Glutaminolysis regulates endometrial fibrosis in intrauterine adhesion via modulating mitochondrial function.

BACKGROUND: Endometrial fibrosis, a significant characteristic of intrauterine adhesion (IUA), is caused by the excessive differentiation and activation of endometrial stromal cells (ESCs). Glutaminolysis is the metabolic process of glutamine (Gln), which has been implicated in multiple types of organ fibrosis. So far, little is known about whether glutaminolysis plays a role in endometrial fibrosis. METHODS: The activation model of ESCs was constructed by TGF-ß1, followed by RNA-sequencing analysis. Changes in glutaminase1 (GLS1) expression at RNA and protein levels in activated ESCs were verified experimentally. Human IUA samples were collected to verify GLS1 expression in endometrial fibrosis. GLS1 inhibitor and glutamine deprivation were applied to ESCs models to investigate the biological functions and mechanisms of glutaminolysis in ESCs activation. The IUA mice model was established to explore the effect of glutaminolysis inhibition on endometrial fibrosis. RESULTS: We found that GLS1 expression was significantly increased in activated ESCs models and fibrotic endometrium. Glutaminolysis inhibition by GLS1 inhibitor bis-2-(5-phenylacetamido-1,2,4-thiadiazol-2-yl) ethyl sulfide (BPTES or glutamine deprivation treatment suppressed the expression of two fibrotic markers, α-SMA and collagen I, as well as the mitochondrial function and mTORC1 signaling in ESCs. Furthermore, inhibition of the mTORC1 signaling pathway by rapamycin suppressed ESCs activation. In IUA mice models, BPTES treatment significantly ameliorated endometrial fibrosis and improved pregnancy outcomes. CONCLUSION: Glutaminolysis and glutaminolysis-associated mTOR signaling play a role in the activation of ESCs and the pathogenesis of endometrial fibrosis through regulating mitochondrial function. Glutaminolysis inhibition suppresses the activation of ESCs, which might be a novel therapeutic strategy for IUA.

New insights on mitochondrial heteroplasmy observed in ovarian diseases.

BACKGROUND: The reportedly high mutation rate of mitochondrial DNA (mtDNA) may be attributed to the absence of histone protection and complete repair mechanisms. Mitochondrial heteroplasmy refers to the coexistence of wild-type and mutant mtDNA. Most healthy individuals carry a low point mutation load (<1 %) in their mtDNA, typically without any discernible phenotypic effects. However, as it exceeds a certain threshold, it may cause the onset of various diseases. Since the ovary is a highly energy-intensive organ, it relies heavily on mitochondrial function. Mitochondrial heteroplasmy can potentially contribute to a variety of significant ovarian disorders. AIM OF REVIEW: In this review, we have elucidated the close relationship between mtDNA heteroplasmy and ovarian diseases, and summarized novel avenues and strategies for the potential treatment of these ovarian diseases. KEY SCIENTIFIC CONCEPTS OF REVIEW: Mitochondrial heteroplasmy can potentially contribute to a variety of significant ovarian disorders, including polycystic ovary syndrome, premature ovarian insufficiency, and endometriosis. Current strategies related to mitochondrial heteroplasmy are untargeted and have low bioavailability. Nanoparticle delivery systems loaded with mitochondrial modulators, mitochondrial replacement/transplantation therapy, and mitochondria-targeted gene editing therapy may offer promising paths towards potentially more effective treatments for these diseases, despite ongoing challenges.

Abnormal expression of fission and fusion genes and the morphology of mitochondria in eutopic and ectopic endometrium.

Mitochondria play a pivotal role in physiological and metabolic function of the cell. Mitochondrial dynamics orchestrate mitochondrial function and morphology, involving fission and fusion as well as ultrastructural remodeling. Mounting evidence unravels the close link between mitochondria and endometriosis. However, how mitochondrial architecture changes through fission and fusion in eutopic and ectopic tissues of women with ovarian endometriosis remains unknown. We detected the expression of fission and fusion genes and the morphology of mitochondria in eutopic and ectopic endometrium in ovarian endometriosis. The results showed that the expression of DRP1 and LCLAT1 was upregulated in eutopic endometrial stromal cells (ESCs), and the expression of DRP1, OPA1, MFN1, MFN2, and LCLAT1 was significantly downregulated in ectopic ESCs, and reduced number of mitochondria, wider cristae width and narrower cristae junction width was observed, but there was no difference in cell survival rate. The altered mitochondrial dynamics and morphology might, respectively, provide an advantage for migration and adhesion in eutopic ESCs and be the adaptive response in ectopic endometrial cells to survive under hypoxic and oxidative stress environment.

The Clinical Picture and Fecundity of Primary and Recurrent Ovarian Endometriosis with Family History: A Retrospective Analysis.

This study aims to evaluate the role of endometriosis family history on the clinical manifestation and fertility performance of primary and recurrent endometriosis. In total, 312 primary and 323 recurrent endometrioma patients with a histological diagnosis were included in this study. Family history was significantly correlated with recurrent endometriosis (adjusted OR: 3.52, 95% CI: 1.09-9.46, p = 0.008). Patients with a family history showed a significantly higher proportion of recurrent endometriosis (75.76% vs. 49.50%), higher rASRM scores, higher incidence of severe dysmenorrhea, and severe pelvic pain than the sporadic cases. Recurrent endometrioma showed statistical increase in rASRM scores, percentage of rASRM Stage IV, dysmenorrhea, dyschezia, those undergoing semi-radical surgery or unilateral oophorosalpingectomy, postoperative medical treatment, e with a positive family history, while a decrease in the incidence of asymptomatic phenomena and those undergoing ovarian cystectomy compared to those with primary endometriosis. The naturally conceived pregnancy rate was higher in primary endometriosis compared to recurrent endometriosis. Compared to recurrent endometriosis with a negative family history, recurrent endometriosis with a positive family history had a higher incidence of severe dysmenorrhea, chronic pelvic pain, a higher spontaneous abortion rate, and a lower natural pregnancy rate. Primary endometriosis with a family history presented a higher incidence of severe dysmenorrhea than those without a family history. In conclusion, endometriosis patients with a positive family history presented a higher pain severity and lower conception probability compared to the sporadic cases. Recurrent endometriosis showed further-exacerbated clinical manifestations, more pronounced familial tendency, and lower pregnancy rates than primary endometriosis.

Risk factors for coexisting deep endometriosis for patients with recurrent ovarian endometrioma.

Aim: The aim of this study was to assess the risk factors for coexisting deep endometriosis (DE) in patients with recurrent ovarian endometrioma (OE). Methods: We retrospectively reviewed 151 recurrent OE patients who had been diagnosed of OE but not DE at the time of their first surgery and then received a second surgery for recurrent endometriosis with or without DE. Their clinical characteristics at the time of the first and second surgeries were collected. Univariate and multivariate logistic regression analyses were conducted to identify potential risk factors for coexisting DE in patients with recurrent OE. Results: Among the 151 recurrent OE patients, 46 were diagnosed of DE during the recurrent surgery and included in the DE group, while the remaining 105 patients were included in the non-DE group. In univariate analysis, there were significant differences in terms of uterine retroversion during the primary surgery and the follow-up time after the primary surgery between the DE and non-DE groups. The multivariate analysis also showed that both uterine retroversion and the follow-up time (≥5 years) were associated with the coexistence of DE during the recurrent surgery. The odds ratio (OR) for uterine retroversion was 3.72 [95% confidence interval (CI) 1.62-8.53], and the OR for follow-up time (≥5 years) was 5.03 (95% CI 2.29-11.02). Conclusions: Our study suggested that for recurrent OE patients, uterine retroversion during the first surgery and a follow-up time of at least 5 years are risk factors for the coexistence of DE in recurrent surgery, early prevention and full preparation before the recurrent surgery should be emphasized in these conditions.

Identification of NFASC and CHL1 as Two Novel Hub Genes in Endometriosis Using Integrated Bioinformatic Analysis and Experimental Verification.

Background: Endometriosis (EMS) is a common and highly recurrent gynecological disease characterized by chronic pain and infertility. There are no definitive therapies for endometriosis since the pathogenesis remains undetermined. This study aimed to identify EMS-related functional modules and hub genes by integrated bioinformatics analysis. Methods: Three endometriosis expression profiling series (GSE25628, GSE23339, and GSE7305) were obtained from Gene Expression Omnibus (GEO). The EMS-related module was constructed by weighted gene co-expression network analysis (WGCNA), followed by Gene Ontology (GO) enrichment analyses. Cytohubba and the MCODE plug-ins of Cytoscape were used to screen out the hub genes, which were verified via receiver operating characteristic (ROC) curves. Immunohistochemistry was performed to verify the protein expression of the hub genes in ectopic endometrial tissues. Moreover, CIBERSORT was used to analyze the relationship between the abundance of immune cells infiltration and the expression of hub genes. Results: Among the 18 modules obtained, the darkmagenta module was identified as the EMS-related module, genes of which were significantly enriched to terms referring to cell migration and neurogenesis. NFASC and CHL1 were screened out and prioritized as hub genes through Cytoscape and confirmed to be differentially upregulated in ectopic endometrial samples. Finally, the expression of hub genes was related to the abundance of immune cells infiltration. The higher expression of NFASC or CHL1 correlated with increased M2 macrophages and decreased natural killer (NK) cells in ectopic lesions. Conclusion: This study provided new insights into the molecular factors underlying the pathogenesis of endometriosis and provided a theoretical basis for the potential that the two hub genes, NFASC and CHL1, might be novel biomarkers and therapeutic targets in the future.

Obstetrical and oncological outcomes of twin pregnancies with hydatidiform mole and coexisting fetus.

OBJECTIVE: To evaluate the obstetrical and oncological progression of twin pregnancies with hydatidiform mole coexisting fetus (HMCF). MATERIALS AND METHODS: Using a retrospective method based on patients from the Women's Hospital, Zhejiang University School of Medicine database between January 1990 and October 2020, 17 patients were histologically confirmed as having HMCF, and the patients' prenatal diagnosis, outcomes and development of gestational trophoblastic neoplasia (GTN) were reviewed. RESULTS: Among these 17 cases, 11 (64.71%) cases were complete hydatidiform mole coexisting fetus (CHMCF), and 6 (35.29%) cases were partial hydatidiform mole coexisting fetus (PHMCF). The gestational age at diagnosis of CHMCF was significantly earlier than that of PHMCF [9 (8-24) vs. 18 (11-32) weeks, respectively, P < 0.05]. The live birth rate of PHMCF was slightly higher than that of CHMCF (33.33%; 18.18%), but this difference was not statistically significant. The overall rate of GTN incidence of HMCF was 47.06% (8/17), and the GTN rates of PHMCF and CHMCF were 33.33% (2/6) and 54.55% (6/11), respectively. There was no significant difference in the GTN rate between patients who chose to continue pregnancy and those who terminated pregnancy before 24 weeks of gestation. The GTN rate of patients with term delivery was not significantly higher than that of preterm delivery. CONCLUSION: In HMCF cases, the incidence rate of CHMCF was higher than that of PHMCF, and PHMCF is more difficult to diagnose in the early stage. Continuing pregnancy does not increase the risk of GTN compared to terminating pregnancy. In cases of HMCF, when the fetal karyotype is normal and maternal complications are controlled, it is safe to continue the pregnancy and extend it to term.

Osteopontin Regulates Endometrial Stromal Cell Migration in Endometriosis through the PI3K Pathway : Osteopontin Regulates Endometrial Cell Migration in Endometriosis.

Endometriosis is generally characterized as a tumor-like disease because of its potential for distant metastasis and local tissue invasion, while whether osteopontin (OPN) plays a role in the pathogenesis of endometriosis has not been thoroughly investigated. We investigated the expression of OPN, urokinase plasminogen activator (uPA), phosphatidylinositol 3 kinase (PI3K), and phospho-PI3 kinase (p-PI3K) in endometrial stromal cells (ESCs). The serum concentration of OPN was determined by enzyme-linked immunosorbent assays (ELISA). OPN was downregulated to explore the corresponding change of uPA, p-PI3K, F-actin, and α-tubulin. The expression of OPN, uPA, PI3K, and p-PI3K was evaluated by western blot and quantitative real-time PCR (RT-qPCR) and the expression of F-actin and α-tubulin was confirmed by immunofluorescence assay. The proliferation and migration abilities of ESCs were investigated by CCK8, transwell, and wound scratch assays. Endometrial OPN, p-PI3K, and uPA expressions and serum OPN levels were increased in patients with endometriosis compared with the control. The expressions of p-PI3K, uPA, and α-tubulin were decreased by siRNA-OPN interference in ectopic ESCs. Activation and inhibition of the PI3K pathway apparently upregulate and downregulate uPA expression. Knockdown of OPN and inhibition of the PI3K pathway remarkably inhibited cell migration in ectopic ESCs. Meanwhile, activation of the PI3K pathway promoted the migration ability of ectopic ESCs. OPN may regulate the expression of uPA through the PI3K signal pathway to affect the migration ability of ESCs, indicating that OPN, uPA, and the PI3K pathway may be potential targets for interrupting development of endometriosis.

Ezrin T567 phosphorylation regulates migration and invasion of ectopic endometrial stromal cells by changing actin cytoskeleton.

AIMS: The enhanced ability of endometrial cell migration and invasion is the foundation for formation of ectopic lesions in endometriosis. Ezrin has been reported to regulate cell motility by remodeling the cytoskeleton. However, little is known about the mechanisms through which ezrin remodels the cytoskeleton and cell structure to promote cell motility in endometriosis. METHODS: In our study, expression and distribution of ezrin, and Rho pathway were detected through immunohistochemical analysis. The effects of inhibiting ezrin T567 phosphorylation on Rho signaling pathway and cytoskeleton were investigated through western blot, transmission electron microscopy and immunofluorescence analysis. KEY FINDINGS: We found that the expression of ezrin and Rho pathway was higher in ectopic endometrium. NSC305787 inhibited the phosphorylation of ezrin T567, resulting in decreased expression of Rho pathway and reduced filopodia formation in ectopic endometrial stromal cells. SIGNIFICANCE: Taken together, our study suggested that ezrin T567 phosphorylation modulated migration and invasion of ectopic ESCs through actin reconstructions, which may serve as a novel therapeutic target in ovarian endometriosis.