Identification of female-enriched and disease-associated microglia (FDAMic) contributes to sexual dimorphism in late-onset Alzheimer's disease.

BACKGROUND: Late-onset Alzheimer's disease (LOAD) is the most common form of dementia; it disproportionally affects women in terms of both incidence rates and severity of progression. The cellular and molecular mechanisms underlying this clinical phenomenon remain elusive and ill-defined. METHODS: In-depth analyses were performed with multiple human LOAD single-nucleus transcriptome datasets to thoroughly characterize cell populations in the cerebral cortex. ROSMAP bulk human brain tissue transcriptome and DNA methylome datasets were also included for validation. Detailed assessments of microglial cell subpopulations and their relevance to sex-biased changes at the tissue level were performed. Clinical trait associations, cell evolutionary trajectories, and transcription regulon analyses were conducted. RESULTS: The relative numbers of functionally defective microglia were aberrantly increased uniquely among affected females. Substratification of the microglia into different subtypes according to their transcriptomic signatures identified a group of female-enriched and disease-associated microglia (FDAMic), the numbers of which were positively associated with disease severity. Phenotypically, these cells exhibit transcriptomic signatures that support active proliferation, MHC class II autoantigen presentation and amyloid-ß binding, but they are also likely defective in phagocytosis. FDAMic are likely evolved from female activated response microglia (ARMic) with an APOE4 background and compromised estrogen receptor (ER) signaling that is deemed to be active among most subtypes of microglia. CONCLUSION: This study offered important insights at both the cellular and molecular levels into how ER signaling affects microglial heterogeneity and function. FDAMic are associated with more advanced pathologies and severe trends of cognitive decline. Their emergence could, at least in part, explain the phenomenon of greater penetrance of the APOE4 genotype found in females. The biases of FDAMic emergence toward female sex and APOE4 status may also explain why hormone replacement therapy is more effective in APOE4 carriers. The pathologic nature of FDAMic suggests that selective modulations of these cells may help to regain brain neuroimmune homeostasis, serving as a new target for future drug development.

High expression of the RET receptor tyrosine kinase and its ligand GDNF identifies a high-risk subset of estrogen receptor positive breast cancer.

PURPOSE: Resistance to endocrine therapy is the primary cause of treatment failure and death in patients with ER-positive (ER +)/luminal breast cancer. Expression and activation of the RET receptor tyrosine kinase may be driving poor outcomes. We aim to identify high-risk patients and druggable pathways for biomarker-based clinical trials. METHODS: We obtained batch-normalized mRNA expression data from Breast Invasive Carcinoma-The Cancer Genome Atlas, PanCancer Atlas (BRCA-TCGA). To determine clinically significant cutoffs for RET expression, patients were grouped at different thresholds for Kaplan-Meier plotting. Differential gene expression (DGE) analysis and enrichment for gene sets was performed. transcriptomic dataset of antiestrogen-treated ER + tumors stratified by clinical response was then analyzed. RESULTS: High RET expression was associated with worse outcomes in patients with ER + tumors, and stratification was enhanced by incorporating GDNF expression. High RET/GDNF patients had significantly lower overall survival (HR = 2.04, p = 0.012), progression-free survival (HR = 2.87, p < 0.001), disease-free survival (HR = 2.67, p < 0.001), and disease-specific survival (HR = 3.53, p < 0.001) than all other ER + patients. High RET/GDNF tumors were enriched for estrogen-independent signaling and targetable pathways including NTRK, PI3K, and KRAS. Tumors with adaptive resistance to endocrine therapy were enriched for gene expression signatures of high RET/GDNF primary tumors. CONCLUSION: Expression and activation of the RET receptor tyrosine kinase may be driving poor outcomes in some patients with ER + breast cancer. ER + patients above the 75th percentile may benefit from clinical trials with tyrosine kinase inhibitors.

2,4-Dichlorophenol Increases Primordial Germ Cell Numbers via ESR2a-Dependent Pathway in Zebrafish Larvae.

Previous studies have reported the feminizing effects of 2,4-dichlorophenol (2,4-DCP) on zebrafish (Danio rerio). However, the effect of 2,4-DCP on the number of primordial germ cells (PGCs), an indicator for early sex differentiation, remains elusive. In the present study, Tg (piwil1:egfp-UTR nanos3) zebrafish (GFP-labeled PGCs) were treated with 2,4-DCP (10, 20, and 40 µg/L) from 5 to 15 days postfertilization to explore the effect on PGC numbers and to elucidate associated molecular mechanisms. The results showed that 2,4-DCP exposure increased PGC numbers, as evidenced by larger GFP fluorescent areas, upregulated expressions of PGC marker genes (vasa and dnd), and raised the female ratio. Notably, the mRNA level of estrogen receptor 2a (esr2a) was also increased subsequently. Moreover, docking studies revealed stable 2,4-DCP interactions with ESR2a, speculating a role of ESR2a signaling pathway in 2,4-DCP toxicity. Furthermore, in esr2a knockout (esr2a-/-) zebrafish, the effects of 2,4-DCP were considerably minimized, proving the involvement of the ESR2a signaling pathway in the 2,4-DCP-mediated increase in PGC numbers. Dual-luciferase reporter gene assay and point mutation studies demonstrated that 2,4-DCP-stimulated promoter activity was mediated by estrogen response element (ERE) located in -686/-674 of the vasa promoter and -731/-719 of the dnd promoter. Overall, 2,4-DCP can potentially enhance the expression of vasa and dnd by binding to zebrafish ESR2a, thus leading to increased PGC numbers and subsequent female-biased sex differentiation.

Transcriptome and Proteome Analysis Revealed Key Pathways Regulating Final Stage of Oocyte Maturation of the Turkey (Meleagris gallopavo).

In birds, the zona pellucida (ZP) matrix that surrounds the ovulated oocyte-called the inner perivitelline layer-is involved in sperm-zona interaction and successful fertilization. To identify the important genes and proteins connected with the final step of egg development, next-generation sequencing and two-dimensional electrophoresis, combined with mass spectrometry, were used for the analysis of mature oocytes at the F1 developmental stage. A total of 8161 genes and 228 proteins were annotated. Six subfamilies of genes, with codes ZP, ZP1-4, ZPD, and ZPAX, were identified, with the dominant expression of ZPD. The main expression site for ZP1 was the liver; however, granulosa cells may also participate in local ZP1 secretion. A ubiquitination system was identified in mature oocytes, where ZP1 was found to be the main ubiquitinated protein. Analysis of transcripts classified in estrogen receptor (ESR) signaling indicated the presence of ESR1 and ESR2, as well as a set of estrogen-dependent genes involved in both genomic and nongenomic mechanisms for the regulation of gene expression by estrogen. Oxidative phosphorylation was found to be a possible source of adenosine triphosphate, and the nuclear factor erythroid 2-related factor 2 signaling pathway could be involved in the response against oxidative stress. Oocyte-granulosa cell communication by tight, adherens, and gap junctions seems to be essential for the final step of oocyte maturation.

Low dose carbendazim disrupts mouse spermatogenesis might Be through estrogen receptor related histone and DNA methylation.

Pesticides, fungicides are reportedly involved in a decline in spermatozoa quality, especially motility, and a consequent increase in the rate of infertility. Fungicide carbendazim (CBZ) is widely used in agriculture and other aspects. Although CBZ is known to disrupt spermatogenesis, causing a decrease in spermatozoa concentration and motility, the mechanisms are not fully understood. We aimed to further explore the underlying mechanisms of CBZ disruption of spermatogenesis. Pubertal mice were exposed to low doses (0.1, 1 and 10 mg/kg body weight) of CBZ for 5 weeks, then many factors related to spermatogenesis have been explored. It was found that 0.1-10 mg/kg body weight of CBZ exposure decreased mouse sperm motility and concentration, diminished the important protein factors (VASA, PGK2, B-Amy and CREM) for spermatogenesis, reduced sperm protein acrosin level, disrupted very vital epigenetic factors H3K27, 5 mC and 5 hmC. Furthermore, CBZ exposure damaged estrogen receptor alpha (ERα) pathway by decreased the protein levels of ERα and its targets PI3K and AKT. In summary low doses of CBZ exposure disrupted mouse spermatogenesis through estrogen receptor signaling; and that histone methylation and DNA methylation might play vital roles in CBZ disturbance of spermatogenesis through intertwining with estrogen signaling pathways. CBZ from the contamination in environment or food chain poses a serious threat to the normal development of spermatozoa. Therefore we strongly recommend to minimise the use of CBZ since it causes the severe issues on spermatogenesis.

Silkworm (Bombyx mori L.) Has Beneficial Effects on Menopausal Symptoms by Enhancing Estrogen Receptor Signaling in Ovariectomized Mice.

Existing hormone replacement therapy for menopause has drawbacks, necessitating new treatment agents. Silkworms have demonstrated estrogenic properties, offering promising alternatives. We assessed the therapeutic effects of freeze-dried silkworm powder (SWP) on menopausal symptoms using an ovariectomized (OVX) mouse model. The experimental design comprised a sham surgery group (Sham), an OVX control group, a low-dose SWP group post-OVX (80 mg/kg, OVX-SWP-L), a high-dose SWP group post-OVX (160 mg/kg, OVX-SWP-H), and an estradiol treatment group post-OVX (OVX-E2). Treatments were administered orally thrice weekly over eight weeks; body weight was monitored weekly. The SWP-treated groups (SWP-L and SWP-H) exhibited less weight gain and increased uterine thickness than the OVX control. Molecular analyses demonstrated that SWP significantly enhanced the phosphorylation of estrogen receptor alpha (ERα), ERK, and AKT. Furthermore, biochemical assays revealed reduced serum neutral lipids across all SWP treatment groups. Notably, HDL-cholesterol levels were significantly increased in the SWP-L group compared to the OVX group. Serum estradiol concentrations were elevated in all the SWP groups, with significant increases in the high-dose group. These findings indicate that SWP may promote the activation of estrogen receptor signaling and improve symptoms associated with estrogen deficiency during menopause.

Eosinophilia in cancer and its regulation by sex hormones.

Gender differences in the functionality of the immune system have been attributed, in part, to direct and indirect effects of sex steroids, especially estrogens, on immune cell repertoire and activity. Notable are studies that have defined roles for estrogens in the regulation of the biology of dendritic cells (DCs), macrophages, T cells and natural killer (NK) cells. Although estrogens can modulate eosinophil function, the mechanisms by which this occurs and how it contributes to the pathobiology of different diseases remains underexplored. Furthermore, although the importance of eosinophils in infection is well established, it remains unclear as to how these innate immune cells, which are present in different tumors, impact the biology of cancer cells and/or response to therapeutics. The observation that eosinophilia influences the efficacy of immune checkpoint blockers (ICBs) is significant considering the role of estrogens as regulators of eosinophil function and recent studies suggesting that response to ICBs is impacted by gender. Thus, in this review, we consider what is known about the roles of estrogen(s) in regulating tissue eosinophilia/eosinophil function and how this influences the pathobiology of breast cancer (in particular). This information provides the context for a discussion of how estrogens/the estrogen receptor (ER) signaling axis can be targeted in eosinophils and how this would be expected to influence the activity of standard-of-care interventions and contemporary immunotherapy regimens in cancer(s).

Activation function 2 (AF2) domain of estrogen receptor-α regulates mechanotransduction during bone fracture healing in estrogen-competent mice.

External mechanostimulation applied by whole-body low-magnitude high-frequency vibration (LMHFV) was demonstrated to cause no or negative effects on fracture healing in estrogen-competent rodents, while in ovariectomized (OVX), estrogen-deficient rodents bone formation after fracture was improved. Using mice with an osteoblast-specific deletion of the estrogen receptor α (ERα), we demonstrated that ERα signaling in osteoblasts is required for both the anabolic and catabolic effects of LMHFV during bone fracture healing in OVX and non-OVX mice, respectively. Because the vibration effects mediated by ERα were strictly dependent on the estrogen status, we hypothesized different roles of ligand-dependent and -independent ERα signaling. To investigate this assumption in the present study, we used mice with a deletion of the C-terminal activation function (AF) domain-2 of the ERα receptor, which mediated ligand-dependent ERα signaling (ERαAF-20). OVX and non-OVX ERαAF-20 animals were subjected to femur osteotomy followed by vibration treatment. We revealed that estrogen-competent mice lacking the AF-2 domain were protected from LMHFV-induced impaired bone regeneration, while the anabolic effects of vibration in OVX mice were not affected by the AF-2 knockout. RNA sequencing further showed that genes involved in Hippo/Yap1-Taz and Wnt signaling were significantly downregulated upon LMHFV in the presence of estrogen in vitro. In conclusion, we demonstrated that the AF-2 domain is crucial for the negative effects of vibration during bone fracture healing in estrogen-competent mice suggesting that the osteoanabolic effects of vibration are rather mediated by ligand-independent ERα signaling.

Postnatal maternal care moderates the effects of prenatal bisphenol exposure on offspring neurodevelopmental, behavioral, and transcriptomic outcomes.

Bisphenols (BPs), including BPA and "BPA-free" structural analogs, are commonly used plasticizers that are present in many plastics and are known endocrine disrupting chemicals. Prenatal exposure to BPA has been associated with negative neurodevelopmental and behavioral outcomes in children and rodent models. Prenatal BPA exposure has also been shown to impair postnatal maternal care provisioning, which can also affect offspring neurodevelopment and behavior. However, there is limited knowledge regarding the biological effects of prenatal exposure to bisphenols other than BPA and the interplay between prenatal BP exposure and postnatal maternal care on adult behavior. The purpose of the current study was to determine the interactive impact of prenatal BP exposure and postnatal maternal care on neurodevelopment and behavior. Our findings suggest that the effects of prenatal BP exposure on eye-opening, adult attentional set shifting and anxiety-like behavior in the open field are dependent on maternal care in the first five days of life. Interestingly, maternal care might also attenuate the effects of prenatal BP exposure on eye opening and adult attentional set shifting. Finally, transcriptomic profiles in male and female medial prefrontal cortex and amygdala suggest that the interactive effects of prenatal BP exposure and postnatal maternal care converge on estrogen receptor signaling and are involved in biological processes related to gene expression and protein translation and synthesis. Overall, these findings indicate that postnatal maternal care plays a critical role in the expression of the effects of prenatal BP exposure on neurodevelopment and adult behavior. Understanding the underlying biological mechanisms involved might allow us to identify potential avenues to mitigate the adverse effects of prenatal BP exposure and improve health and well-being in human populations.

Posttreatment with PaPE-1 Protects from Aß-Induced Neurodegeneration Through Inhibiting the Expression of Alzheimer's Disease-Related Genes and Apoptosis Process That Involves Enhanced DNA Methylation of Specific Genes.

Targeting the non-nuclear estrogen receptor (ER) signaling has been postulated as novel therapeutic strategy for central nervous system pathologies. Recently, we showed that newly designed PaPE-1 (Pathway Preferential Estrogen-1), which selectively activates ER non-nuclear signaling pathways, elicited neuroprotection in a cellular model of Alzheimer's disease (AD) when it was applied at the same time as amyloid-ß (Aß). Since delayed treatment reflects clinical settings better than cotreatment does, current basic study proposes a novel therapeutic approach for AD that relies on a posttreatment with PaPE-1. In this study, mouse neuronal cell cultures treated with preaggregated Aß1-42 (10 µM) showed the presence of extracellular Aß1-42, confirming the adequacy of the AD model used. We are the first to demonstrate that a 24-h delayed posttreatment with PaPE-1 decreased the degree of Aß-induced neurodegeneration, restored neurite outgrowth, and inhibited the expression of AD-related genes, i.e., Rbfox, Apoe, Bace2, App, and Ngrn, except for Chat, which was stimulated. In addition, PaPE-1 elicited anti-apoptotic effects by inhibiting Aß-induced caspase activities as well as attenuating apoptotic chromatin condensation, and in these ways, PaPE-1 prevented neuronal cell death. Posttreatment with PaPE-1 also downregulated the Aß-affected mRNA expression of apoptosis-specific factors, such as Bax, Gsk3b, Fas, and Fasl, except for Bcl2, which was upregulated by PaPE-1. In parallel, PaPE-1 decreased the protein levels of BAX, FAS, and FASL, which were elevated in response to Aß. PaPE-1 elicited a decrease in the BAX/BCL2 ratio that corresponds to increased methylation of the Bax gene. However, the PaPE-1-evoked Bcl2 gene hypermethylation suggests other PaPE-1-dependent mechanisms to control Aß-induced apoptosis.

Membrane estrogen signaling in female reproduction and motivation.

Estrogen receptors were initially identified in the uterus, and later throughout the brain and body as intracellular, ligand-regulated transcription factors that affect genomic change upon ligand binding. However, rapid estrogen receptor signaling initiated outside of the nucleus was also known to occur via mechanisms that were less clear. Recent studies indicate that these traditional receptors, estrogen receptor-α and estrogen receptor-ß, can also be trafficked to act at the surface membrane. Signaling cascades from these membrane-bound estrogen receptors (mERs) not only rapidly effect cellular excitability, but can and do ultimately affect gene expression, as seen through the phosphorylation of CREB. A principal mechanism of neuronal mER action is through glutamate-independent transactivation of metabotropic glutamate receptors (mGluRs), which elicits multiple signaling outcomes. The interaction of mERs with mGluRs has been shown to be important in many diverse functions in females, including, but not limited to, reproduction and motivation. Here we review membrane-initiated estrogen receptor signaling in females, with a focus on the interactions between these mERs and mGluRs.

Regulation of Bcl-2 Family Proteins in Estrogen Receptor-Positive Breast Cancer and Their Implications in Endocrine Therapy.

Estrogen receptor (ER)-positive breast cancer accounts for around two-thirds of breast cancer occurrences, with endocrine therapy serving as first-line therapy in most cases. Targeting estrogen signaling pathways, which play a central role in regulating ER+ breast cell proliferation and survival, has proven to improve patient outcomes. However, despite the undeniable advantages of endocrine therapy, a subset of breast cancer patients develop acquired or intrinsic resistance to ER-targeting agents, limiting their efficacy. The activation of downstream ER signaling pathways upregulates pro-survival mechanisms that have been shown to influence the response of cells to endocrine therapy. The Bcl-2 family proteins play a central role in cell death regulation and have been shown to contribute to endocrine therapy resistance, supporting the survival of breast cancer cells and enhancing cell death evasion. Due to the overexpression of anti-apoptotic Bcl-2 proteins in ER-positive breast cancer, the role of these proteins as potential targets in hormone-responsive breast cancer is growing in interest. In particular, recent advances in the development of BH3 mimetics have enabled their evaluation in preclinical studies with ER+ breast cancer models, and BH3 mimetics have entered early ER+ breast cancer clinical trials. This review summarizes the molecular mechanisms underlying the regulation of Bcl-2 family proteins in ER+ breast cancer. Furthermore, an overview of recent advances in research regarding the efficacy of BH3 mimetics in ER+ breast cancer has been provided.

Intercommunication between Voltage-Gated Calcium Channels and Estrogen Receptor/Estrogen Signaling: Insights into Physiological and Pathological Conditions.

Voltage-gated calcium channels (VGCCs) and estrogen receptors are important cellular proteins that have been shown to interact with each other across varied cells and tissues. Estrogen hormone, the ligand for estrogen receptors, can also exert its effects independent of estrogen receptors that collectively constitute non-genomic mechanisms. Here, we provide insights into the VGCC regulation by estrogen and the possible mechanisms involved therein across several cell types. Notably, most of the interaction is described in neuronal and cardiovascular tissues given the importance of VGCCs in these electrically excitable tissues. We describe the modulation of various VGCCs by estrogen known so far in physiological conditions and pathological conditions. We observed that in most in vitro studies higher concentrations of estrogen were used while a handful of in vivo studies used meager concentrations resulting in inhibition or upregulation of VGCCs, respectively. There is a need for more relevant physiological assays to study the regulation of VGCCs by estrogen. Additionally, other interacting receptors and partners need to be identified that may be involved in exerting estrogen receptor-independent effects of estrogen.

Patient-Derived Breast Cancer Tissue Cultures for Anti-Endocrine Drug Assays.

Breast cancer is a complex and heterogeneous pathology, characterized by a variety of histological and molecular phenotypes. The majority of the breast cancers express the estrogen receptor alpha (ER), which plays a pivotal role in the pathobiology of the disease and are therefore classified as ER-positive (ER+). In fact, targeting of the ER signaling pathway is the main therapeutic strategy for ER+ breast cancer. Despite the success of endocrine therapy, intrinsic and acquired resistance are reported in 30-50% of the ER+ breast cancers. However, the mechanisms underlying ER heterogeneity and therapeutic resistance are far from being fully disclosed, and efficacious clinical strategies to overcome resistance are still pending. One of the hurdles in studying ER+ breast cancer resistance is related with the scarcity of experimental models that can recapitulate ER heterogeneity and signaling. This is the case of ER+ breast cancer cell models, typically based on cells derived from metastasis, which also fail to recapitulate tumor complexity. Primary cultures of patient-derived breast cancer cells are difficult to establish, and generally characterized by stromal fibroblasts overgrowth and rapid loss of phenotypic and molecular traits of the tumor cells, including ER expression. Ex vivo cultures of breast cancer tissue have been reported to retain the tissue architecture, with preservation of the tumor microenvironment (TME) and ER expression for short periods of time.Given the cumulating evidence on the role of the TME in sustaining ER+ tumor cells, we hypothesized that TME preservation in culture would favor the long-term retention of ER expression and signaling. We employed alginate encapsulation to provide a supporting scaffold to breast cancer tissue microstructures, coupled to dynamic culture to improve the lifespan of the culture by avoiding diffusional limitations. In this chapter, we provide a detailed description of this culture methodology, which has been previously published by our group (Cartaxo et al., J Exp Clin Cancer Res 39:161, 2020), based on electrostatically driven breast cancer tissue encapsulation in alginate, coupled to culture under agitation in a defined culture medium. We also describe challenge of the ex vivo model with an ER activator and inhibitors (anti-endocrine drugs) and a gene expression endpoint of drug response using reverse transcription PCR-based analysis of three distinct genes downstream of ER.

Estrogen Receptor α Signaling in Osteoblasts is Required for Mechanotransduction in Bone Fracture Healing.

Biomechanical stimulation by whole-body low-magnitude high-frequency vibration (LMHFV) has demonstrated to provoke anabolic effects on bone metabolism in both non-osteoporotic and osteoporotic animals and humans. However, preclinical studies reported that vibration improved fracture healing and bone formation in osteoporotic, ovariectomized (OVX) mice representing an estrogen-deficient hormonal status, but impaired bone regeneration in skeletally healthy non-OVX mice. These effects were abolished in general estrogen receptor α (ERα)-knockout (KO) mice. However, it remains to be elucidated which cell types in the fracture callus are targeted by LMHFV during bone healing. To answer this question, we generated osteoblast lineage-specific ERα-KO mice that were subjected to ovariectomy, femur osteotomy and subsequent vibration. We found that the ERα specifically on osteoblastic lineage cells facilitated the vibration-induced effects on fracture healing, because in osteoblast lineage-specific ERα-KO (ERαfl/fl; Runx2Cre) mice the negative effects in non-OVX mice were abolished, whereas the positive effects of vibration in OVX mice were reversed. To gain greater mechanistic insights, the influence of vibration on murine and human osteogenic cells was investigated in vitro by whole genome array analysis and qPCR. The results suggested that particularly canonical WNT and Cox2/PGE2 signaling is involved in the mechanotransduction of LMHFV under estrogen-deficient conditions. In conclusion, our study demonstrates a critical role of the osteoblast lineage-specific ERα in LMHFV-induced effects on fracture healing and provides further insights into the molecular mechanism behind these effects.

Coronary Microvascular Dysfunction and Estrogen Receptor Signaling.

Chest pain with non-obstructive coronary artery disease (NOCAD) occurs more frequently in women than in men and is mainly related to coronary microvascular disease (CMD). The majority of CMD patients are postmenopausal women, suggesting a role for lack of estrogens in the development and progression of CMD. Patients are often discharged without a clear treatment plan due to the limited understanding of etiology and diagnostic parameters of CMD and have significantly higher rates of future cardiovascular events. Thus, there is a need for a better understanding of the underlying biology, and CMD-specific diagnostic tests and therapies. In this article, we reviewed recent studies on CMD, estrogen action in coronary microvasculature, and diagnosis and treatment options for CMD in postmenopausal women.

Functional analysis of familial Asp67Glu and Thr1051Ser BRCA1 mutations in breast/ovarian carcinogenesis.

Estrogen is believed to be pre-initiator in the risk of breast cancer. The BRCA1 is a tumor suppressor gene associated with breast and ovarian cancer risk. This report describes functional analysis of two BRCA1 missense mutations (Asp67Glu and Thr1051Ser) observed in the familial breast/ovarian cancer patients in Thailand. Levels of luciferase activity of the two mutations were relatively lower than in the wild-type BRCA1. It is indicated that mutants may fail to promote the estrogen receptor dependent functions. It is presumed that estrogen and insulin/IGF-1 regulate c-Myc and cyclin D1 during breast cancer cell proliferation. It is also likely to affect ubiquitination mechanism. Since three affected cancer families carry the Asp67Glu mutation, it is believed that this type of mutation could have some effect on breast/ovarian cancer progression.

Epigenetic Dysregulation at the Crossroad of Women's Cancer.

An increasingly number of women of all age groups are affected by cancer, despite substantial progress in our understanding of cancer pathobiology, the underlying genomic alterations and signaling cascades, and cellular-environmental interactions. Though our understanding of women's cancer is far more complete than ever before, there is no comprehensive model to explain the reasons behind the increased incidents of certain reproductive cancer among older as well as younger women. It is generally suspected that environmental and life-style factors affecting hormonal and growth control pathways might help account for the rise of women's cancers in younger age, as well, via epigenetic mechanisms. Epigenetic regulators play an important role in orchestrating an orderly coordination of cellular signals in gene activity in response to upstream signaling and/or epigenetic modifiers present in a dynamic extracellular milieu. Here we will discuss the broad principles of epigenetic regulation of DNA methylation and demethylation, histone acetylation and deacetylation, and RNA methylation in women's cancers in the context of gene expression, hormonal action, and the EGFR family of cell surface receptor tyrosine kinases. We anticipate that a better understanding of the epigenetics of women's cancers may provide new regulatory leads and further fuel the development of new epigenetic biomarkers and therapeutic approaches.

Ginsenoside Rg1 activates ligand-independent estrogenic effects via rapid estrogen receptor signaling pathway.

BACKGROUND: Ginsenoside Rg1 was shown to exert ligand-independent activation of estrogen receptor (ER) via mitogen-activated protein kinase-mediated pathway. Our study aimed to delineate the mechanisms by which Rg1 activates the rapid ER signaling pathways. METHODS: ER-positive human breast cancer MCF-7 cells and ER-negative human embryonic kidney HEK293 cells were treated with Rg1 (10-12M, 10-8M), 17ß-estradiol (10-8M), or vehicle. Immunoprecipitation was conducted to investigate the interactions between signaling protein and ER in MCF-7 cells. To determine the roles of these signaling proteins in the actions of Rg1, small interfering RNA or their inhibitors were applied. RESULTS: Rg1 rapidly induced ERα translocation to plasma membrane via caveolin-1 and the formation of signaling complex involving linker protein (Shc), insulin-like growth factor-I receptor, modulator of nongenomic activity of ER (MNAR), ERα, and cellular nonreceptor tyrosine kinase (c-Src) in MCF-7 cells. The induction of extracellular signal-regulated protein kinase and mitogen-activated protein kinase kinase (MEK) phosphorylation in MCF-7 cells by Rg1 was suppressed by cotreatment with small interfering RNA against these signaling proteins. The stimulatory effects of Rg1 on MEK phosphorylation in these cells were suppressed by both PP2 (Src kinase inhibitor) and AG1478 [epidermal growth factor receptor (EGFR) inhibitor]. In addition, Rg1-induced estrogenic activities, EGFR and MEK phosphorylation in MCF-7 cells were abolished by cotreatment with G15 (G protein-coupled estrogen receptor-1 antagonist). The increase in intracellular cyclic AMP accumulation, but not Ca mobilization, in MCF-7 cells by Rg1 could be abolished by G15. CONCLUSION: Ginsenoside Rg1 exerted estrogenic actions by rapidly inducing the formation of ER containing signalosome in MCF-7 cells. Additionally, Rg1 could activate EGFR and c-Src ER-independently and exert estrogenic effects via rapid activation of membrane-associated ER and G protein-coupled estrogen receptor.

Nucleoporin 153 regulates estrogen-dependent nuclear translocation of endothelial nitric oxide synthase and estrogen receptor beta in prostate cancer.

Nucleoporin 153 (Nup153), key regulator of nuclear import/export, has been recently associated to oncogenic properties in pancreatic and breast tumour cells modulating either cell motility and migration or gene expression by chromatin association. In the present work, we have characterized the role of Nup153 in a cellular model of prostate cancer (PCa). The analysis of several immortalized cell lines derived from freshly explants of prostate cancer specimens showed that Nup153 protein was higher and present in multimeric complexes with eNOS and ERß as compared to normal/hyperplastic prostate epithelial cells. This phenomenon was enhanced in the presence of 17ß-estradiol (E2, 10-7M). Further experiments revealed that eNOS and ERß were present in a DNA binding complexes associated with Nup153 promoter as demonstrated by ChIPs. Notably, after Nup153 depletion (siNup153), a reduction of migration capacity and colony formation in primary tumor-derived and metastatic PCa cells was observed. In addition, eNOS and ERß nuclear localization was lost upon siNup 153 regardless of E2 treatment, suggesting that Nup153 is a key regulator of prostate cancer cell function and of the nuclear translocation of these proteins in response to hormone stimulus. Taken altogether our findings indicate that in PCa cells: i. the expression and function of Nup153 is modulated by estrogen signaling; ii. Nup153 contributes to cell migration and proliferation; iii. Nup153 regulates the nuclear translocation of eNOS and ERß by forming a multimeric complex. Our findings unveil Nup153 as a novel component of the estrogen-dependent multimeric complex, thus representing a potential therapeutic candidate in prostate cancer.