Asynchronous Embryo Transfer Followed by Comparative Transcriptomic Analysis of Conceptus Membranes and Endometrium Identifies Processes Important to the Establishment of Equine Pregnancy
Abstract
:1. Introduction
2. Results
2.1. RNA-seq Analysis and Gene Ontology
2.2. Gene Ontology Analysis
2.3. RNA-seq Validation by qRT-PCR
2.4. Cell Proliferation and Cell Attachment in Response to IGFBP3 Stimulation
2.5. Prostaglandin Release after Bradykinin Stimulation
3. Discussion
4. Material and Methods
4.1. Animals and Tissue Collection
4.2. Embryo Collection and Transfer
4.3. RNA Extraction, DNA Library Preparation and Sequencing
4.4. RNA-seq Analysis and Bioinformatics
4.5. Quantitative RT-PCR
4.6. Culture of Conceptus Membrane Cells
4.7. Cell Attachment Assay
4.8. Cell Proliferation Assay
4.9. Bradykinin Stimulation
4.10. Prostaglandin Assay
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
References
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Endometrium | DEGs | Upregulated | Downregulated |
---|---|---|---|
D14–Syn/Asyn | 523 | 174 | 349 |
D19–Syn/Asyn | 715 | 283 | 432 |
D14–Syn/D19–Asyn | 14 | 7 | 7 |
Conceptus | |||
D14–Syn/Asyn | 556 | 285 | 271 |
D19–Syn/Asyn | 3473 | 1870 | 1603 |
D14–Syn/D19–Asyn | 3646 | 1961 | 1685 |
Gene Name | Gene Symbol | D14 RNA-seq | D14 PCR | D19 RNA-seq | D19 PCR | ||||
---|---|---|---|---|---|---|---|---|---|
Log2 FC | P-Adjusted | Log2 FC | p-Value | Log2 FC | P-Adjusted | Log2 FC | p-Value | ||
Interleukin 6 signal transducer | IL6ST | 0.33 | 0.572 | 0.77 | 0.249 | 0.65 | <0.001 | 0.27 | 0.283 |
Insulin receptor | INSR | 0.96 | 0.009 | 0.58 | 0.291 | 0.99 | <0.001 | 0.69 | 0.065 |
Leukemia inhibitory factor (LIF) | LIF | 1.35 | 0.022 | 1.37 | 0.082 | 2.57 | <0.001 | 3.07 | 0.001 |
LIF receptor | LIFR | 0.27 | 0.419 | 1.22 | 0.148 | −0.20 | 0.639 | −0.44 | 0.879 |
Oxytocin receptor | OXTR | 1.51 | 0.007 | 0.74 | 0.226 | 0.63 | 0.071 | 0.04 | 0.711 |
Prostaglandin F receptor | PTGFR | −1.47 | <0.001 | −1.83 | 0.016 | −0.47 | 0.161 | −1.46 | 0.153 |
SLC family 1, member 1 | SLC1A1 | 0.08 | 0.962 | −0.02 | 0.919 | −1.49 | <0.001 | −1.75 | 0.002 |
SLC family 2, member 1 | SLC2A1 | 1.89 | <0.001 | 3.16 | 0.001 | 0.11 | 0.889 | −1.32 | 0.379 |
SLC family 38, member 2 | SLC38A2 | 0.36 | 0.283 | 0.09 | 0.636 | 1.10 | <0.001 | 0.75 | 0.061 |
SLC family 43, member 2 | SLC43A2 | −0.47 | 0.425 | −1.22 | 0.050 | −0.33 | 0.450 | −0.98 | 0.173 |
SLC family 5, member 1 | SLC5A1 | −0.28 | 0.752 | −0.36 | 0.492 | −0.08 | 0.935 | −0.90 | 0.369 |
Gene Name | Gene Symbol | D14 RNA-seq | D14 PCR | D19 RNA-seq | D19 PCR | ||||
---|---|---|---|---|---|---|---|---|---|
Log2 FC | P-Adjusted | Log2 FC | p-Value | Log2 FC | P-Adjusted | Log2 FC | p-Value | ||
DNA-methyltransferase 3B | DNMT3B | −0.71 | <0.001 | −0.64 | 0.048 | 0.12 | 0.613 | 0.66 | 0.018 |
H19, Imprinted maternally expressed transcript | H19 | 0.03 | 0.968 | 0.11 | 0.572 | 0.40 | 0.083 | 0.62 | 0.058 |
Insulin-like growth factor 1 | IGF1 | 0.43 | 0.302 | 2.20 | 0.007 | 1.48 | 0.027 | 1.67 | 0.082 |
Necdin | NDN | −0.17 | 0.729 | −0.53 | 0.405 | 1.42 | 0.001 | 1.57 | 0.019 |
Paternally expressed gene 10 | PEG10 | 0.32 | 0.441 | 0.45 | 0.522 | 2.70 | <0.001 | 2.89 | 0.004 |
SLC family 1, member 4 | SLC1A4 | 0.79 | 0.008 | 1.10 | 0.002 | 0.72 | 0.003 | 0.77 | 0.062 |
SLC family 1, member 5 | SLC1A5 | −1.65 | <0.001 | −2.52 | 0.004 | 2.15 | <0.001 | 3.07 | 0.029 |
SLC family 2, member 3 | SLC2A3 | 0.22 | 0.305 | 0.54 | 0.167 | −0.16 | 0.200 | −0.02 | 0.680 |
SLC family 2, member 5 | SLC2A5 | 0.49 | 0.012 | 1.55 | 0.002 | −1.20 | <0.001 | −1.42 | 0.001 |
SLC family 7, member 5 | SLC7A5 | −0.04 | 0.941 | 0.69 | 0.464 | 1.64 | <0.001 | 2.52 | 0.007 |
Small nuclear ribonucleoprotein-associated protein | SNRPN | −0.04 | 0.914 | −0.03 | 0.772 | −0.05 | 0.866 | −0.40 | 0.542 |
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Gibson, C.; de Ruijter-Villani, M.; Bauersachs, S.; Stout, T.A.E. Asynchronous Embryo Transfer Followed by Comparative Transcriptomic Analysis of Conceptus Membranes and Endometrium Identifies Processes Important to the Establishment of Equine Pregnancy. Int. J. Mol. Sci. 2020, 21, 2562. https://doi.org/10.3390/ijms21072562
Gibson C, de Ruijter-Villani M, Bauersachs S, Stout TAE. Asynchronous Embryo Transfer Followed by Comparative Transcriptomic Analysis of Conceptus Membranes and Endometrium Identifies Processes Important to the Establishment of Equine Pregnancy. International Journal of Molecular Sciences. 2020; 21(7):2562. https://doi.org/10.3390/ijms21072562
Chicago/Turabian StyleGibson, Charlotte, Marta de Ruijter-Villani, Stefan Bauersachs, and Tom A.E. Stout. 2020. "Asynchronous Embryo Transfer Followed by Comparative Transcriptomic Analysis of Conceptus Membranes and Endometrium Identifies Processes Important to the Establishment of Equine Pregnancy" International Journal of Molecular Sciences 21, no. 7: 2562. https://doi.org/10.3390/ijms21072562