improved the intestinal muscular FCD. [PPT: Female; Leukocyte rolling (n/min): V(3) 3.7±0.7 vs 0.8±0.2, Leukocyte adhesion(n/mm(2)): V(3) 131.3±22.6 vs 57.2±13.5, Neutrophil extravasations (n/10000 µm(2)): 3.1±0.7 vs 6 ±1. Male; Leukocyte adhesion (n/mm(2)): V(1) 154.8±19.2 vs 81.3±11.2, V(3) 115.5±23.1 vs 37.8±12]. [DPN: Female; neutrophil extravasations (n/10000 µm(2)) 3.8±0.6 vs 6 ±1. Male; Leukocyte adhesion (n/mm(2)) V(1) 154.8±19.2 vs 70±10.5, V(3) 115.5±23.1 vs 52.8±9.6].Those results suggest that the observed effects of estradiol receptors on different phases of leukocytes recruitment with the improvement of the functional capillary density could partially explain the previous demonstrated salutary effects of estradiol on the intestinal microcirculation during sepsis. The observed activity of this class of compounds could open up a new avenue of research into the potential treatment of sepsis.

Differences in the receptor status between primary and recurrent breast cancer--the frequency of and the reasons for discordance.

OBJECTIVE: Receptor discordances between primary and recurrent breast cancer have been described for years, but only a few analyses have elucidated the factors that influence receptor changes. METHODS: Explorative analyses of prospective data from a breast cancer database of a tertiary breast cancer unit. RESULTS: Recurrent tumours that had expressed oestrogen (ER) and progesterone receptors (PR) and human epidermal growth factor receptor 2 (HER2) as primary tumours were negative for the respective receptor in 22.8, 41.4 and 40.8% of cases. ER, PR and HER2 expression was found in 19.8, 16.7 and 11.5% of recurrent tumours, although no expression had been observed in primary tumours. Receptor discordances in recurrent disease leading to different therapeutic approaches were noted in 126 of 411 patients (30.7%). In patients with tumours expressing primary ER and HER2, independent factors associated with discordance were endocrine therapy and treatment with trastuzumab. CONCLUSION: High rates of receptor discordance were found. The impact of factors that influence receptor changes is small so that no subgroup of patients with recurrent breast cancer should be excluded from biopsy. Whenever possible, a biopsy should be taken to confirm the diagnosis of a possible relapse as well as the receptor status of patients with breast cancer.

Estradiol rapidly rescues synaptic transmission from corticosterone-induced suppression via synaptic/extranuclear steroid receptors in the hippocampus.

We investigated rapid protection effect by estradiol on corticosterone (CORT)-induced suppression of synaptic transmission. Rapid suppression by 1 µM CORT of long-term potentiation (LTP) at CA3-CA1 synapses was abolished via coperfusion of 1 nM estradiol. N-methyl-D-aspartate (NMDA) receptor-derived field excitatory postsynaptic potential (NMDA-R-fEPSP) was used to analyze the mechanisms of these events. Estradiol abolished CORT-induced suppression of NMDA-R-fEPSP slope. This CORT-induced suppression was abolished by calcineurin inhibitor, and the rescue effect by estradiol on the CORT-induced suppression was inhibited by mitogen-activated protein (MAP) kinase inhibitor. The CORT-induced suppressions of LTP and NMDA-R-fEPSP slope were abolished by glucocorticoid receptor (GR) antagonist, and the restorative effects by estradiol on these processes were mimicked by estrogen receptor α (ERα) and ERß agonists. Taken together, estradiol rapidly rescued LTP and NMDA-R-fEPSP slope from CORT-induced suppressions. A GR→calcineurin pathway is involved in these suppressive effects. The rescue effects by estradiol are driven via ERα or ERß→MAP kinase pathway. Synaptic/extranuclear GR, ERα, and ERß probably participate in these rapid events. Mass-spectrometric analysis determined that acute hippocampal slices used for electrophysiological measurements contained 0.48 nM estradiol less than exogenously applied 1 nM. In vivo physiological level of 8 nM estradiol could protect the intact hippocampus against acute stress-induced neural suppression.

Lipopolysaccharide-induced modulation in the expression of progesterone receptor and estradiol receptor leads to early pregnancy loss in mouse.

The objective of the present study was to investigate the effect of Gram-negative bacteria infection on ovarian steroid receptors, i.e. progesterone receptor (PR) and estradiol receptor (ER) during preimplantation days of pregnancy. A well established mouse model of Gram-negative bacteria infection was used to test this objective. Mice were treated with normal saline or lipopolysaccharide (LPS) on day 0.5 of pregnancy and used to collect embryos and uterine horns on day 1.5 to day 4.42 preimplantation day of pregnancy. Total RNA was extracted and reverse-transcription polymerase chain reaction (PCR) was performed to check the expression of PR and ER genes. The mRNA expression of PR and ER was altered in embryos and uterus of LPS-treated animals during preimplantation days of pregnancy studied. These results suggest that PR and ER play an important role in Gram-negative bacteria infection and induced implantation failure in mouse.

Beneficial effects of the phytoestrogen genistein on hippocampal impairments of spontaneously hypertensive rats (SHR).

Hippocampal neuropathology is a recognized feature of the spontaneously hypertensive rat (SHR). The hippocampal alterations associate with cognitive impairment. We have shown that hippocampal abnormalities are reversed by 17ß-estradiol, a steroid binding to intracellular receptors (estrogen receptor α and ß subtypes) or the membrane-located G-protein coupled estradiol receptor. Genistein (GEN) is a neuroprotective phytoestrogen which binds to estrogen receptor ß and G-protein coupled estradiol receptor. Here, we investigated whether GEN neuroprotection extends to SHR. For this purpose, we treated 5-month-old SHR for 2 weeks with 10 mg kg-1 daily s.c injections of GEN. We analyzed the expression of doublecortin+ neuronal progenitors, glial fibrillary acidic protein+ astrocytes and ionized calcium-binding adapter molecule 1+ microglia in the CA1 region and dentate gyrus of the hippocampus using immunocytochemistry, whereas a quantitative real-time polymerase chain reaction was used to measure the expression of pro- and anti-inflammatory factors tumor necrosis factor α, cyclooxygenase-2 and transforming growth factor ß. We also evaluated hippocampal dependent memory using the novel object recognition test. The results showed a decreased number of doublecortin+ neural progenitors in the dentate gyrus of SHR that was reversed with GEN. The number of glial fibrillary acidic protein+ astrocytes in the dentate gyrus and CA1 was increased in SHR but significantly decreased by GEN treatment. Additionally, GEN shifted microglial morphology from the predominantly activated phenotype present in SHR, to the more surveillance phenotype found in normotensive rats. Furthermore, treatment with GEN decreased the mRNA of the pro-inflammatory factors tumor necrosis factor α and cyclooxygenase-2 and increased the mRNA of the anti-inflammatory factor transforming growth factor ß. Discrimination index in the novel object recognition test was decreased in SHR and treatment with GEN increased this parameter. Our results indicate important neuroprotective effects of GEN at the neurochemical and behavioral level in SHR. Our data open an interesting possibility for proposing this phytoestrogen as an alternative therapy in hypertensive encephalopathy.

Stromal Interaction Molecule 1 Maintains ß-Cell Identity and Function in Female Mice Through Preservation of G-Protein-Coupled Estrogen Receptor 1 Signaling.

Altered endoplasmic reticulum (ER) Ca2+ signaling has been linked with ß-cell dysfunction and diabetes development. Store-operated Ca2+ entry replenishes ER Ca2+ through reversible gating of plasma membrane Ca2+ channels by the ER Ca2+ sensor, stromal interaction molecule 1 (STIM1). For characterization of the in vivo impact of STIM1 loss, mice with ß-cell-specific STIM1 deletion (STIM1Δß mice) were generated and challenged with high-fat diet. Interestingly, ß-cell dysfunction was observed in female, but not male, mice. Female STIM1Δß mice displayed reductions in ß-cell mass, a concomitant increase in α-cell mass, and reduced expression of markers of ß-cell maturity, including MafA and UCN3. Consistent with these findings, STIM1 expression was inversely correlated with HbA1c levels in islets from female, but not male, human organ donors. Mechanistic assays demonstrated that the sexually dimorphic phenotype observed in STIM1Δß mice was due, in part, to loss of signaling through the noncanonical 17-ß estradiol receptor (GPER1), as GPER1 knockdown and inhibition led to a similar loss of expression of ß-cell maturity genes in INS-1 cells. Together, these data suggest that STIM1 orchestrates pancreatic ß-cell function and identity through GPER1-mediated estradiol signaling. ARTICLE HIGHLIGHTS: Store-operated Ca2+ entry replenishes endoplasmic reticulum (ER) Ca2+ through reversible gating of plasma membrane Ca2+ channels by the ER Ca2+ sensor, stromal interaction molecule 1 (STIM1). ß-Cell-specific deletion of STIM1 results in a sexually dimorphic phenotype, with ß-cell dysfunction and loss of identity in female but not male mice. Expression of the noncanonical 17-ß estradiol receptor (GPER1) is decreased in islets of female STIM1Δß mice, and modulation of GPER1 levels leads to alterations in expression of ß-cell maturity genes in INS-1 cells.

Oestrogen treatment of experimental autoimmune encephalomyelitis requires 17ß-oestradiol-receptor-positive B cells that up-regulate PD-1 on CD4+ Foxp3+ regulatory T cells.

It is now well accepted that sex hormones have immunoregulatory activity and may prevent exacerbations in multiple sclerosis during pregnancy. Our previous studies demonstrated that oestrogen (17ß-oestradiol; E(2) ) protection against experimental autoimmune encephalomyelitis (EAE) is mediated mainly through oestrogen receptor-α (ERα) and the membrane receptor G-protein-coupled receptor 30 (GPR30) and is abrogated in the absence of B cells and the co-inhibitory receptor, Programmed Death-1 (PD-1). To critically evaluate the cell source of the E2 and PD-1 co-inhibitory pathways in EAE regulation, we assessed the requirement for ERs on transferred B cells and downstream effects on expression of PD-1/PD-ligand on CD4+ Foxp3+ regulatory T (Treg) cells in B-cell-replenished, E2-treated B-cell-deficient (µMT-/-) mice with EAE. The results clearly demonstrated involvement of ERα and GPR30 on transferred B cells that mediated the protective E2 treatment effect on EAE and further showed an E2-mediated B-cell-dependent up-regulation of PD-1 on CD4+ Foxp3+ Treg cells. These findings identify regulatory B-cell populations as key players in potentiating Treg-cell activity during E2-mediated protection against EAE.

Clustering with exclusion zones: genomic applications.

Methods for formally evaluating the clustering of events in space or time, notably the scan statistic, have been richly developed and widely applied. In order to utilize the scan statistic and related approaches, it is necessary to know the extent of the spatial or temporal domains wherein the events arise. Implicit in their usage is that these domains have no "holes"-hereafter "exclusion zones"-regions in which events a priori cannot occur. However, in many contexts, this requirement is not met. When the exclusion zones are known, it is straightforward to correct the scan statistic for their occurrence by simply adjusting the extent of the domain. Here, we tackle the more ambitious objective of formally evaluating clustering in the presence of "unknown" exclusion zones. We develop an algorithm for estimating total exclusion zone extent, the quantity needed to correct scan statistic-based inference, using distributional properties of "spacings," and show how bias correction for this estimator can be effected. Performance of the algorithm is assessed via simulation study. We showcase applications to genomic settings for differing marker (event) types-binding sites, housekeeping genes, and microRNAs-wherein exclusion zones can arise through a variety of mechanisms. In several instances, dramatic changes to unadjusted inference that does not accommodate exclusions are evidenced.

Sex differences in creatine kinase after acute heavy resistance exercise on circulating granulocyte estradiol receptors.

Previous research has shown reduced tissue disruption and inflammatory responses in women as compared to men following acute strenuous exercise. While the mechanism of this action is not known, estrogen may reduce the inflammatory response through its interaction with granulocytes. The purpose of this study was to determine if estrogen receptor ß expression on granulocytes is related to sex differences in tissue disruption in response to an acute heavy resistance exercise protocol. Seven healthy, resistance-trained, eumenorrheic women (23 ± 3 years, 169 ± 9.1 cm, 66.4 ± 10.5 kg) and 8 healthy, resistance-trained men (25 ± 5 years, 178 ± 6.7 cm, 82.3 ± 9.33 kg) volunteered to participate in the study. Subjects performed an acute resistance exercise test consisting of six sets of five squats at 90% of the subject's one repetition maximum. Blood samples were obtained pre-, mid-, post-, and 1-, 6-, and 24-h postexercise. Blood samples were analyzed for 17-ß-estradiol by ELISA, creatine kinase by colorimetric enzyme immunoassay, and estradiol receptors on circulating granulocytes through flow cytometry. Men had higher CK concentrations than women at baseline/control. Men had significantly higher CK concentrations at 24-h postexercise than women. No significant changes in estradiol ß receptors were expressed on granulocytes after exercise or between sexes. While sex differences occur in CK activity in response to strenuous eccentric exercise, they may not be related to estradiol receptor ß expression on granulocytes. Thus, although there are sex differences in CK expression following acute resistance exercise, the differences may not be attributable to estrogen receptor ß expression on granulocytes.

Study of proliferative processes and nuclear estradiol and progesterone receptors in myocytes in pregnant and postpartum mouse uterus.

Numerical densities of the nuclei were morhometrically evaluated in all myocytes and myocytes expressing nuclear estrogen- and progesterone-receptor complexes, which were revealed immunohistochemically with monoclonal antibodies in C57Bl/6 mice. It was shown that the above quantitative parameters of myometrial cells after the first pregnancy were similar to those in nonpregnant mice by day 10 after delivery. In the third pregnancy, especially developed after the second interrupted pregnancy, proliferation processes in the myometrium were not completed by postpartum day 10, but dramatically progressed. It was associated with a significant decrease in the fraction of myocytes carrying nuclear hormone-receptor complexes with estradiol and progesterone and their disturbed physiological relations in the myometrium during and after pregnancy probably due to dedifferentiation of a considerable part of myocytes.

[Estrogens and brain].

Recent data upon molecular mechanisms of pleiotropic action of estrogens in human brain is presented in the article. Given detailed descriptions of properties of classical and membrane bound estradiol receptors, that maintain gene expression regulation, modulation of neurotransmittent systems and signal cascade activation in neuronal cells. Data upon regional distribution of estradiol receptor subtypes in the brain, their participation in main cell population function control (including progenitor cells) is given. Special attention is paid to estrogen participation in neurogenesis, inflammation and apoptosis regulation in central nervous system; in the control of formation and functioning of cerebral vessels.

Genetic variation of putative myokine signaling is dominated by biological sex and sex hormones.

Skeletal muscle plays an integral role in coordinating physiological homeostasis, where signaling to other tissues via myokines allows for coordination of complex processes. Here, we aimed to leverage natural genetic correlation structure of gene expression both within and across tissues to understand how muscle interacts with metabolic tissues. Specifically, we performed a survey of genetic correlations focused on myokine gene regulation, muscle cell composition, cross-tissue signaling, and interactions with genetic sex in humans. While expression levels of a majority of myokines and cell proportions within skeletal muscle showed little relative differences between males and females, nearly all significant cross-tissue enrichments operated in a sex-specific or hormone-dependent fashion; in particular, with estradiol. These sex- and hormone-specific effects were consistent across key metabolic tissues: liver, pancreas, hypothalamus, intestine, heart, visceral, and subcutaneous adipose tissue. To characterize the role of estradiol receptor signaling on myokine expression, we generated male and female mice which lack estrogen receptor α specifically in skeletal muscle (MERKO) and integrated with human data. These analyses highlighted potential mechanisms of sex-dependent myokine signaling conserved between species, such as myostatin enriched for divergent substrate utilization pathways between sexes. Several other putative sex-dependent mechanisms of myokine signaling were uncovered, such as muscle-derived tumor necrosis factor alpha (TNFA) enriched for stronger inflammatory signaling in females compared to males and GPX3 as a male-specific link between glycolytic fiber abundance and hepatic inflammation. Collectively, we provide a population genetics framework for inferring muscle signaling to metabolic tissues in humans. We further highlight sex and estradiol receptor signaling as critical variables when assaying myokine functions and how changes in cell composition are predicted to impact other metabolic organs.

The muscles that are responsible for voluntary movements such as exercise are called skeletal muscles. These muscles secrete proteins called myokines, which play roles in a variety of processes by interacting with other tissues. Essentially, myokines allow skeletal muscles to communicate with organs such as the kidneys, the liver or the brain, which is essential for the body to keep its metabolic balance. Some of the process myokines are involved include inflammation, cancer, the changes brought about by exercise, and even cognition. Despite the clear relevance of myokines to so many physiological outcomes, the way these proteins are regulated and their effects are not well understood. Genetic sex ­ specified by sex chromosomes in mammals ­ contributes to critical aspects of physiology. Specifically, many of the metabolic traits impacted by myokines show striking differences arising from hormonal or genetic interactions depending on the genetic sex of the subject being studied. It is therefore important to consider genetic sex when studying the effects of myokines on the body. Velez, Van et al. wanted to gain a better understanding of how skeletal muscles interact with metabolic tissues such as pancreas, liver and brain, taking genetic sex into consideration. To do this they surveyed human datasets for the correlations between the activity of genes that code for myokines, the composition of muscle cells, the signaling between muscles and metabolic tissues and genetic sex. Their results showed that, genetic sex and sex hormones predicted most of the effects of skeletal muscle on other tissues. For example, myokines from muscle were predicted to be more impactful on liver or pancreas, depending on whether individuals were male or female, respectively. The results of Velez, Van et al. illustrate the importance of considering the effects of genetic sex and sexual hormones when studying metabolism. In the future, these results will allow other researchers to design sex-specific experiments to be able to gather more accurate information about the mechanisms of myokine signaling.