Search Results (173)

Thymic epithelial cells (TECs) provide essential signals for T-cell development, yet the mechanisms governing human TEC differentiation and the acquisition of T-cell-supporting function remain incompletely understood. Using a three-stage system that directs human embryonic stem cells (hESCs) through definitive endoderm and third pharyngeal pouch endoderm (3PPE) toward a thymic epithelial-like lineage, we investigated the role of HOXA3 during thymic epithelial-like cell (TEC-like cell) differentiation. HOXA3 knockdown during 3PPE-to-TEC-like cell transition impaired thymic epithelial differentiation, as indicated by reduced K8, AIRE, and CD205 expression and decreased FOXN1 protein. HOXA3-knockdown TEC-like cells also showed a markedly reduced capacity to support the generation of CD45⁺CD3⁺ cells from cord blood CD34⁺ progenitors. Bulk RNA sequencing identified 1040 upregulated and 774 downregulated genes, revealing broad remodeling of cell-cycle, motility, chemotaxis, angiogenesis, and tissue-remodeling programs. CXCL12 was among the downregulated genes, and its reduction was confirmed at both mRNA and protein levels. Motif analysis identified a putative HOXA3-binding site, and conventional ChIP-PCR showed enrichment of the predicted CXCL12 regulatory region in HOXA3 immunoprecipitates. Recombinant CXCL12 significantly increased CD45⁺CD3⁺ cell generation in HOXA3-knockdown co-cultures. Together, these data support an association between HOXA3 knockdown, reduced acquisition of a thymic epithelial-like phenotype, and diminished early T-lineage support, with CXCL12 partially accounting for the functional phenotype. This study provides a tractable hESC-based model for studying human thymic epithelial development and TEC-mediated early T-lineage differentiation. Full article

Recurrent pregnancy loss (RPL), a multifactorial condition in reproductive medicine, affects approximately 1–3% of couples. Increasing evidence implicates endometrial dysfunction, immune dysregulation, impaired angiogenesis, and oxidative stress (OS) in its pathophysiology, highlighting the need for biologically targeted therapeutic strategies. In this context, platelet-rich plasma (PRP) has emerged as a regenerative approach with the potential to modulate these underlying mechanisms. This narrative review critically evaluates the biological rationale and current clinical evidence supporting PRP as an adjunctive treatment for RPL while proposing a precision medicine framework for biologically guided patient selection. A comprehensive narrative synthesis of the literature was performed, focusing on studies investigating PRP in RPL and related reproductive conditions, including thin endometrium and recurrent implantation failure (RIF). Mechanistically, PRP promotes angiogenesis through vascular endothelial growth factor (VEGF)-mediated pathways, enhances endometrial regeneration by stimulating stromal and epithelial cell proliferation, and modulates immune responses by promoting regulatory T-cell activity while attenuating pro-inflammatory signaling. In addition, PRP-derived extracellular vesicles (EVs) and microRNAs may contribute to the post-transcriptional and epigenetic regulation of implantation-related genes, including leukemia inhibitory factor (LIF) and HOXA10. Clinical studies in RIF and thin endometrium populations suggest that PRP may improve endometrial thickness, implantation, and clinical pregnancy outcomes; however, these findings constitute indirect evidence for RPL. Direct RPL-specific evidence remains very limited and is insufficient to establish a reduction in miscarriage or an improvement in live birth. These biological features instead provide a rationale for investigating PRP in defined patient subgroups characterized by impaired endometrial receptivity, defective angiogenesis, immune dysregulation, or unexplained RPL with suspected endometrial dysfunction. Accordingly, we propose that future clinical investigation of PRP should move beyond empirical use toward biomarker-informed, precision reproductive medicine strategies. Although PRP represents a biologically plausible and promising adjunctive therapy, robust randomized controlled trials incorporating standardized PRP protocols and biologically stratified patient populations are required before its routine clinical use can be recommended. Full article

Exogenous gonadotropin-based estrus synchronization is widely used in livestock reproduction. However, this hormonal treatment has been associated with adverse reproductive outcomes, suggesting that it may alter the uterine environment and endometrial receptivity, although the underlying molecular mechanism remains poorly understood. Here, we investigated the endometrial responses to gonadotropin-induced estrus synchronization using both in vivo and in vitro models. We found that gonadotropin treatment caused structural and inflammatory changes in the endometrium of both pigs and mice, accompanied by elevated circulating estradiol (E₂) levels and enhanced NLRP3 inflammasome-related signaling. In mice, gonadotropin treatment also altered the expression of MUC1, Hand2, and HoxA11, indicating disruption of molecular features associated with endometrial receptivity. In porcine endometrial epithelial cells, supraphysiological E₂ exposure induced mitochondrial oxidative stress, characterized by increased mitochondrial reactive oxygen species production, loss of mitochondrial membrane potential, increased cellular mtDNA abundance, and oxidative DNA damage. E₂ treatment further promoted NLRP3 inflammasome-associated signaling and ASC speck formation, whereas antioxidant N-acetyl-L-cysteine treatment attenuated ASC speck formation. Collectively, these findings support a link between high E₂ exposure, mitochondrial oxidative stress, and enhanced NLRP3 inflammasome signaling in endometrial epithelial cells. Our results suggest that an E₂–mitochondrial oxidative stress–NLRP3 axis may contribute to gonadotropin-associated endometrial dysfunction and provide a basis for further investigation of antioxidant and inflammasome-targeted strategies in livestock reproduction. Full article

Background/Objectives: Genetic and epigenetic alterations accumulate throughout life in hematopoietic stem and progenitor cells (HSPCs), leading to an increased risk of age-related hematological malignancies, such as acute myeloid leukemia (AML). While a few epigenetic drugs have entered clinical practice, studies focusing on histone post-translational modifications (PTMs) altered in AML remain limited. Methods: Here, we used murine HSPCs, murine AML mouse models, human leukemic cell lines, and leukemic patient samples to investigate whether targeting the methylation of histone 3 at lysine 9 (H3K9 methylation) affects HSPCs upon aging and leukemogenesis and might represent a possible target. Results: Our data show that H3K9 methylation changes upon aging in HSPCs and is linked to a pre-malignant phenotype. Low levels of H3K9 methylation in leukemic cells are required to maintain proliferative capacities in vitro. Increasing H3K9 methylation reduces the leukemogenesis of both young and aged murine and patient-derived leukemic cells. Conclusions: Thus, H3K9 methylation might be a potential and selective therapeutic target in AML patients. Full article

Background: Early detection of hepatocellular carcinoma (HCC) is critical for improving patient outcomes; however, ultrasound-based surveillance has limited sensitivity and variable performance across patient populations. We evaluated the performance of a multitarget HCC blood test (mt-HBT), incorporating methylated DNA markers, alpha-fetoprotein (AFP), and patient sex, across clinically relevant subgroups. Methods: We performed a subgroup analysis of a multicenter, prospective case-control study that included 159 patients with early-stage HCC (Barcelona Clinic Liver Cancer Stage 0/A) and 649 control patients with cirrhosis or chronic hepatitis B without HCC. The mt-HBT combined methylated HOXA1, TSPYL5, and B3GALT6 markers with AFP and sex. Sensitivity and specificity were evaluated overall and according to age, sex, obesity, liver disease etiology, Child Pugh class, and tumor size. Performance was compared with AFP and GALAD. Results: Overall sensitivity and specificity of mt-HBT for early-stage HCC detection were 76.7% (95% CI, 69.6–82.6) and 87.5% (95% CI, 84.8–89.8), respectively. Sensitivity was significantly higher than that of AFP (35.2%, p < 0.001) and comparable to that of GALAD (78.6%, p = 0.56), whereas specificity was lower than that of AFP (98.5%, p < 0.001) but higher than that of GALAD (76.9%, p < 0.001). Sensitivity was maintained across key subgroups, including patients with obesity (68.9%), Child Pugh B cirrhosis (75.0%), hepatitis C (80.0%), hepatitis B (72.2%), alcohol-associated liver disease (80.5%), and metabolic dysfunction-associated steatotic disease (69.0%) (all p > 0.05 between subgroups). Specificity exceeded 80% in all examined populations and was significantly higher in women than in men (92.5% vs. 83.9%, p < 0.001). Sensitivity increased with tumor size, ranging from 60.0% for tumors < 2 cm to 100% for tumors > 5 cm. Conclusions: The mt-HBT demonstrated robust, consistent performance for early-stage HCC detection across diverse patient populations, including subgroups in which ultrasound surveillance commonly underperforms. These findings support the prospective validation of mt-HBT as a blood-based surveillance strategy for HCC. Full article

Advances in the molecular characterization of acute myeloid leukemia (AML) in recent decades have driven the development and clinical introduction of several targeted therapies, and menin inhibitors are the most recent additions to the therapeutic arsenal of AML treatments. The KMT2A and NPM1 loci have been the focus of intense investigation over the past two decades because of their well-established roles leukemogenesis, particularly in pre-clinical models of AML involving human xenografts. The ability of KMT2A and NPM1 aberrancies to drive and sustain leukemogenesis has provided a strong rationale for the development of therapeutic strategies to disrupt these molecular pathways. Menin inhibitors preferentially eliminate clones harboring KMT2A rearrangements or NPM1 mutations due to the dependence of these leukemia cells on the menin–KMT2A complex and its downstream HOXA9/MEIS signaling to sustain leukemic self-renewal and disease maintenance. In this review, we discuss the biological rationale for menin–MLL disruption in patients with acute leukemia with KMT2A rearrangement or NPM1 mutation. We explore data from the landmark AUGMENT-101 and KOMET-001 trials, which have led to regulatory approval of revumenib and ziftomenib for select patients. We also explore cutting-edge studies using menin inhibitors in combination strategies, including data from KOMET-007. The evidence supporting combination approaches remains based on early-phase, single-arm trials with relatively short follow-up, and longer-term and comparative data are needed to define their clinical benefit. We highlight resistance mechanisms that have emerged from the use of contemporary menin inhibitors and efforts aimed at addressing this resistance, with a focus on emerging clinical data on enzomenib and bleximenib. Finally, we discuss prospects on the putative role of menin inhibitors in the measurable residual disease (MRD)-positive and maintenance settings in AML. Menin inhibitors have successfully transitioned from biological proof-of-concept to approved targeted therapy, and future advances will depend on rational front line combination strategies, MRD-guided treatment, post-transplant maintenance strategies, and therapeutic sequencing informed by mechanisms of resistance. Full article

As a high-value meat duck breed, Muscovy ducks exhibit marked individual differences in egg production that limit the economic efficiency of commercial farming. To elucidate the underlying mechanisms, we integrated phenotypic observation, ovarian transcriptome sequencing, and cecal 16S rRNA gene amplicon sequencing to compare high- and low-laying black-feathered Muscovy duck populations. We recorded 40-day egg production, assessed ovarian morphology, quantified gene expression profiles, and characterized microbial community structure. High-laying ducks produced significantly more eggs (24.60 ± 9.22) than low-laying ducks (15.80 ± 6.64, p < 0.01), with larger ovaries and a greater number of follicles at all developmental stages. Transcriptomic analysis revealed 823 DEGs, with KEGG enrichment implicating pathways governing ovarian physiology, including MAPK, Calcium, Notch, and Wnt signaling. Microbial profiling demonstrated significant differences in α/β diversity between groups, with high-laying ducks exhibiting an elevated Firmicutes/Bacteroidota ratio and decreased abundance of Ligilactobacillus. Integrative correlation analysis identified significant associations between Ligilactobacillus abundance and the expression of Arhgap27 and HOXA10, two genes linked to reproductive function, and highlighted the glycolysis/gluconeogenesis pathway as a convergently enriched signaling cascade shared between gut microbiome and ovarian transcriptome. This multi-omics study uncovers the molecular and microbial basis of egg production variation in Muscovy ducks, offering potential targets for improving breeding efficiency through microbiota-directed or gene-based interventions. These findings deepen our understanding of host–microbe interactions in poultry reproduction and identify candidate genes and microbial taxa that may serve as potential targets for future nutritional or genetic interventions to enhance laying performance in commercial duck production. Full article

Micro- and nanoplastics (MNPs) are emerging environmental pollutants, but evidence concerning their effects on endometrial injury and adverse pregnancy outcomes remains limited. Therefore, we employed the Targeted Risk Assessment of Environmental Chemicals (TRAEC) strategy to integrate evidence concerning endometrial injury and adverse pregnancy outcomes associated with exposure to MNPs. Following targeted screening, 21 unique studies yielded 24 literature-derived evidence entries. Together with one additional in vitro evidence item from the present study, 25 evidence items were included in the final CES calculation. The available evidence was quantitatively integrated across four predefined TRAEC dimensions: Reliability Scores, Weight of Concentrations, Risk Intensity, and Correlation Scores. Under the tested in vitro conditions, exposure to PS-NPs increased intracellular ROS accumulation, reduced mitochondrial membrane potential, promoted apoptosis, and decreased the expression of key endometrial receptivity proteins, namely HOXA10, FOXO1, and ITGB3, in primary human endometrial stromal cells. Based on this combined evidence pool, the final Comprehensive Evidence Score was 7.67, corresponding to a preliminary moderate-risk classification within the predefined TRAEC framework. In summary, the TRAEC strategy provides a structured approach for integrating multi-level evidence and deriving a framework-specific health-risk classification for exposure to MNPs. Full article

Histone-modifying enzymes and histone post-translational modifications (PTMs) play key roles in the organization (euchromatin versus heterochromatin) and function (active versus silenced genes) of chromatin. The abundance and activity of these enzymes, along with their associated histone PTMs, are often altered in cancer cells, leading to deregulated gene expression. The expression of the KMT2A-MLLT3 protein, resulting from a chromosomal translocation in mixed-lineage leukemia (MLL), a subtype of acute myeloid leukemia, augments transcription elongation, promoting the expression of HOXA9 and MEIS1, genes that play critical roles in MLL development. Bortezomib, a proteasome inhibitor, has been effective at treating various cancers. In this study, we compared the impact of bortezomib on histone PTMs in the MLL cell line MOLM-13 and the chronic myeloid leukemic (CML) cell line K562. We report that MOLM-13 had a greater level of histone H2B monoubiquitinated at lysine 120 (H2BK120ub) and histone H3 dimethylated at lysine 79 (H3K79me2) (modifications involved in elongation) and similar levels of histone H2A monoubiquitinated at lysine 119 (H2AK119ub). Bortezomib treatment resulted in significant reductions in H2BK120ub and H2AK119ub levels, as well as in transcript levels of genes involved in MLL development. Full article

(1) Background: KMT2A (alias MLL) is located at chromosome 11q23 and encodes histone methyltransferase 2A which activates target genes via chromatin methylation of histone H3 lysine K4. Mutations of KMT2A, including partial tandem duplication (PTD) and fusion with partner genes, are present in both lymphoid and myeloid acute leukemia. More than 100 various KMT2A fusion genes have been described, with only a minority represented by cell line models. (2) Methods: Cytogenetic and genomic copy number analyses, PCR, Western blot and RNA-sequencing were performed to characterize aberrations in acute myeloid leukemia (AML) cell line KOPM-88. Bioinformatic analysis of public AML patient data revealed differentially expressed genes. Functional analyses were performed by siRNA-mediated knockdown and live-cell imaging. (3) Results: The AML cell line KOPM-88 is derived from a boy at relapse and has been reported to carry t(X;11)(q24;q23), albeit with uncharacterized breakpoints. In KOPM-88 we identified fusion gene KMT2A::SEPTIN6 generated by this translocation, but excluded KMT2A-PTD. KMT2A::SEPTIN6 activated bone morphogenetic protein (BMP)-signalling and inhibited expression of HOXA7 and HOXA9. BMP signalling in turn activated cell proliferation and inhibited CDKN2B expression. (4) Conclusions: KOPM-88 contains fusion gene KMT2A::SEPTIN6, representing the only cell line model for this rare type of KMT2A rearrangement. KOPM-88 may serve to advance novel therapeutic treatments for KMT2A::SEPTIN6-positive AML. Full article

Sepsis is a life-threatening condition characterized by a dysregulated host response to infection, leading to multi-organ dysfunction. Toll-like receptor signaling via MYD88- and TRIF-dependent pathways plays a central role in this process; however, its temporal and tissue-specific dynamics remain incompletely understood. The aim of this study was to investigate time-dependent transcriptional changes in MYD88- and TRIF-dependent signaling pathways across multiple organs in a murine model of sepsis. mRNA expression of MYD88, IRAK1, IRAK4, NF-kB, CCL4, CCL20, CCR2, IFN-β, IFN-γ, TNF-α, IL-1β, IL-2, IL-4, IL-8, IL-10, IL-18, Klotho, KLF4, HOXA5, NANOG and HIF1α was quantified using qRT-PCR in intestinal, kidney, liver and lung tissues at 24, 48, and 72 h following cecal ligation and puncture-induced sepsis in male C57BL/6J mice. Significant upregulation of innate immune signaling molecules, cytokines, chemokines, and interferon-related genes was observed in all tissues compared with controls. Genes associated with hypoxia and cellular regulation were also increased. These responses were tissue-specific and progressively intensified over time. Sepsis represents a dynamic, time-dependent, and tissue-specific process characterized by sustained activation of immune and hypoxic pathways, providing potential targets for time-stratified therapeutic strategies. Full article

Among the transcriptional dependencies that sustain leukemic identity in acute myeloid leukemia (AML), the menin–KMT2A chromatin complex has emerged as a central regulatory node. The scaffold protein menin, encoded by MEN1, facilitates transcriptional activation of HOX and MEIS family genes during normal hematopoietic development. In AML, this physiologic and developmentally regulated role is co-opted to sustain constitutive HOX/MEIS-driven programs that block differentiation and maintain leukemic potential. Although dependency on menin is most clearly established in KMT2A-rearranged and NPM1-mutated AML, this vulnerability appears to arise from a shared transcriptional state characterized by persistent HOX activation rather than from any single genetic alteration. Pharmacologic disruption of the menin-KMT2A interaction collapses stemness-associated transcriptional networks, promotes myeloid differentiation, and attenuates leukemic self-renewal. Clinical activity observed with menin inhibitors provides translational validation of this dependency and establishes menin inhibition as a differentiation-based therapeutic strategy. In this review, we examine the molecular basis of menin-dependent transcriptional regulation in AML and its implications for therapeutic targeting with menin inhibitors and resistance to therapy. Full article

Microtia–atresia is a rare craniofacial malformation primarily affecting the first and second pharyngeal arches, leading to the deformity of the auricle and atresia of the external ear canal. Its etiology is heterogenous and largely unknown, including both genetic and environmental factors. The HOXA4 gene has been identified as potentially pathogenetic for microtia–atresia in three twin families. A hoxa4a mosaic knockdown zebrafish model was constructed using CRISPR/Cas9. hoxa4a was expressed in the mandible during early development in zebrafish, while the F0 mosaic knockdowns exhibited craniofacial malformations with abnormal chondrocyte morphologies. Specifically, hoxa4a knockdown reduced cranial neural crest cell proliferation while increasing apoptosis, markedly downregulating chondrogenic markers sox9a and col2a1a. Consequently, pharyngeal arch chondrocytes exhibited disorganized arrangement and morphological abnormalities, resulting in mandibular hypoplasia. Our findings provide important insights into the role of hoxa4a in zebrafish mandibular development and the pathology of microtia–atresia caused by HOXA4 gene mutations in humans. Full article

Background/Objectives: Aging is increasingly recognized as a key determinant of changes in human tissue and cellular function. Women’s age, in particular, has been associated with reduced oocyte quality and negatively correlated with the expression of genes involved in endometrial decidualization and cellular function. The ability of endometrial cells to interact and allow the invasion of the growing embryo is defined as endometrial receptivity. Investigating age-related differences in human endometrial receptivity may expand our understanding of factors contributing to infertility. Methods: Stromal cells were isolated and cultured from endometrial pipelle biopsies (n = 28) obtained from female donors at the proliferative phase of the menstrual cycle. Protein and mRNA expression of the receptivity modulators OPN, CD44, and HOXA10 were analyzed by Western blot and real-time PCR, respectively. Results: Data presented a linear decrease in mRNA expression of OPN and HOXA10 (p = 0.0066, R² = 0.3018 and p = 0.0036, R² = 0.529, respectively) with women’s increasing age, and a similar trend was evident at the protein level (OPN, p < 0.05; HOXA10, p < 0.01). Further analysis of the data included separating the samples into three age groups: 25–35 years, 36–40 years, and 41–46 years. ANOVA revealed a significant decrease in OPN and HOXA10 mRNA expression (p = 0.03158 and p = 0.02578, respectively). CD44 expression did not differ with age. Conclusions: OPN and HOXA10 are negatively correlated with increasing maternal age. These findings suggest that age-related alterations in key endometrial receptivity modulators may contribute to impaired implantation and could represent potential targets for diagnostic or therapeutic strategies in human implantation failure. Full article

Background: Chromosomal rearrangements involving lysine methyltransferase 2A (KMT2A) define a genetically distinct subset of acute myeloid leukemia (AML) in 10% of cases in adult patients; the frequency of KMT2A-r is higher in pediatric AML. Translocations involving the KMT2A locus at chromosome 11q23 result in the formation of a chimeric oncogene partner, where the N-terminal part of KMT2A is fused to a variety of translocation partners. The leukemogenic activity of KMT2A-fusion partners is related to their capacity to hyperactivate the expression of HOX-A and MEIS1 target genes, which stimulate the proliferation of hematopoietic stem cells. The oncogenic activity of KMT2A fusion proteins requires the binding with Menin, and this interaction can be targeted pharmacologically by small molecules acting as potent and selective Menin inhibitors. Methods: A search of the literature showed a marked development of experimental studies exploring the molecular pathogenesis of AML with KMT2A-r and of clinical studies evaluating new induction intensive treatments and the development of a targeted therapy based on Menin inhibitors. Results and Conclusions: In the present review article, we summarize our current understanding of the biology of KMT2A-r in AML development and the recent consistent progress made in the treatment of KMT2A-r AML through new chemotherapy regimens and targeted therapy using Menin inhibitors. However, the prognosis of older KMT2A-r AML patients remains poor and could be improved by drug combination studies including Menin inhibitors. Many encouraging observations derived from ongoing clinical trials with Menin inhibitors need to be confirmed through randomized clinical trials. Full article