Estrogenic modulation of inflammation-related genes in male rats following volume overload
ABSTRACT: The purpose of this study was to determine whether estrogen had an effect on inflammation-related genes that contribute to this estrogen-mediated cardioprotection. Volume overload on the heart was induced by aortocaval fistula in 8 week old male Sprague Dawley rats and genes of interest in shams, fistula , and fistula +estrogen were identified using an inflammatory PCR array. Volume overload was induced in male rats treated with estrogen and non treated male rats to determine if inflammation-related genes contributed to estrogen-mediated cardioprotection.
Project description:To find out the potential targets in response to estrogen treatment on ACHN cell, a human renal cell carcinoma (RCC) cell line. Since estrogen can repress ACHN growth in a partly estrogen receptor-dependent manner, it is possible that phosphorylation state in ACHN cells is regulated.
Project description:Abnormalities in metabolism of energetic substrates may play a role in progression of chronic heart failure (HF). The goal of the study was to examine the extent and mechanisms of metabolic alterations in rat model of chronic HF due to volume overload. Volume overload was induced in 3 months old male Wistar rats by aorto-caval fistula. In the phase of symptomatic HF (after 21 weeks), we performed myocardial gene expression analysis. Cardiac tissue gene expression analysis showed downregulation of enzymes of respiratory cycle, mitochondrial fatty acid (FA) oxidation and attenuated expression of proteins responsible for FA translocation/transport (CD36/FAT, FABP3, FATP-1). Simultaneously, we performed gene expression analysis of fat tissue.
Project description:The right ventricle (RV) differs in several aspects from the left ventricle (LV) including its embryonic origin, physiological role and anatomical design. In contrast to LV hypertrophy, little is known about the molecular circuits, which are activated upon RV hypertrophy (RVH). We established a highly reproducible model of RVH in mice using pulmonary artery clipping (PAC), which avoids detrimental RV pressure overload and thus allows long-term survival of operated mice. Magnetic resonance imaging revealed pathognomonic changes with striking similarities to human congenital heart disease- or pulmonary arterial hypertension- patients. Comparative, microarray based transcriptome analysis of right- and left-ventricular remodeling identified distinct transcriptional responses to pressure-induced hypertrophy of either ventricle, which were mainly characterized by stronger transcriptional responses of the RV compared to the LV myocardium. Hierarchic cluster analysis revealed a RV- and LV-specific pattern of gene activity after induction of hypertrophy, however, we did not find evidence for qualitatively distinct regulatory pathways in RV compared to LV. Data mining of nearly three thousand RV-enriched genes under PAC disclosed novel potential (co)-regulators of long-term RV remodeling and hypertrophy. We reason that specific inhibitory mechanisms in RV restrict excessive myocardial hypertrophy and thereby contribute to its vulnerability to pressure overload. Alternative splicing and gene expression analysis during development of the heart and cardiomyoyte differentiation.
Project description:A molecular and bioinformatic pipeline permitting comprehensive analysis and quantification of myocardial miRNA and mRNA expression with next-generation sequencing was developed and the impact of enhanced PI3Kalpha signaling on the myocardial transcriptome signature of pressure overload-induced pathological hypertrophy was explored. miRNA and mRNA-Seq were carried out in four groups of mouse LV samples: WT sham, WT+TAC, caPI3Kalpha sham, caPI3Kalpha+TAC
Project description:Estrogen Receptor subtypes (ERα and ERβ) are transcription factors sharing similar structure, however, they often perform opposite roles in breast cancer’s cell proliferation and tumor progression. Besides the well-characterized genomic actions of ERs upon ligand binding, rapid non-genomic cytoplasmic changes together with the recently discovered ligand-free action of ERs are emerging as key regulators of tumorigenesis. The identification of cytoplasmic interaction partners of unliganded ERα and ERβ may help characterize the molecular basis of the extra-nuclear mechanism of action of these receptors, revealing novel mechanisms to explain their role in breast cancer response or resistance to endocrine therapy. To this aim, in this study, cytoplasmic extracts from stably expressing TAP-ERα and -ERβ MCF-7 cell clones were subjected to interaction proteomics in the absence of estrogen stimulation, leading to the identification of 84 and 142 proteins associated with unliganded ERα and ERβ, respectively. Functional analyses of ER subtype-specific interactomes revealed significant differences in the molecular pathways associated to each receptor in the cytoplasm. This work reports the first identification of the unliganded ERα and ERβ cytoplasmic interactomes in breast cancer cells, providing novel experimental evidence on the non-genomic effects of ERs in the absence of hormonal stimulus.
Project description:The estrogen receptor is the master transcriptional regulator of breast cancer phenotype and the archetype of a molecular therapeutic target. We mapped all estrogen receptor and RNA polymerase II binding sites on a genome-wide scale, identifying the authentic cis binding sites and target genes, in breast cancer cells. Combining this unique resource with gene expression data demonstrates distinct temporal mechanisms of estrogen-mediated gene regulation,particularly in the case of estrogen-suppressed genes. Furthermore, this resource has allowed the identification of cis-regulatory sites in previously unexplored regions of the genome and the cooperating transcription factors underlying estrogen signaling in breast cancer. Experiment Overall Design: This Series currently contains the gene expression data accompaning Carroll JS et al. Nature Genetics 38,1289-1297(2006). MCF7 cells were stimulated with 100 nM estrogen for 0, 3, 6, or 12h. All experiments were performed in triplicate.
Project description:TRIM24 PHD-Bromo domains exhibit preferential binding to unmethylated H3K4 and acetylated H3K27. TRIM24 is a co-activator of estrogen receptor (ER). The results suggest that specific ER-binding sites are depleted of H3K4me2 with estrogen treatment. TRIM24 binds these sites preferentially and facilitates ER-regulated gene expression, cell survival and proliferation. ChIP performed on MCF7 cells +/- estrogen with antibodies against ER, TRIM24 and H3K4me2. ChIP assays of ER, co-activator TRIM24 and H3K4me2 were performed with two concentrations of antibody, without and 6h with estrogen treatment of MCF7 cells. Antibody-enriched samples were sequenced two times, and compared to an IgG negative control and Input. Enriched DNA sequenced by Illumina Solexa.
Project description:The lymphatic system is a common avenue for the spread of breast cancer cells and dissemination through it occurs at least as frequently as hematogenous metastasis. Approximately 75% of primary breast cancers are estrogen receptor (ER) positive and the majority of these maintain receptor expression as lymph node (LN) metastases. However, it is unknown if ER function is equivalent in cancer cells growing in the breast and in the LNs. We have developed a model to assess estrogen responsiveness in ER(+) breast tumors and LN metastases. Fluorescent ER(+) MCF-7 tumors were grown in ovariectomized nude mice supplemented with estradiol. Once axillary LN metastasis arose, estradiol was withdrawn (EWD), for 1 or 4 weeks, or continued, to assess estradiol responsiveness. On EWD, proliferation rates fell similarly in tumors and LN metastases. However, estradiol-dependent ER down-regulation and progesterone receptor induction were deficient in LN metastases, indicating that ER-dependent transcriptional function was altered in the LN. Cancer cells from estradiol-treated and EWD primary tumors and matched LN metastases were isolated by laser capture microdissection. Global gene expression profiling identified transcripts that were regulated by the tissue microenvironment, by hormones, or by both. Interestingly, numerous genes that were estradiol regulated in tumors lost estradiol sensitivity or were regulated in the opposite direction by estradiol in LN metastases. We propose that the LN microenvironment alters estradiol signaling and may contribute to local antiestrogen resistance. Experiment Overall Design: 10 samples, including 3 each of estrogen and estrogen withdrawn axillary lymph nodes and 2 each of estrogen and estrogen withdrawn primary mammary gland tumors.
Project description:Estrogen deprivation using aromatase inhibitors is currently the standard of care for patients with estrogen-receptor (ER)-positive breast cancer. Unfortunately, prolonged estrogen deprivation leads to drug resistance (i.e. hormone-independent growth). We therefore used DNA microarray analysis to study the gene expression profiles of wild-type MCF-7 cells (which are sensitive to antihormone therapy) and long-term estrogen deprived MCF-7:5C and MCF-7:2A breast cancer cells (which are resistance to estrogen-deprivation; aromatase inhibitor resistant). Transcriptional profiling of wild-type MCF-7 cells and estrogen deprived MCF-7:5C and MCF-7:2A cells was performed using Affymetrix Human Genome U133 Plus 2.0 Array. Experiment Overall Design: We wanted to study the gene expression profiles of the different cell lines in their growth media without any drug treatment. Therefore, MCF-7, MCF-7:5C, and MCF-7:2A cells were grown in estrogen-free media (phenol red-free RPMI medium supplemented with 10% 4X dextran-coated charcoal-treated fetal bovine serum) until they were 70-80% confluent then RNA was extracted, labeled, and hybridized to the Affymetrix Human Genome U133 Plus 2.0 Arrays.
Project description:The estrogen receptor-α (ERα) is a transcription factor which plays a critical role in controlling cell proliferation and tumorigenesis by recruiting various cofactors to estrogen response elements (EREs) to induce or repress gene transcription. A deeper understanding of these transcriptional mechanisms may uncover novel therapeutic targets for ERα-dependent cancers. Here we show for the first time that BRD4 regulates ERα−induced gene expression by affecting elongation-associated phosphorylation of RNA Polymerase II (RNAPII P-Ser2) and histone H2B monoubiquitination (H2Bub1). Consistently, BRD4 activity is required for estrogen-induced proliferation of ER+ breast and endometrial cancer cells and uterine growth in mice. Genome-wide occupancy studies revealed an enrichment of BRD4 on transcriptional start sites as well as EREs enriched for H3K27ac and demonstrate a requirement for BRD4 for H2B monoubiquitination in the transcribed region of estrogen-responsive genes. Importantly, we further demonstrate that BRD4 occupancy correlates with active mRNA transcription and is required for the production of ERα-dependent enhancer RNAs (eRNAs). These results uncover BRD4 as a central regulator of ERα function and potential therapeutic target. mRNA expression profiles of MCF7 cells treated with +/- estrogen treatment under negative control siRNA, BRD4 siRNA or JQ1 treatment, in duplicates.