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Outline
To investigate the gene expression profile of cervical exfoliated cells from woman treated by artificial cycle, and their potential association with endometrial receptivity in order to screen specific biomarkers closely related to the receptivity.
A total of 19 female patients were enrolled from those preparing for frozen embryo transfer (FET) at the Reproductive Center of First Medical Center of Chinese PLA General Hospital from February 2024 to October 2024. Under the artificial cycle frozen embryo transfer protocol, the endometrial tissues were collected on the 4th day after progesterone administration (P+4) to verify their endometrial receptivity status. Additionally, cervical exfoliated cells were collected on the 4th day (P+4) and the 6th day (P+6) after progesterone administration. RNA sequencing (RNA-Seq) was used to detect gene expression profiles. Differentially expressed genes (DEGs) were identified using the criteria of |log2fold change| >1 and a false discovery rate (FDR) <0.05, followed by bioinformatics analysis. The protein-protein interaction (PPI) network of DEGs was constructed using R software (4.4.1) and analyzed with gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) analyses. The candidate genes were identified based on the PPI network using Cytoscape software. Quantitative reverse transcription PCR (RT-qPCR) was employed to validate the target candidate genes both in vitro and in vivo.
The rsERT confirmed that all 19 women were in state of endometrial receptivity at P+6. RNA-Seq identified 3 458 DEGs in cervical exfoliated cells between P+4 and P+6. The up-regulated DEGs were mainly enriched in the pathways associated with immune response and cell differentiation, and the down-regulated ones were mainly enriched in the pathways associated with lipid metabolism and cell proliferation. Using maximal clique Centrality (MCC) algorithm in the PPI network, the top 20 genes were selected. Among them, 6 genes, such as IFIT2, OASL, MX1，RSAD2，IFIT1 and IFIT3, tied for the first place, and the 6 genes all belong to interferon-stimulated genes (ISGs). qRT-PCR indicated that the above 6 genes showed significantly higher expression levels in the cervical exfoliated cells at the P+4 stage than the cells at the P+6 stage (P<0.05).
There are changes in the expression levels of the genes related to immunity and cytoskeleton remodeling in cervical exfoliated cells during the endometrial receptivity phase. The decrease in the expression of ISGs may serve as a potential biomarker for endometrial receptivity.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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