Animal models of the placenta accreta spectrum: current status and further perspectives.

Placenta accreta spectrum disorder (PAS) is a kind of disease of placentation defined as abnormal trophoblast invasion of part or all of the placenta into the myometrium, even penetrating the uterus. Decidual deficiency, abnormal vascular remodeling in the maternal-fetal interface, and excessive invasion by extravillous trophoblast (EVT) cells contribute to its onset. However, the mechanisms and signaling pathways underlying such phenotypes are not fully understood, partly due to the lack of suitable experimental animal models. Appropriate animal models will facilitate the comprehensive and systematic elucidation of the pathogenesis of PAS. Due to the remarkably similar functional placental villous units and hemochorial placentation to humans, the current animal models of PAS are based on mice. There are various mouse models induced by uterine surgery to simulate different phenotypes of PAS, such as excessive invasion of EVT or immune disturbance at the maternal-fetal interface, which could define the pathological mechanism of PAS from the perspective of the "soil." Additionally, genetically modified mouse models could be used to study PAS, which is helpful to exploring the pathogenesis of PAS from the perspectives of both "soil" and "seed," respectively. This review details early placental development in mice, with a focus on the approaches of PAS modeling. Additionally, the strengths, limitations and the applicability of each strategy and further perspectives are summarized to provide the theoretical foundation for researchers to select appropriate animal models for various research purposes. This will help better determine the pathogenesis of PAS and even promote possible therapy.

PCSK9 involves in the high-fat diet-induced abnormal testicular function of male mice.

In brief: Impaired spermatogenesis resulting from disturbed cholesterol metabolism due to intake of high-fat diet (HFD) has been widely recognized, however, the role of preprotein invertase subtilin 9 (PCSK9), which is a negative regulator of cholesterol metabolism, has never been reported. This study aims to reveal the role of PCSK9 on spermatogenesis induced by HFD in mice. Abstract: Long-term consumption of a high-fat diet (HFD) is an important factor that leads to impaired spermatogenesis exhibiting poor sperm quantity and quality. However, the mechanism of this is yet to be elucidated. Disrupted cholesterol homeostasis is one of many crucial pathological factors which could contribute to impaired spermatogenesis. As a negative regulator of cholesterol metabolism, preprotein invertase subtilin 9 (PCSK9) mediates low density lipoprotein receptor (LDLR) degradation to the lysosome, thereby reducing the expression of LDLR on the cell membrane and increasing serum low-density lipoprotein cholesterol level, resulting in lipid metabolism disorders. Here, we aim to study whether PCSK9 is a pathological factor for impaired spermatogenesis induced by HFD and the underlying mechanism. To meet the purpose of our study, we utilized wild-type C57BL/6 male mice and PCSK9 knockout mice with same background as experimental subjects and alirocumab, a PCSK9 inhibitor, was used for treatment. Results indicated that HFD induced higher PCSK9 expression in serum, liver, and testes, and serum PCSK9 is negatively correlated with spermatogenesis, while both PCSK9 inhibitor treatment and PCSK9 knockout methodologies ameliorated impaired lipid metabolism and spermatogenesis in mice fed a HFD. This could be due to the overexpression of PCSK9 induced by HFD leading to dyslipidemia, resulting in testicular lipotoxicity, thus activating the Bcl-2-Bax-Caspase3 apoptosis signaling pathway in testes, particularly in Leydig cells. Our study demonstrates that PCSK9 is an important pathological factor in the dysfunction of spermatogenesis in mice induced by HFD. This finding could provide innovative ideas for the diagnosis and treatment of male infertility.

Hyperandrogen-induced polyol pathway flux increase affects ovarian function in polycystic ovary syndrome via excessive oxidative stress.

AIMS: Polycystic ovary syndrome (PCOS) is a common endocrine disorder in the women of childbearing age. It is characterized by hyperandrogenism and abnormal follicular growth and ovulation. The polyol pathway is a glucose metabolism bypass pathway initiated by aldose reductase (ADR). Androgen induces the expression of ADR in the male reproductive tract, which has a general physiological significance for male reproductive function. Here we investigate whether hyperandrogenemia in PCOS leads to increased flux of the polyol pathway in ovarian tissue, which in turn affects follicular maturation and ovulation through oxidative stress. MAIN METHODS: We used clinical epidemiological methods to collect serum and granulosa cells from clinical subjects for a clinical case-control study. At the same time, cell biology and molecular biology techniques were used to conduct animal and cell experiments to further explore the mechanism of hyperandrogen-induced ovarian polyol pathway hyperactivity and damage to ovarian function. KEY FINDINGS: Here, we find that hyperandrogenism of PCOS can induce the expression of ovarian aldose reductase, which leads to the increase of the polyol pathway flux, and affects ovarian function through excessive oxidative stress. SIGNIFICANCE: Our research has enriched the pathological mechanism of PCOS and may provide a new clue for the clinical treatment of PCOS.

The diversity of trophoblast cells and niches of placenta accreta spectrum disorders revealed by single-cell RNA sequencing.

Placenta accreta spectrum disorders (PAS) are severe pregnancy complications that occur when extravillous trophoblast cells (EVTs) invade beyond the uterine inner myometrium and are characterized by hypervascularity on prenatal ultrasound and catastrophic postpartum hemorrhage. The potential mechanisms remain incompletely understood. With single-cell RNA-sequencing analysis on the representative invasive parts and the normal part obtained from the same PAS placenta, we profiled the pathological landscape of invasive PAS placenta and deciphered an intensified differentiation pathway from progenitor cytotrophoblasts (CTBs) to EVTs via LAMB4 + and KRT6A + CTBs. In the absence of the decidua, the invasive trophoblasts of various differentiation states interacted with ADIRF + and DES + maternal stromal cells. The PAS-associated hypervascularity might be due to the enhanced crosstalk of trophoblasts, stromal cells and vascular endothelial cells. Finally, we presented an immune microenvironmental landscape of invasive PAS. The pathogenesis of PAS could be further explored with current resources for future targeted translational studies.

[Progress in the role of endometrial glucose metabolism in embryo implantation].

The synthesis and decomposition of glycogen adjust the blood glucose dynamically to maintain the energy supply required by the cells. As the only hormone that lowers blood sugar in the body, insulin can promote glycogen synthesis by activating the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway and increasing glucose transporter translocation, and inhibit gluconeogenesis to lower blood glucose. In the endometrium, glycogen metabolism is active, but gluconeogenesis does not occur. The glycogen metabolism in the endometrium is controlled not only by the classical glucose regulating hormones, but also by the ovarian hormones. The functional activities related to implantation of the endometrium during the implantation window require glucose as energy source. A large amount of glucose is used to synthesize glycogen in the endometrium before implantation, which could meet the increased energy demand for embryo implantation. In diabetes, glycogen metabolism in the endometrium is impaired, which frequently leads to implantation failure and early abortion. This article reviews the glycogen metabolism in the endometrium and discusses its role in embryo implantation, which provide new ideas for embryo implantation research and infertility treatment.

Consistency and synchronization of AMPK-glycogen in endometrial epithelial cells are critical to the embryo implantation.

Uterine receptivity to the embryo is crucial for successful implantation. The establishment of uterine receptivity requires a large amount of energy, and abnormal energy regulation causes implantation failure. Glucose metabolism in the endometrium is tissue specific. Glucose is largely stored in the form of glycogen, which is the main energy source for the endometrium. AMP-activated protein kinase (AMPK), an important energy-sensing molecule, is a key player in the regulation of glucose metabolism and its regulation is also tissue specific. However, the mechanism of energy regulation in the endometrium for the establishment of uterine receptivity remains to be elucidated. In this study, we aimed to investigate the energy regulation mechanism of mouse uterine receptivity and its significance in embryo implantation. The results showed that the AMPK, p-AMPK, glycogen synthase 1, and glycogen phosphorylase M levels and the glycogen content in mouse endometrial epithelium varied in a periodic manner under regulation by the ovarian hormone. Specifically, progesterone significantly activated AMPK, promoted glycogenolysis, and upregulated glycogen phosphorylase M expression. AMPK regulated glycogen phosphorylase M expression and promoted glycogenolysis. AMPK was also found to be activated by changes in the energy or glycogen of the endometrial epithelial cells. The inhibition of AMPK activity or glycogenolysis altered the uterine receptivity markers during the window of implantation and ultimately interfered with implantation. In summary, consistency and synchronization of AMPK and glycogen metabolism constitute the core regulatory mechanism in mouse endometrial epithelial cells involved in the establishment of uterine receptivity.

Activation of SGLT3a in endometrial epithelial cells induces paracrine stromal cell decidualization.

Endometrial epithelial cells (EECs) and stromal cells (ESCs) have a close functional association. During the peri-implantation period, EECs with enhanced functional activities secrete a variety of paracrine factors to promote the decidualization of ESCs. However, little is known about the specific process by which EECs secrete paracrine factors to induce the decidualization of ESCs. Some evidence suggests that the activation of sodium-glucose cotransporter 3a (SGLT3a) induces the depolarization of ESCs to affect their function. Therefore, SGLT3a acts as a sensor molecule in certain cell types. In this study, the expression of SGLT3a was investigated in EECs to determine whether its levels increased during the peri-implantation period in female mice. The activation of SGLT3a in mouse EECs induced Na+ -dependent depolarization of the cell membrane and an influx of extracellular Ca2+ , which further promoted the expression and release of the paracrine factors prostaglandin E2 (PGE2) and F2-alpha (PGF2α) by upregulating the expression of cyclooxygenase-2. In turn, PGE2 and PGF2α induced the decidualization of ESCs. Importantly, we identified SGLT3a as a key molecule involved in the cross-talk between EECs and ESCs during the process of uterine decidualization.

Contribution of high-fat diet-induced PCSK9 upregulation to a mouse model of PCOS is mediated partly by SREBP2.

The incidence of polycystic ovary syndrome (PCOS) due to high-fat diet (HFD) consumption has been increasing significantly. However, the mechanism by which a HFD contributes to the pathogenesis of PCOS has not been elucidated. Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a key protein that regulates cholesterol metabolism. Our previous study revealed abnormally high PCSK9 levels in serum from patients with PCOS and in serum and hepatic and ovarian tissues from PCOS model mice, suggesting that PCSK9 is involved in the pathogenesis of PCOS. However, the factor that induces high PCSK9 expression in PCOS remains unclear. In this study, Pcsk9 knockout mice were used to further explore the role of PCSK9 in PCOS. We also studied the effects of a HFD on the expression of PCSK9 and sterol regulatory element-binding protein 2 (SREBP2), a regulator of cholesterol homeostasis and a key transcription factor that regulates the expression of PCSK9, and the roles of these proteins in PCOS pathology. Our results indicated HFD may play an important role by inducing abnormally high PCSK9 expression via SREBP2 upregulation. We further investigated the effects of an effective SREBP inhibitor, fatostain, and found that it could reduce HFD-induced PCSK9 expression, ameliorate hyperlipidemia and improve follicular development in PCOS model mice. Our study thus further elucidates the important role of an HFD in the pathogenesis of PCOS and provides a new clue in the prevention and treatment of this disorder.

Expression of SGLT1 in the Mouse Endometrial Epithelium and its Role in Early Embryonic Development and Implantation.

Many functional activities of endometrium epithelium are energy consuming which are very important for maintaining intrauterine environment needed by early embryonic development and establishment of implantation window. Glucose is a main energy supplier and one of the main components of intrauterine fluid. Obviously, glucose transports in endometrium epithelium involve in for these activities but their functions have not been elucidated. In this research, we observed a spatiotemporal pattern of sodium glucose transporter 1 (SGLT1) expression in the mouse endometrium. We also determined that progesterone can promote the expression of SGLT1 in the mouse endometrial epithelium in response to the action of oestrogen. Treatment with the SGLT1 inhibitor phlorizin or small interfering RNA specific for SGLT1 (SGLT1-siRNA) altered glucose uptake in primary cultured endometrial epithelial cells, which exhibited reduced ATP levels and AMPK activation. The injection of phlorizin or SGLT1-siRNA into one uterine horn of each mouse on day 2 of pregnancy led to an increased glucose concentration in the uterine fluid and decreased number of harvested normal blastocysts and decreased expression of integrin αVß3 in endometrial epithelium and increased expression of mucin 1 and lactoferrin in endometrial epithelium and the uterine homogenates exhibited activated AMPK, a decreased ATP level on day 4, and a decreased number of implantation sites on day 5. In embryo transfer experiments, pre-treatment of the uterine horn with phlorizin or SGLT1-siRNA during the implantation window led to a decreased embryo implantation rate on day 5 of pregnancy, even when embryos from normal donor mice were used. In conclusion, SGLT1, which participates in glucose transport in the mouse endometrial epithelium, inhibition and/or reduced expression of SGLT1 affects early embryo development by altering the glucose concentration in the uterine fluid. Inhibition and/or reduced expression of SGLT1 also affects embryo implantation by influencing energy metabolism in epithelial cells, which consequently influences implantation-related functional activities.

GLUT4 in Mouse Endometrial Epithelium: Roles in Embryonic Development and Implantation.

GLUT4 is involved in rapid glucose uptake among various kinds of cells to contribute to glucose homeostasis. Prior data have reported that aberrant glucose metabolism by GLUT4 dysfunction in the uterus could be responsible for infertility and increased miscarriage. However, the expression and precise functions of GLUT4 in the endometrium under physiological conditions remain unknown or controversial. In this study, we observed that GLUT4 exhibits a spatiotemporal expression in mouse uterus on pregnant days 1-4; its expression especially increased on pregnant day 4 during the window of implantation. We also determined that estrogen, in conjunction with progesterone, promotes the expression of GLUT4 in the endometrial epithelium in vivo or in vitro. GLUT4 is an important transporter that mediates glucose transport in endometrial epithelial cells (EECs) in vitro or in vivo. In vitro, glucose uptake decreased in mouse EECs when the cells were treated with GLUT4 small interfering RNA (siRNA). In vivo, the injection of GLUT4-siRNA into one side of the mouse uterine horns resulted in an increased glucose concentration in the uterine fluid on pregnant day 4, although it was still lower than in blood, and impaired endometrial receptivity by inhibiting pinopode formation and the expressions of leukemia inhibitory factor (LIF) and integrin ανß3, finally affecting embryonic development and implantation. Overall, the obtained results indicate that GLUT4 in the endometrial epithelium affects embryo development by altering glucose concentration in the uterine fluid. It can also affect implantation by impairing endometrial receptivity due to dysfunction of GLUT4.

The Effects of Altered Endometrial Glucose Homeostasis on Embryo Implantation in Type 2 Diabetic Mice.

Type 2 diabetes mellitus (T2DM) is a disease characterized by hyperglycemia resulting from insulin resistance. In recent years, the incidence of T2DM has been increasing. Women with T2DM often suffer from infertility and early miscarriage; however, the underlying mechanisms remain unclear. Insulin is the most important regulatory hormone of glycogen metabolism. In addition, 5' adenosine monophosphate-activated protein kinase (AMPK) is an important regulator of glycogen metabolism. Patients with T2DM have inhibited AMPK expression in the liver, which leads to impaired glucose metabolism. However, the role of AMPK in endometrial glycogen metabolism has not been reported. In this study, a mouse model of T2DM was established to investigate whether altered endometrial glucose metabolism affects early embryo implantation. Metformin and insulin were used for therapy; the resulting changes to glycogen metabolism and embryo implantation were examined. The results indicate that the concentrations of glycogen decreased significantly in T2DM mice, resulting in insufficient energy supplies for proper endometrial function, and thereby impeding embryonic implantation. Interestingly, endometrial AMPK was not found to be overactivated. Insulin treatment was found to partially resolve the embryo implantation defects in T2DM mice. Metformin improved blood glucose but did not have a significant effect on local endometrial glucose metabolism. This study explored the changes in endometrial glucose metabolism in T2DM mouse, and the effects of these changes on embryo implantation. We found that insulin, but not metformin, significantly resolved embryo implantation problems. These findings will help to increase our understanding of the pathomechanisms of infertility and early miscarriage in women with T2DM.

Impact of Disturbed Glucose Homeostasis Regulated by AMPK in Endometrium on Embryo Implantation in Diabetes Mice.

The incidence of diabetes in women of childbearing age has been increasing recently and implantation failure and early abortion are important reasons for infertility in diabetic women. Glycogen synthesis and decomposition are the cores of glucose homeostasis in endometrium and AMPK is activated when cellular energy consumption increases. Embryo implantation is a complex process required huge energy. Yet the changes of glucose metabolism in endometrium and its impact on embryo implantation in diabetic women are still unclear. In this research, we established diabetic pregnancy mice model by intraperitoneal injecting streptozotocin on pregnant day 1. We first tested the changes of endometrial glucose homeostasis and embryo implantation. Next, we demonstrated abnormal activation of AMPK in the endometrium of diabetic mice and its affecting endometrial glucose homeostasis. Finally, we compared the endometrial glucose homeostasis and embryo implantation outcome in diabetic pregnant mice treated with insulin or insulin combined with metformin. The results indicated that there was disturbed glucose homeostasis associated with excessive activation of AMPK in endometrium of diabetic pregnant mice. AMPK inhibitor improved the over-activation of AMPK pathway in the endometrium, meanwhile, partially corrected the abnormal glycogen metabolism and improved the implantation. Insulin improved the disorder of endometrial glucose homeostasis and implantation of diabetic mice. Our research explores the causes of high abortion and infertility rate in diabetic women which is to provide a therapeutic reference for patients with diabetes complicated with infertility and early abortion.