Despite usually effective treatments, the occurrence of endometritis in cows continues to exert a detrimental impact on their reproductive capacity, resulting in substantial economic losses within the livestock industry. Endometritis can be caused by various bacterial pathogens, with Escherichia coli (E. coli) being a prominent one [1], [2], [3], [4], [5]. The endometrium, the innermost layer of the uterus, is characterized by a complex network of glands that play a pivotal role in facilitating the intricate interaction between the endometrial cells and an embryo that establishes a successful pregnancy [6,7]. The growth, differentiation, and cyclical changes of the endometrium are regulated by two pivotal ovarian hormones: estrogen, with estradiol (E2) as its primary component and progesterone. Notably, there is predominant expression of E2 during the proliferative phase [8,9]. The role of E2 has been extensively studied in various biological processes, not only reproduction but also as an immunomodulator in regulating lymphocyte expansion and anti-inflammatory responses [[10], [11], [12]].

Wnt/β-catenin plays an important role in E2-mediated uterine development, and E2 promotes endometrial cell proliferation and damage repair through the Wnt/β-catenin pathway [13,14]. The Wnt pathway is intricately associated with the development of glands in the endometrial epithelium and the maintenance of pregnancy. The canonical Wnt pathway relies on β-catenin, a pivotal signaling molecule, which plays a crucial role in cellular proliferation, differentiation, invasion, and adhesion [15,16]. Previous studies have demonstrated that the in vitro growth and development of organoids also depend on the activation of Wnt signaling, while sustained culture of organoids necessitates E2 supplementation. Notably, E2 has been shown to facilitate cilia formation on endometrial organoids, potentially associated with the cyclic changes observed in the endometrium [[17], [18], [19]].

The utilization of 3D organoids derived from cultured endometrial epithelial cells has significantly enhanced the precision of in vitro investigations of human development, diseases and personalized therapies when compared with experiments on conventional 2D cell cultures [20,21]. Moreover, organoids have the potential to simulate organogenesis and pathological processes, thereby broadening their applicability in fundamental research, pharmaceutical exploration, and treatment [[22], [23], [24]]. It has been observed that 2D cultures of endometrial cells have limited longevity and rapidly lose their phenotype and hormone responsiveness, whereas organoids demonstrate a remarkable capacity for expansion while preserving their phenotypic and functional characteristics, overcoming the trend to senescence of most primary cell cultures [[25], [26], [27]]. In this study, we investigated the potential of E2 treatment to ameliorate inflammatory damage in organoids and cells following E. coli infection while also exploring the interplay between E2 and the Wnt pathway, with the aim of providing adjunctive therapy for bovine uterine-related diseases.