115 In Vivo Transcriptomic Response of Bovine Oviduct Epithelial Cells to the Early Embryo

2018 ◽  
Vol 30 (1) ◽  
pp. 197
Author(s):  
B. Rodríguez-Alonso ◽  
M. Hamdi ◽  
J. M. Sánchez ◽  
A. Gutierrez-Adán ◽  
P. Lonergan ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Housekeeping Genes ◽  
Local Effect ◽  
Early Embryo ◽  
Cell Stage ◽  
Transcriptomic Response ◽  
Precise Position ◽  
Horizon 2020 ◽  
Proximal Site

The presence of a single 8-cell embryo does not alter the transcriptome of the cells of the oviducal isthmus, although a local effect at the precise position of the embryo cannot be ruled out. Thus, we aimed to study the local embryo effect on the transcriptomic response of the epithelial cells of the oviduct in vivo. Fifteen heifers were synchronized and all showed standing heat and were artificially inseminated. All heifers were slaughtered on Day 2.5 after oestrus. The oviducts from 13 animals (with a corpus luteum, CL) were isolated, trimmed free of tissue and divided between ampulla and isthmus. The ipsilateral isthmus was then divided into smaller sections (2 cm). Each section was sequentially flushed until the embryo was located and was then opened and scraped longitudinally to obtain the epithelial cells. Cells were snap-frozen in LN2 for gene expression analysis. All recovered embryos were found at the beginning of the isthmus of the oviduct ipsilateral to the CL. Three at 2-cell stage and 1 at 8-cell stage. The recovery rate was 30.8% (4/13) and only samples from these 4 animals were used for analysis. The 2-cm sections selected for the transcriptomic analysis were embryo section (ES), in which the embryo was found; proximal section (PS), through which the embryo had passed; distal section (DS), on the uterine side of the embryo; and contralateral section (CS), section from the contralateral isthmus. The expression pattern of 10 differentially expressed genes between the isthmus of pregnant and cyclic heifers (Maillo et al. 2015 Biol Reprod. 92, 144) were assessed by RTq-PCR relative to 2 housekeeping genes, H2A.Z and ACTG. Five up-regulated genes (STK32A, SLC26A3, KERA, QRFPR, MCTP1) and 5 down-regulated (SOD3, PRELP, VAT1L, SOCS3, CCL20) were analysed. One-way ANOVA and t-test was used for statistical analysis. Comparison between ES and the CS revealed one significantly altered gene (VAT1L). This is in agreement with our in vivo results in which VAT1L was also down-regulated in the presence of embryos. Comparison within the ipsilateral oviduct of ES and PS samples revealed STK32A, SLC26A3, QRFPR, MCTP1, and SOCS3 transcripts significantly down-regulated compared with DS samples, whereas the expression for CCL20 was different between ES and DS but similar to the PS. In conclusion, the fact that 5 out of 10 transcripts were different between the segment where the embryo was collected and other locations in the oviduct suggests the presence of embryo site-specific signal. However, comparison between the ipsilateral embryo site with the contralateral site revealed that only one transcript was different. Moreover, the similarities in the ipsilateral oviduct between embryo and proximal site may be due to the passage of the embryo. Furthermore, the location of the embryo close to the ampullary-isthmic junction may mask the effect due to the spatial differences of the bovine oviduct. Research supported by EU, Horizon 2020 Marie Sklodowska-Curie, REPBIOTECH 675526; Spanish MINECO AGL2015-70140-R & AGL2015-66145-R; OECD-CoOperative Prog TAD/CRP JA00092482.

scholarly journals Gene expression and metabolic response of bovine oviduct epithelial cells to the early embryo

Reproduction ◽  
2019 ◽  
Vol 158 (1) ◽  
pp. 85-94 ◽  
Author(s):  
Meriem Hamdi ◽  
María J Sánchez-Calabuig ◽  
Beatriz Rodríguez-Alonso ◽  
Sandra Bagés Arnal ◽  
Kalliopi Roussi ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Metabolic Response ◽  
Early Embryo ◽  
Bovine Embryo ◽  
Cell Stage ◽  
Transcriptomic Response ◽  
Early Embryos ◽  
Oviduct Epithelial Cells

During its journey through the oviduct, the bovine embryo may induce transcriptomic and metabolic responses, via direct or indirect contact, from bovine oviduct epithelial cells (BOECs). An in vitro model using polyester mesh was established, allowing the study of the local contact during 48 h between a BOEC monolayer and early embryos (2- or 8-cell stage) or their respective conditioned media (CM). The transcriptomic response of BOEC to early embryos was assessed by analyzing the transcript abundance of SMAD6, TDGF1, ROCK1, ROCK2, SOCS3, PRELP and AGR3 selected from previous in vivo studies and GPX4, NFE2L2, SCN9A, EPSTI1 and IGFBP3 selected from in vitro studies. Moreover, metabolic analyses were performed on the media obtained from the co-culture. Results revealed that presence of early embryos or their CM altered the BOEC expression of NFE2L2, GPX4, SMAD6, IGFBP3, ROCK2 and SCN9A. However, the response of BOEC to two-cell embryos or their CM was different from that observed to eight-cell embryos or their CM. Analysis of energy substrates and amino acids revealed that BOEC metabolism was not affected by the presence of early embryos or by their CM. Interestingly, embryo metabolism before embryo genome activation (EGA) seems to be independent of exogenous sources of energy. In conclusion, this study confirms that early embryos affect BOEC transcriptome and BOEC response was embryo stage specific. Moreover, embryo affects BOEC via a direct contact or via its secretions. However transcriptomic response of BOEC to the embryo did not manifest as an observable metabolic response.

scholarly journals Identification of a Novel Role for Endothelins within the Oviduct

Endocrinology ◽  
2010 ◽  
Vol 151 (6) ◽  
pp. 2858-2867 ◽  
Author(s):  
Myoungkun Jeoung ◽  
Sungeun Lee ◽  
Hee-kyung Hawng ◽  
Yong-Pil Cheon ◽  
Youn Kyung Jeong ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Embryo Development ◽  
Early Embryo ◽  
Receptor Subtypes ◽  
Early Embryo Development ◽  
Vitro Fertilization ◽  
Protein Superfamilies ◽  
Endothelin 3

Endothelins were first identified as potent vasoactive peptides; however, diversity in the biological function of these hormones is now evident. We have identified a novel role for endothelins: a requirement for these peptides within the oviduct during fertilization and/or early embryo development. In vivo, treatment after ovulation with a dual endothelin receptor antagonist (tezosentan) decreased the number of two-cell embryos that could be collected from within the oviducts. In vitro fertilization experiments showed that gamete viability and their ability to fertilize were not affected by treatment with this antagonist, suggesting that the effect observed in vivo was mediated by the oviduct itself. Expression of mRNA for all three isoforms of the endothelins and both receptor subtypes was detectable within the oviduct. Expression of mRNA for endothelin-3 was regulated by gonadotropins in epithelial cells of the oviduct and increased specifically within the isthmus of this structure. Immunostaining revealed localization of both endothelin receptors A and B to the columnar epithelial cells within the oviduct, suggestive of a local role for endothelins in the regulation of epithelial function and ultimately oviductal secretions. A microarray analysis revealed three likely endothelin-regulated protein networks for future analysis: the TGFβ, IL-10, and CCAAT/enhancer-binding protein superfamilies. Overall, these results suggest a novel and requisite role for endothelins within the oviduct during fertilization and/or early embryo development.

Epithelial differentiation and intercellular junction formation in the mouse early embryo

Development ◽  
1992 ◽  
Vol 116 (Supplement) ◽  
pp. 105-112
Author(s):  
Tom P. Fleming ◽  
Qamar Javed ◽  
Mark Hay
Keyword(s):  
Tight Junction ◽  
Intercellular Junctions ◽  
Early Embryo ◽  
Control Mechanisms ◽  
Tight Junction Proteins ◽  
Cell Stage ◽  
Tight Junction Formation ◽  
Junction Formation ◽  
Junction Proteins

Trophectoderm differentiation during blastocyst formation provides a model for investigating how an epithelium develops in vivo. This paper briefly reviews our current understanding of the stages of differentiation and possible control mechanisms. The maturation of structural intercellular junctions is considered in more detail. Tight junction formation, essential for blastocoele cavitation and vectorial transport activity, begins at compaction (8-cell stage) and appears complete before fluid accumulation begins a day later (approx 32-cell stage). During this period, initial focal junction sites gradually extend laterally to become zonular and acquire the peripheral tight junction proteins ZO-1 and cingulin. Our studies indicate that junction components assemble in a temporal sequence with ZO-1 assembly preceding that of cingulin, suggesting that the junction forms progressively and in the ‘membrane to cytoplasm’ direction. The protein expression characteristics of ZO-1 and cingulin support this model. In contrast to ZO-1, cingulin expression is also detectable during oogenesis where the protein is localised in the cytocortex and in adjacent cumulus cells. However, maternal cingulin is metabolically unstable and does not appear to contribute to later tight junction formation in trophectoderm. Cell-cell interactions are important regulators of the level of synthesis and state of assembly of tight junction proteins, and also control the tissue-specificity of expression. In contrast to the progressive nature of tight junction formation, nascent desmosomes (formed from cavitation) appear mature in terms of their substructure and composition. The rapidity of desmosome assembly appears to be controlled by the time of expression of their transmembrane glycoprotein constitutents; this occurs later than the expression of more cytoplasmic desmosome components and intermediate filaments which would therefore be available for assembly to occur to completion.

Gene expression, oocyte nuclear maturation and developmental competence of bovine oocytes and embryos produced after in vivo and in vitro heat shock

Zygote ◽  
2016 ◽  
Vol 24 (5) ◽  
pp. 748-759 ◽  
Author(s):  
Krishna C. Pavani ◽  
Erica Baron ◽  
Pedro Correia ◽  
Joana Lourenço ◽  
Bruno Filipe Bettencourt ◽  
...  
Keyword(s):  
Gene Expression ◽  
Heat Shock ◽  
Embryo Development ◽  
Target Genes ◽  
Housekeeping Genes ◽  
Early Embryo ◽  
Developmental Competence ◽  
Nuclear Maturation

SummaryThree assays were performed. In assay 1, oocytes harvested during the winter months were subjected to kinetic heat shock by stressing the oocytes at 39.5°C (HS1) or at 40.5°C (HS2) for either 6, 12, 18 or 24 h and then matured at control temperature (38.5°C). The nuclear maturation rates (NMR) of all oocytes were recorded after 24 h. In assay 2, oocytes collected year-round maturated, were implanted via in vitro fertilization (IVF) and developed for 9 days. Gene expression analysis was performed on target genes (Cx43, CDH1, DNMT1, HSPA14) with reference to the two housekeeping genes (GAPDH and SDHA) in embryos. Similarly, in assay 3, genetic analysis was performed on the embryos produced from heat-stressed oocytes (from HS1 and HS2). In assay 1, the duration of heat stress resulted in a significant decline in NMR (P < 0.05) with HS1 for maturated oocytes at 86.4 ± 4.3; 65.5 ± 0.7; 51.3 ± 0.9; 38.1 ± 1.9 and 36.3 ± 0.9, for control, 6 h, 12 h, 18 h and 24 h, respectively. For assays 2 and 3, results demonstrated that DNMT1, Cx43 and HSPA14 were down-regulated in the embryos produced in the warm with respect to the cold months (P < 0.05). A constant up- and down-regulation of DNMT1 and HSPA14 genes were observed in both HS-treated samples. Also, an inconsistent pattern of gene expression was observed in Cx43 and CDH1 genes (P < 0.05). Targeted gene expression was aberrant in embryo development, which can provide evidence on early embryo arrest and slowed embryo development.

scholarly journals Derivation of Thyroid Follicular Cells From Pluripotent Stem Cells: Insights From Development and Implications for Regenerative Medicine

2021 ◽  
Vol 12 ◽  
Author(s):  
Alberto Posabella ◽  
Andrea B. Alber ◽  
Hendrik J. Undeutsch ◽  
Raoul A. Droeser ◽  
Anthony N. Hollenberg ◽  
...  
Keyword(s):  
Stem Cells ◽  
Epithelial Cells ◽  
Pluripotent Stem Cells ◽  
Early Embryo ◽  
Great Promise ◽  
Normal Humans ◽  
Or Gene ◽  
Follicular Epithelial

Stem cell-based therapies to reconstitute in vivo organ function hold great promise for future clinical applications to a variety of diseases. Hypothyroidism resulting from congenital lack of functional thyrocytes, surgical tissue removal, or gland ablation, represents a particularly attractive endocrine disease target that may be conceivably cured by transplantation of long-lived functional thyroid progenitors or mature follicular epithelial cells, provided a source of autologous cells can be generated and a variety of technical and biological challenges can be surmounted. Here we review the emerging literature indicating that thyroid follicular epithelial cells can now be engineered in vitro from the pluripotent stem cells (PSCs) of mice, normal humans, or patients with congenital hypothyroidism. We review the in vivo embryonic development of the thyroid gland and explain how emerging discoveries in developmental biology have been utilized as a roadmap for driving PSCs, which resemble cells of the early embryo, into mature functional thyroid follicles in vitro. Finally, we discuss the bioengineering, biological, and clinical hurdles that now need to be addressed if the goals of life-long cure of hypothyroidism through cell- and/or gene-based therapies are to be attained.

Measurement of the Platelet Response to Laser- induced Microvascular Injury

1974 ◽  
Vol 32 (02/03) ◽  
pp. 704-713 ◽  
Author(s):  
F. N McKenzie ◽  
K.-E Arfors ◽  
N. A Matheson
Keyword(s):  
Inhibitory Effect ◽  
Microvascular Blood Flow ◽  
Platelet Response ◽  
High Concentration ◽  
Platelet Activity ◽  
Microvascular Injury ◽  
Arterial Blood ◽  
Proximal Site ◽  
Adenosine Phosphates

SummaryA study has been made of the biochemical factors underlying the platelet response to laser-induced microvascular injury. A platelet aggregating substance is produced at sites of laser-induced injury which markedly stimulates platelet activity at a site of injury inflicted a short distance downstream. Distal sites of injury are not similarly influenced if the distance between the injuries is increased or if the proximal site no longer shows platelet-stimulating activity. The stimulating effect of an adjacent proximal injury on platelet activity at a distal site is inhibited by local intra-arterial infusion of adenosine. Measurements of arterial blood pressure and microvascular blood flow velocity during adenosine infusion showed that its inhibitory effect on platelet activity is largely independent of its vasodilator properties. The effect of infusion of different adenosine phosphates (AMP, ADP, ATP) was also studied. Very small amounts of ADP markedly stimulated platelet activity and the emboli formed were similar to those normally produced at sites of laser injury. At high concentration AMP inhibited while ATP stimulated platelet activity in vivo. The results emphasise the fundamental role of ADP as a mediator of the platelet response at sites of laser- induced microvascular injury.

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