Generation of Human Endometrial Assembloids with a Luminal Epithelium using Air-Liquid Interface Culture Methods.

The endometrial lining of the uterus is essential for women's reproductive health and consists of several different types of epithelial and stromal cells. Although models such as gland-like structures (GLSs) and endometrial assembloids (EnAos) are successfully established, they lack an intact luminal epithelium, which makes it difficult to recapitulate endometrial receptivity. Here, a novel EnAo model (ALI-EnAo) is developed by combining endometrial epithelial cells (EnECs) and stromal cells (EnSCs) and using an improved matrix and air-liquid interface (ALI) culture method. ALI-EnAos exhibit intact EnSCs and glandular and luminal epithelia, which recapitulates human endometrium anatomy, cell composition, hormone-induced menstrual cycle changes, gene expression profiles, and dynamic ciliogenesis. The model suggests that EnSCs, together with the extracellular matrix and ALI culture conditions, contribute to EnAo phenotypes and characteristics reflective of the endometrial menstrual cycle. This enables to transcriptionally define endometrial cell subpopulations. It anticipates that ALI-EnAos will facilitate studies on embryo implantation, and endometrial growth, differentiation, and disease.

Role of kisspeptin/GPR54 in the first trimester trophoblast of women with a history of recurrent spontaneous abortion.

Trophoblast migration and invasion during pregnancy are under strict physiological control, both temporally and spatially. The trophoblast's ability to invade the endometrium is regulated by the dynamic interaction between invasion-related genes. Once this dynamic balance is broken, the trophoblast exhibits abnormal invasion ability, often resulting in recurrent spontaneous abortion (RSA). Kisspeptins, products of the KISS1 gene, were originally identified as metastasis suppressor peptides with the ability to bind G protein-coupled receptors (i.e., GPR54). The human placenta expresses both KISS1 and kisspeptin receptor (KISS1R) mRNA within the trophoblast compartment, where it is thought to inhibit physiological invasion. In order to explore the effects of KISS1 on the biological behavior of human trophoblasts and its association with RSA, we used immunohistochemistry to compare the expression of kisspeptin and GPR54 at the maternal-fetal interface in RSA cases and cases with normal pregnancy. Abortion-prone CBA/J×DBA/2 matings were established as a model for spontaneous abortion, while non-abortion-prone CBA/J×BALB/c matings were used as a model for normal pregnancy. The expression of kisspeptin and GPR54 in mice placental tissues was compared by immunohistochemistry. Gene recombination and transfection technology were used to evaluate the effects of the KISS1 gene and kisspeptin on JAR cells in terms of proliferation, colony formation ability, and migration and invasion abilities. Kisspeptin/GPR54 revealed lower levels of expression at the maternal-fetal surface in RSA patients compared to controls (P<0.001). Similarly, the expression of kisspeptin/GPR54 at the maternal-fetal interface of spontaneous-abortion mice (CBA/J×DBA/2) was remarkably lower than the group of mice that experienced normal pregnancy (CBA/J×BALB/c) (P<0.05). Data indicated that the KISS1 gene and kisspeptin play a significant role in the inhibition of trophoblast migration and invasion propensity in vitro without affecting cell growth or proliferation. Moreover, kisspeptin appeared to exert an effect in a dose-dependent manner. These data support the fact that the downregulation of kisspeptin/GPR54 may be related to RSA, and that the abnormal expression of KISS1 acts as an invasion-inhibitor gene. Consequently, KISS1 possesses the ability to interfere with normal homeostasis of trophoblast regulation, ultimately resulting in miscarriage.

Expression of kisspeptin/GPR54 and PIBF/PR in the first trimester trophoblast and decidua of women with recurrent spontaneous abortion.

Kisspeptin and its receptor GPR54 play a major role in trophoblast invasion, and progesterone-induced blocking factor (PIBF) is needed for maintaining pregnancy. The expression of kisspeptin/GPR54 and PIBF/progesterone receptor (PR) in trophoblasts and deciduas and the relationship between kisspeptin and PIBF were investigated in the same women with recurrent spontaneous abortion (RSA). Trophoblastic and decidual tissues were collected from 32 RSA women who miscarried a genetically normal fetus, and 35 women who had voluntary abortion. Kisspeptin, GPR54, PIBF and PR were investigated using immunohistochemistry. Kisspeptin, GPR54 and PIBF expressions in syncytiotrophoblasts and cytotrophoblasts were decreased in RSA women as compared to controls (P<0.05). Kisspeptin, PIBF and PR expressions in deciduas were significantly decreased in RSA women as compared to controls (P<0.01). GPR54 expression in deciduas nearly showed no difference between the RSA group and the control group (P=0.958). Kisspeptin and PIBF expressions in syncytiotrophoblasts, cytotrophoblasts and deciduas were correlated with each other in the RSA group (Kappa=0.602, P=0.001; Kappa=0.590, P=0.001; Kappa=0.392, P=0.011). These data support the hypothesis that decreased kisspeptin and PIBF expressions in trophoblasts and deciduas are associated with RSA.