Project description:Embryo implantation into a receptive endometrium is tightly regulated by a variety of maternal factors, including cytokines, growth factors and transcription factors. Previous studies identified the leukaemia inhibitory factor (LIF), produced in uterine glands, as an essential factor for implantation. It was shown that LIF acts via its cell surface receptor to activate the transcription factor STAT3 in the uterine epithelial cells. However, the mechanisms via which STAT3 promotes uterine receptivity remain unknown. To address the molecular pathways regulated by STAT3 in the uterus, we created mice in which Stat3 gene is conditionally inactivated in uterine epithelium. These mutant mice are infertile due to implantation failure and exhibit a lack of embryo attachment to the luminal epithelium. Gene expression profiling of the epithelial tissue impaired in STAT3 activation revealed dysregulated expression of specific components of junctional complexes, including E-cadherin, M-NM-2-catenin, and claudins, which critically regulate epithelial cell polarity and embryo attachment. Additionally, mice lacking functional epithelial STAT3 showed markedly reduced stromal proliferation and differentiation, indicating that this transcription factor controls stromal function via a paracrine mechanism. The stromal defect arose from a drastic reduction in the production of several members of the epidermal growth factor (EGF) family in luminal epithelium of mutant uteri and consequent lack of activation of EGF receptor signaling and mitotic activity in the stromal cells. Collectively, our results uncovered intricate signaling networks operating downstream of STAT3 in uterine epithelium that regulate epithelial cell polarity, and stromal proliferation and differentiation, which are critical determinants of successful implantation. To identify the downstream targets of STAT3 in mouse uterine epithelial cells during pregnancy, we performed gene expression profling of mouse uterine epithelial cells on day 4 of pregnancy between Stat3 flox control and SW d/d mice. This led to the identification of several junctional molecules (Claudins and Catenins) that are negatively regulated by STAT3 at the time of implantation. Mouse uteirne epithelial cells were isolated from control and knockout mice on the morning of day 4 of pregnancy. (n=3 for each sample), pooled total RNA from these cells was then hybridized to high density affymetrix microarrays according to the Affymetrix protocol (Mouse Genome 430A 2.0 Array) .
Project description:Gene expression profiling of uterine epithelial cells isolated from Msx1Msx2 floxed and Msx1Msx2 ablated mice on day 4 of pregnancy
Project description:Our study revealed that hypoxia inducible factor 2 alpha, Hif2 alpha, is a downstream target of estrogen signaling in mouse uterine stroma at the time of implantation. Further, conditional deletion of Hif2 alpha in mouse uterus leads to infertility due to impaired epithelial remodeling at the time of implantation. To identify the downstream targets of Hif2 alpha in the uterus, we performed gene expression profiling of uterine stromal cells isolated from Hif2 alpha-intact and -null mice on day 5 of pregnancy, overlapping the window of implantation. The microarray results revealed altered expression of mRNAs corresponding to factors involved in protein trafficking in uterine stroma of Hif2 alpha-ablated mice. These factors mediate crosstalk between uterine storma and epithelial cells to promote epithelial remodeling and implantation. Thus, Hif2 alpha regulates embryo implantation by controlled trafficking of secretory granules during early pregnancy.
Project description:PURPOSE: To provide a detailed gene expression profile of the normal postnatal mouse cornea. METHODS: Serial analysis of gene expression (SAGE) was performed on postnatal day (PN)9 and adult mouse (6 week) total corneas. The expression of selected genes was analyzed by in situ hybridization. RESULTS: A total of 64,272 PN9 and 62,206 adult tags were sequenced. Mouse corneal transcriptomes are composed of at least 19,544 and 18,509 unique mRNAs, respectively. One third of the unique tags were expressed at both stages, whereas a third was identified exclusively in PN9 or adult corneas. Three hundred thirty-four PN9 and 339 adult tags were enriched more than fivefold over other published nonocular libraries. Abundant transcripts were associated with metabolic functions, redox activities, and barrier integrity. Three members of the Ly-6/uPAR family whose functions are unknown in the cornea constitute more than 1% of the total mRNA. Aquaporin 5, epithelial membrane protein and glutathione-S-transferase (GST) omega-1, and GST alpha-4 mRNAs were preferentially expressed in distinct corneal epithelial layers, providing new markers for stratification. More than 200 tags were differentially expressed, of which 25 mediate transcription. CONCLUSIONS: In addition to providing a detailed profile of expressed genes in the PN9 and mature mouse cornea, the present SAGE data demonstrate dynamic changes in gene expression after eye opening and provide new probes for exploring corneal epithelial cell stratification, development, and function and for exploring the intricate relationship between programmed and environmentally induced gene expression in the cornea. Keywords: other
Project description:Our preliminary study revealed that the homeobox transcription factors, Msx1 and Msx2, are expressed in the mouse uterus during early pregnancy. Further, conditional deletion of Msx1 and Msx2 in mouse uterus leads to implantation failure due to impaired uterine epithelial receptivity. To identify the downstream targets of Msx1Msx2 in the uterus, we performed gene expression profling of uterine epithelial cells isolated from Msx1Msx2-null mice and the corresponding controls on day4 of pregnancy (the time of implantation). The microarray results revealed elevated expression of mRNAs corresponding to several Wnts in uterine epithelium of Msx1Msx2-ablated mice. We performed conditional ablation of Msx1Msx2 in the mouse uterus using the PRcre mouse model. we isolated uterine epithelial cells from day4 pregnant mice (n=5 for each genotype). Total RNA was purified from these cells to hybridize to high density affymetrix microarrays.
Project description:Uterine receptivity (the window of implantation) is essential for successful implantation. The concept of uterine receptivity was first discovered 50 years ago by Psychoyos. In mice, uterine receptivity begins with the secretion of LIF from uterine glands stimulated by estrogen on the morning of day 4 pregnancies. However, some implantation failure is due to the lack of responses to the estrogen secretion. The current study reveals that uterine glands undergo a differentiation process with increases in more branches during the preimplantation period. The single cell RNA profiling of glandular cells identifies that LIF is expressed exclusively in a Prss29+ subgroup of glands in response to estrogen secretion on day 4 of pregnancy. Previous studies have shown that Foxa2 deficiency in uterine glands results in implantation failure due to lack of LIF production. Interestingly, Foxa2-deficient glands fail to develop branches and the functional Prss29+ subgroup of gland cells, indicating FOXA2 is obligatory for normal glandular differentiation prior to implantation. We found that this group develops by the evening of day 3 of pregnancy prior to estrogen secretion on day 4 mornings. We noticed that Lif mRNA signals are detected in the uterus comprising the Prss29+ subgroup. Our findings indicate that uterine glands undergo a FOXA2-dependent maturation process to acquire the competence, which we have termed the “transitional phase,” necessary to respond to estrogen. This newly identified glandular "transitional phase" represents a landmark concept in the study of uterine receptivity and implantation biology.