Transcriptome landscape of pig embryo implantation site at single-cell resolution.
Ontology highlight
ABSTRACT:
ORGANISM(S): Sus scrofa
PROVIDER: GSE198698 | GEO | 2022/03/18
REPOSITORIES: GEO
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Project description:A novel porcine cDNA microarray, containing PCR products selected for embryonic expression, was used for transcriptional profiling during conceptus development. Total RNA from quadruplicate samples of small spherical (1-5mm), large spherical (6-11mm), tubular (13-45mm), and filamentous (>100mm) was extracted, amplified, and converted to 32P labeled target and hybridized to membranes. Porcine peri-implantation embryo development order is from small spherical to large spherical, then to Tubular, then to filamentous. All this morphological transmission is within two days (day 10 to day 12). In this study, we investigated the gene expression changes by a specifically selected porcine embryo gene array. A number of genes were found to be differentially expressed during those developmental stages and confirmed by quantitative PCR. Keywords = Gene expression Keywords = porcine conceptus Keywords = cDNA microarray Keywords = peri-implantation Keywords = transcriptional profiling Keywords: time-course
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Project description:Endometrial receptivity plays a vital role in the success of embryo implantation. However, the temporal proteomic profile of porcine endometrium during embryo implantation is still unclear. In this study, the expression of proteins in endometrium on days 9, 10, 11, 12, 13, 14, 15 and 18 of pregnancy (D9, 10, 11, 12, 13, 14, 15 and 18) was profiled via iTRAQ technology. The results showed that 25, 55, 103, 91, 100, 120, 149 proteins were up-regulated, and 24, 70, 169, 159, 164, 161, 198 proteins were down-regulated in porcine endometrium on D10, 11, 12, 13, 14, 15 and 18 compared with that on D9, respectively. Among these differentially expressed (DE) proteins, MQM results indicated that S100A9, S100A12, HRG and IFI6 were differentially expressed in endometrial tissues during embryo implantation period. Bioinformatics analysis showed that the proteins differentially expressed in the 8 comparisons (D10 vs D9, D11 vs D9, D12 vs D9, D13 vs D9, D14 vs D9, D15 vs D9 and D18 vs D9) were involved in important processes and pathways related to immunization, endometrial remodeling, which have a vital effect on embryonic implantation. Our results reveal that RBP4 could regulate the cell proliferation, migration and apoptosis of endometrial epithelial cells and endometrial stromal cells to affect embryo implantation. This research also provides resources for studies of proteins in endometrium during early pregnancy.
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Project description:In this study, deep sequencing of endometrial RNA samples during early pregnancy were performed using RNA sequencing (RNA-Seq). The changes of genes expressed among three stages (day 12, 18 and 25) of pregnancy were investigated by the multiple comparisons. Analysis of reads, between 1557 and 8951 differentially expressed genes (DEGs) were revealed among different periods of implantation. Expression of selected genes was validated by the use of quantitative real-time RT-PCR. Bioinformatics analysis of differentially expressed genes in the endometrium revealed a number of biological processes and pathways potentially involved in embryo implantation in the pig, most noticeably cell proliferation, regulation of immune response, interaction of cytokine-cytokine receptor, and cell adhesion. This revealed specific gene expression patterns reflecting the different functions of the endometrium on three stages (maternal recognition, conceptus attachment and embryo implantation). This study identified comprehensive transcriptomic profile in the porcine endometrium, it could be a foundation for targeted studies of genes and pathways potentially involved in abnormal endometrial receptivity and embryo loss in early pregnancy.
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Project description:Litter size is one of the most economically important traits in commercial pig farming. It has been estimated that approximately 30% of porcine embryos are lost during the peri-implantation period. Despite rapid advances over recent years, the molecular mechanism underlying embryo implantation in pigs remains poorly understood. In this study, the conceptus together with a small amount of its surrounding endometrial tissues at the implantation site was collected and subjected to single-cell RNA-seq using the 10x platform. Because embryo and maternal endometrium were genetically different, we successfully dissected embryonic cells from maternal endometrial cells in the data according to single nucleotide polymorphism information captured by single-cell RNA-seq. Undoubtedly, the interaction between trophoblast cells and uterine epithelial cells represents the key mechanism of embryo implantation. Using the CellChat tool, we revealed cell-cell communications between these 2 cell types in terms of secreted signaling, ECM-receptor interaction and cell-cell contact. Additionally, by analyzing the non-pregnant endometrium as control, we were able to identify global gene expression changes associated with embryo implantation in each cell type. Our data provide a valuable resource for deciphering the molecular mechanism of embryo implantation in pigs.
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Project description:In vivo evidence suggests a dynamic change of microRNA expression with the phase of the endometrial cycle to achieve endometrial receptivity. In vivo animal studies outlined post – implantation changes in microRNA profiles. Little information is available on the role of microRNAs in the early stages of the embryonic – endometrial dialogue prior to established implantation .Our aim is to investigate the possible role of microRNAs the early endometrial response to the developing pre- attachment blastocyst prior to established implantation
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Project description:Chemerin (CHEM) is a hormone mainly expressed in adipocytes involved in the regulation of energy homeostasis and inflammatory response. CHEM expression has been demonstrated in the structures of the porcine hypothalamic-pituitary-gonadal axis, as well as in the uterus, trophoblasts and conceptuses of pigs. In this study, we performed high-throughput proteomic analyses (Liquid Chromatography with Tandem Mass Spectrometry, LC-MS/MS) to examine the influence of CHEM (400 ng/ml) on differentially regulated proteins (DRPs) in the porcine endometrial tissue explants during implantation (15 to 16 days of gestation). Among all 352 DRPs, 164 were up-regulated and 188 were down-regulated in CHEM-treated group. DRPs were assigned to 47 gene ontology (GO) terms (p-adjusted < 0.05). Validation of four DRPs (IFIT5, TGFβ1, ACO1 and PGRMC1) by Western blot analysis confirmed the veracity and accuracy of LC-MS/MS method used in the present study. We suggest that CHEM, by modulating various protein expressions takes part in the endometrial cell proliferation, migration and invasion at the time of implantation. It also regulates the endometrial immune response, sensitivity to P4 and formation of new blood vessels. Additionally, CHEM appears to be an important factor involved in endothelial cell dysfunction during the pathogenesis of preeclampsia. The identification of a large number of DRPs under the influence of CHEM provides a valuable resource for understanding the molecular mechanisms of this hormone during implantation, which is a prerequisite for better control of pig reproduction.
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Project description:The female reproductive tract provides a unique environment for a successful pregnancy. It is imperative that the non-pregnant uterus be transformed into a capable environment for the establishment and maintenance of pregnancy. To achieve this, intimate cross-communication between the endometrium and the embryo is necessary at an early stage of life (Almiñana et al., 2012) a dialog which further influences subsequent fetal developmental potential (Fleming et al., 2004) and even post-natal performance. Initially, the trophoblast-derived estrogen is one of the most important embryonic signals to activate the maternal uterus for attachment (Pope & First 1985). Once the conceptus has established its position in the uterus (>d15), its development and growth also require many other maternal-embryonic cellular and molecular interactions to ensure substantial vascular changes in the endometrium and the chorioallantois, mainly development of capillaries under the lamina propriae, essential to provide full function to the pig epiteliochorial placenta required for a successful pregnancy. It is during the peri-attachment window between days 12-30 of gestation when most (20-30%) of the embryos produced in pig natural or artificial breeding die (reviewed by Edwards et al., 2012); embryonic death with a substantial impact on pig production efficiency, especially because it significantly limits litter size. Different studies have focused on the elucidation of the complex embryo-maternal communication network to reduce pregnancy loss. Among these, several studies examined gene expression during the peri-implantation stage, when the majority of embryonic losses occur, or have compared the transcriptomic profiles of pregnant and non- pregnant animals. Interestingly, these studies identified changes in the expression of genes that can -directly or indirectly- contribute to reproductive success such genes related to cell proliferation, hormone synthesis and metabolism, cell adhesion or those related to cytokine production and immune local response (Almiñana et al., 2012, Lin et al, 2015; Smolinska et al., 2019; Yang et al., 2020, Martinez et al., 2020, 2022). However, we should not forget that changes in gene expression do not always lead to a corresponding alteration in the expression of proteins, which are crucial components in all biological processes. The knowledge of the proteome is therefore equally relevant to gene expression changes, being able to help detecting both normal and abnormal physiological conditions. Proteome characterization can led to a better understanding of physiological processes and to identify proteins that may serve as potential biomarkers. To date, only a limited number of studies have unfortunately explored the protein expression profiles of the pig endometrium during the crucial period of maternal recognition of pregnancy, which occurs between day 9 and 13 (Jalali et al., 2015; Kaiser et al., 2006) or during mid/last-gestation, from 40 to 93 days of pregnancy (Chae et al., 2011; Wang et al., 2019). These studies have identified several proteins associated with endometrial function, which could play a role in maternal recognition and progression of pregnancy. Therefore, in order to fully understand the molecular interaction between the conceptus and its chorion with the endometrium, we have aimed to characterize the proteome profile of both, the non-pregnant (control) endometrium compared with that of the “pregnant”endometrium with present extraembryonic membranes between the 3rd and the 4th week period, when the major conceptus loss occurs.
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Project description:Asynchrony of trophectoderm elongation, gestational days 11-12, is evident in porcine concepti and, rapid progression through this phase has been associated with conceptus competency. The goal of the current study was to determine the extent of transcriptomic responses of concepti to developmental delay and their physiological implications. Concepti with the same morphology, ovoid and 7-8 mm, were isolated and designated as normal or developmentally delayed if collected from a homogenous ovoid conceptus population or a heterogeneous conceptus population (ovoid to filamentous), respectively. Total RNA prepared from 4 distinct normal and 4 distinct developmentally delayed concepti, was analyzed using an Agilent high-density custom porcine microarray. Two hundred-nine transcripts were found differentially expressed between normal and developmentally delayed concepti. Functional analysis of these genes indicated that a significant number of the genes regulate signal transduction/transcription, organismal development, metabolism, and cell adhesion and can be modulated by transforming growth factor β1. Ten genes were selected for real-time PCR validation of differential expression based on a known role in steroid synthesis, endometrium receptivity, and modulation of trophoblast differentiation/growth or interaction with transforming growth factor β1. As in the microarray, all except one, achaete-scute complex homolog 2, were preferentially up-regulated in delayed concepti. Overall, the findings suggested that developmentally delayed concepti are distinct from their morphologically similar counterparts and more developmentally advanced concepti. Furthermore, it also highlighted ways by which the conceptus’ microenvironment might be affected and pinpointed developmental factors that can be interrogated further to determine if, and how, they affect embryo competency/elongation.
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Project description:The bovine conceptus elongates near Day 16 of development and releases interferon-tau (IFNT), disrupting the endometrial luteolytic mechanism to sustain luteal P4 and pregnancy. Conceptus factors other than IFNT modify local endometrial activities in support of pregnancy; however, the microenvironment is largely uncharacterized. We utilized a bovine conceptus-endometrial culture system to elucidate the microenvironment in the form of RNA and protein. Estrus synchronized heifers remained cyclic (13) or were inseminated (9) to produce Day 16 cyclic endometrium and elongating conceptuses, respectively. Conceptus sections and cyclic endometrium were then used to generate tissue cultures in 1 mL of medium: (1) no tissue (Control Med; n=7), (2) mono-cultured conceptus (Conceptus; n=9), (3) mono-cultured endometrium (Endo; n=13), or (4) Endo-Conceptus co-culture (n=15). After 12 h, tissue RNA was sequenced (RNA-Seq) and media underwent proteomic analysis (LC-MS/MS). Compared to Conceptus and Endo, co-cultured conceptus and endometrial tissue contained 3,400 and 4,575 differentially expressed genes (DEG), respectively (P ≤ 0.01). Upregulated endometrial DEG, independent of IFNT, were associated with transcription whereas upregulated conceptus DEG were associated with protein homeostasis and metabolism (P ≤ 0.001).
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