ERα Levels Research Articles

The detrimental effects of APOE4 on risk for Alzheimer's disease may result from altered dendritic spine density, synaptic proteins, and estrogen receptor alpha

Women carriers of APOE4, the greatest genetic risk factor for late-onset Alzheimer's disease (AD), are at highest risk of developing AD, yet factors underlying interactions between APOE4 and sex are not well characterized. Here, we examined how sex and APOE3 or APOE4 genotypes modulate object and spatial memory, dendritic spine density and branching, and protein expression in 6-month-old male and female E3FAD and E4FAD mice (APOE+/+/5xFAD+/−). APOE4 negatively impacted object recognition and spatial memory, with male E3FADs exhibiting the best memory across 2 object-based tasks. In both sexes, APOE4 reduced basal dendritic spine density in the medial prefrontal cortex and dorsal hippocampus. APOE4 reduced dorsal hippocampal levels of PDS-95, synaptophysin, and phospho-CREB, yet increased levels of ERα. E4FAD females exhibited strikingly increased GFAP levels, in addition to the lowest levels of PSD-95 and pCREB. Overall, our results suggest that APOE4 negatively impacts object memory, dendritic spine density, and levels of hippocampal synaptic proteins and ERα. However, the general lack of sex differences or sex by genotype interactions suggests that the sex-specific effects of APOE4 on AD risk may be related to factors unexplored in the present study.

Kolaviron pre-treatment suppresses 7, 12 dimethylbenzanthracene-induced alterations in estrogen receptor-α, CYP 1A1, oxidative stress and inflammation in female Wistar rats.

Due to the need to develop locally available, cheaper, and efficacious treatment regimens for breast cancer, the chemopreventive effect of kolaviron (KV), an extract of Garcinia kola seeds was examined. Fifty (50) female Wistar rats (120-180g) were assigned to five groups (control group, 7, 12 dimethylbenzanthracene [DMBA] groups, tamoxifen group) of 10 rats each. They were pre-treated with KV thrice a week for four weeks except control. Estrogen receptor-α (ER-α) levels were determined in the pre-treated rats before induction of mammary carcinogenesis. After the four weeks pre-treatment period, 80mg/kg of DMBA was used for induction. A hundred and fifty (150) days after induction, the rats were sacrificed humanely. Significantly higher levels of ER-α, formation of lobular neoplastic cells, epithelial hyperplasia, lymphocyte infiltration, increased cytokines (interleukin-6 [IL-6] and tumor necrosis factor-α [TNF-α]), CYP1A1 activity and malondialdehyde (MDA) with a corresponding decrease in superoxide dismutase (SOD), catalase and glutathione peroxidase were observed in DMBA-induced rats. Pre-treatment with KV at 200mg/kg body weight significantly (p<.05) decreased ER-α levels by 19.01% and 37.52%, [IL-6] by 36.37% and 20.55%, TNF-α by 42.2% and 12.33% in serum and mammary tissue respectively. Also, a significant (p<.05) decrease in serum CYP1A1 activity, MDA with concomitant increase in SOD, catalase and glutathione peroxidase activities were observed in serum and mammary tissue respectively. Collectively, the results suggest that KV could be further explored in targeting chemoprevention of DMBA-induced mammary damage. PRACTICAL APPLICATIONS: Garcinia kola is widely cultivated in West and Central Africa with kolaviron (KV) as its major constituents. The seeds which have a bitter astringent taste are widely consumed by people in the region. Locals claim that consumption of the seeds provides relief for the management of several ailments including cancer. However, scientific investigations that provide a basis for these claims are still needed. This study provides evidence that points to the ameliorative potential of KV on breast cancer model. The results will be beneficial to local communities who hitherto had no knowledge on the potential of G. kola in chemoprevention. The results from this study will also attract further research attention from the international scientific community to examine the anti-cancer benefits of G. kola. This will also be beneficial to the global community due to the increasing number of breast cancer cases recorded annually.

Association between the rs3802201 polymorphism of the lncRNA MIR2052HG gene and the risk of recurrent miscarriage in a Southern Chinese population.

BackgroundPlenty of studies have indicated that some genetic polymorphisms of the breast cancer which associated with its susceptibility may also be related to the susceptibility of abortion. MIR2052HG plays an important role in the onset and progression of breast cancer by maintaining the level of ERα, but to the best of our knowledge, the correlation between risk of recurrent abortion and MIR2052HG rs3802201 C>G polymorphism is still unclear. Therefore, we conducted this case‐control study to investigate whether MIR2052HG rs3802201 C>G polymorphism is associated with susceptibility of recurrent miscarriage (RM).MethodsWe recruited 392 healthy controls and 248 patients with RM to process this research, the participants were all from southern China, and genotyping was performed by TaqMan method.ResultsOur results showed that there was no evidence indicates the MIR2052HG rs3802201 C>G is related to RM (CG and CC: adjusted OR = 0.970, 95% CI = 0.694–1.355, p = 0.8577; GG and CC: adjusted OR = 0.743, 95% CI = 0.416–1.330, p = 0.3174; dominant model: adjusted OR = 0.925, 95% CI = 0.672–1.272, p = 0.6298; recessive model: adjusted OR = 0.751, 95% CI = 0.430–1.321, p = 0.3233).ConclusionWe verified that the MIR2052HG rs3802201 C>G allele might be uncorrelated to the RM risk, but these findings require further validation in multicenter studies with larger sample size and different ethnicities.

Transcription Regulation and Genome Rewiring Governing Sensitivity and Resistance to FOXM1 Inhibition in Breast Cancer.

Simple SummaryThe oncogenic transcription factor FOXM1 is overexpressed in many cancers and associated with poor patient outcomes. Hence, there is much interest in blocking FOXM1 activity in cancer. We used small molecule inhibitors of FOXM1 to understand how they impact gene regulatory networks in suppressing cancer cell survival and the rewiring of gene networks that occurs when breast cancer cells become resistant to these compounds. Resistant cells showed reversal of the expression of many genes in the FOXM1 network controlling cell cycle progression, DNA damage repair, and apoptosis and also enhanced inflammatory signaling and upregulated HER2 and EGFR pathways. Targeting some of these factors so as to reduce the inflammatory and growth factor-dominant state of the resistant cancer cells should offer promising approaches for suppressing cancer progression and improving treatment of breast cancer.Forkhead box M1 (FOXM1), an oncogenic transcription factor associated with aggressiveness and highly expressed in many cancers, is an emerging therapeutic target. Using novel 1,1-diarylethylene-diammonium small molecule FOXM1 inhibitors, we undertook transcriptomic, protein, and functional analyses to identify mechanisms by which these compounds impact breast cancer growth and survival, and the changes that occur in estrogen receptor (ERα)-positive and triple negative breast cancer cells that acquire resistance upon long-term treatment with the inhibitors. In sensitive cells, these compounds regulated FOXM1 gene networks controlling cell cycle progression, DNA damage repair, and apoptosis. Resistant cells showed transcriptional alterations that reversed the expression of many genes in the FOXM1 network and rewiring that enhanced inflammatory signaling and upregulated HER2 or EGFR growth factor pathways. ERα-positive breast cancer cells that developed resistance showed greatly reduced ERα levels and responsiveness to fulvestrant and a 10-fold increased sensitivity to lapatinib, suggesting that targeting rewired processes in the resistant state may provide benefits and prolong anticancer effectiveness. Improved understanding of how FOXM1 inhibitors suppress breast cancer and how cancer cells can defeat their effectiveness and acquire resistance should be helpful in directing further studies to move these agents towards translation into the clinic.

Effects of Fuke Qianjin Formula on hormones and their receptors and metabonomics study in uterine fibroids model rats

ObjectiveTo investigate the effects of different fractions from Fuke Qianjin Formula (妇科千金方, FKQJF) on uterine leiomyoma (UL) to determine the best fraction. MethodsFKQJF was extracted and isolated to obtain polysaccharides (FKP), flavonoids (FKF), and grease (FKG). 140 female SPF SD rats were divided into 14 groups [model (MOD), normal control (NC), Gouliuqing (GLQ), Mifepristone (MFST), FKQJF, low, medium, and high dose of polysaccharides (l-FKP, m-FKP, and h-FKP), low, medium, and high dose of flavonoids (l-FKF, m-FKF, and h-FKF), low, medium, and high dose of grease (l-FKG, m-FKG, and h-FKG)], and uterine fibroids model rats were treated with drugs for four weeks. Serum levels of estrogen and progesterone were measured using enzyme-linked immunoassay assay (ELISA) kits. The expression of estrogen receptor (ER-α, ER-β) and progesterone receptor (PR) in the uterus was observed using immunohistochemistry (IHC). Serum metabolite profiles and FKG were analyzed using gas chromatography-mass spectrometry (GC-MS). ResultsFKQJF, h-FKF, m-FKG, and h-FKG significantly downregulated the estrogen level in the uterine fibroid model rats (P < 0.01). FKQJF, h-FKF, and h-FKG significantly reduced the level of progesterone in the uterine fibroid model rats (P < 0.01). The levels of ER-α, ER-β, and PR in uterine fibroid model rats were significantly decreased by FKQJF and h-FKG (P < 0.01). The levels of ER-α, ER-β, and PR in the fibroid model rats were decreased by m-FKG (P < 0.05). Additionally, serum metabolism results revealed that h-FKG and FKQJF could regulate related endogenous metabolites and make the pathological indices of uterine fibroids in rats close to the normal group. Forty-six components were identified in the oil, accounting for 91.97% of the total oil components. ConclusionFKQJF and h-FKG showed a significant anti-myoma activity and significantly improved the pathological state of the uterus in rats with hysteromyoma. The mechanism of action may be related to the regulation of estrogen progesterone and its receptor in uterine fibroid model animals. These findings proved the effect of FKQJF on uterine leiomyoma and provided an experimental basis for its clinical research and application.

Effect of 17β-estradiol on the proliferation of condylar chondrocytes.

To study the effects of 17β-estradiol (E2) on the regulation of the proliferation of condylar chondrocytes and provide a preliminary discussion on the role of phosphorylate-mammalian target of rapamycin (p-mTOR) in this regulatory process. Condylar chondrocytes were isolated from 6-week-old female rats for primary culture. Drug treatment with different concentrations of E2 and/or rapamycin (RAPA) was carried out on second-generation cells. Cell Counting Kit 8 was used to measure the cell viability of condylar chondrocytes after culture for 24, 48, or 72 h, and reverse transcription-polymerase chain reaction (RT-PCR) was applied to detect the relative gene expression of estrogen receptor alpha (ERα), estrogen receptor beta (ERβ), collagen type Ⅱ (COLⅡ), autophagy-related gene 6 (Beclin-1), and autophagy-related gene 5 (ATG-5). Western blot was employed to determine the relative protein expression of ERα, ERβ, Beclin-1, lipid-modified light chain 3B (LC3-Ⅱ), and p-mTOR. E2 could significantly promote the proliferation of chondrocytes cultured in vitro, and maximum promotion was achieved at a concentration of 10-8 mol·L-1. RAPA could significantly inhibit cell proliferation. E2 at aconcentration of 10-8 mol·L-1 could greatly improve the gene expression levels of ERα and COLⅡ (P<0.01) with the protein levels of ERα and p-mTOR (P<0.05), and decrease the gene expression levels of Beclin-1 and ATG-5 (P<0.05) with the protein levels of Beclin-1 and LC3-Ⅱ (P<0.05). RAPA could also enhance the relative protein expression of Beclin-1 and LC3-Ⅱ (P<0.01), and reduce the expression of p-mTOR (P<0.01). Treatment with the ERα antagonist significantly reduced the expression of p-mTOR in cells (P<0.01). At a concentration of 10-8 mol·L-1, E2 could effectively activate the phosphorylation of mTOR through the ERα-p-mTOR pathway, inhibit cell autophagy, and promote the proliferation of condylar chondrocytes.

The Role of Estrogen Receptor α in Response to Longitudinal Bone Growth in ob/ob Mice.

The absence of leptin results in contrasting growth pattern of appendicular and axial bone growth in ob/ob mice. Endochondral bone formation is an important procedure of growth plate determining the bone growth, where this procedure is also regulated by estrogen and its receptor (ER) signaling pathway. The present study is undertaken to explore the roles of ERs in regulating the different growth patterns in ob/ob mice. In this study, C57BL/6 female mice were used as wild-type (WT) mice; ob/ob mice and WT mice were age-matched fed, and bone length is analyzed by X-ray plain film at the 12 weeks old. We confirm that ob/ob mice have shorter femoral length and longer spine length than WT mice (p < 0.05). The contrasting expression patterns of chondrocyte proliferation proteins and hypertrophic marker proteins are also observed from the femur and spinal growth plate of ob/ob mice compared with WT mice (p < 0.01). Spearman’s analysis showed that body length (axial and appendicular length) is positively related to the expression level of ERα in growth plate. Three-week-old female ob/ob mice are randomized divided into three groups: 1) ob/ob + ctrl, 2) ob/ob + ERα antagonist (MPP), and 3) ob/ob + ERβ antagonist (PHTPP). Age-matched C57BL/6 mice were also divided into three groups, same as the groups of ob/ob mice. MPP and PHTPP were administered by intraperitoneal injection for 6 weeks. However, the results of X-ray and H&E staining demonstrate that leptin deficiency seems to disturb the regulating effects of ER antagonists on longitudinal bone growth. These findings suggested that region-specific expression of ERα might be associated with contrasting phenotypes of axial and appendicular bone growth in ob/ob mice. However, ER signaling on longitudinal bone growth was blunted by leptin deficiency in ob/ob mice, and the underlying association between ERs and leptin needs to be explored in future work.

Heat shock factor 1 (HSF1) cooperates with estrogen receptor α (ERα) in the regulation of estrogen action in breast cancer cells.

Heat shock factor 1 (HSF1), a key regulator of transcriptional responses to proteotoxic stress, was linked to estrogen (E2) signaling through estrogen receptor α (ERα). We found that an HSF1 deficiency may decrease ERα level, attenuate the mitogenic action of E2, counteract E2-stimulated cell scattering, and reduce adhesion to collagens and cell motility in ER-positive breast cancer cells. The stimulatory effect of E2 on the transcriptome is largely weaker in HSF1-deficient cells, in part due to the higher basal expression of E2-dependent genes, which correlates with the enhanced binding of unliganded ERα to chromatin in such cells. HSF1 and ERα can cooperate directly in E2-stimulated regulation of transcription, and HSF1 potentiates the action of ERα through a mechanism involving chromatin reorganization. Furthermore, HSF1 deficiency may increase the sensitivity to hormonal therapy (4-hydroxytamoxifen) or CDK4/6 inhibitors (palbociclib). Analyses of data from The Cancer Genome Atlas database indicate that HSF1 increases the transcriptome disparity in ER-positive breast cancer and can enhance the genomic action of ERα. Moreover, only in ER-positive cancers an elevated HSF1 level is associated with metastatic disease.

Novel role of estrogen receptor-α on regulating chondrocyte phenotype and response to mechanical loading

In knee cartilage from patients with osteoarthritis (OA), both preserved cartilage and damaged cartilage are observed. In this study, we aim to compare preserved with damaged cartilage to identify the molecule(s) that may be responsible for the mechanical loading-induced differences within cartilage degradation. Preserved and damaged cartilage were harvested from the same OA knee joint. RNA Sequencing was performed to examine the transcriptomic differences between preserved and damaged cartilage cells. Estrogen receptor-α (ERα) was identified, and its function of was tested through gene knockin and knockout. The role of ERα in mediating chondrocyte response to mechanical loading was examined via compression of chondrocyte-laded hydrogel in a strain-controlled manner. Findings from the studies on human samples were verified in animal models. Level of estrogen receptor α (ERα) was significantly reduced in damaged cartilage compared to preserved cartilage, which were observed in both human and mice samples. Knockdown of ESR1, the gene encoding ERα, resulted in an upregulation of senescence- and OA-relevant markers in chondrocytes. Conversely, knockin of ESR1 partially reversed the osteoarthritic and senescent phenotype of OA chondrocytes. Using a three-dimensional (3D) culture model, we demonstrated that mechanical overload significantly suppressed ERα level in chondrocytes with concomitant upregulation of osteoarthritic phenotype. When ESR1 expression was suppressed, mechanical loading enhanced hypertrophic and osteogenic transition. Our study demonstrates a new estrogen-independent role of ERα in mediating chondrocyte phenotype and its response to mechanical loading, and suggests that enhancing ERα level may represent a new method to treat osteoarthritis.

Clinicopathological and Prognostic Significance of Klotho and Estrogen Receptors Expression in Human Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is male-predominant cancer, but the underlying mechanism remains unclear. This study aimed to investigate the expression of the age-suppressing gene klotho and estrogen receptors (ERs) in HCC patients and analyze their association with clinicopathological variables and their effects on prognosis. The expression patterns of klotho, ERα, and ERβ were determined by tissue microarray and immunohistochemical technique, and their correlations with clinicopathological characteristics were investigated using univariate and multivariate analysis. Klotho expression was significantly lower in HCC than in the adjacent noncancerous tissues (52.7% (49/93) vs. 90.8% (79/87), P = .000), and its protein level in HCC tissue was negatively correlated with clinical staging, histological grade, and stage of the primary tumor (T) (P < .05). Whereas the expression of nuclear ERα and ERβ was higher in HCC than their corresponding non-neoplastic tissues (55.9% (52/93) vs. 35.6% (31/87), P = .006; 59.1% (55/93) vs. 43.7% (38/87), P = .038), and the level of nuclear ERα and ERβ in HCC tissue was inversely correlated with T stage, tumor size, and clinical staging (P < .05). Correlation analysis showed the expression level of klotho, which is positively correlated with that of nuclear ERα (r = 0.243, P = .019). Patients with klotho-positive tumors had longer survival than those with klotho-negative tumors (P = .002). Cox proportional hazards model analysis demonstrated that positive expression of klotho was an important factor indicating good prognosis (P = .003). Klotho, partially regulated by ERα-mediated estrogen pathway, acts as a tumor suppressor and might be a novel biomarker candidate for predicting progression and prognosis in HCC patients.

The deep infiltrating endometriosis tissue has lower T-cadherin, E-cadherin, progesterone receptor and oestrogen receptor than endometrioma tissue

ObjectiveTo compare the T-cadherin, E-cadherin, PR and ER staining levels of deep infiltrating endometriosis (DIE) tissue, ovarian endometriomas and normal endometrial tissues. Materials and methodsDIE tissue of 24 cases, endometrioma of 30 cases and normal endometrial tissues of 30 cases were examined. T-cadherin, E-cadherin, ER-α and PR-α staining levels of DIE, endometrioma tissues and endometrial tissues were compared immunohistochemically. H-score was calculated to compare the expression of T-cadherin, E-cadherin, ER-α, PR-α in IHC staining based on the percentage of cells stained at each intensity level. ResultsT-cadherin, E-cadherin, ER and PR H-score were found lowest in DIE tissue and the highest in endometrial tissue (p < 0.0001, <0.0001, <0.0001 and < 0.0001, respectively). In correlation analysis, a positive correlation was found between T-cadherin, E-cadherin, PR and ER H-score (p < 0.0001 for each). No correlation was found between age, body mass index (BMI), visual analog scale (VAS) score, CA125, endometrioma size and the severity of dysmenorrhea, dyspareunia and dystonia (p > 0.05). ConclusionT-cadherin, E-cadherin, ER and PR H-score were found lowest in DIE tissue, the highest in endometrium tissue. The finding of lower expression of PR-α in endometriotic nodule in our study may be related to decrease in progesterone effect which could not inhibit the decrease in the expression of T-cadherin and E-cadherin, thus the invasiveness of DIE tissue. These findings suggest that DIE tissue and ovarian endometrioma tissues have a different biology.

Estrogen Receptor β as a Possible Double-Edged Sword Molecule in Breast Cancer: A Mechanism of Alteration of Its Role by Exposure to Endocrine-Disrupting Chemicals.

Estrogen is essential for the growth and development of mammary glands and its signaling is associated with breast cancer growth. Estrogen can exert physiological actions via estrogen receptors α/β (ERα/β). There is experimental evidence suggesting that in ERα/β-positive breast cancer, ERα promotes tumor cell proliferation and ERβ inhibits ERα-mediated transcriptional activity, resulting in abrogation of cell growth. Therefore, ERβ is attracting attention as a potential tumor suppressor, and as a biomarker and therapeutic target in the ERα/β-positive breast cancer. Based on this information, we have hypothesized that some endocrine-disrupting chemicals (EDCs) that can perturb the balance between ERα and ERβ expression levels in breast cancer cells might have effects on the breast cancer proliferation (i.e., down-regulation of the α-type of ER). We have recently reported that 4-methyl-2,4-bis(4-hydroxyphenyl)pent-1-ene (MBP), an active metabolite of bisphenol A, in ERα/β-positive human breast cancer significantly down-regulates ERα expression, yet stimulates cell proliferation through the activation of ERβ-mediated transcription. These results support our hypothesis by demonstrating that exposure to MBP altered the functional role of ERβ in breast cancer cells from suppressor to promoter. In contrast, some EDCs, such as Δ9-tetrahydrocannabinol and bisphenol AF, can exhibit anti-estrogenic effects through up-regulation of ERβ expression without affecting the ERα expression levels. However, there is no consensus on the correlation between ERβ expression levels and clinical prognosis, which might be due to differences in exposed chemicals. Therefore, elucidating the exposure effects of EDCs can reveal the reason for inconsistent functional role of ERβ in ERα/β-positive breast cancer.

Naringin Improves Osteoblast Mineralization and Bone Healing and Strength through Regulating Estrogen Receptor Alpha-Dependent Alkaline Phosphatase Gene Expression.

Phytoestrogens are strongly recommended for treating osteoporosis. Our previous study showed that naringin, a citrus flavonoid, can enhance the bone mass in ovariectomized rats. In this study, we further elucidated the mechanisms of naringin-induced osteoblast maturation and bone healing. Treatment of human osteoblasts with naringin increased cell viability and proliferation. In parallel, exposure to naringin enhanced translocation of estrogen receptor alpha (ERα) to nuclei and its transactivation activity. Sequentially, naringin induced alkaline phosphatase (ALP) mRNA and protein expression and its enzyme activity. Pretreatment with methylpiperidinopyrazole (MPP), a specific inhibitor of ERα, attenuated naringin-induced augmentations in ERα transactivation activity, ALP gene expression, and cell mineralization. The beneficial effects of naringin were also confirmed in mouse MC3T3-E1 cells. Moreover, administration of mice with a bone defect with naringin increased levels of ERα and ALP in damaged sites and simultaneously enhanced the healing rate and bone strength. Nevertheless, treatment with MPP weakened naringin-triggered expression of ERα and ALP and improved bone healing and mass. Therefore, naringin could improve osteoblast mineralization and bone healing via regulating ERα-dependent ALP gene expression. Naringin can be clinically applied for treatment of osteoporosis-related bone diseases.

Balanced dual acting compounds targeting aromatase and estrogen receptor α as an emerging therapeutic opportunity to counteract estrogen responsive breast cancer

Breast Cancer (BC) is a leading cause of death in women, currently affecting 13% of female population worldwide. First-line clinical treatments against Estrogen Receptor positive (ER+) BC rely on suppressing estrogen production, by inhibiting the aromatase (AR) enzyme, or on blocking estrogen-dependent pro-oncogenic signaling, by targeting Estrogen Receptor (ER) α with selective Modulators/Degraders (SERMs/SERDs). The development of dual acting molecules targeting AR and ERα represents a tantalizing alternative strategy to fight ER+BC, reducing the incidence of adverse effects and resistance onset that limit the effectiveness of these gold-standard therapies. Here, in silico design, synthesis, biological evaluation and an atomic-level characterization of the binding and inhibition mechanism of twelve structurally related drug-candidates enable the discovery of multiple compounds active on both AR and ERα in the sub-μM range. The best drug-candidate 3a displayed a balanced low-nanomolar IC50 towards the two targets, SERM activity and moderate selectivity towards a BC cell line. Moreover, most of the studied compounds reduced ERα levels, suggesting a potential SERD activity. This study dissects the key structural traits needed to obtain optimal dual acting drug-candidates, showing that multitarget compounds may be a viable therapeutic option to counteract ER+BC.

Expression of GALNT8 and O-glycosylation of BMP receptor 1A suppress breast cancer cell proliferation by upregulating ERα levels

BackgroundMucin-type O-glycosylation is one of the most abundant types of O-glycosylation and plays important roles in various human carcinomas, including breast cancer. A large family of polypeptide N-acetyl-α-galactosaminyltransferases (GALNTs) initiate and define sites of mucin-type O-glycosylation. However, the specific mechanisms underlying GALNT8 expression and its roles in tumorigenesis remain poorly characterized. MethodsGALNT8 expression was assessed in 140 breast cancer patients. Immunofluorescence, immunoprecipitation, lectin blot and quantitative real-time PCR were used to investigate the expression of GALNT8 and its role in regulating estrogen receptor α (ERα) via bone morphogenetic protein (BMP) signaling. ResultsThe expression of GALNT8 was associated with breast cancer patient survival. GALNT8 downregulation was associated with a reduction in ERα levels, while GALNT8 overexpression elevated the transcription and protein levels of ERα and suppressed colony formation, suggesting an important role of GALNT8 in cancer cell proliferation. Conversely, GALNT8 knockdown led to the inhibition of BMP/SMAD/RUNX2 axis, which decreased ERα transcription. Further analysis suggested that BMP receptor 1A (BMPR1A) was O-GalNAcylated. Sites mutation of BMPR1A indicated that Thr137 and Ser37/Ser39/Ser44/Thr49 of BMPR1A were the main O-glycosylation sites. Although we cannot exclude the indirect effect of GALNT8, our results demonstrated that the expression of GALNT8 and O-glycosylation of BMPR1A play key roles in regulating the activity of BMP/SMAD/RUNX2 signaling and ERα expression. ConclusionThese findings suggest that GALNT8 expression and abnormal O-GalNAcylation of BMPR1A increase ERα expression and suppress breast cancer cell proliferation by modulating the BMP signaling pathway. General significanceOur results identify the involvement of GALNT8 in regulating ERα expression.

The E3 ligase COP1 promotes ERα signaling and suppresses EMT in breast cancer.

ERα signaling drives proliferation, survival and cancer initiation in the mammary gland. Therefore, it is critical to elucidate mechanisms by which ERα expression is regulated. We show that the tumor suppressor E3 ligase COP1 promotes the degradative polyubiquitination of the microtubule-associated protein HPIP. As such, COP1 negatively regulates estrogen-dependent AKT activation in breast cancer cells. However, COP1 also induces ERα expression and ERα-dependent gene transcription, at least through c-Jun degradation. COP1 and ERα levels are positively correlated in clinical cases of breast cancer. COP1 also supports the metabolic reprogramming by estrogens, including glycolysis. On the other hand, COP1 suppresses EMT in breast cancer cells. COP1 deficiency also contributes to Tamoxifen resistance, at least through protective autophagy. Therefore, COP1 acts as an oncogenic E3 ligase by promoting ERα signaling but also acts as a tumor suppressor candidate by preventing EMT, which reflects a dual role of COP1 in breast cancer.

Knockdown of LMTK3 in the Endometrioid Adenocarcinoma Cell Line Ishikawa: Inhibition of Growth and Estrogen Receptor α.

ObjectiveThe curative effect of high-efficiency progesterone and other therapeutic drugs for endometrioid adenocarcinoma patients with preservation of reproductive capacity has not been satisfactory so far. Novel therapeutic drugs need to be explored.MethodsWe investigated the cytoplastic and nuclear expression levels of LMTK3 between endometrioid adenocarcinoma tissues and adjacent endometrial tissues by immunohistochemistry. We detected the effects of LMTK3 on cell viability of Ishikawa cells by CCK-8. We detected the effects of LMTK3 on cell cycle and apoptosis of Ishikawa cells by flow cytometry. We also detected the effects of LMTK3 knockdown on mRNA and protein levels of ERα by qRT-PCR and western blotting, respectively. We also used the cBioPortal online database to analyze the coexpression of LMTK3 and ESR1 in 1647 UCEC samples.ResultsWe used TMAs to identify that LMTK3 was mainly detected in the cytoplasm of endometrioid tissues, and cytoplasmic LMTK3 expression in endometrioid tissues was higher than that in adjacent endometrial tissues (P < 0.05). LMTK3 knockdown decreased the proliferation of Ishikawa cells through decreasing cell viability (P < 0.01), increasing G1 (P < 0.001) arrest, and promoting apoptosis (P < 0.01). There was a positive correlation between the mRNA expression levels of LMTK3 and ESR1 (Spearman: P=2.011e-5, R=0.13; Pearson: P=7.18e-8, R=0.17). Knockdown of LMTK3 also reduced the mRNA (P < 0.001) and protein (P < 0.001) levels of ERα.ConclusionsInhibitors of LMTK3 may be a possible future treatment for ERα and LMTK3 highly expressed endometrioid adenocarcinoma following appropriate studies.

Exposure of childbearing-aged female to phthalates through the use of personal care products in China: An assessment of absorption via dermal and its risk characterization

Phthalates (PAEs) are widely used in personal care products (PCPs) and skin care packaging materials. Through national representative sampling, 328 childbearing-aged females in China were investigated by questionnaire, whose contact factors for 30 cosmetic products were collected. According to the daily exposure method and adverse cumulative effects of PAE exposure on female reproduction, we derived the ERα, ERβ binding, and AR anti-androgenic effects. The utilization rates of acne cleanser, acne cream, cleanser (non-acne), and cream (non-acne) in volunteers were 21.90%, 22.22%, 51.63%, and 51.96%, respectively. Examining the data for PAEs in PCPs, the content of DBP (dibutyl phthalate) in them was significantly higher for tubes (0.26 ± 0.05 μg/g) and other packaging (pump type and metal tube) (0.25 ± 0.03 μg/g) than bowl (0.17 ± 0.04 μg/g). The DBP content of acne cream (0.27 ± 0.03 μg/g) was significantly higher than that of non-acne cream (0.17 ± 0.03 μg/g); likewise, there was significantly more DEHP (di (2-ethylhexyl) phthalate) in acne cleanser (0.87 ± 0.15 μg/g) than non-acne cleanser (0.64 ± 0.36 μg/g). Students and office worker were the main consumers of PCPs; however, among all occupation groups, the daily exposure dose of PCPs for workers was highest (mean = 0.0004, 0.0002, 0.0009 μg/kg bw/day for DEP (diethyl phthalate), DBP, and DEHP, respectively). The cumulative indices of PAEs' exposure revealed that the level of ERα and ERβ binding and AR anti-androgenic effects in workers was respectively 0.4935, 0.0186, and 0.2411 μg/kg bw/day. The risk index (HITDI and HIRfDs) of DEP, DBP, and DEHP was lower than their corresponding reference value (hazard index <1), but using PCPs may cause potential health risks. Therefore, we should pay attention to the adverse effects of PAEs on female reproductive functioning, especially the cumulative exposure of females of childbearing age.

A Genome-Edited ERα-HiBiT Fusion Reporter Cell Line for the Identification of ERα Modulators Via High-Throughput Screening and CETSA.

The estrogen receptor α (ERα) is a target of intense pharmacological intervention and toxicological biomonitoring. Current methods to directly quantify cellular levels of ERα involve antibody-based assays, which are labor-intensive and of limited throughput. In this study, we generated a post-translational reporter cell line, referred to as MCF7-ERα-HiBiT, by fusing a small pro-luminescent nanoluciferase (NLuc) tag (HiBiT) to the C-terminus of endogenous ERα in MCF7 cells. The tag allows the luminescent detection and quantification of endogenous ERα protein by addition of the complementary NLuc enzyme fragment. This MCF7-ERα-HiBiT cell line was optimized for quantitative high-throughput screening (qHTS) to identify compounds that reduce ERα levels. In addition, the same cell line was optimized for a qHTS cellular thermal shift assay to identify compounds that bind and thermally stabilize ERα. Here, we interrogated the MCF7-ERα-HiBiT assay against the NCATS Pharmacological Collection (NPC) of 2,678 approved drugs and identified compounds that potently reduce and thermally stabilize ERα. Our novel post-translational reporter cell line provides a unique opportunity for profiling large pharmacological and toxicological compound libraries for their effect on ERα levels as well as for assessing direct compound binding to the receptor, thus facilitating mechanistic studies by which compounds exert their biological effects on ERα.

Myocardial Expression of Estrogen Receptor-mRNA Is Associated With Lower Markers of Post-operative Organ Damage in Young Patients With Congenital Cardiac Defect.

Background: Estrogen receptors (ERs) relate to cardio-protection in adults, but their role in younger patients is not known. We aimed to assess the myocardial expression of ERα- and ERβ- mRNA in young patients with congenital cardiac disease and to analyze their putative protective role.Patients and Methods: Twenty children and young adults (seven females and 13 males) with a median age of 13.8 years (interquartile range: 12.3 years) were enrolled in this prospective study. The myocardial expression of ER-mRNA and genes involved in inflammation, growth, and stress response was assessed by real-time PCR and was correlated to post-operative (po) outcome.Results: ER-mRNA was detected in the myocardium of all patients, independently of gender and age. The expression of ER-mRNA correlated with that of mRNA coding for brain natriuretic peptide and for all cytokines tested. A higher ERα-mRNA expression correlated with lower troponin T concentrations at 24 h po (p = 0.032), higher PaO2/FiO2 ratio at 4 h po (p = 0.059), lower fluid retention at 4 h po (p = 0.048), and lower aspartate aminotransferase (AST) levels at 24 h po (p = 0.047). A higher ERβ-mRNA expression was also correlated with lower fluid retention at 24 h po (p = 0.048).Patients in whom the levels of ERα- and ERβ-mRNA were >P50 had lower troponin T (p = 0.003, respectively) and lower AST concentrations at 24 h po (p = 0.043, respectively) than the others.Conclusions: The expression of ERα- and ERβ-mRNA is present in the myocardium of children and young adults with congenital cardiac defect and is associated with lower markers of po organ damage. This suggests that ERs may provide perioperative organ protection in this population.