Altered Gene Expression Research Articles

KAT2B inhibits proliferation and invasion via inactivating TGF-β/Smad3 pathway-medicated autophagy and EMT in epithelial ovarian cancer

Lysine acetyltransferase 2B (KAT2B) plays a crucial role in epigenetic regulation and tumor pathogenesis. Our study investigates KAT2B’s function in epithelial ovarian cancer (EOC) using in vivo and in vitro methods. Immunohistochemistry showed the KAT2B expression in EOC tissues. RNA sequencing (RNA-seq) further identified altered gene expression profiles following KAT2B silencing in EOC cells. Western blot and qRT-PCR were employed to assess the protein and mRNA expression, respectively. KAT2B downregulated in EOC tissues and correlated with both FIGO stage and grade. KAT2B silencing induced autophagy, enhancing cell proliferation and invasion, while overexpression had opposite effects. In vivo, KAT2B silencing increased tumor volume and weight, mitigated by autophagy inhibitor chloroquine. Bioinformatics and co-immunoprecipitation assays identified a KAT2B-SMAD7 interaction. Mechanistic investigations suggested that KAT2B knockdown enhanced autophagy via activation of the TGF-β/Smad3/7 signaling pathway, thereby driving epithelial-mesenchymal transition (EMT), proliferation, and invasion in EOC. Additionally, our data hint at a potential role for the AKT/mTOR pathway. KAT2B acts as a putative tumor suppressor in EOC, where its reduced expression correlates with advanced disease stages. It is implicated in the regulation of autophagy, proliferation, and invasion via the TGF-β/Smad3/7 pathway, positioning KAT2B as a candidate therapeutic target for EOC interventions.

RET inhibition overcomes resistance to combined CDK4/6 inhibitor and endocrine therapy in ER+ breast cancer

BackgroundCombined CDK4/6 inhibitor (CDK4/6i) and endocrine therapy significantly improve the outcome of patients with advanced estrogen receptor-positive (ER+) breast cancer. However, resistance to this treatment and disease progression remains a major clinical challenge. High expression of the receptor tyrosine kinase REarranged during Transfection (RET) has been associated with resistance to endocrine therapy in breast cancer, but the role of RET in CDK4/6i treatment response/resistance remains unexplored.MethodsTo identify gene expression alterations associated with resistance to combined endocrine therapy and CDK4/6i, we performed RNA sequencing of two ER+ breast cancer cell models resistant to this combined therapy. The functional role of RET was assessed by siRNA-mediated RET silencing and targeted inhibition with the FDA/EMA-approved RET-selective inhibitor selpercatinib in resistant breast cancer cells and patient-derived organoids (PDOs). RET silencing was evaluated mechanistically using global gene expression and pathway analysis. The clinical relevance of RET expression in ER+ breast cancer was investigated by gene array analysis of primary tumors treated with endocrine therapy and by immunohistochemical scoring of metastatic lesions from patients who received combined CDK4/6i and endocrine therapy.ResultsWe show that RET is upregulated in ER+ breast cancer cell lines resistant to combined CDK4/6i and fulvestrant compared to isogenic cells resistant to fulvestrant alone. siRNA-mediated silence of RET in high RET-expressing, combined CDK4/6i- and fulvestrant-resistant cells reduced their growth partially by affecting cell cycle regulators of the G2-M phase and E2F targets. Notably, targeting RET with selpercatinib in combination with CDK4/6i inhibited the growth of CDK4/6i-resistant cell lines and resensitized ER+ breast cancer patient-derived organoids resistant to CDK4/6i. Finally, analysis of RET expression in ER+ breast cancer patients treated with endocrine therapy showed that high RET expression correlated with poor clinical outcomes. We further observed a shorter median survival to combined CDK4/6i and endocrine therapy in patients with RET-positive compared to RET-negative tumors, but this difference did not reach statistical significance.ConclusionsOur findings show that RET is overexpressed in ER+ metastatic breast cancer resistant to combined CDK4/6i and endocrine therapy, rendering RET inhibition a promising therapeutic approach for patients who experience disease progression on combined CDK4/6i and endocrine therapy.

Phenotypic and Gene Expression Alterations in Aquatic Organisms Exposed to Microplastics

The use of plastics, valued for its affordability, durability, and convenience, has grown significantly with the advancement of industry. Paradoxically, these very properties of plastics have also led to significant environmental challenges. Plastics are highly resistant to decomposition, resulting in their accumulation on land, where they eventually enter aquatic environments, due to natural processes or human activities. Among these plastics, microplastics, which are tiny plastic particles, are particularly concerning when they enter aquatic ecosystems, including rivers and seas. Their small size makes them easily ingestible by aquatic organisms, either by mistake or through natural feeding behaviors, which poses serious risks. Moreover, microplastics readily adsorb other pollutants present in aquatic environments, creating pollutant complexes that can have a synergistic impact, magnifying their harmful effects compared to microplastics or pollutants acting alone. As a result, extensive research has focused on understanding the effects of microplastics on aquatic organisms. Numerous studies have demonstrated that aquatic organisms exposed to microplastics, either alone or in combination with other pollutants, exhibit abnormal hatching, development, and growth. Additionally, many genes, particularly those associated with the antioxidant system, display abnormal expression patterns in these conditions. In this review, we examine these impacts, by discussing specific studies that explore changes in phenotype and gene expression in aquatic organisms exposed to microplastics, both independently and in combination with adsorbed pollutants.

Efficacy and compatibility mechanism of bear bile powder in Shexiang Tongxin dropping pills for acute myocardial infarction treatment

BackgroundBear bile powder (BBP), a unique animal-derived medicine with anti-inflammatory and antioxidant effects, is used in Shexiang Tongxin dropping pills (STDP), which is applied to treat cardiovascular diseases, including acute myocardial infarction (AMI). The efficacy and compatibility mechanisms of action of BBP in STDP against cardiovascular diseases remain unclear. This study aimed to investigate the compatibility effects of BBP in STDP in rats with AMI.MethodsWe investigated the compatibility effects of BBP in STDP in rats with AMI. Non-targeted metabonomics, 16S rRNA analysis, RNA sequencing, and network pharmacology were performed to explore the underlying mechanisms.ResultsThe combination of BBP and CF (STDP without BBP) significantly reduced AMI-induced infarction size, pathological alterations of cardiac tissues, and serum lactate dehydrogenase and creatine kinase levels in rats, compared with CF or BBP treatment alone. Gut microbiota and metabonomics results revealed that the combination treatment could upregulate the relative abundance of Lactobacillus and downregulate that of Helicobacter, Bilophila, and Butyricimonas, thereby rebalancing the gut microbiota dysbiosis induced by AMI. Consequently, the intestinal metabolite levels of oleoylcholine, glutamylalanine, isokobusone, and hemorphin-4 were altered. However, treatment with CF or BBP alone has a weaker effect on these bacteria. Additionally, the combination treatment induced a 62.34% gene reversion rate compared with 55.56% for BBP and 30.20% for CF treatment alone. Modulation of endothelin 1 and growth factor receptor-bound protein 2 was identified as a key synergistic mechanism underlying the anti-AMI effects of BBP in STDP.ConclusionThis research provides a scientific explanation of the compatibility of BBP in STDP. Our findings suggested that combination treatment with CF and BBP synergistically attenuates AMI by altering gene expression, gut microbiota, and intestinal metabolite profiles.Graphical

MicroRNAs' Significance in Retinoblastoma Diagnosis and Treatment: The Little Heroes.

One in 16, 000 live births is affected by the retinal tumor RB (retinoblastoma), which is frequently found in a child's early years. Both of the RB1 alleles that have been locally mutated in the affected retina are present in 60 percent of cases. Retinoblastoma (RB) can be detected using a variety of techniques, including imaging of the brain and orbits, eye examinations under anesthesia (EUAs), and the discovery of cell-free tumor DNA in samples of aqueous humor or plasma. In addition to the conventional surgical, chemotherapy, and radiotherapy approaches to treating retinoblastoma, new approaches have also been developed. Oncogenes, genes of tumor suppressors, and other molecular elements involved in cell growth and division interact complexly during the pathogenesis of retinoblastoma. The development of new therapies depends on comprehending the function of these molecular components. As a small class of non-coding RNAs capable of altering gene expression, microRNAs (miRNA) are understood to represent potential targets for the treatment of cancer. This study aimed to describe the changes in microRNA expression in some types of cancer, with a particular focus on retinoblastoma.

Single-cell sequencing of human Langerhans cells identifies altered gene expression profiles in patients with atopic dermatitis.

Atopic dermatitis (AD) is characterized by dysregulated T cell immunity and skin microbiome dysbiosis with predominance of Staphylococcus aureus, which is associated with exacerbating AD skin inflammation. Specific glycosylation patterns of S. aureus cell wall structures amplify skin inflammation through interaction with Langerhans cells (LCs). Nevertheless, the role of LCs in AD remains poorly characterized. Here, we performed single cell RNA sequencing of primary epidermal LCs and dermal T cells, isolated from skin biopsies of AD patients and healthy control subjects, alongside specific glycoanalysis of S. aureus strains isolated from the AD lesions. Our findings revealed 4 LC subpopulations ie, 2 steady-state clusters [LC1 and LC1H] and 2 proinflammatory/matured subsets [LC2 and migratory LCs]. The latter 2 subsets were enriched in AD skin. AD LCs showed enhanced expression of C-type lectin receptors, the high-affinity IgE receptor, and activation of prostaglandin and leukotriene biosynthesis pathways, upregulated transcriptional signatures related to T cell activation pathways, and increased expression of CCL17 compared with healthy LCs. Correspondingly, T helper 2 and T regulatory cell populations were increased in AD lesions. Complementary, we performed bulk RNA sequencing of primary LCs stimulated with the S. aureus strains isolated from the AD lesions, which showed upregulation of T helper 2-related pathways. Our study provides proof-of-concept for a role of LCs in connecting the S. aureus-T cell axis in the AD inflammatory cycle.

Enhanced bioaccumulation and toxicity of Fenpropathrin by polystyrene nano(micro)plastics in the model insect, silkworm (Bombyx mori)

BackgroundNano(micro)plastics (NMPs) and agrochemicals are ubiquitous pollutants. The small size and physicochemical properties of NMPs make them potential carriers for pollutants, affecting their bioavailability and impact on living organisms. However, little is known about their interactions in terrestrial ecosystems. This study investigates the adsorption of Fenpropathrin (FPP) onto two different sizes of polystyrene NMPs and examines their impacts on an insect model, silkworm Bombyx mori. We analyzed the systemic effects of acute exposure to NMPs and FPP, individually and combined, at organismal, tissue, cellular, and gut microbiome levels.ResultsOur results showed that NMPs can adsorb FPP, with smaller particles having higher adsorption capacity, leading to size-dependent increases in the bioaccumulation and toxicity of FPP. These effects led to higher mortality, reduced body weight, delayed development, and decreased cocoon production in silkworms. Additionally, the pollutants caused physical and oxidative damage to the midgut and altered gene expression related to juvenile hormone (JH) and silk protein synthesis. The gut microbiome analysis revealed significant changes and reduced abundance of potentially beneficial bacteria. Thus, the aggravated toxicity induced by NMPs was size-dependent, with smaller particles (NPs) having a greater impact.ConclusionsThis study demonstrates the role of NMPs as carriers for contaminants, increasing their bioavailability and toxicity in terrestrial ecosystems. These findings have significant implications for ecosystem health and biodiversity.

Single-cell RNA sequencing highlights the role of distinct natural killer subsets in sporadic amyotrophic lateral sclerosis

BackgroundNeuroinflammation plays a major role in amyotrophic lateral sclerosis (ALS), and cumulative evidence suggests that systemic inflammation and the infiltration of immune cells into the brain contribute to this process. However, no study has investigated the role of peripheral blood immune cells in ALS pathophysiology using single-cell RNA sequencing (scRNAseq).MethodsWe aimed to characterize immune cells from blood and identify ALS-related immune alterations at single-cell resolution. For this purpose, peripheral blood mononuclear cells (PBMC) were isolated from 14 ALS patients and 14 cognitively unimpaired healthy individuals (HC), matched by age and gender, and cryopreserved until library preparation and scRNAseq. We analyzed differences in the proportions of PBMC, gene expression, and cell-cell communication patterns between ALS patients and HC, as well as their association with plasma neurofilament light (NfL) concentrations, a surrogate biomarker for neurodegeneration. Flow cytometry was used to validate alterations in cell type proportions.ResultsWe identified the expansion of CD56dim natural killer (NK) cells in ALS (fold change = 2; adj. p-value = 0.0051), mainly driven by a specific subpopulation, NK_2 cells (fold change = 3.12; adj. p-value = 0.0001), which represent a mature and cytotoxic CD56dim NK subset. Our results revealed extensive gene expression alterations in NK_2 cells, pointing towards the activation of immune response (adj. p-value = 9.2 × 10− 11) and the regulation of lymphocyte proliferation (adj. p-value = 6.46 × 10− 6). We also identified gene expression changes in other immune cells, such as classical monocytes, and distinct CD8 + effector memory T cells which suggested enhanced antigen presentation via major histocompatibility class-II (adj. p-value = 1.23 × 10− 8) in ALS. The inference of cell-cell communication patterns demonstrated that the interaction between HLA-E and CD94:NKG2C from different lymphocytes to NK_2 cells is unique to ALS blood compared to HC. Finally, regression analysis revealed that the proportion of CD56bright NK cells along with the ALSFRS-r, disease duration, and gender, explained up to 76.4% of the variance in plasma NfL levels.ConclusionOur results reveal a signature of relevant changes occurring in peripheral blood immune cells in ALS and underscore alterations in the proportion, gene expression, and signaling patterns of a cytotoxic and terminally differentiated CD56dim NK subpopulation (NK_2), as well as a possible role of CD56bright NK cells in neurodegeneration.

DOP128 Spatial transcriptomic analysis reveals evolutionary dynamics in colitis-associated colorectal cancer

Abstract Background Colitis-associated colorectal cancer (CAC) is a severe complication of inflammatory bowel disease, particularly ulcerative colitis. Despite its clinical significance, the spatiotemporal dynamics of neoplastic progression in CAC remain poorly understood. This study aimed to elucidate the evolutionary trajectory of CAC using spatial transcriptomics. Methods We employed spatial transcriptomics to analyze four tissue samples from three patients with ulcerative colitis at various stages of neoplastic progression. Whole-slide spatial transcriptomics RNA sequencing was performed, enabling simultaneous analysis of gene expression and tissue architecture. Integrated spatial transcriptomic data revealed recurring epithelial programs along the progression of copy number alteration. Results Our analysis using copy number variation (CNV) score and Hematoxylin and Eosin (H&E) staining image revealed progressive changes in gene expression patterns as tissue transitioned from non-tumor to pre-tumor and finally to tumor states. This progression was quantified using beta coefficient values, allowing us to track gradual alterations in gene expression across the disease continuum. We identified a core set of 48 genes that exhibited consistent changes across all four analyzed samples. Among these, 40 genes showed a progressive decrease in expression, while eight genes, including CLDN1, E2F1, and RECQL4, demonstrated a gradual increase in expression as the tissue advanced towards a tumorous state. These upregulated genes pointed to potential oncogenic drivers or compensatory mechanisms activated during tumor development. Pathway analysis of the 48 genes revealed enrichment in key cellular processes, including p53 signaling, transcriptional regulation by E2F factors, and role of PKR in interferon induction and antiviral response. Upstream Regulator analysis revealed YAP1 and TGFB1 as the most significant regulators, highlighting the importance of transcriptional control and growth factor signaling in CAC development. Conclusion This study provides unprecedented insights into the spatiotemporal dynamics of CAC progression. By leveraging spatial transcriptomics, we have mapped the evolutionary trajectory of CAC at high resolution, revealing key molecular events and spatial features associated with disease progression. These findings not only enhance our understanding of CAC pathogenesis but also highlight potential targets for early intervention and personalized treatment strategies.

P0003 Differential Effects of JAK Inhibitors on Epithelial-Immune Interaction in Inflammatory Bowel Disease

Abstract Background Multiple cytokines are involved in the pathogenesis of inflammatory diseases like inflammatory bowel disease (IBD) and affect both immune and epithelial cells. JAK-STAT signaling plays an important role in immune responses by transmitting cytokine signals, resulting in altered gene expression and immune activation. To treat diseases like IBD, compounds that interfere with the JAK-STAT pathway, JAK inhibitors, have been developed to dampen inflammation and reduce cytokine signaling. However, these inhibitors' actual mode of action on the human colon has not been fully elucidated. Therefore, we developed a co-culture system to test the effect of different JAK inhibitors on epithelial-immune interactions. Methods Human colonoids were disrupted into single cells and seeded on Transwell® inserts in an upside-down orientation to form monolayers1,2 (Figure 1). After differentiation of the layer3, epithelial cells were either collected for qPCR analysis or fixed for immunofluorescence microscopy. To study epithelial-immune interactions co-culture experiments were performed using peripheral blood mononuclear cells (PBMCs). Cells were treated with tofacitinib, filgotinib, upadacitinib, and/or different cytokines. Epithelial cells were collected for western blot analysis and PBMCs were collected for flow cytometry analysis. Results Immunofluorescence staining showed a representative number of enterocytes and goblet cells in the upside-down colonoid monolayer which was confirmed by qPCR. Co-culture of epithelial cells with activated PBMCs showed induction of the JAK-STAT pathway; both phosphorylated STAT1 (pSTAT1) and STAT3 (pSTAT3) were induced. Treatment of PBMCs with tofacitinib, upadacitinib, or filgotinib completely diminished pSTAT1 and pSTAT3 induction. In epithelial cells, treatment with tofacitinib and upadacitinib completely diminished pSTAT1 and pSTAT3 induction whereas filgotinib treatment did not. Conclusion We validated an effective method to grow monolayers from colonoid-derived human stem cells, containing differentiated epithelial enterocytes and goblet cells (1). We confirmed that epithelial-immune interactions can be studied with this model as pSTAT1 and pSTAT3 could be induced in the epithelial layer by activated PBMCs (2). pSTAT1 and pSTAT3 were completely inhibited by all inhibitors in PBMCs while epithelial cells didn’t fully respond to filgotinib, indicating that epithelial and immune cells respond differently to the inhibitors (3). We plan to capitalize on this system by unraveling the effects of multiple IBD-related cytokines and JAK inhibitors on healthy and IBD epithelial cells to improve clinical treatment strategies.

P0159 Effects of Transcranial Direct Current Stimulation (tDCS) on enteric nervous system (ENS) gene expression and intestinal barrier in chronic DSS colitis

Abstract Background Chronic visceral pain is a clinical challenge in >50% of Inflammatory Bowel Disease (IBD) cases. A randomized controlled trial (RCT) revealed that transcranial direct current stimulation (tDCS) can modulate brain activity and effectively alleviate chronic abdominal pain associated with IBD. To elucidate the underlying mechanism of tDCS, we implemented this technique in a mouse model of inflammatory, chronic DSS colitis. Methods The tDCS protocol (10 stimulations) was deduced from human tDCS studies. We administered an anodal direct current (0.5 mA, 20 min) to both primary motor cortices over two separate periods (5 consecutive days of stimulation per period separated by a two-day break). The study was controlled by a sham-tDCS group. For determining mucosal barrier integrity murine intestinal mucosa was mounted to Ussing chambers. Histology was done (H&E staining of FFPE colon sections). Murine activity after tDCS was assessed by mouse tracking in home cages (RFID-chipped mice). Myenteric plexus (MP) cells from the murine terminal ileum, were isolated and cultivated for 10 days. Immunostaining and bulk-RNA sequencing (NovaSeq Xplus system, read length 150 bp, read depth 30 M copies/run) were performed. Results DSS caused an inflammation-driven thickening of the colon wall, resulting in an increased transmural resistance. This effect was ameliorated by tDCS (mean±SEM DSS+sham-tDCS: 72.2±8.2 Ω·cm²; DSS+tDCS: 48.1±2.1 Ω·cm²; MWU: p=0.007). Induction of a DSS colitis was associated with a reduced murine activity, which was partially recovered by tDCS (mean±SEM DSS+sham-tDCS: 70.7±15.2 rel.dist.U/day; DSS+tDCS: 205.9 ± 40.4 rel.dist.U/day; MWU: p=0.029). Neuronal cell identity was confirmed in isolated murine MP cells by immunostaining (b3-tubulin, S100B). Bulk-RNAseq significantly separated tDCS- from sham-tDCS treated DSS colitis mice as revealed by PCA. Consecutively, 739 differentially expressed genes were identified in tDCS-treated colitis mice compared to sham-tDCS-treated DSS colitic mice (622 upregulated, 117 downregulated genes; log2FC≥1.5, p-adj<0.05). KEGG and GO pathway analysis included neuroactive ligand-receptor interaction (p=7.94·10-3), gated channel activity (p=2.4·10-7), substrate-specific channel activity (p=1.99·10-7), leukocyte migration (p=3.99·10-7). Conclusion In colitis, tDCS has the capacity to modulate activity of DSS-treated mice and alters gene expression in MP cells. This reveals another layer of evidence for the impact of central pathways on the brain-gut-axis. References Neeb et al., Brain Stimulation, 2019

Hippocampal transcriptome analysis in ClockΔ19 mice identifies pathways associated with glial cell differentiation and myelination.

ClockΔ19 mice demonstrate behavioral characteristics and neurobiological changes that closely resemble those observed in bipolar disorder (BD). Notably, abnormalities in the hippocampus have been observed in patients with BD, yet direct molecular investigation of human hippocampal tissue remains challenging due to its limited accessibility. To model BD, ClockΔ19 mice were employed. Weighted gene co-expression network analysis (WGCNA) was utilized to identify mutation-related modules, and changes in cell populations were determined using the computational deconvolution CIBERSORTx. Furthermore, GeneMANIA and protein-protein interactions (PPIs) were leveraged to construct a comprehensive interaction network. 174 differentially expressed genes (DEGs) were identified, revealing abnormalities in rhythmic processes, mitochondrial metabolism, and various cell functions including morphology, differentiation, and receptor activity. Analysis identified 5 modules correlated with the mutation, with functional enrichment highlighting disturbances in rhythmic processes and neural cell differentiation due to the mutation. Furthermore, a decrease in neural stem cells (NSC), and an increase in astrocyte-restricted precursors (ARP), ependymocytes (EPC), and hemoglobin-expressing vascular cells (Hb-VC) in the mutant mice were observed. A network comprising 12 genes that link rhythmic processes to neural cell differentiation in the hippocampus was also identified. This study focused on the hippocampus of mice, hence the applicability of these findings to human patients warrants further exploration. The ClockΔ19 mutation may disrupt circadian rhythm, myelination, and the differentiation of neural stem cells (NSCs) into glial cells. These abnormalities are linked to altered expression of key genes, including DPB, CIART, NR1D1, GFAP, SLC20A2, and KL. Furthermore, interactions between SLC20A2 and KL might provide a connection between circadian rhythm regulation and cell type transitions.

Sex-specific effects of prenatal bisphenol A exposure on transcriptome-interactome profiles of autism candidate genes in neural stem cells from offspring hippocampus

Bisphenol A (BPA), an endocrine-disrupting chemical, is increasingly linked to the pathogenesis of autism spectrum disorder (ASD). This study investigates the effects of prenatal BPA exposure on neural stem cells (NSCs) from the hippocampi of rat offspring, a brain region critical for neurodevelopment and implicated in ASD. Pregnant rats were administered with BPA or vehicle control once daily via oral gavage from gestational day 1 until parturition. NSCs were isolated from the offspring’s hippocampi on postnatal day 1, and RNA sequencing was performed to examine transcriptomic alterations. Differentially expressed genes (DEGs) were identified through RNA-seq and further analyzed using Ingenuity Pathway Analysis (IPA) to explore disrupted pathways. In addition, in vitro proliferation assays were conducted, utilizing immunofluorescence staining for Sox2, a stem cell marker, and BrdU to quantify proliferating NSCs. Our results revealed that prenatal BPA exposure induced sex-specific alterations in NSC gene expression, with ASD-related genes such as Atp1a3, Nefl, and Grin1 being particularly dysregulated in male offspring. Moreover, sex-specific changes in NSC proliferation were observed. The study underscores BPA’s potential as an environmental risk factor for ASD, emphasizing the need for further research into its role in sex-specific neurodevelopmental effects.

P1301 Bile acid composition and immunological effect in mouse model and IBD patients

Abstract Background Bile acids (BAs) were suggested to have immunomodulatory properties that could influence inflammatory pathways in Inflammatory Bowel Disease (IBD). Dysregulated BA metabolism has been observed in IBD patients, further presented in our study by an analysis of IBD patients BA profiles with association to gene expression and targeted treatment of organoid models to elucidate the specific mechanisms by which BAs may affect intestinal inflammation in IBD. Methods Small intestinal organoids were grown from WT mice and an IBD mouse model with caspase-8 KO in intestinal epithelial cells (IEC). Organoids treated with primary (1°) and secondary (2°) BAs (CDCA, LCA, DCA) and BA + JAK-inhibitor (CDCA + inhibitor) were analysed thoroughly by bulk-RNA sequencing to investigate effects of BA and possible strategies to ameliorate negative effects. BA composition of serum samples from healthy controls (HC) (n=25) and IBD patients (n= 85) was quantified using LC-MS/MS and compared to bulk-RNA sequencing results of the same individuals in order to identify correlations of BA abundancy and gene expression. Results In WT organoids, CDCA treatment induced a significant upregulation of numerous genes associated with key cellular processes such as immunological, cell death but also mitochondrial. In contrast, in genetically-predisposed IBD organoids predominantly displayed gene downregulation in response to CDCA, indicating a divergent transcriptional response between WT and KO genotypes. In IBD organoids, JAK inhibition in conjunction with CDCA significantly altered the expression profile, reversing the expression of several genes that were downregulated by CDCA alone. This reversal was gene-specific and suggests a compensatory or modulatory effect of JAK inhibition on CDCA signalling in a caspase-8 deficient background. Notably, this combinatorial effect was not observed in WT organoids, where CDCA + JAK inhibitor treatment did not substantially alter gene expression compared to CDCA alone. Targeted metabolomic analysis of patient serum samples revealed two distinct bile acid profiles within the IBD cohort. In one subgroup, serum was characterized by elevated levels of the 1°BA CDCA coupled with lower concentrations of both primary and conjugated 2°BAs. In contrast, a second subgroup displayed lower CDCA levels alongside increased concentrations of secondary bile acids. These bile acid composition patterns suggest differing metabolic states within the patient cohort. Conclusion Our findings reveal significant links between BA composition, gene expression, and JAK pathway modulation, offering new insights into BA–driven gene regulatory mechanisms in both murine organoids and an IBD patient cohort. References TRR241 IBDome Consortium, TRR 241 Research Initiative, Berlin-Erlangen, Germany

Unveiling cell-type-specific microRNA networks through alternative polyadenylation in glioblastoma

BackgroundGlioblastoma multiforme (GBM) is characterized by its cellular complexity, with a microenvironment consisting of diverse cell types, including oligodendrocyte precursor cells (OPCs) and neoplastic CD133 + radial glia-like cells. This study focuses on exploring the distinct cellular transitions in GBM, emphasizing the role of alternative polyadenylation (APA) in modulating microRNA-binding and post-transcriptional regulation.ResultsOur research identified unique APA profiles that signify the transitional phases between neoplastic cells and OPCs, underscoring the importance of APA in cellular identity and transformation in GBM. A significant finding was the disconnection between differential APA events and gene expression alterations, indicating that APA operates as an independent regulatory mechanism. We also highlighted the specific genes in neoplastic cells and OPCs that lose microRNA-binding sites due to APA, which are crucial for maintaining stem cell characteristics and DNA repair, respectively. The constructed networks of microRNA-transcription factor-target genes provide insights into the cellular mechanisms influencing cancer cell survival and therapeutic resistance.ConclusionsThis study elucidates the APA-driven regulatory framework within GBM, spotlighting its influence on cell state transitions and microRNA network dynamics. Our comprehensive analysis using single-cell RNA sequencing data to investigate the microRNA-binding sites altered by APA profiles offers a robust foundation for future research, presenting a novel approach to understanding and potentially targeting the complex molecular interplay in GBM.

The Histone Lysine Demethylase KDM7A Contributes to Reward Memory via Fscn1-Induced Synaptic Plasticity in the Medial Prefrontal Cortex.

Lysine demethylase 7A (KDM7A) catalyzes the removal of dimethylation from histone H3 lysine 9 and lysine 27, both of which are associated with transcription repression. Previous study indicates that Kdm7a mRNA in the medial prefrontal cortex (mPFC) increases after drug exposure, yet its role in drug-related behaviors is largely unknown. In a morphine-conditioned place preference (CPP) paradigm, these findings reveal a specific increase of Kdm7a expression in the mPFC 7 days after drug withdrawal. Subsequently, these results demonstrate that knockdown of Kdm7a in the mPFC do not affect the acquisition of morphine-induced CPP, but it attenuate memory consolidation. To further explore Kdm7a-mediated transcriptomic changes, this work employs Nanopore direct RNA sequencing. Transcriptome profiling unveils several gene expression alterations impacted by KDM7A, which are enriched in relevant neural function categories. Notably, this work identifies and validates fascin actin-bundling protein 1 (Fscn1) as a downstream molecular target. Knockdown of Fscn1 has a similar impact on CPP to Kdm7a, along with corresponding decrease of dendritic spine density and neuronal activity in the mPFC. Additionally, silencing Kdm7a decreases enrichment of H3K9me2 and H3K27me2 at the Fscn1 promoter region, suggesting that KDM7A may act as a crucial regulator of transcriptional responses to morphine-related reward memory via Fscn1.

Identification of Potential Intervention Targets Involved in Prior Exercise that Attenuates Peripheral Neuropathic Pain by Integrating Transcriptome and Whole-genome Bisulfite Sequencing Analyses.

Changes in DNA methylation and subsequent alterations in gene expression have opened a new direction in research related to the pathogenesis of peripheral neuropathic pain (PNP). This study aimed to reveal epigenetic perturbations underlying DNA methylation in the dorsal root ganglion (DRG) of rats with peripheral nerve injury in response to prior exercise and identify potential target genes involved. Male Sprague-Dawley rats were divided into three groups, namely, chronic constriction injury (CCI) of the sciatic nerve, CCI with prior 6-week swimming training (CCI_Ex), and sham operated (Sham). Mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were used as the main observation indicators to evaluate behavioral changes associated with pain. In this study, 6-week swimming training before CCI prevented later chronic pain. In particular, CCI rats with prior exercise showed a significant increase in the MWT and TWL of the injured lateral hind paw compared with CCI rats without exercise on days 14, 21, and 28 after CCI. Whole-genome bisulfite sequencing from the injured lumbar (L4-L6) DRGs on the 28th day after surgery was detected. We also generated DNA methylation maps of the two comparisons (sham group vs. CCI and CCI groups vs. CCI_Ex group), and 396 overlapping differentially methylated region-related genes were found between the two comparisons. Moreover, we integrated RNA sequencing to understand the mechanism by which differential DNA methylation after CCI may influence gene expression. Finally, Ryr1 and Xirp2 were identified through association analysis of two omics and quantitative reverse-transcription polymerase chain reaction, respectively. The methylation levels of Ryr1 and Xirp2 were upregulated with a corresponding increase in their mRNA expression in the DRGs of CCI rats, whereas prior exercise downregulated Ryr1 methylation and restore its expression level. Functional enrichment analysis of both omics found that the calcium signaling pathway was significantly enriched. Therefore, the potential intervention targets (Ryr1 and Xirp2) in L4-L6 DRGs may be involved in prior exercise that attenuates PNP induced by CCI. This study provides crucial insights into the epigenetic regulation of PNP responses to prior exercise.

A prospective self-controlled study on the alterations of the ocular surface and conjunctival transcriptomic profile associated with prolonged exposure to video display terminals.

To assess the impact of prolonged and intense exposure to video display terminals (VDTs) on ocular surface homeostasis. 30 subjects limited daily VDT usage to less than 3h for one week, then extended usage to more than 8h/day for the next three weeks. Ocular symptoms and signs were evaluated weekly using the Ocular Surface Disease Index (OSDI) questionnaire and clinical examinations. Eyelid margins and meibomian glands were examined, and ocular surface samples were collected for transcriptomic analysis. Average daily VDT time increased from 2.55±0.46h initially to 11.17±2.45, 11.75±2.63, and 10.89±2.41h over three weeks. The dry eye diagnosis rate rose from 6.67% to 51.67%. Total OSDI score (P=0.008), symptoms score (P=0.014), and visual function score (P=0.002) significantly increased. Mean fluorescein break-up time (FBUT) decreased from 6.46s to 3.08s. Corneal fluorescein staining (CFS) score (P<0.001) and lissamine green conjunctival staining (LCjs) score (P=0.036) worsened. Ocular redness index (RI) increased at 1 week and 3 weeks (P=0.007, P=0.001). Telangiectasia scores of both upper and lower eyelid margins increased at 3 weeks (P=0.002, P<0.001). Meibomian gland orifice blockage worsened (P=0.014, P=0.002). Transcriptomic analysis revealed dynamic alterations in ocular surface gene expression, including inflammatory and hormonal responses. MUC5AC and TFF1 genes showed negative correlations with OSDI and conjunctival staining score, respectively. Prolonged VDT exposure deteriorates ocular surface symptoms and signs, with significant inflammatory responses and hormonal activity indicating an imbalance in ocular surface homeostasis.

The interplay between RNA m6A modification and radiation biology of cancerous and non-cancerous tissues: a narrative review.

The diverse radiation types in medical treatments and the natural environment elicit complex biological effects on both cancerous and non-cancerous tissues. Radiation therapy (RT) induces oncological responses, from molecular to phenotypic alterations, while simultaneously exerting toxic effects on healthy tissue. N6-methyladenosine (m6A), a prevalent modification on coding and non-coding RNAs, is a key epigenetic mark established by a set of evolutionarily conserved enzymes. The interplay between m6A modification and radiobiology of cancerous and non-cancerous tissues merits in-depth investigation. This review summarizes the roles of m6A in the biological effects induced by ionizing radiation and ultraviolet (UV) radiation. It begins with an overview of m6A modification and its detection methods, followed by a detailed examination of how m6A dynamically regulates the sensitivity of cancerous tissues to RT, the injury response in non-cancerous tissues, and the toxicological effects of UV exposure. Notably, this review underscores the importance of novel regulatory mechanisms of m6A and their potential clinical applications in identifying epigenetically modulated radiation-associated biomarkers for cancer therapy and estimation of radiation dosages. In conclusion, enzyme-mediated m6A-modification triggers alterations in target gene expression by affecting the metabolism of the modified RNAs, thus modulating progression and radiosensitivity in cancerous tissues, as well as radiation effects on normal tissues. Several promising avenues for future research are further discussed. This review highlights the importance of m6A modification in the context of radiation biology. Targeting epi-transcriptomic molecules might potentially provide a novel strategy for enhancing the radiosensitivity of cancerous tissues and mitigating radiation-induced injury to normal tissues.

Epigenetic Control of Redox Pathways in Cancer Progression.

Significance: Growing evidence indicates the importance of redox reactions homeostasis, mediated predominantly by reactive oxygen species (ROS) in influencing the development, differentiation, progression, metastasis, programmed cell death, tumor microenvironment, and therapeutic resistance of cancer. Therefore, reviewing the ROS-linked epigenetic changes in cancer is fundamental to understanding the progression and prevention of cancer. Recent Advances: We review in depth the molecular mechanisms involved in ROS-mediated epigenetic changes that lead to alteration of gene expression by altering DNA, modifying histones, and remodeling chromatin and noncoding RNA. Critical Issues: In cancerous cells, alterations of the gene-expression regulatory elements could be generated by the virtue of imbalance in tumor microenvironment. Various oxidizing agents and mitochondrial electron transport chain are the major pathways that generate ROS. ROS plays a key role in carcinogenesis by activating pro-inflammatory signaling pathways and DNA damage. This loss of ROS-mediated epigenetic regulation of the signaling pathways may promote tumorigenesis. We address all such aspects in this review. Future Directions: Developments in this growing field of epigenetics are expected to contribute to further our understanding of human health and diseases such as cancer and to test the clinical applications of redox-based therapy. Recent studies of the cancer-epigenetic landscape have revealed pervasive deregulation of the epigenetic factors in cancer. Thus, the study of interaction between ROS and epigenetic factors in cancer holds a great promise in the development of effective and targeted treatment modalities. Antioxid. Redox Signal. 00, 000-000.