In silico analysis predicts mutational consequences of CITED2, NUDT4, and Ar18B in patients with bipolar disorder.

Bipolar disorder is a mood-related disorder, which can be portrayed as extreme shifts in energy, mood, and activity levels which can also be characterized by manic highs and depressive lows that can be often misdiagnosed as unipolar disorder due to primitive diagnostics techniques based on clinical assessments as well as diagnostic complexities arising due to its heterogeneous nature and overlapping symptoms with conditions like schizophrenia. leading to delays in treatment Strong evidence in support of genetic and epigenetic aspects of bipolar disorder, including mechanisms such as compromised hypothalamic-pituitary-adrenal axis, immune-inflammatory imbalances, oxidative stress, and mitochondrial dysfunction are found. Moreover, some previous research has already stated the role of genes like CITED2, NUDT4, and Arl8B in these processes. The primary goal of this study is to investigate the involvement of the genes in exploring and validating their potential as biomarkers for bipolar disorder. In silico tools like MutationTaster, PolyPhen2, SIFT, GTEx, PhenoScanner, and RegulomeDB were used to perform mutational and gene expression analyses. Results revealed potentially dangerous mutations caused in CITED2, NUDT4, and Arl8B, those which can have diverse outcomes. RegulomeDB, GTEx, and PhenoScanner reveal the involvement of these genes in various brain regions highlighting their relevance to bipolar disorder. This analysis suggests the potential utility of CITED2, NUDT4, and Arl8B as diagnostic markers hence shedding light on their roles to elaborate the molecular range of bipolar disorder. The study also contributes to providing valuable insights into the genetic and molecular basis of bipolar disorders.

Human DCP1 is crucial for mRNA decapping and possesses paralog-specific gene regulating functions.

The mRNA 5'-cap structure removal by the decapping enzyme DCP2 is a critical step in gene regulation. While DCP2 is the catalytic subunit in the decapping complex, its activity is strongly enhanced by multiple factors, particularly DCP1, which is the major activator in yeast. However, the precise role of DCP1 in metazoans has yet to be fully elucidated. Moreover, in humans, the specific biological functions of the two DCP1 paralogs, DCP1a and DCP1b, remain largely unknown. To investigate the role of human DCP1, we generated cell lines that were deficient in DCP1a, DCP1b, or both to evaluate the importance of DCP1 in the decapping machinery. Our results highlight the importance of human DCP1 in decapping process and show that the EVH1 domain of DCP1 enhances the mRNA-binding affinity of DCP2. Transcriptome and metabolome analyses outline the distinct functions of DCP1a and DCP1b in human cells, regulating specific endogenous mRNA targets and biological processes. Overall, our findings provide insights into the molecular mechanism of human DCP1 in mRNA decapping and shed light on the distinct functions of its paralogs.

MYB/MYBL1-altered gliomas frequently harbor truncations and non-productive fusions in the MYB and MYBL1 genes.

Astrocytomas that harbor recurrent genomic alterations in MYB or MYBL1 are a group of Pediatric-type diffuse low-grade gliomas that were newly recognized in the 2021 WHO Classification of Tumors of the Central Nervous System. These tumors are described in the WHO classification as harboring fusions in MYB or MYBL1. In this report, we examine 14 consecutive cases in which a MYB or MYBL1 alteration was identified, each with diagnostic confirmation by genome-wide DNA methylation profiling (6 Angiocentric gliomas and 8 Diffuse astrocytomas, MYB- or MYBL1-altered), for their specific genomic alterations in these genes. Using RNA sequencing, we find productive in-frame fusions of the MYB or MYBL1 genes in only 5/14 cases. The remaining 9 cases show genomic alterations that result in truncation of the gene, without evidence of an in-frame fusion partner. Gene expression analysis showed overexpression of the MYB(L1) genes, regardless of the presence of a productive fusion. In addition, QKI, a recognized fusion partner common in angiocentric glioma, was generally up-regulated in these 14 cases, compared to a cohort comprising >1000 CNS tumors of various types, regardless of whether a genomic alteration in QKI was present. Overall, the results show that truncations, in the absence of a productive fusion, of the MYB(L1) genes can likely drive the tumors and have implications for the analysis and diagnosis of Angiocentric glioma and Diffuse astrocytoma, MYB- or MYBL1-altered, especially for cases that are tested on panels designed to focus on fusion detection.

Interactions between Helcococcus kunzii and Staphylococcus aureus: How a commensal bacterium modulates the virulence and metabolism of a pathogen in a chronic wound in vitro model.

BACKGROUND: Staphylococcus aureus is the predominant pathogen isolated in diabetic foot infections. Recently, the skin commensal bacterium, Helcococcus kunzii, was found to modulate the virulence of this pathogen in an in vivo model. This study aims to elucidate the molecular mechanisms underlying the interaction between these two bacterial species, using a proteomic approach. RESULTS: Our results reveal that H. kunzii can coexist and proliferate alongside S. aureus in a Chronic Wound Media (CWM), thereby mimicking an in vitro chronic wound environment. We noted that the secreted proteome of H. kunzii induced a transcriptional effect on S. aureus virulence, resulting in a decrease in the expression level of agrA, a gene involved in quorum sensing. The observed effect could be ascribed to specific proteins secreted by H. kunzii including polysaccharide deacetylase, peptidoglycan DD-metalloendopeptidase, glyceraldehyde-3-phosphate dehydrogenase, trypsin-like peptidase, and an extracellular solute-binding protein. These proteins potentially interact with the agr system, influencing S. aureus virulence. Additionally, the virulence of S. aureus was notably affected by modifications in iron-related pathways and components of cell wall architecture in the presence of H. kunzii. Furthermore, the overall metabolism of S. aureus was reduced when cocultured with H. kunzii. CONCLUSION: Future research will focus on elucidating the role of these excreted factors in modulating virulence.

Epigenetic regulation of p63 blocks squamous-to-neuroendocrine transdifferentiation in esophageal development and malignancy.

While cell fate determination and maintenance are important in establishing and preserving tissue identity and function during development, aberrant cell fate transition leads to cancer cell heterogeneity and resistance to treatment. Here, we report an unexpected role for the transcription factor p63 (Trp63/TP63) in the fate choice of the squamous versus neuroendocrine lineage in esophageal development and malignancy. Deletion of p63 results in extensive neuroendocrine differentiation in the developing mouse esophagus and esophageal progenitors derived from human embryonic stem cells. In human esophageal neuroendocrine carcinoma (eNEC) cells, p63 is transcriptionally silenced by EZH2-mediated H3K27 trimethylation (H3K27me3). Up-regulation of the major p63 isoform ΔNp63α, through either ectopic expression or EZH2 inhibition, promotes squamous transdifferentiation of eNEC cells. Together, these findings uncover p63 as a rheostat in coordinating the transition between squamous and neuroendocrine cell fates during esophageal development and tumor progression.

Molecular mechanism of proteolytic cleavage-dependent activation of CadC-mediated response to acid in E. coli.

Colonizing in the gastrointestinal tract, Escherichia coli confronts diverse acidic challenges and evolves intricate acid resistance strategies for its survival. The lysine-mediated decarboxylation (Cad) system, featuring lysine decarboxylase CadA, lysine/cadaverine antiporter CadB, and transcriptional activator CadC, plays a crucial role in E. coli's adaptation to moderate acidic stress. While the activation of the one-component system CadC and subsequent upregulation of cadBA operon in response to acid and lysine presence have been proposed, the molecular mechanisms governing the transition of CadC from an inactive to an active state remain elusive. Under neutral conditions, CadC is inhibited by forming a complex with lysine-specific permease LysP, stabilized in this inactive state by a disulfide bond. Our study unveils that, in an acidic environment, the disulfide bond in CadC is reduced by the disulfide bond isomerase DsbC, exposing R184 to periplasmic proteases, namely DegQ and DegP. Cleavage at R184 by DegQ and DegP generates an active N-terminal DNA-binding domain of CadC, which binds to the cadBA promoter, resulting in the upregulated transcription of the cadA and cadB genes. Upon activation, CadA decarboxylates lysine, producing cadaverine, subsequently transported extracellularly by CadB. We propose that accumulating cadaverine gradually binds to the CadC pH-sensing domain, preventing cleavage and activation of CadC as a feedback mechanism. The identification of DegP, DegQ, and DsbC completes a comprehensive roadmap for the activation and regulation of the Cad system in response to moderate acidic stress in E. coli.

The dual-specificity kinase DYRK1A interacts with the Hepatitis B virus genome and regulates the production of viral RNA.

The genome of Hepatitis B virus (HBV) persists in infected hepatocytes as a nuclear episome (cccDNA) that is responsible for the transcription of viral genes and viral rebound, following antiviral treatment arrest in chronically infected patients. There is currently no clinically approved therapeutic strategy able to efficiently target cccDNA (Lucifora J 2016). The development of alternative strategies aiming at permanently abrogating HBV RNA production requires a thorough understanding of cccDNA transcriptional and post-transcriptional regulation. In a previous study, we discovered that 1C8, a compound that inhibits the phosphorylation of some cellular RNA-binding proteins, could decrease the level of HBV RNAs. Here, we aimed at identifying kinases responsible for this effect. Among the kinases targeted by 1C8, we focused on DYRK1A, a dual-specificity kinase that controls the transcription of cellular genes by phosphorylating transcription factors, histones, chromatin regulators as well as RNA polymerase II. The results of a combination of genetic and chemical approaches using HBV-infected hepatocytes, indicated that DYRK1A positively regulates the production of HBV RNAs. In addition, we found that DYRK1A associates with cccDNA, and stimulates the production of HBV nascent RNAs. Finally, reporter gene assays showed that DYRK1A up-regulates the activity of the HBV enhancer 1/X promoter in a sequence-dependent manner. Altogether, these results indicate that DYRK1A is a proviral factor that may participate in the HBV life cycle by stimulating the production of HBx, a viral factor absolutely required to trigger the complete cccDNA transcriptional program.

Whole Exome Sequencing in Children With Type 1 Diabetes Before Age 6 Years Reveals Insights Into Disease Heterogeneity.

Aims: This study is aimed at comparing whole exome sequencing (WES) data with the clinical presentation in children with type 1 diabetes onset ≤ 5 years of age (EOT1D). Methods: WES was performed in 99 unrelated children with EOT1D with subsequent analysis to identify potentially deleterious rare variants in MODY genes. High-resolution HLA class II haplotyping, SNP genotyping, and T1D-genetic risk score (T1D-GRS) were also evaluated. Results: Eight of the ninety-nine EOT1D participants carried a potentially deleterious rare variant in a MODY gene. Rare variants affected five genes: GCK (n = 1), HNF1B (n = 2), HNF4A (n = 1), PDX1 (n = 2), and RFX6 (n = 2). At diagnosis, these children had a mean age of 3.0 years, a mean HbA1c of 10.5%, a detectable C-peptide in 5/8, and a positive islet autoantibody in 6/7. Children with MODY variants tend to exhibit a lower number of pancreatic autoantibodies and a lower fasting C-peptide compared to EOT1D without MODY rare variants. They also carried at least one high-risk DR3-DQ2 or DR4-DQ8 haplotype and exhibited a T1D-GRS similar to the other individuals in the EOT1D cohort, but higher than healthy controls. Conclusions: WES found potentially deleterious rare variants in MODY genes in 8.1% of EOT1D, occurring in the context of a T1D genetic background. Such genetic variants may contribute to disease precipitation by a ß-cell dysfunction mechanism. This supports the concept of different endotypes of T1D, and WES at T1D onset may be a prerequisite for the implementation of precision therapies in children with autoimmune diabetes.

Identification of a novel EYA4 likely pathogenic variant in a Chinese family with postlingual non-syndromic hearing loss and analysis of molecular epidemiology of EYA4 variants.

BACKGROUND: EYA4 variants are responsible for DFNA10 deafness. Due to its insidious onset and slow progression, hearing loss in autosomal dominant non-syndromic hearing loss (ADNSHL) is usually challenging to detect early in clinical settings, with limited intervention options. Genetic testing can aid in early detection of hearing loss, enabling timely intervention to reduce disability rates and improve the quality of life. METHODS: In this study, we report the case of a Chinese family with postlingual and progressive hearing loss that was passed down for four generations. Whole-exome sequencing (WES) was performed on DNA samples from the proband. Candidate variants identified in the proband and family members were confirmed via Sanger sequencing. In silico prediction tools and co-segregation analyses were used to assess the pathogenicity of identified variants. A literature review of known EYA4 variants was performed, analysing variant frequency, distribution characteristics across different populations, and genotype-phenotype correlations. RESULTS: We identified a novel EYA4 variant, c.1745_1748del (p.Glu582ValfsTer6), in a Chinese family with ADNSHL, and co-segregation with the family's phenotype was confirmed. The audiometry showed mid-to-high frequency downsloping hearing loss. To date, 52 pathogenic variants of EYA4 have been reported, with majority identified in Asian populations. Most observed are the missense and frameshift variants. CONCLUSIONS: A novel variant of EYA4 was identified in a Chinese family with postlingual hearing loss, contributing to the expanding spectrum of EYA4 variants. The audiological features of EYA4 variants are highly heterogeneous and often challenging to detect early in clinical settings. Our findings highlight the significance of genetic testing in patients presenting with postlingual hearing loss.

[The rs2564978(T) Allele Associated with Severe Influenza A Disrupts the Binding Site for Myeloid Differentiation Factor PU.1 and Reduces CD55/DAF Gene Promoter Activity in Macrophages].

The complement inhibitor CD55/DAF is expressed on many cell types. Dysregulation of CD55 expression is associated with increased disease severity in influenza A infection and vascular complications in pathologies that involve excessive activation of the complement system. A luciferase reporter system was used to functionally analyze the single nucleotide polymorphism rs2564978 in the U937 human promonocytic cell line. The polymorphism is in the promoter of the CD55 gene, and its minor allele T is associated with a severe course of influenza A(H1N1)pdm09. A decreased activity of the CD55 promoter carrying the minor rs2564978(T) allele was observed in activated U937 cells, which provide a cell model of human macrophages. Using bioinformatics resources, PU.1 was identified as a potential transcription factor that may bind to the CD55 promoter at the rs2564978 site in an allele-specific manner. The involvement of PU.1 in modulating CD55 promoter activity was verified by a PU.1 genetic knockdown with small interfering RNAs under specific monocyte activation conditions.

Upregulation of keratin 15 is required for varicella-zoster virus replication in keratinocytes and is attenuated in the live attenuated vOka vaccine strain.

Varicella-zoster virus (VZV) is the etiological agent of chickenpox and shingles, diseases characterised by epidermal virus replication in skin and mucosa and the formation of blisters. We have previously shown that VZV infection has a profound effect on keratinocyte differentiation, altering the normal pattern of epidermal gene expression. In particular, VZV infection reduces expression of suprabasal keratins 1 and 10 and desmosomal proteins, disrupting epidermal structure to promote expression of a blistering phenotype. Here, we extend these findings to show that VZV infection upregulates the expression of keratin 15 (KRT15), a marker expressed by basal epidermal keratinocytes and hair follicles stem cells. We demonstrate that KRT15 is essential for VZV replication in the skin, since downregulation of KRT15 inhibits VZV replication in keratinocytes, while KRT15 exogenous overexpression supports viral replication. Importantly, our data show that VZV upregulation of KRT15 depends on the expression of the VZV immediate early gene ORF62. ORF62 is the only regulatory gene that is mutated in the live attenuated VZV vaccine and contains four of the five fixed mutations present in the VZV Oka vaccine. Our data indicate that the mutated vaccine ORF62 is not capable of upregulating KRT15, suggesting that this may contribute to the vaccine attenuation in skin. Taken together our data present a novel association between VZV and KRT15, which may open a new therapeutic window for a topical targeting of VZV replication in the skin via modulation of KRT15.

SPI1+CD68+ macrophages as a biomarker for gastric cancer metastasis: a rationale for combined antiangiogenic and immunotherapy strategies.

BACKGROUND: Tumor-associated macrophages (TAMs) have been demonstrated to be associated with tumor progression. However, the different subpopulations of TAMs and their roles in gastric cancer (GC) remain poorly understood. This study aims to assess the effects of Spi-1 proto-oncogene (SPI1)+CD68+ TAMs in GC. METHODS: The distribution of SPI1+CD68+ TAMs in GC tissue was estimated by immunohistochemistry, immunofluorescence, and flow cytometry. Single-cell transcriptome analysis and multiplex fluorescence immunohistochemistry were applied to explore the role of SPI1+CD68+ TAMs in an immune contexture. SPI1 overexpression or knockdown cells were constructed to evaluate its role in macrophage polarization and angiogenesis in vitro and in vivo. Chromatin immunoprecipitation was used to verify the mechanism of SPI1 transcriptional function. The effect of combined antiangiogenic and immunotherapy was further validated using mouse peritoneal metastasis models. RESULTS: Single-cell transcriptome analysis and immunohistochemistry demonstrated that SPI1 was expressed in macrophages, with a higher enrichment in metastatic lesions than in primary tumors. Higher SPI1+CD68+ TAMs infiltration was associated with poor overall survival. Mechanically, SPI1 promoted the M2-type macrophage polarization. SPI1 could bind to the promoter of vascular endothelial growth factor A and facilitate angiogenesis. Moreover, the level of SPI1+CD68+ TAMs infiltration was closely related to the efficacy of immunotherapy, especially when combined with antiangiogenic therapy. CONCLUSIONS: The present study showed that SPI1+CD68+ TAMs are a promising biomarker for predicting prognosis, antiangiogenic drug sensitivity, and combination target of immunotherapy in patients with GC.

Spindle Cell Neoplasm With a Novel MN1::TAF3 Fusion: A Rare Case in a Toddler.

Spindle cell tumors in the pediatric population are uncommonly reported. This case discusses an 18-month-old who presented initially with unilateral ptosis and was found to have an orbital spindle cell tumor. Pathology evaluation of the tissue was extensive with nonspecific morphologic and immunohistochemical features. Molecular testing demonstrated an MN1::TAF3 fusion on RNA sequencing, which has not been previously described in the literature in association with spindle cell neoplasms. This case highlights the challenging nature of classifying and treating a tumor with a novel fusion.

A New Frontier in Tumor Eradication: Harnessing In Vivo Cellular Reprogramming for Durable Cancer Immunotherapy.

Tumors evade immune detection by downregulating antigen presentation and hindering immune responses. Type 1 conventional dendritic cells (cDC1s) are vital in stimulating cytotoxic T cells against tumors. Ascic et al. are now demonstrating the in situ ability of PU.1, IRF8, and BATF3 (PIB) transcription factors to directly reprogram a plethora of tumors bypassing the suppressive effects of the tumor microenvironment, and leading to overall tumor regression while eliciting a systemic immune response that can protect from secondary tumor induction.

Genome-wide identification of CAMTA genes and their expression dependence on light and calcium signaling during seedling growth and development in mung bean.

BACKGROUND: Calmodulin-binding transcription activator (CAMTA) is comprised of a group of transcription factors and plays an important role in the Ca2+ signaling pathway, mediating various molecular responses via interactions with other transcription factors and binding to the promoter region of specific genes. Mung beans (Vigna radiata) are one of the most commonly consumed commodities in Asia. To date, CAMTA proteins have not been characterized in this important crop plant. RESULTS: Eight paralogous VrCAMTA genes were identified and found to be distributed on five of the 11 chromosomes. The proteins possessed CG-1 DNA-binding domains with bipartite NLS signals, ankyrin domains, CaM-binding IQ motifs, and CaM-binding domain (CaMBD). The 2 kb upstream regions of VrCAMTA genes contained sequence motifs of abscisic acid-responsive elements (ABRE) and ethylene-responsive elements (ERE), and binding sites for transcription factors of the bZIP and bHLH domains. Analysis of RNA-seq data from a public repository revealed ubiquitous expression of the VrCAMTA genes, as VrCAMTA1 was expressed at the highest level in seedling leaves, whereas VrCAMTA8 was expressed at the lowest level, which agreed with the RT-qPCR analysis performed on the first true leaves. On day four after leaf emergence, all VrCAMTA genes were upregulated, with VrCAMTA1 exhibiting the highest degree of upregulation. In darkness on day 4, upregulation was not observed in most VrCAMTA genes, except VrCAMTA7, for which a low degree of upregulation was found, whereas no difference was found in VrCAMTA8 expression between light and dark conditions. Treatment with calcium ionophores enhanced VrCAMTA expression under light and/or dark conditions at different times after leaf emergence, suggesting that calcium signaling is involved in the light-induced upregulation of VrCAMTA gene expression. CONCLUSIONS: The expression dependence of nearly all VrCAMTA genes on light and calcium signaling suggests their possible differential but likely complementary roles during the early stages of mung bean growth and development.

Phenotypic diversity of type III secretion system activity in enteropathogenic Escherichia coli clinical isolates.

Introduction. Enteropathogenic Escherichia coli (EPEC) strains pose a significant threat as a leading cause of severe childhood diarrhoea in developing nations. EPEC pathogenicity relies on the type III secretion system (T3SS) encoded by the locus of enterocyte effacement (LEE), facilitating the secretion and translocation of bacterial effector proteins.Gap Statement. While the regulatory roles of PerC (plasmid-encoded regulator) and GrlA (global regulator of LEE-activator) in ler expression and LEE gene activation are well-documented in the EPEC prototype strain E2348/69, understanding the variability in LEE gene expression control mechanisms among clinical EPEC isolates remains an area requiring further investigation.Aim. This study aims to explore the diversity in LEE gene expression control mechanisms among clinical EPEC isolates through a comparative analysis of secretion profiles under defined growth conditions favouring either PerC- or GrlA-mediated activation of LEE expression.Methodology. We compared T3SS-dependent secretion patterns and promoter expression in both typical EPEC (tEPEC) and atypical EPEC (aEPEC) clinical isolates under growth conditions favouring either PerC- or GrlA-mediated activation of LEE expression. Additionally, we conducted promoter reporter activity assays, quantitative real-time PCR and Western blot experiments to assess gene expression activity.Results. Significant differences in T3SS-dependent secretion were observed among tEPEC and aEPEC strains, independent of LEE sequence variations or T3SS gene functionality. Notably, a clinical tEPEC isolate exhibited increased secretion levels under repressive growth conditions and in the absence of both PerC and GrlA, implicating an alternative mechanism in the activation of Ler (LEE-encoded regulator) expression.Conclusion. Our findings indicate that uncharacterized LEE regulatory mechanisms contribute to phenotypic diversity among clinical EPEC isolates, though their impact on clinical outcomes remains unknown. This challenges the conventional understanding based on reference strains and highlights the need to investigate beyond established models to comprehensively elucidate EPEC pathogenesis.

Unveiling signaling pathways inducing MHC class II expression in neutrophils.

Introduction: Gram-negative bacillary bacteremia poses a significant threat, ranking among the most severe infectious diseases capable of triggering life-threatening sepsis. Despite the unambiguous involvement of neutrophils in this potentially fatal disease, there are limited data about the molecular signaling mechanisms, phenotype, and function of human neutrophils during the early phase of gram-negative bacillary bacteremia. Methods: By using an unbiased proteomics and flow cytometry approach, we identified an antigen-presenting cell (APC)-like phenotype in human peripheral blood neutrophils (PMN) with MHC class II molecule expression in the early phase of bacteremia. Using an in-vitro model of GM-CSF-mediated induction of APC-like phenotype in PMN, we investigated downstream signaling pathways leading to MHC class II expression. Results: GM-CSF stimulation of neutrophils leads to the activation of three major signaling pathways, the JAK-STAT, the mitogen-activated protein kinase (MAPK), and the phosphoinositide 3-kinase (PI3K)-Akt-mTOR pathways, while MHC class II induction is mediated by a MAPK-p38-MSK1-CREB1 signaling cascade and the MHC class II transactivator CIITA in a strictly JAK1/2 kinase-dependent manner. Discussion: This study provides new insights into the signaling pathways that induce MHC class II expression in neutrophils, highlighting the potential for therapeutic targeting of JAK1/2 signaling in the treatment of gram-negative bacteremia and sepsis. Understanding these mechanisms may open up novel approaches for managing inflammatory responses during sepsis.

Asiatic acid inhibits HBV cccDNA transcription by promoting HBx degradation.

BACKGROUND: Hepatitis B virus (HBV) infection is a persistent global public health problem, and curing for chronic hepatitis B (CHB) through the application of existing antiviral drugs is beset by numerous challenges. The viral protein HBx is a critical regulatory factor in the life cycle of HBV. Targeting HBx is a promising possibility for the development of novel therapeutic strategies. METHODS: The Nano-Glo® HiBiT Lysis Detection System was used to screen the herbal monomer compound library for compounds that inhibit HBx expression. Western blotting was used to examine proteins expression. Southern blotting or Northern blotting were used to detect HBV DNA or HBV RNA. ELISA was performed to detect the HBsAg level. The effect of asiatic acid on HBV in vivo was investigated by using recombinant cccDNA mouse model. RESULTS: Asiatic acid, an extract of Centella asiatica, significantly reduced the HBx level. Mechanistic studies demonstrated that asiatic acid may promote the degradation of HBx in an autophagy pathway-dependent manner. Subsequently, asiatic acid was found to reduce the amount of HBx bound to covalently closed circular DNA (cccDNA) microchromosomes, and repressive chromatin modifications then occurred, ultimately inhibiting cccDNA transcriptional activity. Moreover, in HBV-infected cells and a mouse model of persistent HBV infection, asiatic acid exhibited potent anti-HBV activity, as evidenced by decreased levels of HBV RNAs, HBV DNA and HBsAg. CONCLUSIONS: Asiatic acid was identified as a compound that targets HBx, revealing its potential for application as an anti-HBV agent.

Hepatitis B Virus X Protein Induces Reactive Oxygen Species Generation via Activation of p53 in Human Hepatoma Cells.

Hepatitis B virus (HBV), particularly through the HBx protein, induces oxidative stress during liver infections. This study reveals that HBx increases reactive oxygen species (ROS) via two distinct mechanisms. The first mechanism is p53-independent, likely involving mitochondrial dysfunction, as demonstrated by elevated ROS levels in p53-deficient Hep3B cells and p53-knocked-down HepG2 cells after HBx expression or HBV infection. The increase in ROS persisted even when p53 transcriptional activity was inhibited by pifithrin-α (PFT-α), a p53 inhibitor. The second mechanism is p53-dependent, wherein HBx activates p53, which then amplifies ROS production through a feedback loop involving ROS and p53. The ability of HBx to elevate ROS levels was higher in HepG2 than in Hep3B cells. Knocking down p53 in HepG2 cells lowered ROS levels, while ectopic p53 expression in Hep3B cells raised ROS. HBx-activated p53 downregulated catalase and upregulated manganese-dependent superoxide dismutase, contributing to ROS amplification. The transcriptional activity of p53 was crucial for these effects, as cells with a p53 R175H mutation or those treated with PFT-α generated less ROS. Additionally, HBx variants with Ser-101 increased p53 and ROS levels, whereas variants with Pro-101 did not. These dual mechanisms of HBx-induced ROS generation are likely significant in the pathogenesis of HBV and may contribute to liver diseases, including hepatocellular carcinoma.

Protective Effect of EBF Transcription Factor 1 (EBF1) Polymorphism in Sporadic and Familial Spontaneous Preterm Birth: Insights from a Case-Control Study.

This study investigated the potential role of specific single-nucleotide polymorphisms (SNPs) in the genes Astrotactin 1 (ASTN1), EBF Transcription Factor 1 (EBF1), Eukaryotic Elongation Factor, Selenocysteine-tRNA Specific (EEFSEC), Microtubule-Associated Serine/Threonine Kinase 1 (MAST1), and Tumor Necrosis Factor Alpha (TNF-α) to assess whether these genetic variants contribute to the risk of spontaneous preterm birth (sPTB). A case-control study was conducted involving 573 women from Croatia and Slovenia: 248 with sporadic sPTB (positive personal and negative family history of sPTB before 37 weeks' gestation), 44 with familial sPTB (positive personal and family history of sPTB before 37 weeks' gestation), and 281 control women. The analysis of ASTN1 rs146756455, EBF1 rs2963463, EBF1 rs2946169, EEFSEC rs201450565, MAST1 rs188343966, and TNF-α rs1800629 SNPs was performed using TaqMan real-time PCR. p-values were Bonferroni-adjusted for multiple comparisons. EBF1 SNP rs2963463 was significantly associated with sPTB (p adj = 0.03). Women carrying the CC genotype had a 3-4-times lower risk of sPTB (p adj < 0.0001). In addition, a significant difference in the frequency of the minor C allele was observed when comparing familial sPTB cases with controls (p adj < 0.0001). All other associations were based on unadjusted p-values. The minor T allele of EBF1 SNP rs2946169 was more frequent in sPTB cases overall than in controls, especially in sporadic sPTB (p = 0.045). Similarly, the CC genotype of ASTN1 SNP rs146756455 was more frequent in sporadic sPTB cases compared to controls (p = 0.019). Finally, the TNF-α SNP rs1800629 minor A allele and AA genotype were more common in the familial sPTB group compared to sporadic sPTB and controls (p < 0.05). The EBF1 SNP rs2963463 polymorphism showed a protective effect in the pathogenesis of sPTB, particularly in women carrying the CC genotype. Moreover, EBF1 SNP rs2946169 and ASTN1 SNP rs146756455, as well as TNF-α SNP rs1800629, were associated with an increased risk of sPTB, representing suggestive potential risk factors for sporadic and familial sPTB, respectively.