Integrating leiomyoma genetics, epigenomics, and single-cell transcriptomics reveals causal genetic variants, genes, and cell types.

Uterine fibroids (UF), that can disrupt normal uterine function and cause significant physical and psychological health problems, are observed in nearly 70% of women of reproductive age. Although heritable genetics is a significant risk factor, specific genetic variations and gene targets causally associated with UF are poorly understood. Here, we performed a meta-analysis on existing fibroid genome-wide association studies (GWAS) and integrated the identified risk loci and potentially causal single nucleotide polymorphisms (SNPs) with epigenomics, transcriptomics, 3D chromatin organization from diverse cell types as well as primary UF patient's samples. This integrative analysis identifies 24 UF-associated risk loci that potentially target 394 genes, of which 168 are differentially expressed in UF tumors. Critically, integrating this data with single-cell gene expression data from UF patients reveales the causal cell types with aberrant expression of these target genes. Lastly, CRISPR-based epigenetic repression (dCas9-KRAB) or activation (dCas9-p300) in a UF disease-relevant cell type further refines and narrows down the potential gene targets. Our findings and the methodological approach indicate the effectiveness of integrating multi-omics data with locus-specific epigenetic editing approaches for identifying gene- and celt type-targets of disease-relevant risk loci.

Hepatocyte-specific regulation of autophagy and inflammasome activation via MyD88 during lethal Ehrlichia infection.

Hepatocytes play a crucial role in host response to infection. Ehrlichia is an obligate intracellular bacterium that causes potentially life-threatening human monocytic ehrlichiosis (HME) characterized by an initial liver injury followed by sepsis and multi-organ failure. We previously showed that infection with highly virulent Ehrlichia japonica (E. japonica) induces liver damage and fatal ehrlichiosis in mice via deleterious MyD88-dependent activation of CASP11 and inhibition of autophagy in macrophage. While macrophages are major target cells for Ehrlichia, the role of hepatocytes (HCs) in ehrlichiosis remains unclear. We investigated here the role of MyD88 signaling in HCs during infection with E. japonica using primary cells from wild-type (WT) and MyD88-/- mice, along with pharmacologic inhibitors of MyD88 in a murine HC cell line. Similar to macrophages, MyD88 signaling in infected HCs led to deleterious CASP11 activation, cleavage of Gasdermin D, secretion of high mobility group box 1, IL-6 production, and inflammatory cell death, while controlling bacterial replication. Unlike macrophages, MyD88 signaling in Ehrlichia-infected HCs attenuated CASP1 activation but activated CASP3. Mechanistically, active CASP1/canonical inflammasome pathway negatively regulated the activation of CASP3 in infected MyD88-/- HCs. Further, MyD88 promoted autophagy induction in HCs, which was surprisingly associated with the activation of the mammalian target of rapamycin complex 1 (mTORC1), a known negative regulator of autophagy. Pharmacologic blocking mTORC1 activation in E. japonica-infected WT, but not infected MyD88-/- HCs, resulted in significant induction of autophagy, suggesting that MyD88 promotes autophagy during Ehrlichia infection not only in an mTORC1-indpenedent manner, but also abrogates mTORC1-mediated inhibition of autophagy in HCs. In conclusion, this study demonstrates that hepatocyte-specific regulation of autophagy and inflammasome pathway via MyD88 is distinct than MyD88 signaling in macrophages during fatal ehrlichiosis. Understanding hepatocyte-specific signaling is critical for the development of new therapeutics against liver-targeting pathogens such as Ehrlichia.

Engineered MED12 mutations drive leiomyoma-like transcriptional and metabolic programs by altering the 3D genome compartmentalization.

Nearly 70% of Uterine fibroid (UF) tumors are driven by recurrent MED12 hotspot mutations. Unfortunately, no cellular models could be generated because the mutant cells have lower fitness in 2D culture conditions. To address this, we employ CRISPR to precisely engineer MED12 Gly44 mutations in UF-relevant myometrial smooth muscle cells. The engineered mutant cells recapitulate several UF-like cellular, transcriptional, and metabolic alterations, including altered Tryptophan/kynurenine metabolism. The aberrant gene expression program in the mutant cells is, in part, driven by a substantial 3D genome compartmentalization switch. At the cellular level, the mutant cells gain enhanced proliferation rates in 3D spheres and form larger lesions in vivo with elevated production of collagen and extracellular matrix deposition. These findings indicate that the engineered cellular model faithfully models key features of UF tumors and provides a platform for the broader scientific community to characterize genomics of recurrent MED12 mutations.

Prevalence of Vancomycin-resistant enterococci (VRE) in Egypt (2010-2022): a systematic review and meta-analysis.

BACKGROUND: Vancomycin-resistant Enterococci (VRE) represent a critical medical and public health concerns due to their association with serious nosocomial infections and a high risk of mortality. We aimed to reveal the pooled prevalence of VRE and antimicrobial resistance profiles among enterococci clinical isolates in Egypt. METHODS: A PubMed, Scopus, Google Scholar, and Web of Science literature search was carried out in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guideline. Only published studies documenting the prevalence of VRE between 2010 and 2022 were included. Using the random effects model and the 95% confidence intervals, the pooled estimate of VRE was calculated by MedCalc Version 20.113. Cochran's Q and I2 tests were used to evaluate the degree of heterogeneity, and publication bias was examined by visually examining the funnel plot and its associated tests (Begg's and Egger's tests). RESULTS: The pooled prevalence of VRE among enterococci clinical isolates in Egypt was estimated to be 26% (95% CI 16.9 to 36.3). E. faecalis had a greater pooled prevalence than E. faecium, with 61.22% (95% CI 53.65 to 68.53) and 32.47% (95% CI 27 to 38.2), respectively. The VanA gene is more frequent than the VanB gene among VRE, with a pooled prevalence of 63.3% (95% CI 52.1 to 73.7) and 17.95% (95% CI 7.8 to 31), respectively. The pooled resistance rate of linezolid was substantially lower than that of ampicillin and high-level gentamicin (HLG) 5.54% (95% CI 2.33 to 10%), 65.7% (95% CI 50.8 to 79.2%), and 61.1% (95% CI 47.4 to 73.9), respectively. CONCLUSION: The prevalence of VRE is alarmingly high in Egypt. It is imperative that antimicrobial stewardship activities and infection control programs are strictly adhered to and implemented to prevent further escalation of the problem.

p120-Catenin suppresses NLRP3 inflammasome activation in macrophages.

Inflammasome activation is of central importance for the process of generation of overwhelming inflammatory response and the pathogenesis of sepsis. The intrinsic molecular mechanism for controlling inflammasome activation is still poorly understood. Here we investigated the role of p120-catenin expression in macrophages in regulating nucleotide-binding oligomerization domain (NOD) and leucine-rich repeat (LRR)- and pyrin domain-containing proteins 3 (NLRP3) inflammasome activation. Depletion of p120-catenin in murine bone marrow-derived macrophages enhanced caspase-1 activation and secretion of active interleukin (IL)-1ß in response to ATP stimulation following LPS priming. Coimmunoprecipitation analysis showed that p120-catenin deletion promoted NLRP3 inflammasome activation by accelerating the assembly of the inflammasome complex comprised of NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), and pro-caspase-1. Depletion of p120-catenin also increased the production of mitochondrial reactive oxygen species. Pharmacological inhibition of mitochondrial reactive oxygen species nearly completely abolished NLRP3 inflammasome activation, caspase-1 activation, and the production of IL-1ß in p120-catenin-depleted macrophages. Furthermore, p120-catenin ablation significantly disrupted mitochondrial function, evidenced by decreased mitochondrial membrane potential and lower production of intracellular ATP. In alveolar macrophage-depleted mice challenged with cecal ligation and puncture, pulmonary transplantation of p120-catenin-deficient macrophages dramatically enhanced the accumulation of IL-1ß and IL-18 in bronchoalveolar lavage fluid. These results demonstrate that p120-catenin prevents NLRP3 inflammasome activation in macrophages by maintaining mitochondrial homeostasis and reducing the production of mitochondrial reactive oxygen species in response to endotoxin insult. Thus, inhibition of NLRP3 inflammasome activation by stabilization of p120-catenin expression in macrophages may be a novel strategy to prevent an uncontrolled inflammatory response in sepsis.

Comprehensive Review of Uterine Fibroids: Developmental Origin, Pathogenesis, and Treatment.

Uterine fibroids are benign monoclonal neoplasms of the myometrium, representing the most common tumors in women worldwide. To date, no long-term or noninvasive treatment option exists for hormone-dependent uterine fibroids, due to the limited knowledge about the molecular mechanisms underlying the initiation and development of uterine fibroids. This paper comprehensively summarizes the recent research advances on uterine fibroids, focusing on risk factors, development origin, pathogenetic mechanisms, and treatment options. Additionally, we describe the current treatment interventions for uterine fibroids. Finally, future perspectives on uterine fibroids studies are summarized. Deeper mechanistic insights into tumor etiology and the complexity of uterine fibroids can contribute to the progress of newer targeted therapies.

Gut and genital tract microbiomes: Dysbiosis and link to gynecological disorders.

Every year, millions of women are affected by genital tract disorders, such as bacterial vaginosis (BV), endometrial cancer, polycystic ovary syndrome (PCOS), endometriosis, and uterine fibroids (UFs). These disorders pose a significant economic burden on healthcare systems and have serious implications for health and fertility outcomes. This review explores the relationships between gut, vaginal, and uterine dysbiosis and the pathogenesis of various diseases of the female genital tract. In recent years, reproductive health clinicians and scientists have focused on the microbiome to investigate its role in the pathogenesis and prevention of such diseases. Recent studies of the gut, vaginal, and uterine microbiomes have identified patterns in bacterial composition and changes across individuals' lives associated with specific healthy and diseased states, particularly regarding the effects of the estrogen-gut microbiome axis on estrogen-driven disorders (such as endometrial cancer, endometriosis, and UFs) and disorders associated with estrogen deficiency (such as PCOS). Furthermore, this review discusses the contribution of vitamin D deficiency to gut dysbiosis and altered estrogen metabolism as well as how these changes play key roles in the pathogenesis of UFs. More research on the microbiome influences on reproductive health and fertility is vital.

Endocrine-Disrupting Chemicals and Vitamin D Deficiency in the Pathogenesis of Uterine Fibroids.

Uterine fibroids (UFs) are the most prevalent gynecologic neoplasm, affecting 70-80% of women over their lifespan. Although UFs are benign they can become life-threatening and require invasive surgeries such as myomectomy and hysterectomy. Notwithstanding the significant negative influence UFs have on female reproductive health, very little is known about early events that initiate tumor development. Several risk factors for UFs have been identified including vitamin D deficiency, inflammation, DNA repair deficiency, and environmental exposures to endocrine-disrupting chemicals (EDCs). EDCs have come under scrutiny recently due to their role in UF development. Epidemiologic studies have found an association between increased risk for early UF diagnosis and in utero EDC exposure. Environmental exposure to EDCs during uterine development increases UF incidence in a UF animal model. Notably, several studies demonstrated that abnormal myometrial stem cells (MMSCs) are the cell origin for UFs development. Our recent studies demonstrated that early-life EDC exposure reprogrammed the MMSCs toward a pro-fibroid landscape and altered the DNA repair and inflammation pathways. Notably, Vitamin D3 (VITD3) as a natural compound shrank the UF growth concomitantly with the reversion of several abnormal biological pathways and ameliorated the developmental exposure-induced DNA damage and pro-inflammation pathway in primed MMSCs. This review highlights and emphasizes the importance of multiple pathway interactions in the context of hypovitaminosis D at the MMSCs level and provides proof-of-concept information that can help develop a safe, long-term, durable, and non-surgical therapeutic option for UFs.

Nutrition in Gynecological Diseases: Current Perspectives.

Diet and nutrition are fundamental in maintaining the general health of populations, including women's health. Health status can be affected by nutrient deficiency and vice versa. Gene-nutrient interactions are important contributors to health management and disease prevention. Nutrition can alter gene expression, as well as the susceptibility to diseases, including cancer, through several mechanisms. Gynecological diseases in general are diseases involving the female reproductive system and include benign and malignant tumors, infections, and endocrine diseases. Benign diseases such as uterine fibroids and endometriosis are common, with a negative impact on women's quality of life, while malignant tumors are among the most common cause of death in the recent years. In this comprehensive review article, a bibliographic search was performed for retrieving information about nutrients and how their deficiencies can be associated with gynecological diseases, namely polycystic ovary syndrome, infertility, uterine fibroids, endometriosis, dysmenorrhea, and infections, as well as cervical, endometrial, and ovarian cancers. Moreover, we discussed the potential beneficial impact of promising natural compounds and dietary supplements on alleviating these significant diseases.

Exosomes as Biomarkers for Female Reproductive Diseases Diagnosis and Therapy.

Cell-cell communication is an essential mechanism for the maintenance and development of various organs, including the female reproductive system. Today, it is well-known that the function of the female reproductive system and successful pregnancy are related to appropriate follicular growth, oogenesis, implantation, embryo development, and proper fertilization, dependent on the main regulators of cellular crosstalk, exosomes. During exosome synthesis, selective packaging of different factors into these vesicles happens within the originating cells. Therefore, exosomes contain both genetic and proteomic data that could be applied as biomarkers or therapeutic targets in pregnancy-associated disorders or placental functions. In this context, the present review aims to compile information about the potential exosomes with key molecular cargos that are dysregulated in female reproductive diseases which lead to infertility, including polycystic ovary syndrome (PCOS), premature ovarian failure (POF), Asherman syndrome, endometriosis, endometrial cancer, cervical cancer, ovarian cancer, and preeclampsia, as well as signaling pathways related to the regulation of the reproductive system and pregnancy outcome during these pathological conditions. This review might help us realize the etiology of reproductive dysfunction and improve the early diagnosis and treatment of the related complications.

Vitamin D3 Ameliorates DNA Damage Caused by Developmental Exposure to Endocrine Disruptors in the Uterine Myometrial Stem Cells of Eker Rats.

Early-life exposure of the myometrium to endocrine-disrupting chemicals (EDCs) has been shown to increase the risk of uterine fibroid (UF) prevalence in adulthood. Vitamin D3 (VitD3) is a unique, natural compound that may reduce the risk of developing UFs. However, little is known about the role and molecular mechanism of VitD3 on exposed myometrial stem cells (MMSCs). We investigated the role of, and molecular mechanism behind, VitD3 action on DNA damage response (DDR) defects in rat MMSCs due to developmental exposure to diethylstilbestrol (DES), with the additional goal of understanding how VitD3 decreases the incidence of UFs later in life. Female newborn Eker rats were exposed to DES or a vehicle early in life; they were then sacrificed at 5 months of age (pro-fibroid stage) and subjected to myometrial Stro1+/CD44+ stem cell isolation. Several techniques were performed to determine the effect of VitD3 treatment on the DNA repair pathway in DES-exposed MMSCs (DES-MMSCs). Results showed that there was a significantly reduced expression of RAD50 and MRE11, key DNA repair proteins in DES-exposed myometrial tissues, compared to vehicle (VEH)-exposed tissues (p < 0.01). VitD3 treatment significantly decreased the DNA damage levels in DES-MMSCs. Concomitantly, the levels of key DNA damage repair members, including the MRN complex, increased in DES-MMSCs following treatment with VitD3 (p < 0.01). VitD3 acts on DNA repair via the MRN complex/ATM axis, restores the DNA repair signaling network, and enhances DDR. This study demonstrates, for the first time, that VitD3 treatment attenuated the DNA damage load in MMSCs exposed to DES and classic DNA damage inducers. Moreover, VitD3 targets primed MMSCs, suggesting a novel therapeutic approach for the prevention of UF development.

Diet-induced vitamin D deficiency triggers inflammation and DNA damage profile in murine myometrium.

BACKGROUND: Previously, we reported a significantly higher prevalence of uterine fibroids (UFs) in African American women. This minority group also commonly suffers from vitamin D deficiency. We have demonstrated that 1,25(OH)2D3 attains a fibroid growth inhibitory impact through its ability to block the G1/S (gap 1/synthesis) phase of the cell cycle. Vitamin D is involved in DNA damage as well as in immune response regulation, anti-inflammation, autoimmunity and cancer. Since most of the prior data on vitamin D and UF were generated in vitro via established cell lines, it was necessary to verify and validate this observation in vivo using a diet-induced vitamin D-deficient mouse model. MATERIALS AND METHODS: Our model of vitamin D lacking function was established using 8-week exposure of C57/BL6 mice to vitamin D-deficient diet provides evidence of different functions accomplished by vitamin D in the regulation of myometrium homeostasis disrupted in the context of uterine fibroid. RESULTS: We found that vitamin D deficiency was associated with increased expression of sex steroid receptors in murine myometrium, increased expression of proliferation related genes, the promotion of fibrosis and enhanced inflammation, and promoted immunosuppression through Tregs expansion in murine myometrium. We also showed that a vitamin D deficient diet enhanced DNA damage in murine myometrium. CONCLUSION: Our model mimics the effects in humans that a lack of vitamin D has and propels the study of in vivo interaction between inflammation, genomic instability and cell proliferation in the myometrium.