International Journal of Molecular Sciences doi: 10.3390/ijms27219435

Authors: Thi Be Ta Truong Thuat T. Trinh

Reactive molecular dynamics (ReaxFF MD) simulations of polyethylene (PE) oxidative gasification generate large trajectory datasets suited to machine learning (ML). We recast H2 production monitoring as a leakage-free forecasting task: given the simulated thermochemical state at time t, predict the time-weighted cumulative H2 production cumH2(t+h) a fixed horizon h=25 ps ahead. Data from 48 trajectories spanning six O2 loadings (1–900 molecules per 50 PE chains) and eight temperatures (1500–2900 K) yield 72,117 forecast pairs (45 simulations; three truncated trajectories excluded). Eight tabular regression models are benchmarked against persistence and physics-informed baselines under simulation-grouped nested cross-validation, a held-out test partition, temperature- and loading-holdout stress tests, and a paired ablation removing all H2-derived inputs. With the H2 history available, the best test model (random forest) attains R2=0.9999 at RMSE 121.8 molecules·ps, a 93.0% RMSE reduction relative to persistence, robust to unseen O2 loadings and largely robust to unseen temperatures. Without it, no model beats persistence in any evaluation: the best ML model (an artificial neural network) is 26.0% worse, and accuracy decreases substantially under temperature extrapolation. The ablation attributes 93.3–98.3% of the tree and kernel models’ RMSE reduction to the H2-derived features, quantifying how much forecasting skill derives from the target history rather than instantaneous observables.

International Journal of Molecular Sciences doi: 10.3390/ijms27219434

Authors: Fengxiao Guo Shenghong Ge Mingguang Lei

BZR1 (Brassinazole-resistant 1) and BES1 (bri1-EMS suppressor1) are the principal transcription factors that convert brassinosteroid (BR) perception into genome-wide transcriptional programs governing plant growth, development, and stress responses. Once regarded as end-effectors of a linear receptor-to-nucleus relay, they are now recognized as central hubs that integrate diverse endogenous and environmental cues into context-specific transcriptional outputs. We synthesize recent progress across four connected levels. Structurally, a modular architecture enables simultaneous target recognition, signal input, and output control. Mechanistically, the canonical BIN2-mediated phosphorylation switch is expanded by upstream regulators that couple membrane dynamics, carbon status, protein homeostasis, and nuclear import to BZR1/BES1, while ubiquitination, SUMOylation, redox modification, and acetylation form an interconnected post-translational network. During transcriptional execution, dephosphorylated BZR1/BES1 partition targets into activated and repressed sets through cis-element preference, corepressor recruitment, chromatin modifiers, and partnerships with factors such as PIF4 and ARF6. Physiologically, BZR1/BES1 coordinate anutrient-responsive metabolism, yield-related traits, and abiotic-stress tolerance, with marked functional divergence among crop orthologs. We highlight open questions in signal-integration specificity and discuss how BZR1/BES1 modules could be engineered for climate-resilient crops.

International Journal of Molecular Sciences doi: 10.3390/ijms27219432

Authors: Bikash KC Nirav Modha Pitambar Poudel Shailesh Kumar Panday Emil Alexov

Monogenic disorders present a compelling target for drug discovery due to their clear, single-gene etiology. Yet, effective therapies remain scarce, largely because diverse mutation types scattered across a protein obstruct targeted drug design. In this study, we leverage MOGEDO, a curated database of monogenic disorders, to test whether different pathogenic genotypes within a single protein converge on a common molecular mechanism. By clustering MOGEDO proteins into eight functional families and analyzing representative proteins carrying both benign and pathogenic mutations, we integrated molecular dynamics, network analysis, and side-chain entropy metrics. Our findings reveal that distinct pathogenic mutations disrupt the structural and dynamic properties of the wild-type and benign protein through common pathways. By demonstrating that diverse disease-causing mutations manifest a unified pathogenic phenotype, this work provides a framework for designing broad-spectrum therapeutics that target common putative intervention sites rather than individual mutations.

International Journal of Molecular Sciences doi: 10.3390/ijms27219431

Authors: Yuchi Liu Cheng Guo Xiaodan Yu

Food allergy (FA) is associated with anxiety, but the underlying mechanisms remain incompletely understood. An ovalbumin (OVA)-induced FA model was established in female BALB/c mice to evaluate anxiety-like behavior and characterize molecular alterations in the prefrontal cortex (PFC). We assessed the FA model by anaphylactic scores, OVA-specific IgE and IgG1 levels, serum cytokines, and intestinal histology, and evaluated anxiety-like behavior using the open field test (OFT) and elevated plus maze (EPM). Mice with OVA-induced food allergy exhibited pronounced allergic responses and intestinal mucosal injury, together with reduced center-zone exploration in the OFT and reduced open-arm exploration in the EPM. In the PFC, inflammatory and chemotactic mediators were increased, accompanied by enlarged Iba1-positive microglial soma area. Bulk RNA sequencing identified 183 differentially expressed genes, including upregulated genes involved in immune and inflammatory responses and downregulated genes associated with neuronal activity and plasticity. Western blotting further showed an increased p-Akt (Ser473)/Akt ratio. These findings reveal that OVA-induced food allergy in female BALB/c mice is associated with neuroimmune dysregulation and broader transcriptional alterations in the PFC involving immune and inflammatory responses and processes related to neuronal activity and plasticity, and suggest that altered Akt-related signaling may provide a molecular link between these changes.

International Journal of Molecular Sciences doi: 10.3390/ijms27219433

Authors: Chenfan Geng Hailiang Song Yinglan Xia Xin Zhang Shuisheng Li Shuang Liu Jinmeng Teng Yong Zhang Hongxia Hu

Large variation in body weight poses a major challenge to the development and industrialization of formulated-feed aquaculture in Siniperca chuatsi. Breeding varieties that combine formulated-diet adaptation, rapid growth, and uniform body weight is therefore important for improving production efficiency. However, the genetic architecture of body weight and optimal genomic selection (GS) strategies remain poorly understood. Here, whole-genome resequencing was performed on 522 formulated-diet-adapted individuals. Genome-wide association study (GWAS) identified 21 significant loci, with candidate genes associated with lipid metabolism, endocrine regulation, and neuroendocrine signaling. Genomic prediction was evaluated using genome-wide single-nucleotide polymorphisms (SNPs) and GWAS-prioritized SNP panels with Bayesian and machine-learning models. Prediction accuracy increased with marker number and generally plateaued at approximately 30,000 SNPs. Random forest using GWAS-prioritized markers achieved the highest accuracy, improving prediction accuracy by 4.8% over genome-wide genomic best linear unbiased prediction (GBLUP) while using fewer markers. These findings demonstrate the potential of GWAS-informed marker prioritization to reduce marker requirements and improve GS efficiency for body weight in formulated-diet-adapted S. chuatsi.

International Journal of Molecular Sciences doi: 10.3390/ijms27219430

Authors: Yanbin Zhang Wenbin Ji Liang Cao Yan Wang Wei Wang Zaiqi Zhang

Madeng’ai, the dried root of Potentilla freyniana Bornm., is used in Dong medicine for infections and wound care. This study examined the chemical composition and prophylactic protective effects of the ethyl acetate fraction of its ethanolic root extract (referred to as MEA) in lipopolysaccharide (LPS)-induced acute lung injury (ALI). Ultra-high-performance liquid chromatography coupled to Q-Exactive Orbitrap mass spectrometry detected 77 constituents, comprising 13 compounds authenticated with reference standards and 64 constituents tentatively annotated from accurate mass and tandem mass spectrometry (MS/MS) fragmentation. Network pharmacology prioritized six candidate compounds, 120 overlapping targets, and nine hub targets, and molecular docking explored theoretical interactions with inflammation-related proteins. MEA was evaluated in LPS-stimulated MH-S alveolar macrophages and in male C57BL/6J mice receiving oral pretreatment for seven days prior to intraperitoneal LPS challenge. In MH-S cells, MEA pretreatment reduced nitrite, interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA)-detectable intracellular oxidative fluorescence, and Nlrp3, Pycard, Casp1, and Gsdmd mRNA levels. In mice, seven-day oral pretreatment with MEA attenuated qualitative histological lung injury, pulmonary edema, and serum cytokine elevation, accompanied by lower lung protein levels of NOD-like receptor family pyrin domain-containing 3 (NLRP3), apoptosis-associated speck-like protein containing a CARD (ASC), pro-caspase-1, and the ~30 kDa N-terminal fragment of gasdermin D (GSDMD-N), together with related transcripts. These findings indicate that prophylactic MEA administration attenuates LPS-induced macrophage inflammation and experimental lung injury, accompanied by reduced intracellular oxidative stress and decreased expression of NLRP3–GSDMD pathway-associated markers.

International Journal of Molecular Sciences doi: 10.3390/ijms27219429

Authors: Mesut Cerit Aysberg Şamil Önlü Ildus I. Ahmetov Özlem Özge Yılmaz Hızarcı Beste Tacal Aslan Metin Dalip Korkut Ulucan

This cross-sectional study examined ACTN3 rs1815739 (C/T; R577X) genotype distribution and its associations with physical performance and biological maturation in male youth football players. A total of 274 players (11–14 years, n = 166; 15–18 years, n = 108) from a professional club in North Macedonia completed Yo-Yo Intermittent Recovery, linear sprint, and jump tests, including the drop jump, countermovement jump (CMJ), and squat jump (SJ). Genotype frequencies were compared with the 1000 Genomes European reference population (n = 503). ACTN3 CC (RR) genotype was more frequent in players than in the reference population (48.2% vs. 32.0%; OR = 1.97, 95% CI 1.46–2.67, p < 0.0001), as was the C (R) allele (OR = 1.34, 95% CI 1.08–1.66, p = 0.009). However, the player genotype distribution deviated from Hardy–Weinberg equilibrium (p < 0.001). CMJ height increased by approximately 2.3 cm per one-year increase in maturity offset (p < 0.001). In the younger cohort, TT (XX) carriers had lower Yo-Yo distance and estimated VO2max than CC and CT (RX) carriers (p = 0.007). Exploratory whole-sample models provided limited evidence that the association between maturity offset, and vertical-jump performance (CMJ, SJ) might differ by genotype; however, these interactions were not statistically significant after correction for multiple testing. No significant genotype differences were observed for sprint performance or most other jump-related outcomes. These findings indicate that ACTN3–performance associations in youth football players are modest and outcome-specific and should be interpreted cautiously given the external reference population and deviation from Hardy–Weinberg equilibrium.

International Journal of Molecular Sciences doi: 10.3390/ijms27219427

Authors: François Chauvigné Daniel Toala-Franco Àngel Rey-Alfonso Marc Catalán-García Roderick Nigel Finn Joan Cerdà

In marine teleosts, the neuroendocrine regulation of aquaporin intracellular trafficking in osmoregulatory organs such as the intestinal rectum remains underexplored. By using paralog-specific antibodies, here we determined the subcellular localization of seven aquaporin and aquaglyceroporin paralogs in the rectal enterocytes of the marine teleost gilthead seabream (Sparus aurata) and investigated the neuropeptinergic effects of arginine vasopressin (Avp) and oxytocin (Oxt) on channel trafficking. The data reveal that Aqp1aa, -1ab2 and -3a are mainly localized in the cytoplasm and basolateral membranes of the enterocytes, while Aqp1ab1, -10bb and -14 are cytoplasmatic and also present in the apical plasma membrane microvilli. In contrast, Aqp0a localizes in the enterocyte subapical cytoplasm. Treatment of rectal explants with Avp in vitro increases the targeting of Aqp1ab1, -1ab2, -10bb and -14, but not of Aqp0a, -1aa and -3a, to the enterocyte apical microvilli, which involves protein kinase A (PKA)-mediated phosphorylation of the channel subdomains. In addition, we find that the differential interaction of the Aqp1ab-type paralogs with teleost-specific 14-3-3ζ-like binding proteins is associated with the trafficking of the channels to the apical microvilli membranes. Conversely, Oxt treatment does not affect the subcellular distribution of any of the investigated aquaporins, except Aqp14, which is internalized in response to the neuropeptide by a partially independent protein kinase C pathway. These data suggest that Avp-PKA signaling controls plasma membrane trafficking of several seabream rectal aquaporins, possibly to facilitate water and solute fluxes across the enterocyte membrane.

International Journal of Molecular Sciences doi: 10.3390/ijms27219428

Authors: Vitaliy V. Chagovets Natalia A. Frankevich Alisa O. Tokareva Anastasia V. Novoselova Marina N. Yushina Fadi Hajjar Tamara E. Karapetyan Irina I. Ryumina Vladimir E. Frankevich Gennady T. Sukhikh

Multiple pregnancy is associated with a high risk of perinatal complications, among which twin-to-twin transfusion syndrome (TTTS) and selective fetal growth restriction (sFGR) are of particular importance. These conditions are specific to monochorionic twins, yet their metabolic consequences in newborns remain poorly understood. Dried blood spots (DBSs) represent a convenient and non-invasive biomaterial for metabolomic analysis. This study included 7 twin pairs with sFGR, 10 pairs with TTTS, and 16 pairs from uncomplicated pregnancies. The amino acid profile in DBSs was determined by HPLC–MS/MS. Statistical significance of variation in normalized intra-pair difference in amino acid levels (ΔS_twins) was assessed using the Mann–Whitney U test with Benjamini–Hochberg false discovery rate correction. The dynamics of ΔS_twins between birth and the corrected full-term age were analysed. In comparing sFGR group with controls, intra-pair differences for glycine and alanine were altered at birth, and for β-alanine and leucine at the second time point. In the TTTS group, deviations from controls were observed for glutamine (at the second time point) and for alanine, α-aminobutyric acid, and aspartic acid (in terms of dynamics). Direct comparison between the two pathologies revealed differences in citrulline (TTTS vs. sFGR at birth) and in several amino acids over time (valine, leucine, ornithine). Dynamic analysis further showed that placental type significantly influenced the postnatal metabolic reorganisation of valine and alanine. These preliminary observations require independent validation and should not be considered validated clinical biomarkers.

International Journal of Molecular Sciences doi: 10.3390/ijms27219423

Authors: Sofia A. Korneva Alexander P. Chernyaev Oleg V. Belov Anna Chigasova Andrey А. Osipov Maxim Ignatov Elizaveta Yashkina Vyacheslav Saburov Natalia Vorobyeva Nikolay Zyuzikov Andreyan N. Osipov

Although ionizing radiation quality shapes the spatial structure and complexity of primary DNA lesions, its contribution relative to absorbed dose in the late DNA damage response (DDR) remains poorly quantified. We analyzed γH2AX and pATM foci kinetics in human mesenchymal stromal/stem cells exposed to 60Co γ-rays, 150 MeV protons, and 14.1 MeV neutrons at doses of 0.25, 0.5, 1.0, and 2.0 Gy, delivered at dose rates of 0.5 Gy/min for γ-rays, 1 Gy/min for protons, and 46 mGy/min for neutrons, respectively. An effective kinetic model was used to describe the time course of the DDR and incorporated an ultraslow component as a model-based index of long-lasting repair burden. Radiation types differed less in early signal amplitude than in decay rate and late persistence. At every investigated dose, the normalized ultraslow fraction followed the order γ-rays < protons < neutrons. This ordering was preserved in per-dose fits, while parametric bootstrapping supported the γ–neutron separation, although the absolute values of the internal fractions were only partially identifiable. RAD51, analyzed independently from model fitting, showed the same qualitative ordering at late time points, supporting the interpretation that radiation quality is more associated with persistent DDR signaling than absorbed dose within the investigated range. These findings suggest that absorbed dose alone may not fully characterize the late cellular response to different radiation qualities and highlight the importance of radiation quality in comparative radiobiology, radiation protection, and the interpretation of normal-tissue responses to therapeutic irradiation.

International Journal of Molecular Sciences doi: 10.3390/ijms27219426

Authors: Prabhu Manickam Hanna Alhoraibi Alaguraj Veluchamy Marilia Almeida-Trapp Naganand Rayapuram Heribert Hirt

In eukaryotes, mitogen-activated protein kinase (MAPK) cascades are highly conserved signaling modules in transducing extracellular signals perceived by receptors/sensors. MAPKs phosphorylate cytoplasmic and nuclear target proteins, modifying their stability, protein–protein interaction, enzyme activity, DNA-binding affinity, and subcellular localization. A comparative phosphoproteomics analysis of wild-type and mapk mutant plants identified a previously uncharacterized protein, USP1, as a novel MAPK target. In this work, we investigated the role of USP1 in pathogen responses. CRISPR-Cas9-generated knockout usp1 mutants displayed enhanced resistance to the bacterial pathogen P. syringae pv. tomato DC3000, reduced apoplast reactive oxygen species (ROS) accumulation, and upregulated defense marker genes such as WRKY29 and FRK1. Transcriptome analyses revealed that immune hormone signaling genes, such as salicylic acid (SA) and jasmonic acid (JA), are differentially regulated. These hormones are responsible for primary defense responses against biotrophic and necrotrophic pathogens, respectively. Accordingly, usp1 mutants show enhanced and reduced resistance to bacterial and fungal infection by Pst DC3000 and Botrytis cinerea, respectively. Biochemical analyses showed that USP1 exhibits chaperone-like activity that stabilizes various proteins, including the MAPKs MPK4 and MPK6. Furthermore, usp1 mutants were defective in flg22-mediated immune priming, indicating that USP1 is required for the establishment of PAMP-induced immune memory. Collectively, our findings identify USP1 as a MAPK-regulated chaperone that plays an important role in coordinating plant immune responses.

International Journal of Molecular Sciences doi: 10.3390/ijms27219424

Authors: Annika Sundlof Michiko Taniguchi Rohit Gupta Mehri Mollaee Francis Cordova Kartik Shenoy Steven Geier Abdelhamid Liacini

The presence of HLA donor-specific antibodies (DSAs), both before and after transplantation, is a well-established risk factor for antibody-mediated rejection (AMR) and poor graft survival. However, emerging evidence suggests that non-HLA antibodies may also provoke immune responses, contributing to rejection. Among these, antibodies targeting the angiotensin II type-1 receptor (AT1R) have been linked to adverse clinical outcomes in kidney and orthotopic heart transplant recipients. This case presents an investigation of non-HLA antibodies in a lung transplant recipient diagnosed with AMR in the absence of HLA DSAs, as well as the use of belatacept as part of a combination immunosuppressive regimen to treat AMR in a lung transplant recipient.

International Journal of Molecular Sciences doi: 10.3390/ijms27219425

Authors: Weishan Zhao Lei Fang Weitian Zhou Xialian Bu Shangong Wu Hong Zou Wenxiang Li Guitang Wang Ming Li

Balantidium ctenopharyngodoni Chen, 1955 is the only ciliate species known to inhabit the digestive tract of grass carp, where it predominantly resides in the spaces between mucosal folds and within the intestinal contents of the hindgut with a high prevalence. Despite its high prevalence, the pathogenicity remains controversial. The intestinal bacterial microbiota interacts closely with enteric protozoa, and such interactions may alter host–parasite dynamics, with consequences for host susceptibility and overall health. In this study, we conducted a feeding experiment with three different dietary regimes to explore the effects of B. ctenopharyngodoni colonization on grass carp, using an integrative approach that combined microbial amplicon sequencing, metabolomics, and intestinal transcriptomics. Our results showed that balantidia colonization was associated with slightly lower body length and body weight, although these differences were not statistically significant. Balantidia colonization was also related to altered microbiota composition and diversity: the relative abundance of Bacteroidetes was more abundant in the balantidia-positive fish with a plant-based diet, while alphaproteobacteria and betaproteobacteria were enriched in the balantidia-positive fish with commercial feed. Although B. ctenopharyngodoni moderately perturbed the microbial profiles and metabolic processes of the grass carp hindgut, intestinal transcriptomic analysis did not detect significant transcriptional changes in immune-related genes between balantidia-positive and balantidia-negative groups. Collectively, under the tested dietary and physiological conditions, our findings provide no evidence of overt pathogenicity, further supporting the view that B. ctenopharyngodoni acts as a commensal ciliate in the grass carp hindgut.

International Journal of Molecular Sciences doi: 10.3390/ijms27219421

Authors: Yuna Wang Di Zhao Huangjin Xiao Leyin Gao Liwen Huang Dan Bai Deming Gou Shuya Zhang

Pulmonary hypertension (PH) is a progressive cardiopulmonary disease characterized by pulmonary vascular remodeling and dysregulated angiogenesis. However, the angiogenesis-related molecular mechanisms underlying PH remain incompletely understood, and reliable diagnostic biomarkers are still lacking. Therefore, To identify angiogenesis-related key molecules in PH through integrated transcriptomic analyses and experimental validation, we aimed to establish a candidate molecular signature and evaluate the discriminative potential of PIK3C2A. Specifically, GSE15197 was used as the primary bulk-transcriptomic discovery dataset, whereas GSE1909 for cross-species validation, and human pulmonary artery single-cell RNA sequencing data (GSE210248) for cellular characterization. Moreover, Functional enrichment, Weighted Gene Co-expression Network Analysis (WGCNA), immune infiltration analysis, and three machine learning algorithms—logistic regression, random forest (RF), and Least Absolute Shrinkage and Selection Operator (LASSO) were performed to identify model genes. In addition, Single-cell analyses were performed using GSE210248, followed by quantitative real-time polymerase chain reaction (qRT-PCR) validation of candidate-gene expression in monocrotaline (MCT) and Sugen5416 plus hypoxia (SuHx) rat models. As a result, Forty-three angiogenesis-related differentially expressed genes (ARDEGs) were identified and were enriched in terms and pathways related to angiogenesis, vascular development, extracellular matrix organization, and HIF-1 signaling. Four model genes (DCN, DKK1, PIK3C2A, and UNC5B) were identified and showed moderate-to-high discrimination between PH and control samples in GSE15197. Among these genes, PIK3C2A showed its highest expression in endothelial cells, while complementary immune-infiltration and CellChat (version 2.1.1) analyses characterized the broader immune–stromal context of the angiogenesis-related signature. qRT-PCR further confirmed significant downregulation of PIK3C2A in the lungs and pulmonary arteries of MCTand SuHxinduced PH models (p < 0.05). In conclusion, An angiogenesis-related candidate molecular signature comprising DCN, DKK1, PIK3C2A and UNC5B was identified in PH. Among these genes, PIK3C2A showed preferential endothelial expression and was downregulated in both the MCT and SuHx rat PH models, supporting its further evaluation as a candidate molecule associated with PH.

International Journal of Molecular Sciences doi: 10.3390/ijms27219422

Authors: Yana Todorova Radostina Dimitrova Teodora Kalfova Steliyan Petrov Martina Bozhkova Mariya Ivanovska Dobrina Dudova Mariana Murdjeva Hristo Taskov Michael Maes Maria Nikolova

Chronic immune dysregulation is the possible pathophysiological mechanism in myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) and long coronavirus infectious disease (long COVID, LC). However, a detailed characterization of regulatory and effector T-cell subsets in these conditions remains limited. To assess and compare T-cell subsets in relation to serum biomarkers of chronic activation and oxidative/nitrosative stress in an attempt to differentiate between LC and ME/CFS patients. Peripheral blood mononuclear cells (PMNC) from LC patients (n = 89), ME/CFS patients (n = 125), and healthy controls (HC, n = 80) were analyzed using multiparameter flow cytometry. The frequencies of naïve (CD45RA+), effector (CD45RA-), CD31+, CD39+, CD73+, GARP+, PI16+, CD15s/CD161+, and CD147+ Treg subsets were determined in relation to CD4+ and CD8+ T-cell differentiation, activation, and effector subsets. Catalase (CAT), superoxide dismutase 1 (SOD1), glutathione peroxidase 1 (GPX1), malondialdehyde (MDA), nitric oxide synthase 1 and 3 (NOS1, NOS3), zonulin, lipopolysaccharide (LPS), immunoglobulin G (IgG) against lipopolysaccharide (IgG-LPS), Toll-like receptor 4 (TLR4), indoleamine 2,3-dioxygenase (IDO), and Dickkopf-related protein 1 (DKK1) were measured in serum by ELISA. Clinical symptom scores (HAMA, HAMD, FF, dePol) were additionally assessed. The immune-serum biomarkers distinguishing LC, ME/CFS, and HC comprised Th17, CD15s+CD161−eTreg, and CD161+ Treg. The five-marker signature retained significance after adjustment for age, BMI, and sex. Altered T-cell subsets were strongly associated with markers of oxidative (GPX1) and nitrosative (NOS3) stress but not directly with symptom severity. ME/CFS showed a strong separation from HC across Random Forest (AUC = 0.907) and logistic regression (AUC = 0.924). The modest distinction of LC from HC (AUC = 0.727 and 0.577 respectively) possibly reflected important endotypic diversity, though differences in disease duration, vaccination history, biomarker selection, and other cohort characteristics might add to it. MDA was an independent predictor for three-class classification, ME/CFS vs. HC, and LC vs. HC pairwise comparisons, while GPX1 discriminated between ME/CFS and LC. In spite of shared biomarkers of oxidative stress and T-cell response dysregulation, our results support the hypothesis that LC and ME/CFS differ in the extent of cellular damage and the exhaustion of compensatory mechanisms.

International Journal of Molecular Sciences doi: 10.3390/ijms27219419

Authors: Daniela Braconi Simona Guerrini Francesca Girolamo Simone Costagli Alessandro Paffetti Alessandro Armini Sara Draghi Gabriella Jacomelli Andrea Bernini Daniela Grasso Viola Galli Donatella Ganucci Lisa Granchi Annalisa Santucci

Indigenous Saccharomyces cerevisiae strains persist within individual cellars and contribute to the character of spontaneously fermented wines, yet the molecular basis of their oenological differences remains largely unknown. In this work, eight predominant strains isolated from spontaneous fermentations in four Tuscan wineries were characterized by technological traits of oenological relevance and laboratory-scale fermentations of both a synthetic must and Sangiovese grape must. In each must, the strains reached similar final ethanol concentrations, although lag phase and maximum fermentation rate differed significantly, whereas the resulting wines differed significantly in glycerol, acetic acid and phenolic alcohol content and in the scores assigned by a trained sensory panel. Four strains spanning the observed range of behaviors were hence analyzed by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and Matrix-Assisted Laser Desorption/Ionization Time-Of-Flight Mass Spectrometry (MALDI-ToF MS) at an early and a mid-fermentation stage, yielding 30–46 differentially abundant proteins identified per strain. Strain-specific abundance of proteins involved in glycerol formation, acetaldehyde oxidation and sulfur amino acid metabolism paralleled the corresponding differences in glycerol, acetic acid and hydrogen sulfide production, whereas glycolytic enzyme abundance varied without any corresponding variation in ethanol. Proteomic analysis thus identified candidate molecular correlates of the traits that distinguish strains, suggesting that it could complement traditional screening in the selection of indigenous starter cultures.

International Journal of Molecular Sciences doi: 10.3390/ijms27219420

Authors: Ayşegül Sümer

Green tea (Camellia sinensis) contains bioactive catechins, particularly epigallocatechin-3-gallate (EGCG), but traditional infusions and concentrated green tea extracts differ in EGCG content, dosing pattern, product matrix, systemic exposure, and safety. This structured narrative review evaluates how fasting affects EGCG pharmacokinetics and the relevance of increased systemic exposure to hepatotoxicity. Its distinctive contribution is not a further summary of green tea safety but an explicit appraisal of the evidence chain that is generally assumed to connect the two: each link is assessed separately, and the steps that are demonstrated in humans are separated from those that rest on biological inference. Standardized catechin preparations or purified EGCG taken after an overnight fast produce substantially higher peak plasma concentrations and total systemic exposure than when taken with food, although the two randomized within-subject comparisons that establish this effect were small, differed in dose and formulation, and assessed neither hepatic exposure nor liver injury. Evidence linking high EGCG exposure to prooxidant effects, mitochondrial dysfunction, and hepatocellular injury derives mainly from cellular and animal models at concentrations that exceed reported human peak plasma concentrations by roughly one order of magnitude; in mice, caloric restriction renders otherwise non-hepatotoxic EGCG doses hepatotoxic. In humans these outcomes must be kept apart. High-dose, long-term extract use raises serum aminotransferases in a susceptible minority, a laboratory abnormality demonstrated in controlled trials; hepatic adverse events cluster among catechin-rich bolus preparations rather than brewed beverages; and clinically apparent liver injury, documented in registry cases and associated with HLA-B*35:01, is idiosyncratic rather than dose-determined. Progression from the laboratory abnormality to clinically apparent injury has not been established. Fasting therefore increases systemic EGCG exposure, particularly from concentrated preparations, but has not been established as an independent cause of hepatotoxicity in humans. Treating fasting as an exposure-modifying condition rather than a hepatotoxic factor and nutritional state as a graded variable rather than a single category clarifies which precautions are evidence-based and which research questions remain genuinely open. Safety assessments should consider product type, dose, bolus administration, fed or fasting state, concomitant exposures, and individual susceptibility.

International Journal of Molecular Sciences doi: 10.3390/ijms27219417

Authors: Yan Wang Bowei Cao Huili Lu Zhicheng Wang Yanli Qin

Tumor cells possess nucleic-acid sensing systems that detect aberrant DNA and RNA arising from genomic and transcriptomic dysregulation. Cytosolic DNA and RNA sensors, together with endosomal nucleic acid sensors, can induce type I interferons, inflammatory chemokines, antigen presentation programs, and immune cell recruitment. Yet many tumors remain immunologically silent despite harboring potentially immunostimulatory nucleic acids, indicating that malignant cells can restrain these pathways at multiple levels. Nucleic acid clearance, selective RNA editing and remodeling, extracellular second messenger degradation, and epigenetic silencing can suppress these responses. These regulatory layers may favor immune-poor tumor states and limit responsiveness to immune checkpoint blockade. Therapeutic reactivation must be spatially and temporally controlled, because persistent signaling can undergo maladaptive rewiring. We review these molecular brakes, therapeutic strategies to restore innate sensing, and a proposed biomarker framework integrating sensor competence, suppressor activity, and functional immune output. We distinguish the effects arising within tumor cells from those mediated by sensing mechanisms in neighboring host cells and discuss cellular and spatial context, signaling dynamics, and safety. The proposed framework for matching treatments to patients remains investigational and requires prospective clinical validation.

International Journal of Molecular Sciences doi: 10.3390/ijms27219418

Authors: Xiaofei Cui Yijun Ren Guohua Ding Sen Chen

Evaluating candidate molecules in diabetic kidney disease (DKD) requires evidence for both the reproducibility of expression changes and their cellular origin. Using the historically fixed SERINC1/TDO2 panel, we reappraised how these candidates were derived, evaluated both genes in human glomerular transcriptomes and kidney-cortex single-cell data, and examined their relationship with renal injury in a mouse model. The historical WGCNA, LASSO, and SVM-RFE-labelled frozen lists intersected at SERINC1 and TDO2, although the legacy RFE implementation could not be confirmed. In the current 83-sample analysis, 584 differentially expressed genes were detected; intersecting the cross-cohort direction-consistent differentially expressed genes with the disease-associated WGCNA module retained 26 candidates that did not include the two targets, whereas machine learning applied to a topology-derived subset of these candidates identified DUSP1. TDO2 was increased in both cohorts (Hedges’ g = 2.07 and 0.714; the latter Holm-adjusted p = 0.0182), whereas the SERINC1 reduction was supported only in the discovery cohort (g = −1.90) and not in GSE96804 (g = −0.252; Holm-adjusted p = 0.348). Single-cell analysis showed widespread SERINC1 distribution without donor-level significance after Benjamini–Hochberg correction, and TDO2 was detected in only 50 of 37,331 cells. In streptozotocin-induced mice, SERINC1 decreased and TDO2 increased alongside renal pathology and podocyte ultrastructural changes. Both genes remain candidates for further study, but clinical diagnostic value and shared pathogenic mechanisms are not yet established.

International Journal of Molecular Sciences doi: 10.3390/ijms27219416

Authors: Waed Mohammad Alnsour Xenie Johnson Münevver Aksoy

Microalgae are some of the organisms that produce carotenoids, which are high value metabolites. However, algae produce carotenoids under stress conditions and there is a need for more practical and economical ways to increase carotenoid accumulation in algae. Haematococcus lacustris has been one of the most important species for the production of industrial-scale carotenoids in biotechnology. In recent years, new algal species have been shown to accumulate carotenoids as alternative resources of carotenoids; Dysmorphococcus globosus is one of them. In a previous study, we isolated and identified for the first time a D. globosus strain in Türkiye. In the current study, we tested different stress conditions on carotenoid accumulation on D. globosus-KINIK and a Haematococcus sp.-KINIK strain that was isolated along with it from the same water sample. We partially sequenced the 18S rRNA gene of the newly isolated Haematoccoccus strain; it is 96% identical to Haematococcus lacustris. Further studies are necessary to determine the species of our strain. We tested three stress conditions: (i) salt stress by adding 2 g L−1 NaCl, (ii) fresh water stress by replacing growth media with fresh water, and iii) sea water stress by replacing growth media with sea water. Salt stress and fresh water stress have been shown to increase carotenoids in Haematococcus strains in the literature; our Haematococcus strain also responded well to these stresses and accumulated carotenoids. To our knowledge, none of these stresses have been tested for Dysmosphococcus strains in the literature. Our results show that D. globosus-KINIK also accumulates carotenoids under salt stress and fresh water, with salt stress being more effective. We also tested different growth media on the growth of D. globosus-KINIK and showed that it can grow without vitamin supplementation. Our study reveals local strains of carotenogenic algae in Türkiye for consideration for industrial-scale carotenoid production.

International Journal of Molecular Sciences doi: 10.3390/ijms27219414

Authors: Yoshino Suzuki Ujjal K. Bhawal Aman Chowdhry Takao Watanabe Hiroyuki Okada Kazunari Kimoto

Hydroxyapatite (HA) is widely used as an osteoconductive bone substitute, and fluoride can influence bone-forming cells in a concentration-dependent manner. This study examined how pretreating resorbable HA with low or high concentrations of sodium fluoride (NaF) affects early mandibular bone healing in beagle dogs. Twelve beagle dogs were used in the project, including three in a pilot study and nine in the main experiment. At each observation time point (Days 4, 7, and 14), one dog was assigned to each of the three HA-based treatment conditions (HA, HA + 0.2% NaF, or HA + 4% NaF). Four experimental defects in each dog received the same assigned treatment, while four contralateral defects were left unfilled as controls. The four defects were considered within-animal subsamples rather than independent biological replicates, and all treatment comparisons were descriptive and exploratory. Before implantation, HA was exposed to the designated NaF solution for 3 min and then washed with 100 mL of distilled water. Healing was evaluated by H&E, Azan, Masson’s trichrome and toluidine blue staining, immunohistochemistry, and quantitative real-time RT-PCR. Histological examination showed progressive tissue and matrix organization around the implanted material, with more evident new bone and osteoblast-associated changes in the low-NaF group at Days 7 and 14. Runx2 immunoreactivity was more prominent in the early healing phase, and osteocalcin was more prominent in the later healing phase. Osteocalcin mRNA gradually increased until Day 14, whereas Runx2 mRNA was maximally expressed at Day 7. Both responses were strongest with low-NaF-pretreated HA. High-NaF-pretreated HA retained osteogenic activity but did not produce a response of the same magnitude. These exploratory findings suggest that NaF pretreatment concentration may influence early HA-associated bone healing, warranting confirmation in studies with independent animal-level replication.

International Journal of Molecular Sciences doi: 10.3390/ijms27219415

Authors: Husam Qanash Aisha M. H. Al-Rajhi Abdulrahman S. Bazaid Fahad Almarshadi Nourah M. Almimoni Hussam A. Althagafi Abdullah M. Almotayri Reham Yahya Sohaila Fathi El-Hawary Abdu Aldarhami

Jojoba oil (Simmondsia chinensis) is widely used in cosmetic and pharmaceutical formulations, yet approaches to enhance its biological activity remain insufficiently explored. This study investigated whether infrared (IR) treatment could modify the chemical profile and in vitro biological properties of jojoba oil. A thermal control matched to the IR temperature–time profile was included to separate bulk heating from IR-specific effects. Crude (untreated) and IR-treated oil (30 min, 50 mW/cm2) were characterized by Fourier-transform infrared spectroscopy (FTIR) and gas chromatography-mass spectrometry (GC-MS), followed by evaluation of antimicrobial, antibiofilm, inhibition of protein denaturation, fibroblast scratch-closure, and antiproliferative activity against SK-OV-3 cells. IR treatment altered the FTIR profile and produced substantial changes in the GC-MS-detectable composition, including the detection of a peak tentatively assigned to 9-hexadecenoic acid, 9-octadecenyl ester (27.45% relative peak area), and 9-eicosyne (8.97% relative peak area); these values represent relative chromatographic abundance, not absolute concentrations. Compared with crude oil, IR-treated jojoba oil exhibited two- to fourfold lower minimum inhibitory concentrations (MIC), reaching 7.8 µg/mL against Candida albicans, and inhibited Staphylococcus aureus biofilm formation by 96.8%. It also enhanced fibroblast scratch closure (84.1% vs. 67.0%) and showed greater inhibition of protein denaturation (IC50, 4.43 vs. 6.52 µg/mL). Against SK-OV-3 ovarian cancer cells, IR-treated oil reduced the IC50 from 114.5 to 78.7 µg/mL and produced marked G0/G1 accumulation (97.9%). This response was accompanied by increased BAX (3.67-fold) and CASP3 (5.73-fold) expression and decreased BCL2 expression (0.61-fold). Collectively, these findings demonstrate that IR treatment, beyond a matched thermal control, substantially modifies jojoba oil and enhances multiple in vitro biological activities. The novelty is the linkage between IR-induced chemical alteration and functional biological changes, not the mere combination of assays.

International Journal of Molecular Sciences doi: 10.3390/ijms27219412

Authors: Wendi Xu Lan Luo Jiaqi Wang Ruirui Ren Jun Zhou Cuiping Wang Ken Qin Yanlong Li Yunxiang Li Xin Zhang Jing Wang Binbin Si Linyuan Duan Guoli Dai

Powdery mildew severely constrains the production of Lycium barbarum L., yet the molecular mechanisms underlying its response to infection remain poorly understood. Here, ‘Ningqi-1’ plants were inoculated with the powdery mildew pathogen and sampled at 0, 12, 24, and 48 hpi for integrated transcriptomic, proteomic, and metabolomic analyses. Quantitative PCR and CRISPR/Cas9-mediated gene editing were further used to evaluate the candidate susceptibility gene LbMLO6. Infection induced extensive molecular reprogramming, yielding 1299 differentially expressed genes with sustained changes, 75 DAPs shared across all three time points, and 127 shared differentially accumulated metabolites. These molecules were mainly enriched in plant–pathogen interaction, plant hormone signal transduction, MAPK signaling, phenylpropanoid biosynthesis, and α-linolenic acid metabolism pathways, suggesting coordinated roles for Ca2+ signaling, reactive oxygen species, defense-related proteins, and secondary metabolism in the host response. Integrated multi-omics analysis identified LbMLO6 as a candidate gene associated with powdery mildew susceptibility, its expression was markedly induced from 24 hpi onward. CRISPR/Cas9-edited LbMLO6 lines showed reduced fungal development compared with the control, particularly during the early stages of infection. These findings provide preliminary evidence that LbMLO6 may be associated with powdery mildew susceptibility in L. barbarum.

International Journal of Molecular Sciences doi: 10.3390/ijms27219409

Authors: Junyu Liu Yan Xia Pengcheng Li Mianyu Liu Xianhong Meng Kun Luo Juan Sui Ping Dai Qiang Fu Jian Tan Jiawang Cao Baolong Chen Xupeng Li Jie Kong Sheng Luan

The Pacific white shrimp (Penaeus vannamei) exhibits pronounced sexual dimorphism in growth, making early and accurate sex identification crucial for targeted monitoring of sexual development and in-depth research into monosex aquaculture. However, morphological sex identification is unreliable at early developmental stages, and single-marker molecular assays are prone to diagnostic failures caused by genotyping errors or missing data. In this study, we developed a multi-SNP framework for sex prediction by integrating genome-wide association studies (GWAS) with Random Forest (RF) modeling. Analyzing 1244 individuals with a 55K SNP array, we identified 18 genome-wide significant SNPs and 10 candidate genes, including SRRM1 and SPSB3, that may be associated with sexual differentiation. Subsequent multi-marker comparative analyses demonstrated that although the top-performing single marker yielded high accuracy, simulated genotyping errors and missing data caused significant drops in accuracy and elevated no-call rates, whereas multi-SNP panels exhibited substantially improved robustness. A stable performance plateau was reached when expanding the panel to 5 SNPs. The model achieved 100% classification accuracy in the two mixed-sex independent populations, ZJ and HN, and correctly identified all females in the all-female MK cohort. Overall, this study provides a robust multi-SNP tool for sex identification in P. vannamei and highlights the practical utility of machine learning in aquaculture.

International Journal of Molecular Sciences doi: 10.3390/ijms27219410

Authors: Dool-Ri Oh Hwan Lee Yonguk Kim Yujin Kim Donghyuck Bae

This study investigated the cognitive-enhancing and neuroprotective effects of a mixed extract of Atractylodes macrocephala Koidzumi (AM) and Rumex acetosa L. (RA) in models of scopolamine (SCO)-induced toxicity and memory impairment. In vitro, the 2:1 ratio maximized neuroprotection and p-CREB/CREB expression in SCO-treated SH-SY5Y cells. In vivo, rats received the AR mixture orally for 29 days, with SCO challenges introduced from Day 22. In the Morris water maze, the AR mixture reduced escape latency during the 3-day spatial training period and increased target quadrant exploration during the probe trial. In the passive avoidance test, it significantly prolonged step-through latency, confirming enhanced memory retention. Hippocampal biochemical assessments revealed that the AR mixture attenuated acetylcholinesterase activity, decreased malondialdehyde accumulation, and replenished acetylcholine levels. Furthermore, the formulation suppressed the pro-inflammatory cytokines IL-1β, TNF-α, and IL-6. AR mixture treatment attenuated the SCO-associated reductions in BDNF, p-TrkB, p-CREB, and PSD-95 expression and in ERK1/2 and Akt phosphorylation in the hippocampus. These findings suggest that the AR mixture improves cognitive and biochemical outcomes in SCO-treated rats, accompanied by changes in cholinergic function, lipid peroxidation, inflammation, and BDNF/TrkB-related signaling.

International Journal of Molecular Sciences doi: 10.3390/ijms27219411

Authors: Iuliana Samoilă Cristian George Țieranu Anca Hermenean Sorina Dinescu

Inflammatory bowel diseases (IBDs) are chronic immune-mediated disorders of the gastrointestinal tract resulting from a complex interplay between genetic susceptibility and dysregulated immune responses. To improve the understanding of the molecular mechanisms’ underlying disease pathogenesis, mouse models with experimental colitis and patients with ulcerative colitis (UC) were investigated for the expression of genes and small non-coding RNAs involved in key pathogenic pathways. In experimental colitis, the potential modulatory effects of STAT6 knockout and Corylus avellana treatment were also evaluated. Our findings revealed significant transcriptional alterations in molecular markers associated with inflammation, epithelial barrier integrity, autophagy, and endoplasmic reticulum stress, highlighting their association with the development and persistence of chronic intestinal inflammation. Both STAT6 deletion and Corylus avellana extract treatment modulated several of these alterations, with STAT6 knockout showing a more pronounced effect on the molecular profile associated with experimental colitis. In addition, miRNA expression profiling revealed a panel of dysregulated miRNAs, while bioinformatic analysis identified potential miRNA-gene regulatory interactions, supporting their possible involvement in the post-transcriptional regulation of these biological processes. Overall, the combined analysis of gene and miRNA expression profiles highlights molecular alterations associated with experimental colitis and human UC and suggests potential regulatory interactions that require further experimental validation.

International Journal of Molecular Sciences doi: 10.3390/ijms27219413

Authors: Hui Yuan Yuxin Zhu Kaibo Jin Yuan Liu Jun Lu Runxiang Zhang

Excessive deposition of abdominal fat in broilers compromises feed efficiency, carcass yield, processing efficiency, and inflates production costs. Because this trait is governed by multi-tissue crosstalk coupling hepatic lipid synthesis to adipose lipid storage, its coordinated regulation has seldom been addressed. We combined three abdominal fat transcriptomic datasets (GSE42980, GSE49121, GSE104039) with one hepatic dataset (GSE104038), obtaining 2520 meta-differentially expressed genes (meta-DEGs). Functional enrichment revealed a prominent enrichment of lipid-metabolism terms, including peroxisome proliferator-activated receptor (PPAR) signaling. Fatty acid-binding protein 3 (FABP3) appeared repeatedly within the Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and gene set enrichment analysis (GSEA) lipid-metabolism categories and was therefore taken forward for functional work. Overexpressing FABP3 enhanced proliferation, lipid accumulation and lipogenic marker expression, whereas knockdown reversed these effects. In both adipocytes and hepatocytes, FABP3 raised peroxisome proliferator-activated receptor gamma (PPARγ) and fatty acid-binding protein 4 (FABP4) at mRNA and protein levels. Pharmacological intervention showed that the PPARγ antagonist GW9662 suppressed the pro-lipogenic action of FABP3 overexpression, while the PPARα agonist GW7647 reinstated lipid accumulation in FABP3-silenced cells, indicating a reliance on PPAR-pathway activity. FABP3 therefore drives lipid deposition in adipocytes and hepatocytes via PPAR signaling, offering a framework for how the liver and abdominal adipose tissue jointly govern abdominal fat deposition in broilers.

International Journal of Molecular Sciences doi: 10.3390/ijms27219405

Authors: Lyudmil Tonchev Nikola Staykov Tsanko Gechev

Cytoplasmic vitrification is the standard explanation for how resurrection plants endure drying, yet the glass is rarely examined in tissue—its composition, glass transition and residual water remain unresolved. We characterize the glassy matrix of Haberlea rhodopensis by integrating transcriptomic and metabolomic data, biophysical modeling and targeted experiments. Sugar-metabolism genes, such as SUCROSE-PHOSPHATE SYNTHASES and SUCROSE SYNTHASES, respond to desiccation with strong concordance across three studies (Kendall’s W = 0.872; 81 genes measured in all three) and, with the metabolites, define a four-component glass-former set—sucrose plus the raffinose family oligosaccharides raffinose, stachyose and verbascose, with sucrose dominant (~92%). Apparent contradictions are plausibly explained: INVERTASE induction is consistent with compartmental separation from accumulating sucrose and declining trehalose with trehalose-6-phosphate (T6P) signaling, not vitrification. A parameterized state diagram traces the desiccation–rehydration path through vitrification and predicts that unfreezable water is locked throughout the glass. Desiccated tissue survived liquid nitrogen (−196 °C), suggesting that freezing tolerance reflects vitrification. Rewetting dried tissue was exothermic, consistent with an amorphous glass. We argue the residual water is functionally bound, that late embryogenesis abundant (LEA) proteins help shape the glass, and hypothesize that natural deep eutectic solvent (NaDES)-forming metabolites provide a transient reaction medium.

International Journal of Molecular Sciences doi: 10.3390/ijms27219396

Authors: Vishit Patel Asmita Raja Madeha Khan Alan A. Melcher Irene Chong Adam Young

Radiotherapy remains a cornerstone of cancer treatment, yet treatment response continues to be limited by tumour radioresistance and considerable biological heterogeneity. Although advances in radiation delivery have substantially improved dose conformity and reduced toxicity, increasing evidence suggests that optimising therapeutic outcomes will require greater integration of tumour biology into treatment strategies. This review examines emerging molecular approaches that seek to enhance anti-tumour responses to radiotherapy beyond direct tumour cell killing. We discuss the DNA damage response as a therapeutic target for radiosensitisation, highlighting the evolving role of DNA damage response inhibitors in promoting both tumour cytotoxicity and immune activation. We further examine the mechanisms linking radiation-induced DNA damage to anti-tumour immunity, including immunogenic cell death, cGAS–STING signalling, and remodelling of the tumour microenvironment, together with the opportunities and challenges of combining radiotherapy with immune checkpoint inhibition. Finally, we review emerging concepts in precision radiation oncology, including spatially tailored irradiation strategies, genomically informed radiation prescription, radiogenomics, and the influence of the gut microbiome on treatment response. Collectively, these advances support a shift from anatomically guided radiotherapy towards biologically personalised treatment, integrating molecular, immunological and genomic information with normal-tissue susceptibility to improve the therapeutic ratio and enable more precise, patient-centred cancer care.

International Journal of Molecular Sciences doi: 10.3390/ijms27219406

Authors: Nargiz Sultanova Elshan Musazade Serife Topkaya

Sharka disease, induced by the Plum pox virus (PPV; genus Potyvirus, family Potyviridae), represents one of the most serious viral threats to stone fruit production worldwide. This study assessed PPV occurrence in major fruit-growing areas in Azerbaijan and characterized the physiological and oxidative stress responses of infected Prunus species. Symptomatic leaves from orchards in the Quba district were screened by immunochromatographic strips, DAS-ELISA, and RT-PCR. Of 60 trees surveyed, 22 (36.7%) tested positive across all three hosts, confirmed by RT-PCR amplification of a 243 bp coat protein (CP) gene fragment in peach (Prunus persica L.), plum (P. domestica L.), and apricot (P. armeniaca L.). Infection was associated with pronounced oxidative stress: hydrogen peroxide (H2O2) and malondialdehyde (MDA) contents were higher in infected trees (6.80–10.60 µmol g−1 FW and 8.0–15.2 nmol g−1 FW) than in healthy trees (3.20–3.90 µmol g−1 FW and 4.5–8.2 nmol g−1 FW). Antioxidant defenses were activated, with increased ascorbate peroxidase (APX) and catalase (CAT) activities and higher ascorbic acid (AsA) and reduced glutathione (GSH) levels. Photosynthetic performance declined markedly: total chlorophyll fell by 17.3–30.8% (greatest in apricot), maximum photosystem II quantum efficiency (Fv/Fm) was lower in infected trees (0.408–0.561) than in healthy trees (0.716–0.728), and total soluble protein was lower in infected trees (7.88–8.45 mg g−1 FW) than in healthy trees (9.38–9.58 mg g−1 FW). Collectively, PPV infection was associated with oxidative stress and lipid peroxidation, with activation of enzymatic and non-enzymatic antioxidant defenses, and with reduced photosynthetic performance in Prunus species. These findings advance understanding of host responses to PPV and support sustainable monitoring and management strategies for stone fruit production, aligned with SDG 2 (Zero Hunger) and SDG 15 (Life on Land).

International Journal of Molecular Sciences doi: 10.3390/ijms27219408

Authors: Michał Bednarski Robert Kupczyński

Cytotoxic necrotizing factor (CNF)-associated Escherichia coli (E. coli) occur in enteric and extraintestinal disease in calves, but the distribution of CNF1 and CNF2 and their virulence profiles remain incompletely defined. We analysed E. coli from 544 calves: 131 healthy, 210 diarrhoeic, 173 with septicaemia, and 30 with central nervous system (CNS) infection. Isolates were classified by PCR detection of cnf1 or cnf2, and only CNF-positive isolates were screened for eight accessory virulence genes. In age-adjusted models restricted to calves aged 2–19 days, overall CNF positivity (aOR 2.83, 95% CI 1.25–6.44; Holm-adjusted p = 0.039) and CNF2 positivity (aOR 3.14, 95% CI 1.18–8.31; adjusted p = 0.043) were associated with diarrhoea, whereas CNF1 was not. CNF2-positive isolates were enriched for f17A, afa8E, and cdtIII, whereas CNF1-positive isolates more often carried papC, sfa/focDE, hlyA, and kpsMT K1. In invasive cases, CNF1 was more frequent in CNS infection than in septicaemia (33.3% vs. 12.7%; adjusted p = 0.034). Among CNF1-positive invasive isolates, kpsMT K1 occurred in 90.0% of CNS versus 9.1% of septicaemia isolates. These findings support distinct CNF2-associated enteric and CNF1-associated invasive virulence profiles; PCR detection does not establish expression, phenotype, causality, or genomic linkage.

International Journal of Molecular Sciences doi: 10.3390/ijms27219407

Authors: Javier Crespo Marta Alonso-Peña Carolina Jiménez-González Lorena Cayón-González Paula Iruzubieta

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a systemic disorder in which cardiovascular, hepatic and oncological outcomes shape long-term prognosis. Statins are particularly relevant because their cardiovascular benefit is firmly established, yet they remain underused in MASLD, while their potential liver-related effects remain uncertain. Observational studies consistently associate statin exposure with slower fibrosis progression, fewer major adverse liver outcomes, lower hepatocellular carcinoma incidence and reduced liver-related mortality. However, direct randomized evidence in MASLD remains limited, and available studies differ substantially in disease stage, exposure, endpoint and follow-up. In this narrative review, we critically examine the cardiovascular, hepatic, oncological, randomized, observational, genetic and pharmacological evidence on statin therapy in MASLD. We aim to clarify the apparent discordance between evidence sources, assess whether statins may modify liver-related outcomes beyond their established cardiovascular benefit, and propose a framework for evaluating multidomain therapeutic effects. We propose outcome convergence as a framework for assessing whether a therapy can improve more than one clinically relevant disease domain while preserving independent causal evaluation of each outcome. Statins should therefore be regarded as a candidate, rather than an established, outcome-convergent therapy.

International Journal of Molecular Sciences doi: 10.3390/ijms27219403

Authors: Bing Bing Fan An Rong Zhang Zhen Long Wu Ying Yang

To broaden the antibacterial spectrum of yak gastric lysozyme (YGL) against Gram-negative bacteria and improve its secretory expression in Pichia pastoris, this study employed a combined strategy of functional short-peptide fusion and host engineering. Four hydrophobic peptides, FFVAP, GVSLANWVCLAK, VVYP, and AWVAWK, were individually fused to the C-terminus of YGL, and the enzymatic activity and antibacterial performance of the modified proteins were compared. AW-YGL was subsequently selected for sequential optimization of the host strain, secretion signal peptide, endoplasmic-reticulum-associated molecular chaperones, and clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9)-mediated host-gene knockout, followed by physicochemical stability evaluation of the optimized product. The results showed that AWVAWK modification produced the most pronounced enhancement of YGL activity against Gram-negative bacteria. AW-YGL exhibited inhibition rates of 65%, 68%, and 67% against Escherichia coli, Pseudomonas aeruginosa, and Salmonella enteritidis, respectively. During expression-system optimization, the GS115 host significantly improved AW-YGL secretion, with total enzyme activity and specific activity in the culture supernatant reaching 3.1 × 105 U/L and 983 U/mg, respectively. Replacement with the HSS signal peptide further increased total enzyme activity to 4.3 × 105 U/L and specific activity to 1264 U/mg. Co-expression of the endoplasmic reticulum molecular chaperone binding immunoglobulin protein (BIP) further promoted functional AW-YGL expression, increasing the specific activity to 1560.3 U/mg. On this basis, knockout of the cell-wall-related gene FKS1 further increased the total enzyme activity in the culture supernatant to 6.9 × 105 U/L and the specific activity to 1638.3 U/mg, indicating that stepwise optimization of the signal peptide, folding assistance, and host-cell engineering effectively enhanced AW-YGL secretory expression and enzymatic activity. The optimized AW-YGL retained relatively high activity over defined pH and temperature ranges and showed tolerance to several metal ions and digestion-related proteases. Collectively, C-terminal fusion with hydrophobic short peptides enhanced the inhibitory activity of YGL against Gram-negative bacteria, while the combined strategies of signal-peptide optimization, folding assistance, and cell-wall engineering further improved its secretory expression in P. pastoris. These findings provide an experimental basis for the engineering and scalable production of novel broad-spectrum lysozymes.

International Journal of Molecular Sciences doi: 10.3390/ijms27219404

Authors: Jong-Woan Kim Woo Sub Yang Minji Park Hyun Jung Koh

Chronic active Epstein–Barr virus disease (CAEBV) is a rare, potentially life-threatening EBV-positive T/NK-cell lymphoproliferative disorder characterized by persistent systemic inflammation and multiorgan involvement. Although hemophagocytic lymphohistiocytosis and progression to lymphoid malignancy are well-recognized complications, vascular manifestations—including vasculitis, arterial dilatation and aneurysm formation, intracranial hemorrhage, and vascular involvement of multiple organ systems—remain poorly characterized. Available evidence suggests that persistent activation of EBV-infected T/NK cells, cytokine-mediated hyperinflammation, endothelial injury, and coagulation abnormalities may contribute to CAEBV-associated vasculopathy, whereas the roles of complement activation and thrombotic microangiopathy remain less well established. This review synthesizes the clinical spectrum and proposed mechanisms of vascular complications in CAEBV, with particular attention to the strength of evidence supporting individual pathogenic pathways. Improved recognition of vascular manifestations, application of contemporary CAEBV diagnostic criteria, and further mechanistic and multicenter studies are needed to improve risk stratification and management.

International Journal of Molecular Sciences doi: 10.3390/ijms27219402

Authors: Susmi Varghese Prathik Basthikoppa Shivamurthy Vinitha Ramanath Pai Sowmya Soman Poornima Ramesh Rajesh Raju

Glucose metabolism is organized through interconnected metabolic pathways whose enzymes determine whether glucose is utilized for energy production, storage, or biosynthesis. Although protein kinases modulate metabolic enzymes through phosphorylation, how kinase signaling collectively interfaces with the glucose metabolic network remains unclear. In addition, many reported kinase–substrate relationships are derived from prediction-based resources and lack experimental validation. Here, we systematically mapped kinase associations for 144 enzymes across six glucose metabolic pathways by integrating kinase–substrate databases, quantitative phosphoproteomics from 154 human studies, and protein–protein interaction networks. We developed a multi-layer prioritization framework combining kinase–substrate annotations, phosphoproteomic co-regulation, and interaction-supported evidence to rank kinase–enzyme associations and reconstruct a phosphoproteomic-supported kinase–metabolism network. Prioritization reduced a large initial association space to a refined set of candidate kinase–enzyme associations while retaining broad coverage across glucose metabolism. The resulting network revealed an uneven distribution of kinase associations across metabolic pathways and enzymes, with the highest kinase association density observed at metabolic control points, particularly enzymes integrating branch-point and flux-control functions. Pathway-level analysis further showed that glycogen metabolism exhibited the greatest kinase association density despite comprising relatively few enzymes, while futile-cycle pairs displayed distinct patterns of shared and reaction-specific kinase associations. Network analysis identified a small subset of highly connected kinase hubs together with bridge proteins linking the kinase and metabolic enzyme layers, providing a systems-level view of kinase association patterns across glucose metabolism. Together, these findings indicate that kinase associations are not uniformly distributed across glucose metabolism but preferentially concentrate at metabolic control points that govern pathway routing and glucose allocation. This study provides a systems-level framework for prioritizing candidate kinase–enzyme associations and a resource for investigating signaling–metabolism interactions in metabolic disease and cancer.

International Journal of Molecular Sciences doi: 10.3390/ijms27219401

Authors: Gaoyang Qu Mingwan Sun Shixiang Wei Jiaming Yang Hongbo Li Jiushi Liu

Schisandra chinensis (Turcz.) Baill. (S. chinensis) is a traditional Chinese medicinal plant whose fruits are rich in bioactive constituents, including lignans, organic acids, and flavonoids. Nevertheless, the biochemical dynamics underlying fruit development remain insufficiently characterized, limiting the effective cultivation, harvest, and utilization of S. chinensis. Here, we applied widely targeted metabolomics, together with a dynamic assessment of fruit growth, to profile temporal changes in major metabolites and to identify the optimal harvest window. The results show that the morphological changes in fruit mainly occurred from 70 to 110 days after fruit set; fresh fruit mass peaked at 110 days after fruit set. During the harvest period, the fruits are deep red and firm, showing no signs of softening. Meanwhile, fruit cluster weight, single fruit weight, and fruit longitudinal diameter all reached their peak values. Over the course of maturation, lignan levels declined markedly, whereas terpenoids, organic acids and flavonoids increased significantly. The most pronounced shifts were observed for schizandrin, gomisin N, shikimic acid, citric acid, gamma-terpinene and cyanidin 3-sophoroside, key marker compounds of S. chinensis fruit. Network pharmacology predicts that ESR1, RXRA, GATM, NCOA2, and SONEA are core targets. These findings provide a basis for determining timely harvest and for establishing quality evaluation criteria for S. chinensis.

International Journal of Molecular Sciences doi: 10.3390/ijms27219400

Authors: Andrea Guala Fabio Tognon Giuseppe Minniti Fabio Sirchia Silvia Kalantari Michela Malacarne Chiara Baldo Alessandro Ferretti Carla Liaci Davide Politano Roberto De Icco Giorgio Roberto Merlo Enza Maria Valente Cesare Danesino

Cri du Chat syndrome (CdCS) results from a variable-sized deletion of the short arm of chromosome 5 (5p) and is characterized by marked phenotypic heterogeneity. Previous genotype–phenotype correlation studies have primarily focused on deletion size, an approach inherently limited by the large number of genes within the deletion. A gene-centered strategy may therefore offer a better framework for interpreting specific clinical features. Among candidate genes on 5p, TRIO plays a critical role in RAC1-mediated neurodevelopment; furthermore, in related neurodevelopmental disorders, loss-of-function variants are associated with microcephaly, whereas gain-of-function variants are associated with macrocephaly. Clinical, cytogenetic, and molecular data were retrieved from the main database of the Italian charity A.B.C. (Associazione Bambini Cri du Chat). Fifty-two patients, classified as carriers of one or two copies of the TRIO gene, were included based on the availability of head circumference (HC) measurements, developmental milestones (sitting, walking, and first words) data, and a CGH array. All data were analyzed in relation to TRIO allele dosage, deletion size, and sex. All patients showed a reduced HC compared with reference curves. However, HC measurements were consistently higher in patients carrying two TRIO alleles versus those carrying only one. Deletion size did not correlate with HC or developmental milestones. Patients with two TRIO alleles generally achieved motor and language milestones earlier, and some differences between males and females, suggested the possibility of sex-specific differences. These findings indicate that TRIO allele dosage might contribute to head size and developmental outcomes in CdCS independent of deletion size. A gene-centered approach could thus improve genotype–phenotype correlations in chromosomal disorders and highlight the importance of considering sex-specific effects in clinical and rehabilitative management.

International Journal of Molecular Sciences doi: 10.3390/ijms27219399

Authors: Maja A. Zaczek-Moczydłowska Bartosz Kopyciński Izabella Biesiada Purusottam Mishra Krzysztof Matus Saeid Ghavami Ewa Langer Marek J. Łos

Bioactive antiviral biomaterial coatings capable of rapidly inactivating respiratory viruses while preserving cytocompatibility are highly desirable for development protective and high-contact biomedical materials. Here, we engineered black (Bl) organic coatings functionalized with two silver (Ag)-based inorganic additives—nano (n) Ag supported on colloidal silica (SiO2) and Ag-substituted zirconium phosphate—to develop multifunctional antiviral biomaterial surfaces. Their antiviral efficacy was evaluated against human coronavirus (HCoV) 229E, a model enveloped respiratory coronavirus, together with their cytocompatibility toward human keratinocytes (HaCaT) and human lung fibroblasts (MRC-5) cells. The morphology and elemental composition of the coatings were characterized using scanning electron microscopy (SEM) coupled with energy-dispersive X-ray spectroscopy (EDS). Antiviral performance was quantified by a neutral red uptake cytopathic (NRU-CPE) infectivity assay following 30 and 60 min of virus–surface contact. Coatings functionalized with nAg supported on SiO2 reduced infectious virus recovery by 2.2–5.3 log10, whereas coatings containing Ag-substituted zirconium phosphate achieved reductions of 1.9–4.7 log10, depending on additive loading and exposure time. Interestingly, the unmodified black organic coating also reduced viral infectivity, indicating that both the coating matrix and Ag-based functionalization contribute to the overall antiviral performance. Importantly, all functionalized coatings maintained cell viability above the ISO 10993-5 acceptance threshold (≥70%), demonstrating excellent cytocompatibility.

International Journal of Molecular Sciences doi: 10.3390/ijms27219398

Authors: Paweł Zieliński Magdalena Frąszczak Bartosz Czech Paula Dobosz Kinga Adamiak Patryk Mierzejewski Rafał Stępień Magda Mielczarek

Reference genome and transcriptome are fundamental components of RNA-seq analysis and can influence transcript quantification and downstream differential expression results. Although GRCh38 is the current human reference build, GRCh37 remains in use in research and clinical bioinformatics workflows. The extent to which reference transcriptome choice affects transcriptomic inference in breast cancer remains insufficiently characterized. We performed a comparative analysis of publicly available RNA-seq data comprising two MCF7 breast cancer cell-line samples and two healthy breast tissue samples from the European Nucleotide Archive (PRJEB4829). Identical preprocessing, transcript quantification, and differential expression workflows were applied using GRCh37- and GRCh38-based reference transcriptomes. Reference transcriptome-dependent variability was further examined by comparing the same samples quantified under the two reference transcriptomes. Concordance of differentially expressed genes and pathway enrichment was assessed, and sensitivity to reference transcriptome choice was evaluated for genes included in the MammaPrint and Naderi expression signatures and selected hereditary cancer gene panels. The MCF7-versus-normal-tissue comparison identified 15,950 differentially expressed genes using GRCh37 and 16,348 using GRCh38, with 13,899 genes shared between the two analyses. The most significantly enriched Hallmark and KEGG pathways were also concordant between annotations. Principal component analysis showed that the dominant pattern of variation in the MCF7-versus-normal-tissue comparison was associated with biological status. In contrast, when the same MCF7 samples were quantified using different reference transcriptomes, reference transcriptome choice represented a major source of variation. Expression of genes comprising the evaluated prognostic signatures and hereditary cancer panels showed no significant differences in median expression between reference transcriptomes. However, individual cancer-relevant genes, including PTEN, RAD51D, NANOG, and SOX11, demonstrated reference transcriptome-dependent expression differences. In this proof-of-concept analysis, reference transcriptome choice produced measurable differences in RNA-seq quantification and differential expression, while broad biological patterns and the aggregate expression of established gene signatures were relatively stable. Reference transcriptome-dependent variability became particularly apparent when biological heterogeneity was limited, suggesting that reference transcriptome choice can act as a hidden analytical variable in transcriptomic studies. These findings support explicit reporting and version control of reference annotations and highlight the need for validation in larger, independently replicated breast cancer cohorts before drawing clinical conclusions.

International Journal of Molecular Sciences doi: 10.3390/ijms27219397

Authors: Kristen A. Marcellus Michal Scur Shahad Abdulmawjood Gen-Sheng Wang David Prescott Jith Thomas Nikia Ross Santokh S. Gill

The degradation of plastic in the environment can result in the generation of micro- and nano-plastics. As one of the common types of plastics used, polyethylene terephthalate (PET) microplastics are pervasive in the environment. Yet, their potential toxicity remains poorly understood. In this study, we addressed this gap by examining systemic toxicity in C57BL/6 mice following 28-day oral gavage with irregularly shaped PET microplastics at human-relevant exposure levels of 0, 0.3, 1.5, 7.5, or 37.5 mg/kg bodyweight/day. No significant changes were seen in bodyweights, food consumption, organ weights, clinical biochemistry, or hematological parameters. Histological changes were seen in the kidneys of male mice with minimum to mild apoptosis in the outer strip of the outer medulla. Additionally, decreased goblet cells were observed in both male and female mice, albeit in different intestinal sections. However, this change in goblet cell population did not result in overt epithelial damage. Overall, sub-chronic PET microplastic exposure did not cause overt systemic toxicity, but sex-specific minor histological alterations in the kidney and gut were observed. Our findings show only minor effects following short-term exposure, but real-world microplastic exposure is likely to be chronic, thus a longer exposure period may be necessary to fully elucidate the potential impact of PET microplastic-induced toxicity.

International Journal of Molecular Sciences doi: 10.3390/ijms27219394

Authors: Ellen Hertz Gani Perez Krystyna Rytel Martha Kirby Stacie Anderson Stephen Wincovitch Tim Ahfeldt Joel Blanchard Ying Hao Ziyi Li Kate Andersh Yue Andy Qi Ellen Sidransky Yu Chen

Comparative studies of dopaminergic (DA) neurons differentiated from induced pluripotent stem cells (iPSCs) derived from patients discordant for parkinsonism, especially siblings, provide a valuable model to explore factors underlying Parkinson disease (PD) pathogenesis. However, such studies are complicated by the inherent heterogeneity in differentiation efficiency among iPSC lines derived from different individuals. To address this technical challenge, we devised a selection strategy to enrich DA neurons expressing tyrosine hydroxylase (TH). A neomycin resistance gene (neo) was inserted at the C-terminus of the TH gene, placing its expression under the endogenous promoter. Using iPSC-derived neurons from a healthy individual (HT711), we achieved selective enrichment of TH+ neurons through geneticin selection without compromising differentiation capacity, as demonstrated by Western blots, immunocytochemistry and flow cytometry. We then applied the method to iPSCs derived from two siblings of HT711, one with Gaucher disease (GD) alone and one with both PD and GD. Quantification of multiple DA neuron markers through whole-cell proteomics confirmed their comparable enrichment in the three lines, demonstrating that the new method can be applied to iPSC lines from different individuals to standardize DA neuron cultures. While we focus on methodological developments here, a companion analysis of the same whole-cell proteomics dataset, examining molecular features potentially associated with disease statuses, has been reported separately.

International Journal of Molecular Sciences doi: 10.3390/ijms27219395

Authors: Nadica S. Đorđević Ilija M. Dragojević Milica Mijović Jelena T. Todić Dragan Marjanović Aleksandar M. Đorđević Zvezdan Milanović Julijana Rašić Bojana Kisić Zlatibor Anđelković Miljana Z. Stašević Milena M. Kostić

Photobiomodulation (PBM) may improve diabetic wound repair, but the temporal relationship between local angiogenic signaling and tissue-level healing remains incompletely defined. We examined previously unreported tissue vascular endothelial growth factor (VEGF) and histopathological outcomes from 72 ulcer-bearing male Wistar rats from a previously reported experimental cohort, using a balanced 2 × 2 × 3 design comprising metabolic status, PBM treatment, and healing day (3, 7, or 10; n = 6 per cell). Tissue VEGF was quantified by ELISA and normalized to total protein; histopathological outcomes included exploratory inflammation/injury and tissue repair composite scores. In the factorial log-VEGF model, diabetes × day (p = 0.0017) and LLLT × day (p = 0.0010) interactions demonstrated phase-dependent responses. On day 3, PBM-treated animals had an adjusted VEGF ratio of 2.13 versus untreated animals (95% CI 1.63–2.78; Holm-adjusted p < 0.001), whereas diabetic animals had lower VEGF than healthy animals (ratio 0.50, 95% CI 0.39–0.66; adjusted p < 0.001). VEGF concentrations converged by day 10. PBM remained associated with a higher exploratory tissue repair score after adjustment for healing day, diabetes status, and contemporaneous VEGF, whereas VEGF itself was not independently associated with repair. These findings demonstrate phase-dependent differences in tissue VEGF and suggest that contemporaneous VEGF concentrations do not fully explain treatment-associated variation in exploratory histopathological repair scores.

International Journal of Molecular Sciences doi: 10.3390/ijms27219393

Authors: Laísa M. Rocha Danyel F. Contiliani Vanessa Thomé Claudia B. Monteiro-Vitorello Silvana Creste

Sugarcane and energy cane (Saccharum spp. hybrids) yields are significantly threatened by smut disease, caused by the fungus Sporisorium scitamineum. Gene expression studies are essential for understanding molecular responses in this pathosystem; however, reference gene stability may vary among genotypes, plant organs, and across infection stages, highlighting the need for systematic validation under the specific conditions of each experiment. In this study, eight candidate genes were assessed in the buds of sugarcane IACSP02-1064 (intermediate resistance) and energy cane Vertix 10 (susceptible) during the course of the infection (12, 24, 48, and 120 hpi). Expression stability was assessed using geNorm, NormFinder, and BestKeeper algorithms. For IACSP02-1064, ACAD (Acyl-CoA dehydrogenase family member 10) and GAPDH (Glyceraldehyde-3-phosphate dehydrogenase) were the most stable genes, whereas GAPDH/ACAD and ACAD/UBQ2 (Ubiquitin 2)were identified as optimal pairs. For Vertix 10, GAPDH, UBQ2, and ACAD showed the highest stability, while GAPDH/UBQ2, ACAD/GAPDH, and ACAD/SARMp1 were recommended for normalization. The selected reference genes were further evaluated by analyzing ScSPL19 (SQUAMOSA promoter-binding protein-like 19 from Saccharum spp.) expression, a member of the SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE (SPL) transcription factor family, whose members have previously been reported to respond to smut infection, using the most and least stable reference gene pairs. This analysis revealed differential temporal regulation of ScSPL19 when normalized with the most stable reference gene pair (GAPDH/ACAD). These results support the suitability of this reference gene combination for accurate relative gene expression analysis under the evaluated conditions.

International Journal of Molecular Sciences doi: 10.3390/ijms27209392

Authors: Esra Tekin Meliha Koldemir Gündüz Zeynep Yığman Beyza Aydoğan Kara Ayşegül Küçük Mustafa Arslan

Metal dyshomeostasis contributes to Alzheimer’s disease (AD) by influencing amyloid aggregation, oxidative imbalance, and impaired clearance mechanisms. Safranal, a bioactive constituent of Crocus sativus, exhibits antioxidant properties, but its metal-modulating actions remain insufficiently understood. This study investigated whether safranal modulates metal (Fe, Cu, Zn) levels in an AD rat model while evaluating its cytoprotective rescue capacity against Aβ1-42 induced neurotoxicity in differentiated SH-SY5Y cells. AD was induced in male rats via intracerebroventricular streptozotocin, followed by a four-week safranal administration (0.5 mg/kg/day, ip). Behavioral performance, plasma metal levels (ICP-MS), and cerebral total antioxidant capacity (FRAP) were evaluated alongside histological assessment. Post-treatment protocols were employed to evaluate the effects of safranal on cell viability and apoptosis in differentiated SH-SY5Y cells exposed to Aβ1-42. Longitudinal analyses revealed that safranal attenuated exploratory deficits and recognition memory impairments in AD rats. Decreased plasma iron levels in the AD group were restored by safranal, accompanied by a return of cerebral antioxidant capacity to normal. Although overt hippocampal histological alterations were absent in this early phase, post-treatment with safranal successfully offered a therapeutic rescue window, preserving cell viability and attenuating Aβ1-42-induced apoptotic features in vitro. Safranal demonstrates beneficial neuroprotective effects in AD models by improving behavioral outcomes, partially correcting plasma iron levels, and mediating cytoprotective rescue windows against Aβ1-42-induced cellular toxicity. These findings suggest that safranal represents a promising natural candidate for reducing behavioral deficits, supporting peripheral iron status, and offering functional cell survival in AD models.

International Journal of Molecular Sciences doi: 10.3390/ijms27209391

Authors: Teodora Bianca Tirla-Sirca Mariana Eugenia Mureșan Luciana Dobjanschi Tamás Ilyés Rodica Anamaria Negrean Laura Grațiela Vicaș Corina Cinezan Elisabeta Cristina Crăciun (Rusu) Camelia Mraz Alexandra M. Crăciun

Vascular calcification and atherosclerotic arterial disease are complex processes involving multiple inhibitory and pro-calcific mechanisms. Matrix Gla protein (MGP) is an important inhibitor of vascular calcification, and its inactive dephosphorylated–uncarboxylated form (dp-ucMGP) has been proposed as a biomarker of cardiovascular risk. The present study aimed to evaluate the association of circulating dp-ucMGP, total MGP (tMGP), and Fetuin-A concentrations as markers for calcification associated with the presence of fibrocalcific plaques and/or arterial stenosis. A total of 160 patients were included in this observational study. Patients were classified according to the presence of fibrocalcific plaques and/or arterial stenosis. The study group included 136 patients, whereas 24 patients without fibrocalcific plaques or stenosis served as the control group. dp-ucMGP concentrations were significantly higher in the study group [1.29 vs. 1.07, p = 0.047]. In contrast, no significant differences were observed for tMGP [15.79 vs. 12.67, p = 0.532] or Fetuin-A [388.50 vs. 403.05, p = 0.894]. Glucose concentrations were significantly higher in the study group (p < 0.001), together with significant differences in fibrinogen, urea, creatinine, LDL cholesterol, and uric acid. Spearman correlation analysis showed weak positive correlations between dp-ucMGP and urea (ρ = 0.175, p = 0.034), creatinine (ρ = 0.183, p = 0.026), and triglycerides (ρ = 0.197, p = 0.017). Higher circulating dp-ucMGP concentrations were associated with the presence of fibrocalcific plaques and/or arterial stenosis, whereas tMGP and Fetuin-A did not differ significantly between groups. These findings support the potential relevance of dp-ucMGP as a biomarker associated with fibrocalcific and stenotic arterial disease. Further studies are required to determine its independent predictive value and clinical utility.

International Journal of Molecular Sciences doi: 10.3390/ijms27209390

Authors: Jilei Qiao Shuran Chen Shuo Li Wenbo Shao Yiran Zhang Ying Huang Yangjing Xie

The molecular features shared by non-alcoholic fatty liver disease (NAFLD) and myocardial infarction (MI) remain incompletely understood. We analyzed six microarray cohorts from the GEO database to identify shared disease-associated transcriptomic features across independent NAFLD and MI populations. Weighted gene co-expression network analysis, differential expression analysis, and fuzzy c-means clustering across ordered cross-sectional NAFLD groups were used to define candidate genes. LASSO regression, random forest, and SVM-RFE identified consensus computational candidates, which were examined using exploratory classification, receiver operating characteristic curves, calibration curves, decision curve analysis, and SHapley Additive exPlanations. IDI1 was selected for exploratory expression assessment by qPCR in patients with established MI and in a mouse model of acute ischemic myocardial injury. Fifty-three shared candidate genes were identified, and five genes were retained by all three feature-selection methods. Single-gene ROC analyses showed SULF2, GADD45A, LIN7A, and IDI1 within the analyzed case-control datasets. IDI1 expression was higher in patients with MI than in healthy controls (p = 0.004), as well as in mice blood (p = 0.034) and infarcted myocardium (p = 0.0072) following LAD ligation. These findings indicate shared transcriptomic links between NAFLD and MI, supporting an association between IDI1 and myocardial injury.

International Journal of Molecular Sciences doi: 10.3390/ijms27209388

Authors: Esra Kaya Elif Okumuş Elif Gülsüm Torun

Carbapenem-resistant Klebsiella pneumoniae (CRKP) causes serious infections in intensive care units (ICUs), but the relationship between carbapenemase gene profiles and mortality remains uncertain. This prospective two-center cohort included 94 adult ICU patients with clinical CRKP infection from April to September 2026. The first clinical isolate from each patient was included. blaKPC, blaNDM, blaVIM, blaIMP, blaOXA-48, and blaCTX-M genes were investigated by polymerase chain reaction. The primary outcome was 28-day ICU mortality; 14-day and final ICU mortality were secondary outcomes. blaOXA-48, blaNDM, blaKPC, and blaCTX-M were detected in 65 (69.1%), 49 (52.1%), 19 (20.2%), and 73 (77.7%) of isolates, respectively; blaNDM and blaOXA-48 co-occurred in 39 (41.5%). Fourteen-day, 28-day, and final ICU mortality rates occured in 38 (40.4%), 48 (51.1%), and 53 (56.4%), respectively. No statistically significant associations were observed between individual genes, the presence of two carbapenemase genes, or carbapenemase profiles and any mortality endpoint; adjusted analyses of 28-day ICU mortality yielded similar findings. In the exploratory Firth model, the profile effect was not significant (p = 0.198), whereas a higher age-adjusted Charlson Comorbidity Index and bloodstream infection or ventilator-associated pneumonia, compared with urinary tract infection, were associated with higher odds of 28-day ICU mortality. Frequent gene co-occurrence was observed; however, clinically meaningful gene–mortality associations cannot be excluded because of imprecise estimates.

International Journal of Molecular Sciences doi: 10.3390/ijms27209387

Authors: Enxhi Shaba Claudia Landi Ilaria Vicenti Lorenza Vantaggiato Francesco Saladini Camilla Biba Niccolò Bartolini Maurizio Zazzi Luca Bini

West Nile Virus (WNV) is a significant global health threat, and the molecular mechanisms of its pathogenic activity in human hosts remain partially understood. This study applies an extensive proteomic approach combining “top-down” (2DE/MS) and “bottom-up” (LC-MS/MS) technologies to investigate protein changes in the Huh7 human hepatoma cellular model at 48 h post-infection with WNV. Our results demonstrate that WNV infection induces a massive downregulation of host proteins, particularly those involved in cell–cell adhesion and extracellular matrix (ECM) integrity (FN1, VTN, desmosome proteins and tight junctional proteins), which supports theories of blood–brain barrier disruption. Moreover, we highlighted a modulation of iron homeostasis (FTH1, IRP1, and IREB2) reflecting a “nutritional immunity” host response aimed at limiting host enzyme functions essential for virus replication. In particular, the top-down approach demonstrated a significant protein species modulation related to the unfolded protein response (UPR) (HSP90, HSPA8), as well as the hijacking of translation initiation factors (EIF1, eIF3 complex, and EIF4A1), which suggests viral strategies for immune evasion and efficient replication. Taken together, these findings offer a comprehensive overview of WNV-induced perturbations, revealing the quantitative modulation of proteins and potential post-translational signatures, thereby providing a robust foundation for identifying new antiviral drug targets and therapeutic strategies.

International Journal of Molecular Sciences doi: 10.3390/ijms27209389

Authors: Hayato Oku Yusuke Nishiguchi Tomofumi Yamamoto Jun-Ichiro Jo Yoshiya Hashimoto Hirohito Kubo Toshihiko Takenobu

Temporomandibular joint osteoarthritis (TMJ-OA) involves inflammation and degeneration of mandibular condylar cartilage and subchondral bone. Dedifferentiated fat cells (DFATs) were established from buccal fat pad tissue obtained from a single human donor. We examined the effects of DFAT-derived extracellular vesicle preparations (DFAT-EVs) on inflammatory macrophage responses in vitro and disease-related outcomes in TMJ-OA in vivo. DFAT-EVs were applied to lipopolysaccharide- and interferon-γ-stimulated RAW264.7 macrophages and administered intra-articularly in rats with monosodium iodoacetate-induced TMJ-OA. Fluorescence signals from labeled DFAT-EVs were observed in RAW264.7 cells. DFAT-EVs reduced nitric oxide production and modulated nitric oxide synthase 2, interleukin (IL)-1β, IL-6, IL-10, IL-4, and tumor necrosis factor-α responses, although effects varied among inflammatory mediators. Head withdrawal threshold was significantly higher in the DFAT-EVs group than in the OA group at several time points in the 6-week cohort, but not in the 4-week cohort. Administration of DFAT-EVs attenuated the reduction in mandibular condylar bone volume fraction and was associated with preservation of cartilage architecture and matrix. Osteoarthritis Research Society International scores were significantly lower in the DFAT-EVs group than in the OA group at 4 weeks but not at 6 weeks. These in vitro and in vivo findings support further evaluation of DFAT-EVs for TMJ-OA.

International Journal of Molecular Sciences doi: 10.3390/ijms27209385

Authors: Xuyang Chen Xuejing Li Yiyang Zhu Yantao Liang Xing Ren Shumeng Zhang Junyi Tao Kailiang Zou Longfei Lu

A novel lytic bacteriophage, designated vB_ValS-L9, infecting the potentially pathogenic bacterium Vibrio alginolyticus L4, was isolated from a coastal aquaculture area in Beihai, China. Transmission electron microscopy revealed that vB_ValS-L9 possesses an icosahedral head (56.50 ± 4.19 nm in diameter) and a long tail (143.03 ± 5.32 nm in length), exhibiting a typical siphovirus-like morphology within the class Caudoviricetes. The phage demonstrated stability across various temperatures, pH levels, and salinities. Genomic sequencing showed that vB_ValS-L9 has a circular double-stranded DNA genome of 44,025 bp with a G + C content of 43.56% and 61 predicted ORFs. No tRNA, virulence, or antibiotic resistance genes were identified. Genomic and phylogenetic analyses, including ANI, VICTOR, VIRIDIC, and conserved protein phylogenies, identified vB_ValS-L9 as a novel species that, together with Vibrio phages Artemius and VapS30, forms a new genus in the class Caudoviricetes, for which the name Artemiovirus is proposed. Furthermore, in vivo trials using juvenile pearl oysters (Pinctada martensii) revealed that vB_ValS-L9 provides substantial prophylactic and therapeutic protection against V. alginolyticus L4 infection, especially when the phage was administered before bacterial challenge. Collectively, these findings suggest that vB_ValS-L9 shows potential as a safe, environmentally robust, and promising biocontrol candidate for managing vibriosis in aquaculture.

International Journal of Molecular Sciences doi: 10.3390/ijms27209386

Authors: Elif Tuğçe Güner Ayfer Bakır Cemal Çiçek Murat Aral

Carbapenem-resistant Klebsiella pneumoniae (CRKP) is a major healthcare threat, and carbapenemase genotype may influence ceftazidime–avibactam (CZA) susceptibility. This study evaluated the association between carbapenemase gene profiles and CZA resistance in CRKP isolates. In this retrospective laboratory-based study, 184 CRKP isolates (92 CZA-susceptible and 92 CZA-resistant) were analyzed. Carbapenemase genes were detected by multiplex real-time PCR, and gene distributions and profiles were compared according to CZA susceptibility. NDM was detected in 82.6% of CZA-resistant isolates but in none of the susceptible isolates (q < 0.001). All CZA-susceptible isolates carried a single carbapenemase gene, whereas 68.5% of resistant isolates carried two or more genes. NDM + OXA-48-like was the predominant profile among resistant isolates (63.0%), while OXA-48-like alone predominated among susceptible isolates (92.4%) (both q < 0.001). NDM showed 82.6% sensitivity and 100% specificity for discriminating CZA resistance, with an AUC of 0.913. Amikacin resistance was also higher among CZA-resistant isolates (60.9% vs. 31.5%; q < 0.001). NDM, particularly when co-occurring with OXA-48-like, was strongly associated with CZA resistance. Rapid carbapenemase genotyping may complement phenotypic susceptibility testing and support antimicrobial treatment decisions.

International Journal of Molecular Sciences doi: 10.3390/ijms27209384

Authors: Roksana Dwornik Dagmara Cylwik Katarzyna Białkowska

Cervical cancer (CxCa) remains one of the most common malignancies in women worldwide. Although almost all cases are associated with chronic infection with oncogenic human papillomavirus (HPV) types, not every infected woman develops the disease, indicating a significant role of genetic factors. This study reviewed 131 studies on genes related to immune system function and their impact on the risk of developing cervical cancer. The majority of the studies analyzed were conducted in Asian populations, primarily Chinese and Indian. The analysis focused in particular on polymorphisms in genes involved in the antiviral response, inflammatory processes, antigen presentation, and regulation of lymphocyte activity. Most of the evidence points to the importance of human leukocyte antigens (HLA) genes, Toll-like receptors (TLRs), interleukins (ILs), tumor necrosis factor alpha (TNF-α), and interferons (IFNs). Variants in these genes may influence the effectiveness of HPV clearance, susceptibility to persistent infection, and the risk of progression of precancerous lesions to invasive cancer. It is worth emphasizing that identifying genetic risk factors may, in the future, support the development of personalized prevention strategies, screening tests, and the assessment of the individual risk of disease. Nevertheless, most available findings warrant further validation in large and diverse populations. In summary, cervical cancer is the result of a complex interaction between HPV infection and the host’s genetic predisposition. Research on immune response genes provides valuable information about the mechanisms of disease development, and may contribute to improved methods of prevention and early detection.

International Journal of Molecular Sciences doi: 10.3390/ijms27209382

Authors: Ziyi Qi Chenyang Wu Qiheng Qian Ziyang Zeng Jingfa Xiao Dongyu Zhao

Spatial transcriptomics enables investigation of cell–cell communication (CCC) while preserving spatial context, yet large-scale resources that organize spatial CCC results across studies and make them readily searchable, visualizable, and comparable remain limited. Here, we present CCCView, a knowledge platform integrating 699 spatial transcriptomics samples and 7,473,444 inferred CCC interactions across 24 tissues, 182 annotated cell types, and 27 biological conditions in human and mouse. CCCView uses a unified cell-type annotation workflow and organizes results from three complementary CCC-inference approaches—COMMOT, CellNEST, and SpatialDM—while preserving method-specific outputs. The platform provides interactive search, browsing, visualization, disease-oriented exploration, data download, and online analysis functions. For cross-sample exploration, CCCView also includes MuCIA, a Tweedie regression-based statistical module for comparing inferred interaction strength across user-defined sample groups. A glioblastoma use case illustrates how CCCView can connect spatial cell-type context, inferred communication patterns, and condition-associated signals to generate biologically interpretable hypotheses. CCCView provides an integrated resource for systematic exploration of spatial transcriptomic CCC and for reuse of precomputed communication results across heterogeneous datasets.

International Journal of Molecular Sciences doi: 10.3390/ijms27209380

Authors: Yue Cai Ryu Yamada Yuki Marugame Satoshi Yano Fumihiko Okumura Haruhisa Kikuchi Yuzuru Kubohara Taichi Hara

Novel anticancer agents are needed to overcome therapeutic resistance, and autophagy represents a promising target. The chlorinated alkylphenones Dictyostelium differentiation-inducing factors 1 and 3 (DIF-1 and DIF-3) and their derivatives exert anti-proliferative and anti-metastatic effects; however, their molecular mechanisms remain incompletely understood. Using the GFP-LC3-RFP-LC3ΔG autophagic flux reporter, we identified Bu-DIF-3 and CP-DIF-3 as autophagy-promoting derivatives and characterized their actions principally in HeLa cells, with selected autophagy and AMP-activated protein kinase (AMPK) measurements in HAP1 cells. Both suppressed mTORC1 signaling, activated AMPK, and reduced mitochondrial membrane potential (MMP). Bu-DIF-3 additionally induced reactive oxygen species and redox stress response genes, including SLC7A11 and CHAC1. SLC7A11 induction may reflect a compensatory antioxidant response. Unlike CP-DIF-3, Bu-DIF-3 increased the FerroOrange-detectable labile Fe2+ signal, and deferoxamine partially restored the reduced MTT signal, supporting possible iron involvement in reduced metabolic activity. Thus, modification of the DIF-3 methoxy group modulates autophagy-promoting activity and associated stress responses. Bu-DIF-3 was associated with oxidative stress and altered labile Fe2+ homeostasis; whether a ferroptosis-related mechanism contributes remains unresolved. These findings provide a cellular basis for further evaluation of Bu-DIF-3 and related DIF derivatives.

International Journal of Molecular Sciences doi: 10.3390/ijms27209381

Authors: Dana Dildabayeva Diana Samarkhanova Larissa Lezina

Clinical use of PARP inhibitors (PARPi), including olaparib, has improved outcomes for patients with BRCA1/2-mutant and homologous recombination (HR)-deficient tumors. However, acquired resistance remains a major limitation to their efficacy. CRISPR-Cas9 screening approaches have been developed to identify genetic drivers of PARPi resistance. However, differences in experimental conditions, library coverage, and selection design complicate comparisons across studies. In this review, we present a harmonized reanalysis of CRISPR knockout olaparib resistance screens to identify recurrent resistance-associated genes and pathways. Datasets with raw data were collected from the BioGRID ORCS database. Resistance hits identified in at least two screens were subjected to Reactome pathway-enrichment analysis and organized into functional modules. Using this approach, we identified eight modules: DNA-damage repair and replication stress; cell-cycle regulation; chromatin remodeling and histone modification; post-translational regulation; mitochondrial function; TGF-β and mTOR signaling; RNA processing and nucleocytoplasmic transport; and emerging adaptive mechanisms. Importantly, mitochondrial function genes were observed as recurrent hits, but their relation to olaparib resistance is underinvestigated. Our integrative analysis suggests that PARPi resistance is a complex, adaptive phenotype shaped by recurrent but context-dependent vulnerabilities across multiple cellular programs. This framework prioritizes candidate resistance drivers for validation and highlights directions for the design of combination therapies aimed at overcoming PARPi resistance.

International Journal of Molecular Sciences doi: 10.3390/ijms27209383

Authors: Elena N. Subcheva Alexandra A. Martynova Darya S. Volkova Andrey V. Zvyagin Alessandro Parodi Alexey V. Ermakov

Unique physicochemical properties of ferritin nanoparticles, such as high biocompatibility, precise structural assembly, and modular surface chemistry make them a promising biomimetic platform for the delivery of therapeutic and diagnostic agents, including radioactive isotopes. However, the translation of ferritin from laboratory-scale synthesis to clinical applications remains hindered by the lack of standardized analytical workflows and requires precise qualitative and quantitative analysis of core loading, structural integrity, and long-term stability. This review systematically examines current characterization techniques ranging from morphological evaluation and quantitative elemental analysis to advanced spectroscopic and functional performance assays for evaluating metal and non-metal encapsulation efficiency, release kinetics, stability in complex biological media, and functional performance of ferritin-based nanocarriers. We highlight the correlation between the analytical metrics and biomedical outcomes, addressing the critical challenges in standardization necessary for clinical translation. The discussed methodologies establish a methodological roadmap for the rational design and quality control of ferritin-based theranostic systems, facilitating their path toward clinical implementation.

International Journal of Molecular Sciences doi: 10.3390/ijms27209379

Authors: Marcos González-Pérez Citlali Gómez-Márquez Miguel Ortiz-Flores Marycarmen Godínez-Victoria Nayelli Nájera Guillermo Ceballos

Several molecules have been investigated for their potential to improve muscle health in models of muscle damage simulating sarcopenia, injury, or aging. Some small molecules induced beneficial effects on skeletal muscle by stimulating the apelin receptor (APLNR). This receptor has emerged as a promising pharmacological target for modulating muscle homeostasis. Using an in silico approach, we recently identified 11β-hydroxypregnenolone (11β-OHP5) as a potential skeletal muscle (SKM) modulator. We hypothesized that 11β-OHP5 interacts with and activates APLNR. This study employed several research approaches: (1) in silico molecular docking and molecular dynamics to explore the feasibility of 11β-OHP5 interacting with APLNR; (2) in vitro activation analysis to APLNR; (3) in vitro analysis C2C12 myoblast differentiation; (4) analysis of the molecule’s presence in mouse skeletal muscle and its differential presence between young and older mice. The results show that 11β-OHP5, acting through APLNR, positively affects myoblast differentiation and may participate in the exercise-induced benefits in aging mice. We show that 11β-OHP5 interacts with APLNR, induces myogenic responses, is found in SKM; its concentration decreases with age, and exercise restores it. More work is necessary to understand the physiology of this possible endogenous molecule.

International Journal of Molecular Sciences doi: 10.3390/ijms27209377

Authors: Paul-Ștefan Panaitescu Vlad-Alexandru Toma Claudia-Andreea Moldoveanu Remus Moldovan Ioana Bâldea Irina Camelia Haranguș Carmen Costache Simona Clichici Gabriela-Adriana Filip

Parkinson’s disease (PD) remains incompletely understood despite its high social and economic burden, with oxidative stress, neuroinflammation, and mitochondrial dysfunction recognized as central pathogenic mechanisms. 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a complex I inhibitor, is widely used as a model of dopaminergic neurodegeneration, while growing evidence highlights the gut–liver–brain and kidney–brain axes as important contributors to PD progression. Within this framework, trimethylamine N-oxide (TMAO) has emerged as a molecule of interest, linked to dementia risk and proinflammatory effects. Our study aimed to evaluate the peripheral oxidative and inflammatory consequences of acute and semi-acute MPTP exposure, alone and in combination with TMAO. In total, 96 male CD-1 mice were equally divided across two experimental settings: a semi-acute protocol (30 mg/kg i.p., MPTP once daily for five consecutive days) and an acute protocol (20 mg/kg i.p., four administrations of MPTP at two-hour intervals), with TMAO administration remaining identical across both experimental settings. The animals were randomized into four groups (control group, a TMAO-only group (40 mg/kg/day, gavage, days 0–42), an MPTP-only group, and a combined TMAO + MPTP group), with MPTP showing consistently depleted liver SOD activity across both models. In the liver, MPTP showed consistently depleted SOD activity in both models, whereas glutathione disruption and lipid peroxidation were most evident in the combined TMAO + MPTP group. These effects were explained mainly by the additive main effects of MPTP and TMAO, as no significant MPTP × TMAO interaction was found for any oxidative stress marker. Nonetheless, a significant MPTP × TMAO interaction was confirmed for liver and kidney IL-1β in the acute model. IL-1β and IL-6 were significantly elevated in the liver and kidney by both MPTP alone and its combination with TMAO, with the combined groups consistently showing the largest increases (TMAO + MPTP semi-acute vs. control and vs. TMAO, p < 0.001–0.01; TMAO + MPTP acute vs. all groups, p < 0.0001–0.01). In the kidney, the contribution of MPTP and TMAO to oxidative stress was model-dependent: in the semi-acute protocol, both MPTP and TMAO contributed to increased MDA and glutathione ratio disruption, while in the acute protocol, TMAO alone was the dominant driver of SOD depletion and MDA elevation, with their combination further amplifying IL-6 secretion. TMAO also independently drove lipid peroxidation in the heart and intestine. The acute MPTP model produced more extensive redox imbalance independently of TMAO, whereas in the semi-acute model, a comparable degree of redox disruption was only achieved through combined TMAO + MPTP exposure.

International Journal of Molecular Sciences doi: 10.3390/ijms27209378

Authors: Ewelina Zielińska Anna Jakubczyk Kamila Jaglińska Beata Polak

Edible insects are increasingly recognized as a viable alternative food source, highlighting the urgent need for systematic research into their integration into conventional food products. This study examined the effect of adding mealworm powder (Tenebrio molitor) to crackers, focusing on factors such as color, texture, nutritional value, antioxidant properties, starch digestibility, and molecular structure. As the level of mealworm powder increased (from 2% to 10%), the crackers became darker, while instrumental hardness and fracture time decreased, indicating that a more brittle structure formed. Nutritionally, the enriched crackers contained higher protein, fat, and mineral content, while their carbohydrate content decreased; moreover, their protein profile was modified. Antioxidant activity and total phenolic content also rose in proportion to the level of mealworm enrichment. FTIR analysis revealed differences in spectral regions associated with proteins, lipids, and carbohydrates, indicating compositional and local molecular-environment changes following mealworm powder incorporation. Mealworm powder incorporation markedly decreased rapidly digestible starch and increased slowly digestible starch, whereas the predicted glycemic index showed only a modest but statistically significant reduction at higher supplementation levels. Furthermore, sensory evaluation indicated that consumer acceptance was optimal at a low inclusion level of 2%. Overall, mealworm powder represents a promising ingredient for improving selected nutritional and functional characteristics of crackers; however, its inclusion level should be optimized to maintain sensory quality.

International Journal of Molecular Sciences doi: 10.3390/ijms27209374

Authors: Roisin McMorrow Henriette S. de Bruijn Ivo Que Karishma Lila Thierry P. P. van den Bosch Michail Doukas Clemens W. G. M. Löwik Laura Mezzanotte Dominic J. Robinson

Photodynamic therapy (PDT) is a minimally invasive treatment whose efficacy is strongly influenced by tumor oxygenation and the tumor microenvironment (TME). Pancreatic ductal adenocarcinoma (PDAC) is characterized by severe hypoxia, dense stroma, and immune suppression, limiting therapeutic responsiveness. In this study, we investigated how Bremachlorin-based PDT remodels the TME in two murine PDAC models with distinct immune phenotypes (T-cell–high and T-cell–low) and compared them with the immunogenic MC38 colon carcinoma model. Single-fiber reflectance and fluorescence spectroscopy were used to monitor tumor physiology and photosensitizer dynamics during PDT, while multiplex immunohistochemistry enabled spatial profiling of stromal and immune components post-treatment. PDAC tumors exhibited significantly lower baseline oxygenation than MC38 tumors, which was not substantially improved by reducing the PDT fluence rate from 116 to 50 mW·cm−2. Nevertheless, PDT reduced collagen type I content in both PDAC models, indicating stromal remodeling. Importantly, illumination with a moderate fluence rate of 50 mW·cm−2 significantly increased CD8+ T-cell infiltration and improved the CD8+/neutrophil ratio in T-cell–high PDAC tumors, suggesting a shift toward a less immunosuppressive immune landscape. These findings demonstrate that Bremachlorin-PDT can modulate the PDAC tumor microenvironment and highlight the importance of immune context in shaping PDT outcomes, supporting rational optimization of PDT-based combination therapies.

International Journal of Molecular Sciences doi: 10.3390/ijms27209376

Authors: Abhishek Mishra Md. Abdul Hye Khan Ripa Jamal Ilham Kadhim John D. Imig

Vascular endothelial growth factor (VEGF) signaling is a central regulator of tumor angiogenesis and a major therapeutic target in oncology. VEGF-neutralizing agents and VEGF receptor tyrosine kinase inhibitors (VEGFR-TKIs) confer clinical benefit in multiple solid malignancies, but resistance and cardiovascular and renal toxicities can limit durable treatment. This review summarizes VEGF ligands, VEGF receptors, and the principal signaling mechanisms relevant to angiogenesis and lymphangiogenesis, with emphasis on VEGFR2 and VEGFR3 biology. We then examine established VEGF-targeted therapies, representative clinical evidence for VEGFR-TKIs, mechanisms of therapeutic resistance, and VEGF inhibitor-associated hypertension, endothelial dysfunction, proteinuria, and kidney injury. Particular attention is given to strategies for improving the therapeutic index of VEGF pathway inhibition, including risk assessment and toxicity monitoring, immunotherapy combinations, and emerging mechanism-based approaches. Modulation of the epoxyeicosatrienoic acid (EET)/soluble epoxide hydrolase axis is discussed as a promising experimental strategy for cardiorenal protection; however, current support is predominantly preclinical and requires clinical validation. Integrating tumor vascular biology with mechanisms of normal vascular and renal homeostasis may facilitate safer and more effective VEGF-directed cancer therapy.

International Journal of Molecular Sciences doi: 10.3390/ijms27209375

Authors: Zihan Wang Junchen He Ting Zhang Jiayu Peng Lulu Xie Bingrun Li Cunhao Shan Litao Zhang Zuotao Zhao

Bruton tyrosine kinase (BTK) is a central signaling molecule that links the B-cell receptor (BCR), Fc receptors, and Toll-like receptors (TLRs). By coordinating signaling in B cells, mast cells, macrophages, and other immune cells, BTK occupies an important position in cutaneous immune responses. BTK inhibitors (BTKis) can inhibit B-cell and Fc-receptor-dependent effector-cell signaling, providing a mechanistic rationale for several immune-mediated skin diseases. However, these cellular effects should not be equated with suppression of established autoantibody production or durable disease modification. This review summarizes the biological functions of BTK and critically evaluates clinical and translational evidence in chronic spontaneous urticaria (CSU), atopic dermatitis (AD), pemphigus, psoriasis, and hidradenitis suppurativa (HS). Evidence is strongest for short-term symptom control in CSU cases. Clinical value remains uncertain in pemphigus and HS, rilzabrutinib did not meet its primary AD efficacy endpoint and topical atuzabrutinib showed no exploratory efficacy benefit, and psoriasis evidence is preclinical. Priorities include prospective validation of predictive biomarkers, disease-specific confirmatory trials, agent- and setting-specific safety assessments, and long-term and real-world follow-up.

International Journal of Molecular Sciences doi: 10.3390/ijms27209372

Authors: Georgiana-Mihaela Cebuc Simeanu Ioana-Gabriela Dragne Adina Mitrea Diana Clenciu Diana-Cristina Protasiewicz-Timofticiuc Alina-Liliana Popa Denisa-Maria Mitroi Bianca-Florentina Pîrvu Ion-Cristian Efrem Adina Dorina Glodeanu Adina-Andreea Mirea Adela-Gabriela Ștefan Maria-Magdalena Roșu Theodora-Claudia Radu-Gheonea Beatrice-Elena Vladu Ana-Maria Efrem Lorena-Anda Dijmarescu Mircea-Catalin Fortofoiu

Obstructive sleep apnea (OSA) and metabolic dysfunction-associated steatotic liver disease (MASLD) frequently coexist and share a common cardiometabolic substrate through a shared cardiometabolic substrate. This narrative review critically appraises the epidemiological, experimental, and interventional evidence proposed to link chronic intermittent hypoxia (CIH) and sleep fragmentation to hepatic steatosis, steatohepatitis, and fibrosis. We summarize the candidate mechanisms, including mitochondrial oxidative stress, hypoxia-inducible factor 1-alpha signaling, sympathetically driven lipolysis, and Kupffer and stellate cell activation and then examine the limits of the evidence supporting them: hypoxia-induced hypophagia and the scarcity of pair-fed controls in rodent models, species differences in the physiological response to hypoxia, publication bias, common-cause confounding by age and systemic inflammation, and the rarity of progressive liver disease in lean patients with anatomically driven OSA. We further review non-invasive diagnostic tools, polysomnographic hypoxemia indices, transient elastography, METS-IR, FIB-4, and emerging biomarkers, such as omentin-1 and follistatin-like 1, that enable bidirectional screening between sleep and hepatology clinics. Finally, we discuss therapeutic strategies, including continuous positive airway pressure, structured weight loss, and incretin-based pharmacotherapy (GLP-1 and dual GIP/GLP-1 receptor agonists), highlighting recent phase 3 evidence for tirzepatide in OSA and semaglutide in metabolic dysfunction-associated steatohepatitis. Current evidence supports CIH as a context-dependent amplifier of liver injury in patients with pre-existing metabolic dysfunction rather than as an established independent cause of MASH. This interpretation still justifies integrated screening and treatment pathways, while identifying the study designs needed to test causality directly.

International Journal of Molecular Sciences doi: 10.3390/ijms27209370

Authors: Ismail Alsalloum Ekatherine I. Terentieva Vitalii S. Moskaliuk Sophia M. Antonova Tatiana V. Ilchibaeva Alexander V. Kulikov

Tyrosine hydroxylase (TH) catalyzes the hydroxylation of L-tyrosine to L-3,4-dihydroxyphenylalanine (L-DOPA), which is the rate-limiting step in dopamine (DA) synthesis in the brain. Certain mutations in the human TH gene are associated with severe neurological and psychiatric disorders. The c.886C>T mutation in the mouse Th gene, which results in the R278H substitution, decreases TH activity in the mouse midbrain. Here, we compared the molecular characteristics of the wild-type (TH-278R) and mutant (TH-278H) recombinant forms of TH. Both recombinant proteins were expressed in Escherichia coli BL21 cells and purified by immobilized metal affinity chromatography using Ni-NTA agarose resin. TH activity was assayed by HPLC, and the thermal stability of the recombinant proteins was assessed by differential scanning fluorimetry. The R278H substitution markedly decreased Vmax and increased the KM for the cofactor 6,7-dimethyltetrahydropterin (DMPH4), but had no effect on the KM for the substrate L-tyrosine. The substitution did not affect the T50 of thermal inactivation but markedly increased the Tm of thermal unfolding of the TH-278H protein. The substitution decreased the IC50 values for dopamine and 1,2-dihydroxybenzene several-fold, both of which contain a catechol moiety that is also present in L-DOPA. These results indicate that the R278H substitution alters the sensitivity of TH to catechol-containing compounds. Although direct evidence is lacking, endogenous L-DOPA produced during the reaction may interact differently with the catalytic center of the TH-278H mutant than with that of the wild-type TH-278R.

International Journal of Molecular Sciences doi: 10.3390/ijms27209373

Authors: Sayeed Khan Ashtyn Vogt Osman Khan Jeremy Tan Sanober Nusrat

Plasminogen deficiency is an ultra-rare disorder of fibrinolysis that most commonly presents as ligneous conjunctivitis. Ophthalmic manifestations often precede systemic diagnosis, making ophthalmologists essential to early detection and management. We report two contrasting cases illustrating the spectrum of disease and therapeutic response. A 65-year-old woman with untreated type 1 plasminogen deficiency developed recurrent ligneous conjunctivitis leading to enucleation and multisystem morbidity, while a 4-year-old girl with autosomal dominant type 1 disease achieved sustained remission following systemic plasminogen replacement therapy (Ryplazim). These cases highlight the evolution of management from supportive care to disease-modifying therapy and the critical importance of early recognition. Long-term outcomes and treatment durability remain poorly defined, underscoring the need for registry-based collaboration to better characterize disease progression, therapeutic response, and quality-of-life impact in plasminogen deficiency.

International Journal of Molecular Sciences doi: 10.3390/ijms27209369

Authors: Senwen Yao Zhenyuan Lu Guosheng Zhao Yang Liu Shan Wu Jiaxuan Zhang Renjie Dong Yangming Hu Ye Zhang Yang Wang

Acute skeletal muscle repair depends on a finely tuned balance between myogenic regeneration and fibrotic remodeling. Macrophages are essential to this process, and specific subsets have been linked to fibrosis in some organs. The M4-like macrophage, a distinct polarization subset implicated in fibrotic diseases, has not been well characterized in the context of injured muscle, particularly regarding its impact on fibroblast activation and myogenic suppression. The study aims to explore the effects of M4-like macrophages in the repair of muscle injury and possible mechanism. We analyzed a murine single-cell RNA-sequencing dataset from cardiotoxin (CTX)-injured skeletal muscle to identify injury-expanded macrophage subsets and their molecular programs. Immunohistochemistry and immunofluorescence staining of muscle sections were used to assess CXCL4 expression and platelet (CD41+) infiltration in injured versus uninjured tissues. CXCL4-induced M4-like macrophages were established as an in vitro model to examine their effects on L929 fibroblasts and C2C12 myoblasts. The MMP12–SPP1 interaction was predicted by AlphaFold3 and validated by co-immunoprecipitation and immunofluorescence colocalization. Loss-of-function and rescue experiments targeting MMP12 and SPP1 were performed to determine their causal roles in fibroblast activation and myogenic suppression. Transcriptomic and biochemical analyses were used to define downstream macrophage-intrinsic NF-κB/TNF-α signaling. Finally, AAV-mediated macrophage-targeted SPP1 knockdown and pharmacological NF-κB inhibition were evaluated in the CTX injury model. Using a BALB/c mouse model of cardiotoxin (CTX)-induced acute skeletal muscle injury and a publicly available murine scRNA-seq dataset from CTX-injured skeletal muscle, we observed an injury-induced MMP12/SPP1-positive macrophage subset with an M4-like transcriptional signature. Mechanistically, co-immunoprecipitation assays and immunofluorescence colocalization supported an association between MMP12 and SPP1. MMP12 knockdown attenuated the pro-fibrotic and anti-myogenic effects of M4-like macrophages, whereas recombinant SPP1 largely restored these phenotypes. Furthermore, transcriptomic and biochemical analyses revealed that autocrine SPP1 activated NF-κB signaling in M4-like macrophages, and increased TNF-α secretion. Pharmacological inhibition of NF-κB signaling with QNZ reduced fibroblast activation and restored the expression of myogenic markers. However, QNZ is not TNF-α specific and may affect multiple cell types. In vivo, AAV-mediated macrophage-targeted SPP1 knockdown driven by the F4/80 promoter, as well as QNZ treatment in the CTX-induced muscle injury model, markedly decreased collagen deposition, enhanced myogenic regeneration, and improved functional recovery. These findings identify an MMP12/SPP1/NF-κB signaling pathway in M4-like macrophages, with TNF-α as a potential downstream mediator, that may contribute to an imbalance between regeneration and fibrosis and may represent a therapeutic target for enhancing skeletal muscle repair after acute injury.

International Journal of Molecular Sciences doi: 10.3390/ijms27209371

Authors: Hua Zhang Zhengxu Cai

MicroRNAs (miRNAs) are a class of short single-stranded non-coding RNAs widely present in multicellular eukaryotes, including animals, plants, and fungi. Typically, approximately 22 nucleotides in length (with some ranging from 22 to 25 nucleotides), miRNAs are predominantly localized to non-coding genomic regions and exhibit high evolutionary conservation. As key post-transcriptional modulators of gene expression, miRNAs are non-protein-coding molecules. They bind complementarily to the 3′UTR of target mRNAs. This interaction triggers mRNA breakdown or inhibits protein translation. In this way, miRNAs regulate nearly 30% of cellular protein expression. MiRNA biogenesis is tightly regulated. Abnormal miRNA expression alters disease-related genes and signaling pathways. It further drives the onset, development, and pathological processes of various human diseases. This review outlines recent progress in research on miRNA functions across multiple illnesses. These illnesses cover neurological, metabolic, tumor, and cardiovascular disorders.

International Journal of Molecular Sciences doi: 10.3390/ijms27209368

Authors: Weijie Liao Naihan Xu

Ubiquitin-specific protease 30 (USP30) is a mitochondrial outer membrane-associated deubiquitinase best known for antagonizing PTEN-induced kinase 1 (PINK1)/Parkin-dependent ubiquitination and suppressing mitophagy. This established role has linked USP30 to mitochondrial quality control and neurodegenerative diseases. However, emerging studies in cancer have expanded the biological scope of USP30, revealing functions that cannot be explained by mitophagy regulation alone. USP30 regulates diverse cancer-associated substrates through deubiquitination, thereby affecting protein abundance, subcellular localization, protein interactions, and downstream transcriptional programs. These substrate-dependent effects influence mitochondrial dynamics, lipid synthesis, glucose and amino acid metabolism, redox homeostasis, apoptosis, ferroptosis, epithelial–mesenchymal transition, cancer stemness, T-cell exhaustion, and therapy resistance. Although most studies indicate tumor-promoting roles of USP30 in hepatocellular carcinoma, breast cancer, colorectal cancer, oral squamous cell carcinoma, ovarian cancer, bladder cancer, prostate cancer, and leukemia models, context-dependent tumor-restraining activities have also been reported. In this Review, we examine cancer-associated functions of USP30 that extend beyond its established role in PINK1/Parkin-dependent mitophagy, discuss the spatial and substrate-dependent mechanisms underlying these activities, and highlight opportunities and challenges for therapeutic targeting.

International Journal of Molecular Sciences doi: 10.3390/ijms27209364

Authors: Qingxian Wang Shixue Wang Zhenbiao Xu Linxia Song Kexin Yang Chunmei Zhang Xiaorui Cheng

The prevalence of Alzheimer’s disease has significantly increased, which will place a heavy burden on families and society. Cognitive impairment is a typical feature of Alzheimer’s disease. Rodent models play a key role in AD research, but their considerable expense and slow development pose challenges to the pace of therapeutic discovery. Here, we established a new model of cognitive dysfunction in planarians using AlCl3. After exposure to 50 μM AlCl3 for 48 h, the planarians exhibited cognitive dysfunction in the phototaxis test, dual-stimulus decision test and T-maze test. Immunohistochemical and immunofluorescence experiments confirmed that AlCl3 damaged the neural structure of planarians. Moreover, AlCl3 disrupted the balance of neurotransmitters, leading to cell apoptosis and oxidative stress. Finally, we confirmed that donepezil alleviated the cognitive impairment caused by AlCl3. This indicates that the model we established for cognitive dysfunction in planarians was feasible. However, planarians lack the amyloid-β plaques and tau neurofibrillary tangles typical of AD. Therefore, the planarian model can be considered a promising complementary platform for high-throughput screening of candidate drugs for cognitive impairment.

International Journal of Molecular Sciences doi: 10.3390/ijms27209365

Authors: Amelia Tero-Vescan Ruxandra Ștefănescu Bianca Eugenia Ősz Mădălina Buț Amalia Pușcaș Ylenia Pastorello Mark Slevin

Esports and gaming resemble traditional sports and physical activity because both involve sustained cognitive and emotional stress; however, this stress occurs against a background of prolonged sedentary behavior. This narrative review examines the neuroendocrine, metabolic, and endothelial consequences of this “stress without movement” phenotype and its potential implications for long-term cardiovascular health. Activation of the hypothalamic–pituitary–adrenal (HPA) axis and sympathetic nervous system disrupts sleep and circadian rhythms. Continuous neuroendocrine activation may impair muscle–adipose tissue crosstalk and nitric oxide-dependent endothelial function. Although direct longitudinal evidence linking gaming to cardiovascular disease is currently lacking, these convergent mechanisms identify prolonged sedentary gaming as a potential cardiovascular risk amplifier, particularly when combined with occupational sitting, sleep disruption, adiposity, and other cardiometabolic risk factors.

International Journal of Molecular Sciences doi: 10.3390/ijms27209366

Authors: Jasmine Covarelli Elisa Vinciarelli Alessandra Mirarchi Paolo Prontera Cataldo Arcuri

The journal retracts the article titled, “Retinoic Acid Induced 1 and Smith–Magenis Syndrome: From Genetics to Biology and Possible Therapeutic Strategies” [...]

International Journal of Molecular Sciences doi: 10.3390/ijms27209367

Authors: Xzaviar K. Solone Md-Selim Reza Paul Renauer Kalpana Subedi Nirmal Parajuli Li Zhou Qing-Sheng Mi

Malignant cells hijack epigenetic processes to sustain proliferation and to tolerate stressors that would otherwise limit cell growth, yet the chromatin remodeling activities that establish and maintain these programs are poorly defined. H2A.Z is a histone variant deposited into nucleosomes by the SRCAP and TIP60 chromatin-remodeling complexes, which share VPS72 as a chaperone subunit. VPS72 is located at 1q21, a region recurrently gained in human cancers, yet its contribution to tumor biology remains largely unexplored. Here, we show that VPS72 copy-number gain occurs in more than one-third of tumors and its expression is enriched beyond that expected from neighboring 1q21 genes and remains elevated over normal tissue in copy-neutral tumors. Across cancer types, higher VPS72 expression was associated with proliferative transcriptional programs, cell-cycle–related chromatin accessibility, and genomic instability. Single-cell analyses localized these associations primarily to malignant cells. Higher VPS72 expression and copy-number gain were associated with shorter overall survival in several solid tumors, although these relationships varied by lineage. VPS72 copy-number gain was also associated with shorter overall survival after adjustment for age and sex. Cancer-cell screens further identified recurrent VPS72 dependency, and VPS72 depletion suppressed proliferative programs while inducing stress, inflammatory, and apoptotic responses that were only partly shared with H2A.Z depletion. Together, these findings identify VPS72 as a recurrently altered component of tumor chromatin regulation associated with proliferative and stress-response states and support further investigation of its therapeutic significance.

International Journal of Molecular Sciences doi: 10.3390/ijms27209363

Authors: Ashley D. Clifton Micah J. Jeppesen Shohaib Shaffiey Arpita Chatterjee Kia T. Liermann-Wooldrik Oluwaseun Adebisi Simran Takkar Naresh Mutha Rebecca E. Oberley-Deegan Chittibabu Guda Massimiliano Pierobon James W. Checco Robert Powers

Large-scale radiation catastrophes and localized exposure incidents remain major threats to public health. Effective treatment following radiation exposure is challenging because radiation dose, timing, and affected tissues can vary widely between individuals. To better understand the biological response to radiation injury, we employed a multi-omics approach integrating transcriptomics, proteomics, and metabolomics. In this study, mouse bone marrow was analyzed at 1, 6, and 24 h after whole-body irradiation and compared with bone marrow from non-irradiated mice. Multivariate and univariate analyses revealed time-dependent changes across transcriptomic, proteomic, and metabolomic profiles. We further applied machine learning-based integrative analysis and biomarker discovery approaches to identify key molecular responses associated with radiation exposure. Integrated analysis showed that the acute radiation response in bone marrow included immune-related processes, followed by intermediate metabolic reprogramming, and later disruptions in cell cycle regulation. Biomarker analysis identified several promising candidates, including the transcripts Sapcd2 and Cd28, proteins FRIL1 and CDK1, and the metabolites 3-hydroxyisobutyrate and taurine. Overall, our findings demonstrate that bone marrow from irradiated mice undergoes dynamic and coordinated molecular changes over time. These responses include activation of ferroptosis defense mechanisms, mitotic dysfunction, and pathways associated with programmed cell death.

International Journal of Molecular Sciences doi: 10.3390/ijms27209357

Authors: Francesco Calì Antonino Musumeci Mirella Vinci Simona L’Episcopo Massimiliano Bartolone Angelo Gloria Raffaele Ferri Simone Treccarichi Antonio Fallea

Tooth agenesis (TA) is a common developmental abnormality that may occur as an isolated trait or as part of a multisystem disorder. Its genetic architecture remains incompletely characterized, particularly in individuals with neurodevelopmental disorders (NDDs). This descriptive case series aimed to investigate potentially relevant genetic variants in a clinically selected cohort of patients with TA using whole-exome sequencing (WES). Ten patients with TA and neurological or neurodevelopmental conditions were recruited at the IRCCS Oasi Research Institute of Troina between January 2024 and July 2026. The cohort included eight males and two females (age range: 5–50 years). The associated clinical spectrum included autism spectrum disorder (ASD), intellectual disability (ID), language impairment, behavioral abnormalities, and Down syndrome (DS). WES was performed in the examined cohort and variants were prioritized according to population frequency, predicted functional impact, inheritance pattern, and phenotypic concordance. The cohort comprised nine patients with hypodontia and one with oligodontia. Thirty variants involving 28 genes were identified. A homozygous GLI3 p.Gly727Arg variant, inherited from two heterozygous parents, was detected in an eight-year-old boy with ASD, global developmental delay, and hypodontia. Pathogenic or likely pathogenic loss-of-function variants in BCL11B were identified in two girls with ID and hypodontia. A likely pathogenic NIPBL splice-site variant was identified in a 15-year-old boy with hypodontia. His similarly affected father carried a novel FGFR1 p.Asn185His variant that was absent in the son, excluding it as the sole explanation for the familial dental phenotype. A 50-year-old man with DS and oligodontia carried the recurrent pathogenic BBS1 p.Met390Arg allele in the heterozygous state, together with variants of uncertain significance in PTH1R and STAT3. However, none of these variants independently explained his dental phenotype. The remaining variants were primarily relevant to the associated neurological phenotypes or represented secondary findings. This descriptive case series illustrates the clinical and genetic heterogeneity of TA in individuals with NDDs. Although the findings do not establish causal relationships with TA, they highlight the importance of distinguishing the general clinical significance of a variant from its specific relevance to dental development. Replication in larger cohorts and functional validation will be required to clarify the contribution of the identified candidate variants.

International Journal of Molecular Sciences doi: 10.3390/ijms27209348

Authors: Natalya Risinskaya Sofia Starchenko Abdulpatakh Abdulpatakhov Yulia Chabaeva Olga Aleshina Ekaterina Kotova Tatiana Obukhova Anna Yushkova Olga Maiorova Nikolay Kapranov Irina Galtseva Valeria Surimova Sergey Kulikov Andrey Sudarikov Elena Parovichnikova

Chromosomal microarray analysis (CMA) detects copy-number alterations and copy-neutral loss of heterozygosity (cnLOH), but the CMA landscape of adult T-ALL has been described in few cohorts. Day +70 measurable residual disease (MRD) is a surrogate marker of early treatment response, available only after induction. Thirty-nine adults with Ph-negative T-ALL on a single protocol underwent diagnostic CMA. Mosaic events were catalogued per chromosome and against a 57-gene panel and described in relation to day +70 MRD status (13 MRD-positive, 25 MRD-negative). The analysis is presented as descriptive without formal inference. Mosaic alterations were present in almost all patients, so groups differed in the amount rather than the presence of genomic change. Complex profiles (≥3 affected chromosomes) were seen in 46.2% of MRD-positive and 20.0% of MRD-negative patients. Biallelic 9p21.3 deletion occurred in both groups, whereas 9p cnLOH without deletion occurred predominantly in MRD-negative patients. Total cnLOH burden did not track with copy-number burden. Copy-number and copy-neutral events are mechanistically distinct and were not interchangeable here: 9p deletion and 9p cnLOH are separable by SNP array and should be reported separately rather than pooled. No association reached significance, although the cohort was too small to detect all but very large effects.

International Journal of Molecular Sciences doi: 10.3390/ijms27209362

Authors: Shinji Miwa Katsuhiro Hayashi Satoru Demura

Although the developments in multidisciplinary treatment have significantly improved clinical outcomes in bone and soft-tissue sarcomas, the outcomes of patients with metastatic/recurrent sarcoma remain poor [...]

International Journal of Molecular Sciences doi: 10.3390/ijms27209361

Authors: Federico Vello Sara Fortuna Federico Fogolari

Driven by the common entropy formalism in statistical physics and information theory, here we show that methods developed for the analysis of molecular configurational ensembles may be successfully used for classification. Entropy is estimated using the k-th nearest neighbor method combined with the Maximum Information Spanning Tree method to account for the mutual information between variables. This formalism is used to classify new samples using their estimated cross-entropy with the training set. The method is tested using the hand-written digits MNIST and the functional genomics CuMiDa datasets. The results obtained on these diverse datasets are comparable to those obtained by machine learning methods, or better when the number of samples is small. The methods described here have the advantage, compared to other classification methods, of directly linking variables and pairs of variables to the assigned class, a valuable feature when aiming at identifying causal relationships between features and classes (e.g., the relationship between genes and diseases).

International Journal of Molecular Sciences doi: 10.3390/ijms27209359

Authors: Hiruni Sandunika Kumarasinghe Ducdat Le Ji-Hyang Kim Su-Lim Kim Kyeoung Cheol Kim Rambukka-na Maggonage Thiruni Dananjana Perera Yaejun Kim Mina Lee Dong-Sun Lee

Triple-negative breast cancer (TNBC) is highly aggressive and lacks targeted therapies. This study investigated the anti-cancer mechanisms of sinapaldehyde (SNAH). The biological effects of SNAH were evaluated in vitro using MDA-MB-231 TNBC cells. Cellular responses, including autophagy, mitochondrial function, apoptosis, cell-cycle progression, migration, and colony formation were investigated. In silico studies explored associated regulatory networks and binding targets. Sinapaldehyde (SNAH) treatment was associated with mitochondrial dysfunction, characterized by alterations in mitochondrial structural and functional markers, increased reactive oxygen species(ROS) production, and loss of mitochondrial membrane potential. SNAH treatment also altered the expression of PINK1 and Parkin and modulated autophagy-associated markers, including LC3B, p62, LAMP1, Beclin-1, and ATG proteins. Time-dependent changes in autophagy- and apoptosis-associated markers were accompanied by increased Annexin V-positive cells and activation of cleaved caspase-9 and caspase-3. SNAH also altered FTH1 and NCOA4 expression, suggesting modulation of ferritinophagy-associated processes, and inhibited migration and colony formation while inducing G2/M cell-cycle arrest. In silico analyses identified potential breast cancer-related molecular targets and signaling pathways associated with SNAH. SNAH induces mitochondrial dysfunction and is associated with alterations in autophagy/mitophagy-associated signaling and activation of apoptosis in MDA-MB-231 TNBC cells. These findings provide insight into the cellular responses to SNAH treatment, while further functional studies are required to establish changes in mitophagic and ferritinophagic flux and the causal relationship between autophagy and apoptosis.

International Journal of Molecular Sciences doi: 10.3390/ijms27209351

Authors: Mohsen Nemat-Gorgani Michael Anthony Jensen Ronald Wayne Davis

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) and Post-Acute Sequelae of SARS-CoV-2 infection (Long COVID) are complex multisystem disorders affecting 60–400 million individuals globally. They are considered post-infectious illnesses and share similar biological mechanisms, including immune dysregulation, orthostatic intolerance and cognitive dysfunction, highlighting a substantial overlap between the two conditions. Persistent fatigue, sleep dysfunction, post-exertional malaise and brain fog are among some of their most common and debilitating symptoms. Despite extensive research, they remain incurable, with no effective treatments, or simple diagnostic tests. They are increasingly understood as heterogeneous, multisystem diseases, involving neurological symptoms, with varying severities and diverse underlying biological abnormalities. More recently, the possibility of glymphatic dysfunction and autophagy impairment have been proposed as parts of the mechanistic features of the pathophysiology of these conditions, which are discussed more fully in the present perspective. With the glymphatic and meningeal lymphatic systems being operationally integrated, we will try to delineate how the metabolic waste removal mechanism afforded by this system may work together with autophagy to maintain brain homeostasis. We will provide some of the established mechanistic features of toxicity of misfolded proteins, explore their role in causing glymphatic dysfunction and impairing autophagy, and discuss the potential for therapeutic interventions.

International Journal of Molecular Sciences doi: 10.3390/ijms27209358

Authors: Kazuki Nakamura Senri Takenouchi Yurika Nakagawa-Saito Yasufumi Ito Yuta Mitobe Shuhei Suzuki Keita Togashi Asuka Sugai Yukihiko Sonoda Chifumi Kitanaka Masashi Okada

Glioma stem cells (GSCs) are cancer stem cells in glioblastoma (GBM) that are resistant to both radiotherapy and chemotherapy and contribute to tumor recurrence. Therefore, the development of GSC-targeted therapies is urgently needed to improve the poor prognosis of GBM patients. Given the growing interest in redox homeostasis, which is closely linked to mitochondrial metabolism supporting the unique properties of GSCs, we focused on capsaicin, a widely used exogenous calcium channel agonist, and its structural analogs as candidate compounds capable of disrupting redox homeostasis in GSCs. Based on a comprehensive evaluation of the effects of the candidate compounds, we found that the capsaicin analog arvanil selectively induced cell death in GSCs with high selectivity over differentiated GSCs at concentrations that were non-toxic to normal cells. Arvanil-induced cell death was associated with a preferential elevation in intracellular reactive oxygen species (ROS) levels in GSCs over those in differentiated GSCs and was suppressed by a Ca2+ chelator and a mitochondria-targeted antioxidant. These results suggest that arvanil preferentially induced GSC death in association with intracellular Ca2+ dysregulation and mitochondria-related oxidative stress, highlighting potential vulnerabilities in the redox homeostasis of GSCs and supporting further investigations on arvanil as a feasible therapeutic approach for GBM.

International Journal of Molecular Sciences doi: 10.3390/ijms27209360

Authors: Ayaka Yasuda Natsuko Saito-Sasaki Reiko Hara Yu Sawada

Epidermal barrier dysfunction and cutaneous inflammation are closely interconnected in inflammatory skin diseases. However, the molecular effects of heparinoid cream on inflamed skin remain incompletely understood. In this study, we investigated the effects of heparinoid cream on epidermal barrier homeostasis and cutaneous inflammation using an MC903-induced dermatitis model. Eight-week-old female BALB/c mice were treated topically with MC903 for 7 consecutive days, with or without concomitant application of heparinoid cream. Skin barrier function, histological changes, inflammatory markers, and gene expression were evaluated by transepidermal water loss measurements, histology, immunohistochemistry, quantitative PCR, and RNA sequencing. Therapeutic effects were also assessed by applying heparinoid cream after dermatitis had been established. RNA sequencing identified 816 differentially expressed genes following heparinoid cream treatment, including 146 upregulated and 670 downregulated genes. Upregulated genes were enriched in epidermal differentiation- and barrier-related pathways, whereas downregulated genes were enriched in immune and inflammatory pathways. Heparinoid cream significantly reduced transepidermal water loss and epidermal thickness, suppressed Il13 and Tslp expression, and reduced epidermal KLK5 and TSLP expression. In parallel, expression of multiple epidermal differentiation- and barrier-associated genes, including members of the Lce family, Flg, and Flg2, was restored. Therapeutic application of heparinoid cream after establishment of dermatitis was also associated with improved barrier function, increased Flg expression, reduced Tslp mRNA expression, and decreased epidermal KLK5 and TSLP immunohistochemical staining. Secondary reanalysis of a previously published human keratinocyte RNA-seq dataset revealed changes in selected barrier- and innate immunity-associated genes following MPS treatment under non-inflammatory monolayer culture conditions. These findings demonstrate that heparinoid cream treatment is associated with concurrent improvements in epidermal barrier-related parameters and attenuation of type 2 inflammatory responses in MC903-induced dermatitis. These findings provide molecular insights into the changes associated with topical application of a heparinoid-containing formulation in inflamed skin, although the specific contribution of MPS remains to be established.

International Journal of Molecular Sciences doi: 10.3390/ijms27209353

Authors: Miliausha Galimova Oleg Pimenov Ilia Teplov Alexey Berezhnov Vladislav Starkov Victor Tsetlin Yuri Utkin

Snake venom phospholipase A2 (PLA2) affect, among others, the cardiovascular system; however, data on their effect on isolated cardiomyocytes are practically absent. In this work, we investigated the influence of HDP-1 and HDP-2, heterodimeric PLA2s from the venom of viper Vipera nikolskii, on isolated rat cardiomyocytes. Cardiomyocytes were stained with fluorescent probes and fluorescence was measured using Cell Observer fluorescent station. After PLA2 application, there was a prolonged lag phase with no changes in cell morphology or probe signal, followed by fluctuations in the cytosolic calcium level and cell hypercontracture with subsequent disruption of plasma membrane. In individual cardiomyocytes, time from the toxin addition until the hypercontracture varied by more than two-fold. In experiments with electrical stimulation, at HDP-2 application, a “pause” in contractions was observed, characterized by a decrease in the amplitude of the fluorescent response. Then, the contraction rhythm recovered, and later, hypercontracture was observed followed by cell destruction. Depending on times to these events, all cardiomyocytes studied were classified into two groups with fast and slow response to PLA2 action. The observed differences in the sensitivity of cardiomyocytes may be explained by unequal concentrations of intracellular calcium due to the different states of the cells at isolation.

International Journal of Molecular Sciences doi: 10.3390/ijms27209355

Authors: Mateen A. Khan Raja Chinnappan Sowmya Andole Mohd. Akif Tanveer Ahmad Mir Ahmed Yaqinuddin Phuong-T T. Pham Mamoun M. Bader

Amyloid-β (Aβ) aggregation and associated structural changes are central features of Alzheimer’s disease (AD) pathology. In this study, we investigated the binding of α-halogenated curcumins, α-bromocurcumin and α-chlorocurcumin, to Aβ1-42 and examined their effects on the peptide secondary structure using fluorescence spectroscopy and circular dichroism (CD). Both α-halogenated curcumins interacted with Aβ1-42, but their binding properties differed from those of the parent curcumin. α-Chlorocurcumin exhibited a higher binding affinity for Aβ1-42 than curcumin, whereas α-bromocurcumin showed comparatively weaker binding. Temperature-dependent binding studies revealed that the association of α-halogenated curcumins with Aβ1-42 was influenced by temperature, with reduced binding affinity at higher temperatures. Thermodynamic analysis demonstrated favorable free-energy changes for the interactions and revealed that α-halogenation altered the relative enthalpic and entropic contributions to Aβ1-42 binding. CD analysis further showed that binding of the curcumin altered the secondary structure of Aβ1-42. These structural changes were more pronounced in the presence of the α-halogenated derivatives, indicating that halogen substitution influences the interaction and conformational response of Aβ1-42. Molecular docking analysis provided additional insight into the possible binding modes and predicted intermolecular contacts between the α-halogenated curcumin and Aβ1-42. Overall, the findings demonstrate that α-halogenation modulates the binding and structural effects of curcumin derivatives on Aβ1-42, with α-chlorocurcumin exhibiting particularly favorable binding characteristics. These findings provide a basis for further investigation of α-halogenated curcumins as potential modulators of Aβ-related molecular processes associated with AD.

International Journal of Molecular Sciences doi: 10.3390/ijms27209354

Authors: Jaishriram Rathored Sandesh Shende

Coronavirus disease 2019 (COVID-19) has caused global morbidity and mortality. Vaccination remains the most effective preventive strategy. However, variation in immune response after immunization has been observed among individuals. vitamin D plays a crucial role in immune regulation through activation of the VDR, which is expressed on various immune cells. The role of vitamin D status in post-vaccination immune responses among COVID-19 patients has gained increasing scientific interest. This narrative review evaluates the immunomodulatory roles of vitamin D receptors (VDRs) and serum vitamin D status in COVID-19 patients following vaccination, with particular emphasis on immune mechanisms, inflammatory regulation, and vaccine-induced immune responses. The relevant literature concerning vitamin D, VDR signaling, COVID-19 infection, vaccination, and immune responses was comprehensively reviewed. Emphasis was placed on studies discussing immune modulation, cytokine regulation, and antibody responses. Current evidence suggests that adequate serum vitamin D status may support both innate and adaptive immune responses following COVID-19 vaccination. Activation of the VDR regulates T-cell differentiation, modulates pro-inflammatory cytokine production, and supports humoral immune responses. Vitamin D deficiency has been associated with impaired vaccine responsiveness and persistent systemic inflammation. Vitamin D and VDR-mediated signaling pathways appear to play important roles in regulating immune responses following COVID-19 vaccination. Maintaining optimal vitamin D levels may help improve vaccine-induced immunity and reduce excessive inflammatory responses.

International Journal of Molecular Sciences doi: 10.3390/ijms27209356

Authors: Anna Grudniewska Beata Mrozikiewicz-Rakowska Kacper Wiszniewski Piotr Dudek Alina Kuryłowicz

Obesity should no longer be regarded as a disease confined to adipose tissue. Contemporary evidence indicates that adipose tissue dysfunction can initiate a cascade of metabolic and inflammatory responses that progressively involve multiple peripheral organs. Drawing on current molecular and translational evidence, we delineate mechanisms through which lipotoxicity, oxidative and endoplasmic reticulum stress, NLRP3 inflammasome activation and adipokine dysregulation contribute to inflammatory and metabolic dysfunction in the liver, skeletal muscle, pancreatic β-cells, vascular endothelium, perivascular adipose tissue and tumour microenvironment. Once established, organ-specific processes may become partially self-reinforcing and reciprocally amplify systemic immunometabolic dysfunction. We propose the Hydra of Inflammation as an integrative framework in which peripheral organs function not only as targets of adipose tissue-derived signals but also as secondary contributors to a progressively interconnected inflammatory network. This network-based perspective may help explain the heterogeneity of obesity-associated metabolic phenotypes and supports a shift from isolated organ-centred models towards systems-level approaches. The framework integrates the adipose tissue mechanisms discussed in Part I with the therapeutic perspectives developed in Part III of the Prometheus’s Fire series.

International Journal of Molecular Sciences doi: 10.3390/ijms27209352

Authors: Kyuri Lee Jeung hi Han Hairu Zhao Young Eun Kim Yang Gyu Choi Hosam Ki Jaehong Kim Young Jin Choi Hye Yoon Park Areum Jang Min Hyuk Park Hang Eui Cho Jae Sung Hwang

Skin photoaging is largely driven by ultraviolet (UV) irradiation, which contributes to chronic inflammation and extracellular matrix (ECM) degradation. This study investigated the protective effects of paeonol against UVB-induced cellular senescence in human dermal fibroblasts (HDFn). Repeated UVB irradiation was used to induce premature senescence. Paeonol treatment reduced the senescence-associated β-galactosidase (SA-β-gal) positive cells and intracellular reactive oxygen species (ROS) accumulation. Paeonol also reduced the expression of senescence-associated cell cycle regulators, including p21 and p16 at the mRNA level and the protein level. In addition, paeonol suppressed matrix metalloproteinase-1 (MMP-1) production but did not significantly restore type I procollagen secretion. Paeonol also attenuated the expression and secretion of the senescence-associated secretory phenotype (SASP) factors inter-leukin-6 (IL-6) and interleukin-8 (IL-8). These effects were accompanied by reduced phosphorylation of JNK and c-Jun. Furthermore, activation of JNK signaling with anisomycin partially reversed the paeonol-mediated suppression of MMP-1 mRNA expression, supporting the involvement of JNK signaling in the regulation of MMP-1. Collectively, these findings suggest that paeonol attenuates multiple features of UVB-induced cellular senescence in HDFn cells, with JNK signaling potentially contributing to its inhibitory effect on MMP-1 expression. Further studies are needed to clarify the mechanisms underlying these effects and establish their physiological relevance in UVA-based and in vivo models.

International Journal of Molecular Sciences doi: 10.3390/ijms27209349

Authors: Manna Huang Kexin Zhang Tingting Wang Peiyi Li Yue Xin Baoyin Chen

Mitochondrial complex I (CI) is a major source of reactive oxygen species (ROS) in plant cells, and iron–sulfur (Fe-S) cluster proteins are critical for maintaining CI structure and electron transport. C1-FDX, a mitochondrion-localized ferredoxin containing a [2Fe-2S] cluster, acts as a structural component and assembly factor of mitochondrial CI and complex V (CV) in Arabidopsis. In this study, we further characterized the physiological and molecular functions of C1-FDX in regulating ROS production, NADH/NAD+ homeostasis, and iron metabolism. Histochemical staining demonstrated that loss of function of C1-FDX in Arabidopsis c1-fdx mutants accumulated excessive superoxide anions (O2•−) but exhibited reduced hydrogen peroxide (H2O2) relative to the wild type (WT). Biochemical assays further confirmed that there was an elevated NADH/NAD+ ratio in mutant seedlings, which facilitated O2•− overproduction at mitochondrial CI. Loss of C1-FDX also led to Fe3+ accumulation in mutant tissues. Immunoblot and qRT-PCR results showed that the abundance of MnSOD markedly increased, while other antioxidant proteins, including FeSOD, CAT1, and GR, were downregulated in c1-fdx mutants of Arabidopsis. The enhanced MnSOD activity accelerated the conversion of O2•− into H2O2, and the subsequently accumulated Fe3+ may be derived from the Fenton reaction, which consumes H2O2, ultimately resulting in decreased H2O2 levels. Additionally, deficiency of C1-FDX disturbed the abundance of mitochondrial chaperones, proteases, and alternative respiratory pathway proteins. Collectively, our findings demonstrate that C1-FDX participates in the regulation of mitochondrial ROS balance and iron homeostasis and suggest there is a novel regulatory link between ferredoxin, the electron transport chain, and the Fenton reaction in plant mitochondria.

International Journal of Molecular Sciences doi: 10.3390/ijms27209350

Authors: Sherin Ramadan Emanuele Panza Antonio Orlacchio

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by marked genetic and phenotypic heterogeneity. Although pathogenic variants in major ALS genes, including C9ORF72, SOD1, TARDBP, and FUS, have provided fundamental insights into disease pathogenesis, many familial and most sporadic cases remain without a molecular diagnosis, suggesting a genetic architecture extending beyond coding variants and monogenic inheritance. Advances in genomic technologies have revealed contributions from deep-intronic and regulatory variants, structural rearrangements, repeat expansions, and other forms of genomic variation incompletely captured by standard diagnostic approaches. Evidence also indicates that combinations of rare variants with moderate or incomplete penetrance may contribute to disease susceptibility and clinical variability, supporting a multilayer model of ALS genetics. Population-specific variation and convergence of genetically diverse defects on common molecular pathways further complicate variant interpretation and disease classification. This review examines emerging components of ALS genetic architecture, including non-coding variation, structural variants, repeat expansions, oligogenic burden, ancestry-related gaps, and convergent neurodegenerative pathways. Integrating these genomic layers with functional and transcriptomic evidence may improve variant interpretation, diagnostic yield, patient stratification, and the development of mechanism-based precision therapies.

International Journal of Molecular Sciences doi: 10.3390/ijms27209347

Authors: Sara A. Gaballah Ahmed I. Abulsoud Asmaa Saleh Noha A. Eldesoky

Hepatocellular carcinoma (HCC) is a life-threatening malignancy. Although regorafenib is used for HCC treatment, its clinical utility is limited by significant adverse effects. This study evaluated whether combining regorafenib with naringenin could enhance its anticancer activity against HepG2 cells. This study included seven groups of HepG2 cancer cells: an untreated control group, a naringenin-treated group (100 μM), a regorafenib-treated group (1 μM), and four groups receiving naringenin (100 μM) combined with decreasing concentrations of regorafenib (1, 0.75, 0.5, and 0.25 μM). Cell viability and the expression of E-cadherin and MRP1 were assessed. All treatments significantly reduced cell viability compared with the control group, with greater inhibition observed in the combination-treated groups than in the monotherapy groups. Combination treatment also increased E-cadherin expression and decreased MRP1 expression. The combination of regorafenib (1 μM) and naringenin (100 μM) exhibited the greatest growth inhibition, the highest E-cadherin, and the lowest MRP1 expression levels. Notably, combining the IC50 of naringenin with one-quarter of the regorafenib IC50 significantly enhanced E-cadherin expression and reduced MRP1 expression compared with regorafenib alone. The present findings suggest that naringenin enhances the anticancer effects of regorafenib in HepG2 cells and modulates E-cadherin and MRP1 gene expression.

International Journal of Molecular Sciences doi: 10.3390/ijms27209345

Authors: Maren Rehders Helena Kerp Robert Nica Rupert Ecker Dagmar Führer Klaudia Brix

Adequate thyroid hormone concentrations are essential for tissue homeostasis, including in the thyroid gland. Thyrocyte function is regulated by the hypothalamic–pituitary–thyroid axis and by the coordinated action of thyroglobulin-processing cathepsins and thyroid hormone transporters such as monocarboxylate transporter 8 (Mct8). Here, we investigated the effects of hypothyroidism and thyroxine-induced exogenous hyperthyroidism on thyroid proteases and Mct8 expression in male mice. Thyroid follicular architecture was largely preserved in hyperthyroidism, whereas hypothyroidism resulted in increased thyrocyte extensions, collapsed follicular lumina, and enhanced thyroglobulin turnover. Cathepsin B was reduced in both conditions, pro-cathepsin D declined only in hyperthyroidism, and cathepsin L was upregulated in hypothyroidism. Consistently, thyroglobulin fragments were approximately two-fold more abundant in hypothyroid tissue. Mct8 expression remained unchanged in hyperthyroidism but was nearly absent in hypothyroidism. Hypothyroid tissue also exhibited markedly increased thyrocyte proliferation, while cell death rates were not altered across thyroid dysfunction. In addition, alterations in laminin indicated disbalanced extracellular matrix remodeling in hypothyroidism. Together, these findings demonstrate profound changes in thyroid architecture, proteolytic activity, and thyroid hormone transport under chronic thyroid dysfunction, particularly hypothyroidism. Further studies are needed to define the specific role of individual proteases in these processes.

International Journal of Molecular Sciences doi: 10.3390/ijms27209346

Authors: Andrei Pascu Victor Ianole Mihai Danciu

Digestive malignancies remain a major cause of cancer-related mortality, with tumor progression (TP) being strongly influenced by the complex interactions between tumor cells (TCs) and their microenvironment. This review summarizes recently described mechanisms of interactions between tumor cells, macrophages and CD8+ T lymphocytes in colorectal and gastric cancer. PubMed and Web of Science were searched for studies published between 2020 and 2026, with the final search being conducted on 8 July 2026. Experimental and clinical studies, together with relevant reviews, were considered to examine these interactions and their therapeutic implications. The evidence shows a bilateral crosstalk in which tumor cells influence immune cell phenotype and function, while immune cells can further support or restrict tumor progression. These interactions involve receptor-ligand systems and paracrine signaling, followed by changes in intracellular pathways regulating proliferation, invasion, macrophage polarization and CD8+ T-cell exhaustion. Considerable methodological heterogeneity among studies limits a direct comparison of the reported mechanisms. Several therapeutic approaches targeting these interactions are currently being investigated in early-phase clinical trials. However, many components of tumor–immune crosstalk and their connections remain incompletely understood. A further characterization of these mechanisms may help identify therapeutic strategies able to interfere with the immunosuppressive microenvironment of colorectal and gastric cancers.

International Journal of Molecular Sciences doi: 10.3390/ijms27209344

Authors: Delf-Magnus Kummerfeld Tasneem Khanam Anna Wolters Boris V. Skryabin Timofey S. Rozhdestvensky

Prader–Willi syndrome (PWS) is a neurodevelopmental disorder characterized by distinctive developmental transition from neonatal hypotonia and poor feeding to progressive weight gain and, later, hyperphagia and obesity. The paternally expressed SNORD116 cluster is critical for postnatal growth, but how its deficiency impacts the progression of PWS remains unclear. Here, we analyzed the day-over-day growth rates for 351 mice with Snord116 deletion or altered Snord116 dosage, including their wild-type littermates, across both sexes and two genetic backgrounds. Snord116 expression increased markedly during early brain development. The Snord116-deficient mice exhibited a distinct three-stage growth trajectory: impaired growth until approximately P16, normalization between P17 and P21, followed by transient accelerated weight gain until approximately P30. Although growth rates subsequently normalized, the accumulated deficit was not fully compensated, and the adult Snord116KO mice remained smaller than the wild-type controls. These developmental transitions broadly parallel the early nutritional phases of human PWS, although the mice did not progress to exhibit persistent obesity-associated growth. Reactivation of the maternal Snord116 cluster partially rescued the growth phenotype, whereas activation of both parental alleles produced a modest growth advantage, which was most evident in the males. These findings identify early postnatal development as a critical window of Snord116-dependent growth regulation and provide a framework for investigating the mechanisms underlying PWS’s progression.

International Journal of Molecular Sciences doi: 10.3390/ijms27209343

Authors: Andreea-Ramona Treteanu Ana Maria Alexandra Stănescu Alexandru Constantinescu Ovidiu Lucian Băjenaru Cătălina Raluca Nuță Gratiela Gradisteanu Pircalabioru Octavian Andronic

Digestive neoplasms represent a major global health burden, accounting for a substantial proportion of cancer incidence and mortality. Increasing attention has been directed toward modifiable factors that may influence both the onset and progression of these malignancies. Among them, vitamin D has emerged as a potential protective factor, with deficiency frequently associated with elevated risk and unfavorable prognosis in gastrointestinal cancers. This review critically examines the current evidence regarding the role of vitamin D and its receptor (VDR) in digestive tumorigenesis. Beyond its classical function in calcium and bone homeostasis, vitamin D exerts pleiotropic effects on cellular processes fundamental to carcinogenesis, including regulation of proliferation, differentiation, apoptosis, angiogenesis, immune modulation and genomic stability. Experimental studies demonstrate that adequate vitamin D status or administration of its analogues may suppress tumor growth and enhance responsiveness to conventional therapies. Epidemiological and clinical data, however, remain inconsistent across cancer subtypes, geographic populations and methods of vitamin D assessment. The analysis underscores the need for standardized evaluation of vitamin D levels, elucidation of dose–response relationships and clarification of interactions with genetic polymorphisms and environmental exposures. Future large-scale, randomized controlled trials are essential to establish causality and to define the optimal role of vitamin D in prevention and treatment strategies. Taken together, current findings support the hypothesis that vitamin D may serve not only as a biomarker of prognosis but also as a promising adjunctive approach in the management of digestive neoplasms.

International Journal of Molecular Sciences doi: 10.3390/ijms27209342

Authors: Xinchao Zhang Mengjie Ren Anke Xu Jintian Liu Dongran Liang Fang Kuang Mingmei Wu

Diosmin is a citrus-derived flavone glycoside with anti-inflammatory activity in preclinical models. This study aimed to investigate the effects of diosmin on PTSD-like behaviors, astrocyte morphology, and the expression of STAT1 and CXCL9 in a single prolonged stress and shock (SPS&S) mouse model, and to assess whether Stat1 knockdown could mimic diosmin’s effects in primary astrocytes. Male C57BL/6J mice were assigned to the control, SPS&S, and SPS&S + diosmin groups and underwent 14 days of treatment. Behavioral assessments included open-field and elevated plus-maze tests, alongside cue- and context-associated fear tests, complemented by GFAP-based morphometry, immunofluorescence, Western blotting, serum cytokine assays, hippocampal RNA sequencing, and quantitative reverse-transcription PCR (qRT-PCR). The results indicated that diosmin significantly enhanced exploratory behavior and reduced fear responses compared to SPS&S alone, alongside notable alterations in hippocampal astrocyte morphology and a shift in cytokine profiles, with decreased pro-inflammatory cytokines and increased anti-inflammatory markers. RNA sequencing revealed 297 differentially expressed genes, particularly highlighting the roles of Stat1 and Cxcl9, both of which were significantly downregulated upon diosmin treatment. Furthermore, diosmin modulated astrocyte responses to inflammatory stimuli in primary cultures. These findings suggest that diosmin exerts protective effects against PTSD-like symptoms by regulating astrocytic function and immune-related pathways, with STAT1 signaling serving as a key but non-exclusive downstream mechanism underlying its therapeutic actions. Future studies should further elucidate the full spectrum of associated pathways to advance the development of targeted interventions for PTSD.

International Journal of Molecular Sciences doi: 10.3390/ijms27209338

Authors: Jie Liu Chunyang Liu Linhuan Wang Zhenli Zhao Yabing Cao

Heat shock transcription factors (HSFs) act as crucial regulatory components in plant stress regulatory network and play essential roles in abiotic stress adaptation, particularly drought tolerance. Here, we identified 36 PfHSF gene family members in Paulownia fortunei through genome-wide analysis. Their chromosomal distribution, physicochemical properties, phylogenetic relationships, gene structures, conserved motifs, cis-acting elements, and post-translational modification sites were systematically characterized. Transcriptome-based expression profiling demonstrated distinct tissue-specific and drought-responsive expression patterns among PfHSF members, with PfHSF6 being strongly up-regulated under drought. Subcellular localization assays confirmed nuclear and plasma membrane localization of PfHSF6. Transient overexpression of PfHSF6 in tobacco significantly enhanced drought tolerance, as evidenced by alleviated wilting, a ~60% reduction in MDA content, ~2-fold increases in SOD and POD activities, and a ~67% reduction in electrolyte leakage relative to the control (p < 0.01); consistently, stress-responsive genes such as NbRD29A and NbLEA5 were up-regulated by ~6- and ~3-fold, respectively. This study provides a comprehensive characterization of the PfHSF family and preliminary functional evidence for PfHSF6, offering candidate genes for further functional validation and potential drought-tolerance breeding in paulownia.

International Journal of Molecular Sciences doi: 10.3390/ijms27209341

Authors: Aqsa Shahid Amy Scott-Thomas Stephen Chambers Andrew Miller Madhav Bhatia

Sepsis is a life-threatening syndrome characterized by a dysregulated host immune response to infection. Sepsis development usually results in organ dysfunction and multiple organ failure, including the liver and lungs. Alpha-lipoic acid (ALA) is a well-known anti-inflammatory agent. However, its effects on the endogenous cystathionine γ-lyase/hydrogen sulfide (CSE/H2S) pathway during cecal ligation and puncture (CLP)-induced sepsis remain unclear. In this study, the protective effects of ALA in wild-type (WT) and CSE knockout (CSE−/−) mice were investigated. Mice were randomly divided into six experimental groups, including (a) WT sham (b) CSE−/− sham (c) WT CLP (d) CSE−/− CLP (e) WT CLP + ALA.P and (f) CSE−/− CLP + ALA.P (n = 8 in each group). ALA (100 mg/kg) was administered intraperitoneally one hour before CLP surgery. Eight hours after CLP induction, mice were euthanized, and blood, liver and lung tissues were collected. Body weight changes, proinflammatory cytokine and chemokine concentrations (including tumor necrosis factor-alpha (TNF-α), monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-2 alpha (MIP-2α), myeloperoxidase (MPO) activity, histopathological changes, plasma H2S concentrations, and CSE expression in the liver and lung were measured. Severe liver and lung damage was characterized by markedly increased levels of MCP-1, MIP-2 α, TNF-α, and myeloperoxidase (MPO) activity, compared to WT sham control and CSE−/− groups (p < 0.0001). WT CLP-induced septic mice showed liver and lung injury as evidenced by hepatocyte swelling and ballooning degeneration with liver sinusoidal congestion, as well as pulmonary intra-alveolar and interstitial edema associated with thickening of alveolar septa and leucocyte infiltration, as compared to WT sham control and CSE−/− groups (p < 0.001). The WT-CLP group also showed a significant increase in hepatic and pulmonary expression of the H2S-producing enzyme (CSE) with elevated levels of plasma H2S concentrations, as compared to the WT sham group (p < 0.0001). Prophylactic ALA administration markedly improved these changes in the WT group. ALA also significantly decreased plasma H2S concentrations and hepatic and pulmonary CSE expression as compared to the WT CLP group (p < 0.01). However, ALA administration did not confer additional significant protective effects in CSE−/− mice, which supports the hypothesis that the effects of ALA are mediated through the modulation of the CSE/H2S pathway. In conclusion, this is the first experimental evidence showing that ALA prevents sepsis-associated organ injury at least partly by the downregulation of hepatic and pulmonary cystathionine γ-lyase expression.

International Journal of Molecular Sciences doi: 10.3390/ijms27209340

Authors: Jingyu Yang Yuehong Chen Yulin Zhang Qibing Xie Geng Yin

Interferon (IFN)-induced protein 6 (IFI6) is an important member of the IFN-stimulated gene (ISG) family. It plays significant roles in anti-apoptosis, immune regulation, maintenance of mitochondrial homeostasis, antiviral defense, and tumor progression. At the molecular mechanism level, IFI6 inhibits mitochondrial membrane depolarization, cytochrome c release, and caspase activation. It also regulates the balance between B-cell lymphoma 2 (Bcl-2)/Bcl-2 interacting mediator of cell death (Bim), activates the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) and phosphatidylinositol 3-kinase-protein kinase B (PI3K-AKT) pathways, and upregulates the ferroptosis-related proteins solute carrier family 7 member 11 (SLC7A11) and glutathione peroxidase 4 (GPX4), thereby enhancing cell survival and proliferation through multiple pathways. In terms of immune regulation, IFI6 is associated with immune cells (such as T/B cells, etc.) and the tumor microenvironment. IFI6 regulates mitochondrial reactive oxygen species (mtROS) and mitochondrial calcium homeostasis, influencing respiratory chain assembly and metabolism. In terms of viral infection, IFI6 may inhibit the replication of multiple viruses (such as hepatitis B virus (HBV), hepatitis C virus (HCV), SARS-CoV-2, etc.) by inhibiting flavivirus-induced endoplasmic reticulum (ER) invagination, interfering with retinoic acid-inducible gene I (RIG-I)-like receptor signaling pathways, and inhibiting epidermal growth factor receptor (EGFR) kinase activity. In addition, IFI6 is highly expressed in autoimmune diseases and malignant tumors and is associated with proliferation, metastasis, and poor prognosis. In this review, we aim to highlight IFI6 as a potential key target in disease diagnosis and treatment.

International Journal of Molecular Sciences doi: 10.3390/ijms27209339

Authors: Jovana Duric Frederick E. Williams Isaac T. Schiefer

The worldwide opioid epidemic continues to claim thousands of lives each year and remains one of the most urgent global public health challenges, driven by the high abuse potential and overdose risk of potent, fully synthetic μ-opioid receptor (MOR) agonists such as fentanyl. While fentanyl is highly effective in pain management, it also carries severe risks including respiratory depression, addiction, and fatal overdose. Understanding the neurobiological pathways underlying these risks is essential for developing safer analgesics and effective interventions. The MOR, a G-protein-coupled receptor (GPCR) that modulates neuronal excitability and synaptic transmission in important brain circuits like the ventral tegmental area (VTA), nucleus accumbens (NAc), and prefrontal cortex (PFC), is the primary receptor through which opioids exert their therapeutic effects. Dopaminergic dysregulation, transcriptional remodeling, and altered synaptic plasticity are neuroadaptations brought on by long-term opioid exposure that contribute to dependence and relapse. Life-threatening opioid toxicity is primarily driven by MOR-mediated suppression of respiratory networks, with risk shaped by dose, pharmacokinetics, receptor efficacy, and circuit sensitivity. Zebrafish (Danio rerio) provide a powerful complementary system for dissecting opioid-related neurocircuitry and screening for potential therapeutics. Zebrafish share conserved opioid-signaling components with mammals, and larval transparency facilitates in vivo imaging of neuronal activity at cellular resolution. Behavioral paradigms capture reward-related and drug-seeking phenotypes, while CRISPR-based approaches offer a means to investigate receptor-specific functions in validated models. In addition, zebrafish larvae provide an efficient platform for high-throughput screening of compounds that modulate opioid signaling and respiratory-related phenotypes. Together, insights from human, rodent, and zebrafish studies provide a more integrated understanding of opioid overdose and addiction neurobiology. This review incorporates current knowledge on the molecular and circuit-level mechanisms driving these processes and discusses how zebrafish models accelerate the discovery of safer analgesics and novel treatments for opioid use disorder and overdose.

International Journal of Molecular Sciences doi: 10.3390/ijms27209337

Authors: Yuka Shiotani Jun-ichiro Jo Sara Takasugi Takeryo Adachi Kenichirou Yasui Yoshiya Hashimoto Aki Nishiura

Owing to its excellent mechanical properties and biocompatibility, polyetheretherketone (PEEK) is a promising biomaterial in dental and medical fields. However, PEEK lacks osteoconductive and osteoinductive properties for efficient bone regeneration. In the present study, we aimed to develop a porous PEEK scaffold integrating gelatin hydrogel as a carrier for the sustained delivery of bone morphogenetic protein-2 (BMP-2) and evaluate its bone regeneration potential. The pores in the three-dimensional printed porous PEEK were filled with an aqueous solution of gelatin, followed by freeze-drying and dehydrothermal crosslinking to obtain a porous PEEK scaffold with gelatin hydrogel (Gel-PEEK). BMP-2 dissolved in phosphate-buffered saline was impregnated into Gel-PEEK to obtain Gel-PEEK incorporating BMP-2 (BMP-2@Gel-PEEK). After confirming the sustained release of BMP-2 from Gel-PEEK in vitro, the bone regeneration potential of BMP-2@Gel-PEEK was evaluated using a model of critical-sized calvarial defects in rats. Micro-computed tomography and histological analyses demonstrated augmented bone regeneration by BMP-2@Gel-PEEK compared with PEEK only and Gel-PEEK without incorporating BMP-2. These findings suggest that through the sustained release of BMP-2, the porous PEEK with gelatin hydrogel is a promising platform for bone regeneration.

International Journal of Molecular Sciences doi: 10.3390/ijms27209335

Authors: Necmiye Şengel Mustafa Arslan

Ischemia–reperfusion (I/R) injury is a complex pathological process involving adenosine triphosphate (ATP) depletion, calcium overload, excessive reactive oxygen species generation, mitochondrial dysfunction, inflammatory activation, and regulated cell death. Volatile anesthetics, including sevoflurane, desflurane, and isoflurane, may modulate several of these processes through antioxidant and prosurvival signaling, suppression of inflammatory pathways, and regulation of apoptosis. Because I/R injury involves interconnected mechanisms, combining interventions that act on different targets may offer additional protection. Experimental studies have investigated volatile anesthetics together with fullerenol C60, ischemic conditioning, and selected pharmacological interventions. Other combination strategies involving ozone conditioning and boric acid have also been evaluated in experimental models, although these have not established the effects of combining either intervention with a volatile anesthetic. This narrative review examines the molecular mechanisms and organ-specific evidence underlying these approaches, with particular attention to study design, treatment timing, and the interpretation of combined effects. While some combinations have shown greater protection than individual interventions, the findings remain heterogeneous, and formal analyses distinguishing additivity from synergy are generally lacking. Clinical evidence is limited, and further studies are needed to establish whether these experimental findings can be translated into meaningful patient outcomes.