Search Results (317)

Metabolic dysfunction-associated steatotic liver disease (MASLD) is one of the leading causes of chronic liver disease worldwide, and effective pharmacological therapies remain limited. Pemafibrate (Pema), a selective peroxisome proliferator-activated receptor alpha (PPARα) modulator (SPPARMα), has shown promising therapeutic potential in patients with MASLD and hypertriglyceridemia; however, its underlying mechanisms remain incompletely understood. Here, we investigated the therapeutic effects and molecular mechanisms of Pema using in vitro and in vivo MASLD models and evaluated its clinical relevance in patients with MASLD. In a choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD)-induced MASLD mouse model, Pema dose-dependently ameliorated hepatic steatosis and fibrosis and significantly suppressed the hepatic expression of inflammatory and fibrogenic genes, including TLR4, TNFα, αSMA, and Col1A1. Pema also favorably altered the gut microbiota by increasing the abundance of the phylum Verrucomicrobia, suggesting modulation of the gut–liver axis. In differentiated 3T3-L1 adipocyte-like cells, Pema induced PPARα expression, inhibited insulin-mediated EGR-1 induction, and significantly reduced leptin secretion. Consistent with these findings, serum insulin and leptin levels, as well as the hepatic accumulation of both molecules, were markedly decreased in Pema-treated MASLD mice. Furthermore, in a clinical cohort of 192 patients with MASLD and hypertriglyceridemia, long-term Pema treatment significantly improved liver-related biochemical parameters, insulin resistance, surrogate markers of liver fibrosis, and serum leptin levels. Collectively, these findings demonstrate that Pema attenuates MASLD progression by suppressing leptin-mediated metabolic and inflammatory signaling while improving the gut microenvironment. These results highlight SPPARMα as a promising therapeutic strategy for MASLD and support further clinical investigation of Pema as a disease-modifying treatment targeting the gut–liver axis. Full article

The growing demand for high-quality plasmid DNA (pDNA) in cell and gene therapy, including chimeric antigen receptor T-cell therapy (CAR-T) manufacturing, is constrained by the limited availability of transfection-grade plasmids. This study describes a practical, scalable, and cost-effective acidic phenol extraction method based on a modified alkaline lysis protocol. This approach efficiently removes endotoxins and residual genomic DNA without requiring chromatography or ultracentrifugation. When benchmarked against CsCl density gradient ultracentrifugation and a commercial midiprep kit, the acidic phenol method delivered comparable plasmid purity and functional performance while providing superior scalability, yielding 5–10 mg of pDNA per 500 mL culture. Notably, residual endotoxin levels were negligible for downstream applications such as transfection and lentiviral vector packaging. Lentiviral vectors produced with these plasmids reached titers exceeding 5 × 10⁶ TU/mL, with transduction efficiencies statistically indistinguishable from those obtained with CsCl-purified DNA. The acidic phenol extraction method is a robust, scalable, and cost-effective alternative to industry-standard methods, matching them in yield and purity while being readily scalable, making it a practical tool for both academic research and preparative laboratory production. Full article

mRNA plays a pivotal role in cellular processes of genetic information transfer, and post-transcriptional modifications of its nucleotides enable regulation of these processes with each particular mRNA. m5C methylation is a specific RNA modification that is rather common in tRNA, rRNA, lncRNA and other types of non-coding RNAs, whereas in mRNA it is found not very often. However, the functioning of m5C-methylated mRNAs differs from the non-methylated ones quite dramatically. Of the eight m5C RNA methyltransferases in humans, only two, NSUN2 and NSUN6, were found to be capable of modifying the main portion of cellular mRNAs. A deficiency of NSUN2 and NSUN6 causes global changes in the transcriptome and translatome, and a corruption of the NSUN2 gene in humans is associated with neurodegenerative diseases and intellectual disability. Despite intensive research of m5C mRNA methylation in recent years, many aspects of this phenomenon and its significance remain problematic and far from understood. In this review, we discuss the available information on mRNA methylome biogenesis, methods for its study and analysis, effects of m5C modification on mRNA life, and place it in the general context of cell biology, attempting to draw the biological relevance of m5C mRNA methylation. We highlight the main inconsistencies and difficulties that arise, analyze their possible causes, and propose potential directions for further research that could clarify the controversial issues and provide a link between the NSUN2 deficiency and neurodegenerative diseases. Full article

Short-chain fatty acids (SCFAs), acetate, propionate, and butyrate, are typically produced in a 3:1:1 ratio, respectively, via microbial fermentation of non-digestible carbohydrates, and to a lesser degree, from undigested protein. The effects of individual SCFAs on adipocyte function have been described; however, the effects of SCFAs in combination on adipocyte function remain unknown. Mature 3T3-L1 adipocytes were treated with a 1 mM total dose of acetate, propionate, and butyrate combined in a 3:1:1 ratio, respectively, for 24 h ± lipopolysaccharide (LPS, 10 ng/mL) under both normoxic and hypoxic (via the addition of 100 µM cobalt chloride) environmental conditions. In both normoxic and hypoxic LPS-stimulated conditions, SCFAs increased the secretion of adiponectin and reduced the secretion of resistin, interleukin (IL)-6, monocyte chemoattractant protein (MCP)-1/C-C motif chemokine ligand (CCL)2, and RANTES/CCL5, in addition to reducing intracellular protein levels of activated (i.e., the ratio of phosphorylated-to-total) nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) p65 and signal transducer and activator of transcription 3 (STAT3) (p < 0.05). Additionally, SCFA treatment reduced leptin secretion only in LPS-stimulated normoxic environmental conditions compared to control (p < 0.05). In normoxic conditions, SCFA + LPS increased mRNA expression of genes involved in fat storage), fatty acid recycling, and lipolysis, whereas in hypoxic conditions, SCFA + LPS decreased mRNA expression of genes involved in fat storage and triglyceride synthesis (p < 0.05), indicating different effects of SCFAs on adipocyte metabolic function depending on hypoxia status. Collectively, combined SCFAs in a 3:1:1 ratio beneficially modify the adipocyte adipokine secretory profile under both normoxic and hypoxic environmental conditions. Full article

Background/Objective: Accumulation of amyloid-beta (Aβ) senile plaques in the human brain is a major hallmark of Alzheimer’s disease (AD), which manifests as progressive decline in memory and cognitive functions and currently lacks effective disease-modifying therapies. Emerging evidence demonstrates that polyphenol-rich plant foods are potential complementary therapies for AD. Methods: In this study, we investigated crude polyphenol extracts (CPEs) and purified polyphenol extracts (PPEs) from three sorghum genotypes for their ability to inhibit Aβ₄₂-induced toxicity in MC-65 cells. Thioflavin T fluorescence, cell viability, mitochondrial function, oxidative stress assays, and Western blotting, along with RNA sequencing and computational analyses, were used to characterise both functional and transcriptomic responses of the cells to polyphenol treatments. Results: CPEs and PPEs inhibited Aβ₄₂ aggregation by 67–76% and significantly reduced Aβ oligomer species. The extracts increased cell viability against Aβ-induced toxicity by more than 70%, decreased intracellular oxidative stress, and enhanced mitochondrial activity by over 80%. Transcriptomic profiling revealed differential modulation of genes associated with ferroptosis and MAPK/NF- κB signalling pathways, indicating regulation of inflammatory and oxidative-stress responses are mechanisms underlying the observed neuroprotection. Conclusions: This study demonstrates that polyphenol extracts from black and red sorghum genotypes exert strong multitarget neuroprotection against Aβ₄₂ toxicity in MC-65 cells. These findings support further evaluation of sorghum-derived polyphenols as complementary therapeutic candidates for AD, with in vivo studies required to establish efficacy and translational potential. Full article

Ataxia-Telangiectasia (A-T) is a multisystem disorder caused by loss of A-T mutated (ATM) protein activity, characterized clinically by immunodeficiency and cerebellar ataxia. ATM is a master regulator of DNA damage responses and loss of ATM function is accompanied by persistent activation of PARP1 leading to depletion of intracellular NAD⁺ and dysfunction of a series of cellular signalling pathways dependent on NAD⁺, providing a mechanistic rationale for NAD⁺ augmentation therapy. We performed a clinical trial of NAD⁺ augmentation with nicotinamide riboside (NR) over 24 months in A-T patients where we observed improved coordination and eye movements in A-T patients. Here, by using peripheral blood mononuclear cells, we performed longitudinal transcriptome profiling to define molecular signatures of A-T and to assess pathway-level responses to NR supplementation. A-T patients exhibited reproducible transcriptomic alterations involving immune, vascular, and inflammatory pathways. NAD⁺ augmentation was associated with suppression of interferon response genes and modulation of networks correlated with neurological improvement. These findings establish systemic molecular signatures of A-T and identify potential blood-based biomarkers that reflect disease processes and therapeutic response, supporting the use of NAD⁺ augmentation as a disease-modifying strategy in A-T by dampening interferon signalling. Full article

Neuroinflammation in multiple sclerosis (MS) is driven by the infiltration of myelin-reactive T cells into the central nervous system (CNS). Interferon-β (IFN-β) is one of the earliest disease-modifying treatments (DMTs) approved for MS and remains widely used in special populations (pregnant and elderly patients) owing to its favorable safety profile. However, the exact mechanism of action of this drug and reliable biomarkers of treatment response remain unclear. Transcriptomic profiling and data integration approaches offer powerful tools for investigating complex patterns of regulation and molecular mechanisms underlying therapeutic efficacy. In this study, we performed an integrative analysis of openly available transcriptomic datasets to characterize IFN-β-induced gene expression changes in MS patients. By combining data from large independent cohorts, we identified a 43-gene transcriptional signature consistently associated with IFN-β treatment across disease stages, including progressive MS. To explore the relevance of this signature, we cross-referenced the 43-gene signature with publicly available expression quantitative trait loci (eQTL) datasets to determine whether these genes could be influenced by known MS-associated risk variants highlighting Interferon-Induced Transmembrane Protein 3 (IFITM3) as a candidate molecular mediator of MS. This integrative approach provides new insights into IFN-β-driven immune modulation and supports the development of therapeutic strategies for MS. Full article

Background: Concomitant HIV/HBV infection results in worse health outcomes, with HBV reactivations being observed in immunocompromised individuals. However, data on HBV infection in people with advanced HIV disease (AHD) remains sparse in Botswana. We aimed to determine the prevalence and molecular characteristics of HBV in people living with HIV (PLHIV) with CD4+ T-cell counts ≤100 cells/µL in Botswana. Methods: Plasma samples (n = 1097) of PLHIV with CD4+ T-cell count ≤100 cells/uL collected between 2014 and 2016 were screened for hepatitis B surface antigen (HBsAg) and HBV core antibodies (anti-HBc). A 415bp region of the HBV surface gene was amplified and sequenced using Sanger sequencing. Genotypic and mutational analysis was performed using Geno2pheno. Adjusted prevalence ratios (aPRs) were estimated from a modified Poisson regression model to explore factors associated with HBV infection. p-values < 0.05 indicated statistical significance. Results: The median age was 37 years (IQR: 32–43), and 565/1097 (51.5%) were male. HBsAg prevalence was 10.6% (95%CI: 8.8–12.5%) and anti-HBc prevalence was 50.0% (95%CI:46.9–52.9%). Factors associated with HBV infection were male sex [aPR: 1.6 (p < 0.01)] and those that were ART-experienced [aPR: 1.43 (p = 0.04). Eighteen samples were successfully genotyped. The prevalence of genotype A was (12/18, 66.7%) and D (6/18, 33.3%). Sixty-three mutations were identified as associated with drug resistance and immune and diagnostic escape. Highly prevalent immune escape mutations in the surface region were S207N (12/63, 19%) and A194V (9/63, 14.3%). V163I (12/63, 19%) and M129L (12/63, 19%) were highly prevalent in the reverse transcriptase region. Two classical lamivudine-associated drug resistance mutations were observed, each occurring in one participant (L180M and V173L). Conclusions: The prevalence of HBV in people with AHD is high, highlighting the importance of HBV screening and HIV/HBV co-management in this population. Full article

Zearalenone (ZEA) is a common contaminant in crops and animal feed. However, research on the effects of ZEA on animal mammary tissue is relatively limited. Sulforaphane (SFN) is a naturally active compound mainly derived from cruciferous vegetables (such as broccoli), with significant antioxidant and cytoprotective effects. The purpose of this study is the effect of SFN on ZEA-induced toxicity in bovine mammary epithelial cells (MAC-T). By treating MAC-T cells with different concentrations of ZEA and SFN for 24 h, the results showed that different concentrations of ZEA (10, 20, 40, 60, 80, or 100 μM) could inhibit MAC-T cell viability. Treatment with SFN at concentrations of 1, 2.5, and 5 μM had no significant effect on cell viability. The results of combined treatment with 10 μM ZEA and 1, 2.5, or 5 μM SFN showed that SFN could significantly reverse the decrease in cell viability caused by ZEA; reduce the increase in lactate dehydrogenase (LDH) release, reactive oxygen species (ROS), and malondialdehyde (MDA) content induced by ZEA; and increase the levels of glutathione (GSH), superoxide dismutase (SOD), and mitochondrial membrane potential that were decreased by ZEA. SFN can significantly inhibit the upregulation of interleukin 6 (IL-6), tumor necrosis factor alpha (TNF-α), and interleukin 1 beta (IL-1β) induced by ZEA exposure and markedly reverse the increase in cell apoptosis rate caused by ZEA. Compared with the control group, the expression of genes nuclear factor erythroid 2–related factor 2 (Nrf2), heme oxygenase 1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO1), glutamate-cysteine ligase modifier subunit (GCLM), and glutathione peroxidase 1 (GPX1) was significantly reduced in the ZEA group, while the addition of SFN effectively increased the expression levels of these genes. Corresponding protein detection results were consistent with the trends in gene expression. This study demonstrated that SFN alleviates ZEA-induced damage to MAC-T cells by activating the Nrf2 pathway, providing a theoretical basis for the subsequent application of SFN in dairy farming to prevent and control breast health risks related to mycotoxins. Full article

Background: Currently, increasing attention is being paid to the role of genes other than CFTR and their variants as factors modifying the course of cystic fibrosis (CF). One such gene is SLC26A9, which encodes a protein involved in chloride and bicarbonate transport across the epithelial cell membrane. Variants of SLC26A9, such as c.229G>A (p.Gly77Ser) and c.1885C>T (p.Pro629Ser), have been described in patients with severe and rapidly progressive CF. The aim of this study was to identify SLC26A9 variants in a group of 20 patients with CF. Methods: DNA was isolated from blood samples and collected from all patients. Fragments of exons 3 and 17 of the SLC26A9 gene were amplified by PCR and sequenced using the Sanger method. Results: An SLC26A9 variant was identified in two siblings. These patients were diagnosed with CF in adulthood and presented with moderate pulmonary symptoms without exocrine pancreatic insufficiency. In both siblings carrying the CFTR variants p.Phe508del and c.3140-26A>G, the SLC26A9 variant c.1847C>T (p.Pro616Leu) was detected. This variant has not been widely described in the literature and has not previously been associated with CF. Conclusions: The c.1847C>T (p.Pro616Leu) variant is located near a domain that may affect the transport function of the SLC26A9 protein. However, patients in whom the variant was identified did not present a severe disease phenotype. Further studies on larger patient cohorts are required, and at present this variant should be considered of uncertain significance in CF. Full article

Curcumin is a polyphenolic bio-compound derived from the rhizomes of the turmeric plant (Curcuma longa) that has proven anti-carcinogenic properties but poor bioavailability. By modifying its chemical structure, the monocarbonyl analogs of curcumin (MACs) possess improved stability, resorption, and circulation. This dataset presents RT-qPCR array analysis of 84 genes associated with Epithelial to Mesenchymal Transition (EMT), a key early event in cancer progression and metastasis, in human MCF-7 breast cancer cells. Cells were stimulated toward EMT reprogramming by treatment with a combination of EMT-inducing factors and co-treated with two experimental MACs, C66 or B2BrBC. Gene expression was measured using the human EMT QIAGEN RT² Profiler kit, and results were obtained from three independent experiments. Gene expression changes are presented as both fold regulation and fold change values, with statistical significance determined by Student’s t-test (p < 0.05). This comprehensive dataset enables investigation into how MACs modulate the EMT transcriptome in breast cancer cells, with potential applications for understanding EMT mechanisms. The raw and processed data are publicly available and can be used for comparative analyses, validation studies, and bioinformatic analyses of EMT-related signaling pathways. Full article

This article discusses rheumatoid arthritis (RA) as a chronic, systemic autoimmune disease leading to progressive joint damage and multi-organ complications. The complex pathogenesis of the disease is presented, involving the interaction of environmental, genetic, and immunological factors, including the role of autoantibodies and proinflammatory cytokines. Particular attention is paid to leflunomide, a disease-modifying antirheumatic drug (DMARD), which primarily works by inhibiting the DHODH enzyme, leading to reduced T and B cell proliferation. The additional anti-inflammatory properties of the drug’s active metabolite, teriflunomide, and its impact on signaling pathways related to the immune response are also discussed. This article examines the variability in patient responses to leflunomide treatment in terms of both efficacy and toxicity, with particular emphasis on the potential role of pharmacogenetic factors. It was pointed out that polymorphisms in genes related to drug metabolism, transport, and mechanism of action may influence the pharmacokinetics and safety of the therapy. It was also emphasized that the available data are primarily derived from observational studies and small cohorts, and the results are often inconsistent. Although some genetic variants and plasma teriflunomide concentrations show potential as predictors of treatment response, the current level of evidence does not support the routine use of pharmacogenetic testing in clinical practice. The article emphasizes that the pharmacogenetics of leflunomide represents a promising, yet still exploratory, avenue of research in the context of personalized RA therapy. It emphasizes the need for larger, well-designed clinical trials and the development of standardized guidelines, which would be necessary before the potential implementation of such strategies in routine clinical practice. Full article

Phosphatidylinositol-3-kinases (PI3Ks) are heterodimers of catalytic and regulatory subunits that regulate cell metabolism, activation, and survival. PI3K, particularly the p110α catalytic isoform, is frequently mutated in cancer, and highly specific inhibitors such as alpelisib are currently used in oncology and in PIK3CA-related overgrowth disorders. Given the relevance of macrophages in anti-tumor immunity, we examined the impact of alpelisib on murine monocytes’ intracellular signaling and on in vitro differentiation, polarization, and effector functions of macrophages. Real-time qPCR (RT-qPCR) showed comparable relative expression of PI3K isoforms (p110α, p110β, p110δ, p110γ and p85) in bone marrow monocytes and in macrophages differentiated with macrophage colony-stimulating factor (M-CSF) or granulocyte-macrophage colony-stimulating factor (GM-CSF). However, alpelisib increased p110α, p110β, and p85 relative gene expression (2–3-fold) during M-CSF-dependent differentiation. Functionally, alpelisib-treated M-CSF macrophages displayed enhanced interleukin (IL)-6 and tumor necrosis factor alpha (TNF-α) secretion and reduced IL-10 production after lipopolysaccharide (LPS) plus interferon gamma (IFN-γ) or LPS stimulation. In contrast, GM-CSF macrophages differentiated with alpelisib secreted lower levels of IL-6 and TNF-α and reduced inducible nitric oxide synthase (iNOS) and arginase-1 (Arg-1) gene expression. Additionally, cytokine profiles (IL-2, IL-6, IFN-γ and IL-10) were altered when alpelisib-treated macrophages were cocultured with CD4⁺ T cells under either antigen-specific or polyclonal activation conditions, indicating that the inhibitor modifies both differentiation and subsequent effector interactions of the macrophages. Thus, alpelisib induces lasting effects on macrophage differentiation and function, with potential implications in tumor-associated macrophages that develop under M-CSF or GM-CSF-rich cancer microenvironments. Full article

Diabetic peripheral neuropathy (DPN) is a common and debilitating complication of diabetes mellitus which affects individuals with both type 1 and type 2 diabetes mellitus (T2DM), presenting with sensory loss, pain, and progressive nerve dysfunction. DPN pathogenesis is multifactorial: chronic hyperglycemia activates the polyol, hexosamine, and protein kinase C (PKC) pathways, increases advanced glycation end-products, and drives oxidative stress, mitochondrial dysfunction, inflammation, and impaired neurotrophic signaling. In addition to hyperglycemia-driven mechanisms, dyslipidemia and microvascular insufficiency exacerbate neural ischemia and metabolic stress. Recent mechanistic, animal, and associative human studies further implicate amyloidogenic toxicity, particularly from human islet amyloid polypeptide (hIAPP), as a plausible contributory factor in peripheral nerve degeneration in T2DM, linking protein misfolding and aggregation to axonal damage and demyelination in DPN. Despite increased understanding of these mechanisms, current treatments remain mainly symptomatic. Emerging therapeutic strategies, including antioxidants, anti-inflammatory agents, modulators of mitochondrial function, amyloid oligomer modulators, neurotrophic enhancers, and regenerative approaches such as stem cells and gene-based therapies, offer potential to modify disease progression. The strength of evidence across these methods varies, ranging from mechanistic and animal studies to early human research and, in some cases, randomized clinical trials. Therefore, although several candidates show potential to alter the disease, few have demonstrated consistent benefits on objective measures of nerve structure or function in large clinical trials. This review summarizes the key mechanisms driving DPN in T2DM and highlights promising therapeutic innovations poised for clinical translation. Full article

Background/Objectives: The large size of commonly used regulatory elements such as the cytomegalovirus (CMV) immediate-early promoter imposes a significant burden on the already restricted payload capacity of first-generation adenoviral vectors, potentially hindering the development of multi-antigen vaccine candidates. To address this limitation, we have engineered a panel of novel, small, semi-synthetic promoters designed to leverage the changes in transcriptomic milieu following adenoviral vector entry. Methods: Eight synthetic enhancer modules (SE1–SE8) were designed in silico, each composed of transcription factor binding sites (TFBSs) previously found in host genes that are upregulated during early adenoviral infection. These synthetic enhancers were coupled with a minimal CMV core promoter to generate a panel of compact semi-synthetic promoters (cSE1–cSE8), and their activity was evaluated in the context of ChAdOx1 viral vectors expressing GFP or a modified Plasmodium falciparum circumsporozoite (CSN) antigen. Promoter performance was characterised in vitro via flow cytometry, RT-qPCR, and Western blotting, and in vivo by quantifying antigen-specific T-cell (IFN-γ ELISpot) and IgG antibody (ELISA) responses in BALB/c mice. Results: In vitro characterisation revealed a wide range of promoter activity across the panel, with cSE3 and cSE5 driving transgene expression levels comparable to the benchmark CMV promoters despite their markedly reduced genomic footprint. In vivo, ChAdOx1 vectors incorporating cSE3 and cSE5 elicited potent antigen-specific T-cell and IgG responses that were comparable to those induced by the larger CMV control promoters. Conclusions: We have successfully developed semi-synthetic promoters that match the potency of the much larger, frequently used CMV promoters whilst simultaneously reducing genomic footprint. These novel regulatory elements will facilitate the design of next-generation vaccines, particularly those requiring large antigens or multi-antigen cassettes. Full article