Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of mouse mammary gland responses to estrogen and progesterone

ABSTRACT: Epidemiological studies have shown that a full-term pregnancy at early age can decrease the breast cancer risk up to one-half. Pregnancy has been shown to prevent carcinogen-induced mammary tumors in rodents as well. The protective effect of pregnancy can be mimicked by administration of estrogen and progesterone to nulliparous rodents in amounts that are similar to those during pregnancy and alters responsiveness of p53 to DNA damage. Ovariectomized mice were treated with estrogen (E), progesterone (P), both estrogen and progesterone (E + P), or vehicle (V) alone and global expression profiles was analyzed to identify the mechanisms by which estrogen and progesterone combine to sensitize p53 function. Experiment Overall Design: Twenty ovariectomized animals were used for examine transcriptional effects of hormones by microarray. The hormones were administered by daily i.p. injection for 4 days.Treatment groups included: 4 animals receiving 2ug 17-beta-estrogen (E), 4 animals receiving 1mg progesterone (P), 5 animals receiving both estrogen and progesterone (E+P) and 4 animals receiving 100ul sesame oil (V). Epithelial-free fat pads from 3 animals receiving E+P were also analyzed to distinguish responses to E+P in the stroma (E+P CFP). Lymph nodes were removed from mammary glands at the time of collection.

ORGANISM(S): Mus musculus

SUBMITTER: D Joseph Jerry

PROVIDER: E-GEOD-5483 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Synchrony between embryo competency and uterine receptivity is essential for a successful implantation. Mice with ablation of COUP-TFII in the uterus (PRCre/+;COUP-TFIIflox/flox), exhibit implantation defects and increased ER activity in the luminal epithelium, suggesting the high ER activity may disrupt the window of uterine receptivity. In order to determine if the increased ER activity in PRCre/+;COUP-TFIIflox/flox mutant is the cause of the defective implantation, we inhibited of ER activity in order to rescue the implantation defect in mutant mice. ICI 182,780 (ICI), a pure ER antagonist, was administered to PRCre/+;COUP-TFIIflox/flox mutant and COUP-TFIIflox/flox control mice during receptive period and the number of implantation sites were examined. COUP-TFIIflox/flox control mice treated with oil or ICI showed the normal number of implantation sites. As expected no implantation sites were observed in PRCre/+;COUP-TFIIflox/flox mutant mice treated with oil, consistent with previous observation. However, implantation sites were detected, albeit at a reduced number in comparison to the control in PRCre/+;COUP-TFIIflox/flox mutant mice upon ICI treatment.. ICI treatment was also able to rescue the expression of WNT4 and BMP2, genes important for endometrial decidualization in the PRCre/+;COUP-TFIIflox/flox mutant mice. To ensure the rescue of embryo attachment and decidualization is a consequence of a reduction of estrogen receptor activity with ICI treatment of the mutants, we examined the expression of ER target gene, such as lactoferrin, in PRCre/+;COUP-TFIIflox/flox mutant mice. Having shown that ICI could rescue the implantation and decidualization defects of the PRCre/+;COUP-TFIIflox/flox mutant mice, the ability of ICI treatment to rescue pregnancy in these mice was assayed. While mice were born in COUP-TFIIflox/flox control mice given ICI, no pups were born in the PRCre/+;COUP-TFIIflox/flox mutant mice, with the loss in pregnancy in the PRCre/+;COUP-TFIIflox/flox mutant mice treated with ICI being due to defects in placentation. These results demonstrate that during the peri implantation period, COUP-TFIIâs role in regulating embryo attachment and decidualiton is through the reduction of ER activity. However COUP-TFII expression is still required in the post implantation period to facilitate placentation. Experiment Overall Design: Murine uteri were isolated at 6 hour after oil or ICI 182,780 injection for RNA extraction and hybridization on Affymetrix microarrays. To do this mice were ovariectomized and treated hormones (estrogen and progesterone) to mimic the early pregnant condition.

Project description:The growth and development of the uterus in response to 17β-estradiol (E2) is genetically controlled, with marked variation observed depending on the mouse strain studied. Previous work from our laboratory using inbred mice that are high (C57BL6/J; B6) or low (C3H/HeJ; C3H) responders to E2 has led to the identification of quantitative trait loci (QTL) associated with phenotypic variation in uterine growth and eosinophil infiltration. The mechanisms underlying differential responsiveness to E2, and the genes involved, are unknown. Therefore, we used a microarray approach to show association of distinct E2-regulated transcriptional signatures with genetically controlled high and low responses to E2. Among the 6,664 E2-responsive uterine transcripts, several reside within our previously identified QTL, including the ERα-tethering factor Runx1, demonstrated to enhance E2-mediated transcript regulation. The level of RUNX1 in uterine epithelial cells was shown to be 3.5-fold greater in B6 compared to C3H. Analysis of cellular functions in sets of strain-dependent E2-responsive transcripts indicated C3H-selective enrichment of apoptosis, consistent with a 7-fold increase in the apoptosis indicator CASP3, and a 2.4-fold decrease in the apoptosis inhibitor Naip1 in C3H vs. B6 following treatment with E2. Our novel insights into the mechanisms underlying the genetic control of tissue sensitivity to estrogen have great potential to advance understanding of individualized effects in physiological and disease states. Eight-week-old female B6, C3H, and (B6 × C3H)F1 (B6C3) hybrid mice were purchased from The Jackson Laboratory (Bar Harbor, ME). Animals were ovariectomized at NIEHS, rested for 1 to 2 wk, and then subjected to treatment with either E2 (40.0 μg/kg BW injected i.p.) in 0.1 ml saline containing 0.25% ethanol, or ethanol/saline vehicle. Animals were euthanized and tissue collected at 2 or 24 h after injection. Uterine tissue from 3 animals per treatment group was collected, and snap-frozen in liquid nitrogen for subsequent RNA isolation. Frozen uterine tissue from three animals per treatment group was pulverized, then homogenized in Trizol (Invitrogen, Carlsbad, CA), and RNA was prepared according to the manufacturer's protocol. Isolated RNA was then further purified using the QIAGEN (Valencia, CA) RNeasy mini prep kit clean-up protocol. Gene expression analysis was conducted using Agilent Whole Mouse Genome 4x44 Multiplex format oligo arrays (014868; Agilent Technologies, Santa Clara, CA) following the Agilent 1-color microarray-based gene expression analysis protocol. Starting with 500 ng of total RNA, Cy3-labeled cRNA was produced according to the manufacturer's protocol. For each sample, 1.65 ug of Cy3-labeled cRNAs were fragmented and hybridized for 17 hours in a rotating hybridization oven. Slides were washed and then scanned with an Agilent Scanner.

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Transcription profiling of mouse mammary gland responses to estrogen and progesterone

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