Study of the molecular mechanisms involved in the development of adenomyosis and related infertility using endometrial organoids as a preclinical model

Adenomyosis, a prevalent uterine disease affecting approximately 30% of women of reproductive age, is associated with chronic pelvic pain, dysmenorrhea, and infertility. Adenomyosis patients not only have infertility but also have higher implantation failure, higher miscarriage rates, and lower live birth rates when undergoing assisted reproductive technologies (ART) than infertile patients without the disease. This study aimed to elucidate the molecular mechanisms underlying adenomyosis-related infertility, focusing on the dysregulated pathways in the mid-secretory and gestational eutopic endometrium, as well as the role of extracellular vesicles (EVs) secreted by this endometrium. This was achieved by generating endometrial organoids serving as a preclinical platform. As a first step, transcriptomic signature analysis of the eutopic endometrium from adenomyosis patients revealed a higher prevalence of non-receptive endometrial states, indicating altered expression of genes involved in decidualization. In these patients, attempts to synchronize the endometrium by adjusting progesterone based on endometrial have proven ineffective, suggesting the involvement of other dysregulated pathways besides progesterone in reported implantation failures. We successfully established a human organoid model recapitulating specific tissue traits and disease characteristics to address the lack of robust in vitro models to study these molecular mechanisms. Next-generation sequencing of adenomyosis patient-derived organoids identified dysregulated genes and pathways during mid-secretory and gestational phases. In the mid-secretory endometrium, several critical genes associated with non-receptive endometrium, epithelial-mesenchymal transition, endometriosis-related infertility, chronic inflammatory responses, and hypoxia-related complications in pregnancy were dysregulated. In the gestational phase, the study highlighted the dysregulation of genes crucial for pregnancy-related processes and genes related to pregnancy disorders such as preeclampsia and recurrent pregnancy loss. Furthermore, this study investigated the role of microRNA cargo of EVs released by adenomyosis eutopic endometrium. EVs secreted by adenomyosis organoids were isolated, and microRNA cargo was evaluated, showing potential roles in regulating molecular mechanisms related to endometrial receptivity, embryo implantation, and embryo early development. In conclusion, this study establishes an adenomyosis organoid model that provides a valuable tool for studying adenomyosis-related infertility, advancing the understanding of molecular mechanisms dysregulated in the adenomyosis eutopic endometrium and contributing to the identification of potential biomarkers for adenomyosis and predictive indicators for reproductive outcomes.