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pmid:35147192
Uterine fluid microRNAs are dysregulated in women with recurrent implantation failure.
Carolina von Grothusen · Caroline Frisendahl · Vijayachitra Modhukur · Parameswaran Grace Lalitkumar · Maire Peters · Omid R Faridani · Andres Salumets · Nageswara Rao Boggavarapu · Kristina Gemzell-Danielsson
Human reproduction (Oxford, England) · 2022 doi:10.1093/humrep/deac019
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Experiment
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Complete file
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Endometrial miRNAs are dysregulated in women with RIF.\nSTUDY DESIGN, SIZE, DURATION: In this descriptive laboratory case-control study, miRNA abundancy was compared between UF collected during implantation phase from healthy fertile women (n = 17) and women with RIF (n = 34), which was defined as three failed IVF cycles with high-quality embryos.\nPARTICIPANTS/MATERIALS, SETTING, METHODS: Recruitment of study subjects and sampling of UF were performed at two university clinics in Stockholm, Sweden and Tartu, Estonia. The study participants monitored their menstrual cycles using an LH test kit. The UF samples were collected on Day LH + 7-9 by flushing with saline. Samples were processed for small RNA sequencing and mapped for miRNAs. The differential abundance of miRNAs in UF was compared between the two groups using differential expression analysis (DESeq2). Further downstream analyses, including miRNA target gene prediction (miRTarBase), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis (g:Profiler) and external validation using relevant published data, were performed on the dysregulated miRNAs. Two miRNAs were technically validated with quantitative real-time PCR (RT-PCR).\nMAIN RESULTS AND THE ROLE OF CHANCE: After processing of the sequencing data, there were 15 samples in the healthy fertile group and 33 samples in the RIF group. We found 61 differentially abundant UF miRNAs (34 upregulated and 27 downregulated) in RIF compared to healthy women with a false discovery rate of <0.05 and a fold change (FC) of ≤-2 or ≥2. When analyzed with published literature, we found that several of the differentially abundant miRNAs are expressed in endometrial epithelial cells and have been reported in endometrial extracellular vesicles and in association with endometrial receptivity and RIF. Their predicted target genes were further expressed both in the trophectodermal cells of blastocyst-stage embryos and endometrial mid-secretory epithelial cells, as assessed by publicly available single-cell transcriptome-sequencing studies. Pathway analysis further revealed that 25 pathways, having key roles in endometrial receptivity and implantation, were significantly enriched. Hsa-miR-486-5p (FC -20.32; P-value = 0.004) and hsa-miR-92b-3p (FC -9.72; P-value = 0.004) were successfully technically validated with RT-PCR.\nLARGE SCALE DATA: The data are available in Gene Expression Omnibus (GEO) at https://www.ncbi.nlm.nih.gov/geo/ with GEO accession number: GSE173289.\nLIMITATIONS, REASONS FOR CAUTION: This is a descriptive study with a limited number of study participants. Moreover, the identified differentially abundant miRNAs should be validated in a larger study cohort, and the predicted miRNA target genes and enriched pathways in RIF need to be confirmed and further explored in vitro.\nWIDER IMPLICATIONS OF THE FINDINGS: RIF is a major challenge in the current IVF setting with no diagnostic markers nor effective treatment options at hand. For the first time, total miRNAs have been extensively mapped in receptive phase UF of both healthy women with proven fertility and women diagnosed with RIF. Our observations shed further light on the molecular mechanisms behind RIF, with possible implications in future biomarker and clinical treatment studies.\nSTUDY FUNDING/COMPETING INTEREST(S): This work was financially supported by the Swedish Research Council (2017-00932), a joint grant from Region Stockholm and Karolinska Institutet (ALF Medicine 2020, FoUI-954072), Estonian Research Council (PRG1076), Horizon 2020 innovation (ERIN, EU952516) and European Commission and Enterprise Estonia (EU48695). The authors have no competing interests to declare for the current study.", "claims": [{"id": "claim-method", "statement": "The differential abundance of miRNAs in UF was compared between the two groups using differential expression analysis (DESeq2).", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:35147192", "locator": "sections[0] \"Abstract\" paragraph 5", "quote": "The differential abundance of miRNAs in UF was compared between the two groups using differential expression analysis (DESeq2)."}], "state": "complete"}, {"id": "claim-data-access", "statement": "The data underlying this article are available in Gene Expression Omnibus (GEO) at https://www.ncbi.nlm.nih.gov/geo/ and can be accessed with GEO accession number: GSE173289.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:35147192", "locator": "sections[26] \"Data availability\" paragraph 2", "quote": "The data underlying this article are available in Gene Expression Omnibus (GEO) at https://www.ncbi.nlm.nih.gov/geo/ and can be accessed with GEO accession number: GSE173289."}], "state": "complete"}, {"id": "claim-target-result", "statement": "We found 61 differentially abundant UF miRNAs (34 upregulated and 27 downregulated) in RIF compared to healthy women with a false discovery rate of <0.05 and a fold change (FC) of ≤−2 or ≥2.", "kind": "qualitative", "target": null, "evidence": [{"asset": "source:pmid:35147192", "locator": "sections[0] \"Abstract\" paragraph 6", "quote": "We found 61 differentially abundant UF miRNAs (34 upregulated and 27 downregulated) in RIF compared to healthy women with a false discovery rate of <0.05 and a fold change (FC) of ≤−2 or ≥2."}], "state": "complete"}], "gaps": []}, "experiment": {"state": "complete", "objective": "We found 61 differentially abundant UF miRNAs (34 upregulated and 27 downregulated) in RIF compared to healthy women with a false discovery rate of <0.05 and a fold change (FC) of ≤−2 or ≥2.", "steps": [{"id": "read_published_estimate", "order": 1, "name": "Read the published estimate from the source", "kind": "other", "state": "complete", "blocked_on": null, "description": "The claim, method and evidence span published with the source article.", "duration_hours": null, "parameters": {}, "inputs": ["source:pmid:35147192"], "outputs": ["the published estimate and its evidence"], "instruments": [], "reagents": []}, {"id": "obtain_source_data", "order": 2, "name": "Obtain the source data from the named repository", "kind": "data-preparation", "state": "complete", "blocked_on": null, "description": "The GEO series accession resolves and its sample accessions are listed. 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