ALTERATIONS IN ENDOMETRIAL PROGRAMMING LEAD TO RECURRENT IN VITRO FERTILIZATION FAILURES

MARCELA IRIGOYEN; LARA CASTAGNOLA; SOLEDAD GORI; ESTEBAN GRASSO; DIEGO GNOCCHI; ANTONIO CATTANEO; CLAUDIA PÉREZ LEIRÓS; ROSANNA RAMHORST

Copenhagen

Congreso; ESHRE 39th Annual Meeting; 2023

ESRHE

Study question:Could patients with recurrent implantation failures (RIF) display alterations on thedecidualization program conditioning decidual cells’ functions and the inflammatoryresponse during peri-implantation period?Summary answer:Endometrium from RIF-patients displays alterations into the decidualization programand into the induction of a physiological inflammation preventing the attachment andcell-migration for embryo implantation.What is known already:The decidualization program starts on each menstrual cycle and implies not onlyphenotypical changes on the endometrial stromal cells, but also in their secretoryprofile. Moreover, this process induces a physiological and sterile inflammatoryresponse associated with inflammatory mediators production, such as IL-1β. At thesame time, several molecules associated with receptivity are modulated; increasingthe expression of adhesion molecules and metalloproteinases, while decreasing MUC-1 to allow embryo attachment and implantation. Even nowadays with differenttechniques that can identify a competent embryo and the implantation window, RIF-patients cannot reach implantation and the mechanisms involved have not beenelucidated yet.Study design, size, duration:First, a bioinformatic analysis was performed using standardized pathways involved inangiogenesis, placentation, decidualization, inflammation and immune regulation frompublic databases (Reactome, Gene Ontology Biological Process, WikiPathways, KEGG).Based on arrays that compare gene expression in endometrial biopsies from RIF orfertile women, we selected those that connect two or more processes and wevalidated their modulation in endometrial samples. Functional processes, such asmigration and inflammatory production were also evaluated in primary cultures.Participants/materials, setting, methods:Endometrial samples were obtained from fertile and RIF patients during the secretoryphase. The Investigation and Ethics Committee from San Isidro, Argentina approvedthis study. RT-qPCR was performed to test decidualization markers: IGFBP1, PRL, PGR;inflammation: IL-1β, NLRP3, PTGS1; angiogenesis balance: CXCL14, ARG2, VEGFA;adhesion molecules: ITGA8 and MUC1. IL-1β production was also quantified by FlowCytometry. Wound healing assay was performed in human endometrial stromal cellsderived from cell line and RIF patient’s biopsy.Main results and the role of chance:First, we performed bioinformatic analysis based on standardized pathways in publicdatabases. We focused on genes that connected and were modulated in processesinvolved in implantation, angiogenesis, placentation, decidualization, inflammationand immune regulation. Then, we validated the expression of 15 genes with thehighest score in biopsies from RIF or fertile women taken on LH+7. We founddecreased IGFBP1 expression, an early decidualization marker, and the progesteronereceptor (p<0.0001), suggesting alterations in the decidualization program. Sinceinflammation is associated with decidualization, we evaluated the IL-1β pathway. RIFbiopsies had reduced expression of NLRP3, associated with inflammasome assembly,accompanied by a reduction in PTGS1 and IL-1β production in comparison withendometrial samples from fertile women. Moreover, when we evaluated genesinvolved in processes regulated by inflammation, we observed increased expression ofMUC1, avoiding blastocyst adhesion, and decreased ITGA8 preventing its attachmentaccompanied by decreased MMP9 levels (p<0.005). Also, we observed an imbalancebetween pro and anti-angiogenic factors (VEGFA vs CXCL14). Finally, we isolated andcultivated stromal cells from RIF samples to analyze migration ability, a process thatmediates blastocyst inclusion into the decidua, by wound healing assay. These primarycultures showed a differential migration pattern compared with decidualized stromalcells.Limitations, reasons for caution:The present results were obtained using endometrial samples from RIF patients andfertile women obtained during the secretory phase of the menstrual cycle. Eventhough the samples represent the endometrium after in vivo decidualization, furtherstudies are necessary to elucidate whether these mechanisms operate similarly in vivo.Wider implications of the findings:Decidualization process induces a physiological and sterile inflammatory responseassociated with the modulation of several adhesion molecules and MMP to allow theattachment and embryo implantation. However, this initial inflammatory response isdifferentially induced in RIF patients in comparison with fertile women, suggesting itsrelevance as a potential pharmacological target.Study funding and competing interest(s):This work was funded by the National Agency of Sciences and Technology ANPCyT(PICT 2018-4715 and PICT 2028-2461 to RR) and University of Buenos Aires (UBACyTUBACyT 20020090200034 to RR). The authors have no conflicts of interest to disclose.