Canine endometrial organoids respond to exogenous steroid hormones and are an in vitro model for cystic endometrial hyperplasia.

Abstract

ABSTRACT: The etiology, epidemiology, and pathogenesis of many uterine diseases, including endometritis, cystic endometrial hyperplasia (CEH), and pyometra, remain poorly understood in canids, hindering the advancement of effective preventative therapeutics to ultimately improve fertility. The development of an in vitro endometrial cell culture system that mimics in vivo architecture, cellular morphology, and physiological responses would facilitate a greater understanding of the mechanisms of complex canine uterine diseases and reduce the need for in vivo research. Three-dimensional organoid cell cultures may provide an innovative approach for studying uterine pathologies in the bitch. Unlike traditional two-dimensional monolayer cell culture systems, organoids can be grown long-term while maintaining the structure, phenotype, and function of their organ of origin. Here, we report the generation and characterization of canine endometrial organoids via immunohistochemistry (IHC), transmission electron microscopy, and RT-qPCR, and demonstrate their responsiveness to exogenous steroid hormones mimicking the estrous cycle. Furthermore, gene expression of key inflammatory markers was upregulated in endometrial organoids derived from bitches with CEH compared to organoids derived from healthy tissue. Findings highlight canine endometrial organoids as a valuable model to study both normal uterine physiology and disease-related processes in dogs. LAY SUMMARY: Organoids are cells that are grown three-dimensionally in the laboratory and retain similar functions to the organ from which the cells were collected. Organoids were used here to study female dog reproductive tissue without the need for live animals, which saves time and money and improves animal welfare. We developed a three-dimensional model of the dog uterus that responds to reproductive hormones and maintains its normal structure. In addition, we used diseased canine tissue to replicate CEH, a condition that affects pregnancy and can lead to the serious disease of pyometra, in which the uterus is infected and filled with pus. In future studies, this validated model can be used to study how CEH develops and to test new treatments for the disease.