Biology and Life Sciences

Abstract: Antimicrobial resistance (AMR) in ESKAPE pathogens represents a major threat to infectious disease therapy and has stimulated interest in metal-based antibacterial agents as complements or alternatives to conventional organic antibiotics. Their diverse coordination geometries, tunable redox properties, ligand-exchange behaviour, and capacity to interact with multiple biological targets offer distinctive opportunities for antibacterial drug development. However, despite the potent in vitro activity reported for numerous compounds, progression towards clinical application remains limited. This critical narrative review examines the factors separating antibacterial activity from therapeutic development across six representative metal classes: gold, bismuth, gallium, copper, silver, and ruthenium compounds. Three recurrent limitations are identified: insufficient characterisation of metal-complex speciation under biologically relevant conditions; reliance on minimum inhibitory concentration (MIC) measurements and indirect assays without rigorous validation of molecular targets or causal mechanisms; and the scarcity and poor integration of pharmacokinetic, safety, and in vivo efficacy data. The available evidence reveals compound-specific advances, including the repurposing potential of auranofin, bismuth-mediated inhibition of New Delhi metallo-β-lactamase 1 (NDM-1), and the clinical evaluation of gallium nitrate in chronic Pseudomonas aeruginosa airway infection in people with cystic fibrosis. Nevertheless, these findings cannot be extrapolated to entire metal classes. Progress towards clinically viable metalloantibiotics requires an integrated development strategy combining biologically relevant speciation studies, causal mechanistic validation, standardised antibacterial testing, and early assessment of exposure, safety, and efficacy in indication-appropriate models.

Abstract: Actin remodeling accompanies myoblast proliferation, lineage commitment, elongation, fusion, and myofibril assembly, but these changes do not constitute a single linear pathway. This review separates three experimentally distinguishable scales of actin function in skeletal myogenesis. First, the balance between monomeric and filamentous actin controls myocardin-related transcription factor–serum response factor (MRTF–SRF) signaling and thereby couples cytoskeletal state to gene expression. Second, integrin–FAK–Rho signaling and actomyosin tension influence YAP/TAZ activity, with outcomes that depend on cell state, density, mechanical context, and the non-equivalent functions of YAP and TAZ. Third, spatially restricted branched actin networks generate protrusive force at the fusogenic synapse, while a mechanically reinforced receiving-cell cortex provides counterforce and cooperates with membrane fusogens to form and enlarge fusion pores. We compare evidence from cultured myoblasts, developmental models, and adult muscle regeneration, with attention to what each assay measures. We propose that bulk actin abundance, filament turnover, mechanical tension, and local actin architecture should be treated as separate variables. This framework reconciles apparently conflicting results and identifies experiments that can distinguish correlation from causal actin-to-nucleus signaling during muscle formation and repair.

Abstract: Hormone-driven cancers, including prostate, breast, endometrial and ovarian cancers are a leading cause of cancer-related mortality. Endocrine therapies targeting androgen receptor and oestrogen receptor/progesterone receptor signalling have improved outcomes, but acquired resistance arising through genetic and epigenetic mechanisms that bypass receptor dependence remains a major challenge. BORIS (Brother of the Regulator of Imprinted Sites), a CTCF paralogue, is a germline-restricted regulator that becomes aberrantly reactivated in cancer through promoter hypomethylation, p53 loss, and CTCF displacement. Once expressed, BORIS displaces CTCF at shared genomic binding sites, disrupts TAD boundaries and reprograms CTCF binding sites into alternative transcriptional start sites, through SRCAP-mediated H2A.Z deposition, to activate germline genes that sustain proliferation, and therapeutic resistance. BORIS expression correlates with poor prognosis in ovarian and endometrial cancers, associates with ER/PR levels in breast cancer, and correlates with Gleason score in prostate cancer. BORIS represses AR signalling in ovarian cancer and inversely correlates with receptor expression in endometrial cancer. Its cancer-testis antigen properties position BORIS as an immunotherapy target, with preclinical vaccine efficacy demonstrated in breast cancer models. This review synthesises disease-specific evidence positioning BORIS as a driver of transcriptional reprogramming and endocrine resistance, identifies current knowledge gaps, and discusses translational implications of targeting this pathway.

Abstract: Nanobodies (VHHs, ~12-15 kDa) are single-domain antibody fragments valued for diagnostic applications, yet their engineering is frequently limited by poor solubility and aggregation. This study used the aggregation-prone anti-VirB4 nanobody a_b5 as a model to establish a two-stage solubility optimization strategy: rational single-point mutagenesis, then AI-driven de novo design. Consensus sequence analysis and CamSol prediction converged on glycine 54; G54A and G54D mutations reduced the Z-average hydrodynamic diameter by ~31% and ~32%, respectively, but particle sizes remained in the hundreds of nanometers, revealing an inherent ceiling for single-point mutagenesis. To surpass this limit, a modular AI pipeline integrating AlphaFold3 hotspot analysis, RFdiffusion backbone generation, dual-pathway ProteinMPNN sequence design (Vanilla and Soluble), AlphaFold3 ipTM screening, and 100-ns molecular dynamics was constructed. Among 780 designed sequences across 10 nanobody-antigen pairs, B5S achieved a CamSol solubility score of 3.007 (480% increase over wild-type 0.518). DLS confirmed a Z-average diameter of 6.084 nm with 43.4% monomer peak; B5S was solubly expressed in E. coli while the Vanilla counterpart B5V precipitated. Direct ELISA confirmed dose-dependent binding of B5S to VirB4 with apparent K_D ~856.2 nmol/L. These results demonstrate that AI-driven holistic redesign can simultaneously optimize binding interface quality and macromolecular solubility.

Abstract: The methylxanthine caffeine is one of the most widely consumed neuroactive substances globally. Caffeine in the context of coffee consumption has been associated with diverse effects on human health and has been widely reviewed. Caffeine has drawn considerable interest due to its effects on cellular signalling, DNA damage sensitivity, Target of Rapamycin (TOR) activity, activation of AMP-activated Protein Kinase (AMPK), and extension of chronological lifespan in fission yeast and other model organisms. The cell cycle effects of caffeine have overshadowed its modulation of DNA damage and repair. Caffeine provides an interesting example of how scientific knowledge evolves and is influenced by trends within epochs. Despite recent advances in our understanding of how caffeine modulates cell cycle progression, these findings raise many new questions. These include its structural similarity to adenine and the mechanisms by which it affects DNA damage and repair, and its impact on TOR Complex 2 (TORC2) activity. Herein, I review our current knowledge on the cellular effects of caffeine and focus primarily on studies in fission yeast. It proposes that caffeine be considered a radiomimetic compound, outlining our current state of knowledge and future directions in applying its pharmacology in the prevention and treatment of disease.

Abstract: The spliceosome became very complex in eukaryogenesis. The Group IIA intron progenitor was associated with a single protein, homologous to spliceosomal Prp8, but the LECA spliceosome included ~140 proteins. The acquisition of proteins was a neutral process, providing a pool of factors for the development of a coordinated assembly process, where proteins act as scaffold and chaperones, supporting RNA moieties. The RNA component of spliceosomal complexes is tiny. Structural studies offer us snapshots of protein re-arrangements remodelling the RNA. The spliceosome is commonly described as a ‘protein directed ribozyme’. What does this mean? Just how much control ribozymes can delegate to proteins? Spliceosomal ribozymes never lost their primary function of guiding catalysis by RNA base-pairing. To help with alternative splice site choices and to enforce precision, the spliceosome recruited another two small RNAs, U1 and U4, and still employs base-pairing. We discuss RNA structures central in spliceosomal and Group IIA intron ribozymes. Spliceosomal introns preserve protosplice site repeats CAG|GU at 5’ss and 3’ss that dictate a strict order of ribozyme folding. The demarcation of the 5’ss must involve the 3’ss in the downstream repeat. The distinct 5’-3’ss pair of spliceosomal introns serves to reconstruct the correct splice junction between the two repeats, preventing the intron ends from binding U5 snRNA Loop1. Although modern Group IIA introns never splice within repeats, structural and biochemical studies confirm 3’ss involvement at pre-catalytic stage. The exact configuration of the 5’-3’ss pair is different in Group IIA introns, but the parallel strands orientation is conserved. Our updated U5 model, that includes the 5’-3’ss pair, shows that the pre-mRNA strand flipping to achieve the local parallel orientation occurs after the short 3’exon duplex. This asymmetric 5’ and 3’exon binding with the recognition loop is shared with Group IIA introns. The exons are aligned for ligation on U5 Loop1, guided by Watson-Crick pairs as we have previously concluded based on positional dependencies at human splice sites. Our U5 model shows that exon duplexes of stacked pairs are demarcated from the 5’-3’ss pair by a gap, which is how the ribozyme structurally defines the cleavage sites.

Abstract: Interest in the properties of miRNAs has significantly declined in recent years, largely due to inadequate conceptualizations of their interactions with mRNA. Recent publications have shown a growing tendency to consider fully complementary miRNA interactions with mRNA of transcription factor genes. In the present study, we examined the miRNAs identified by Backes et al. which we have designated as BmiRNA to distinguish them from those in the NCBI database and other standard miRNAs. The MirTarget software was utilized to determine the quantitative physicochemical characteristics of the interactions between BmiRNAs and target mRNAs. BmiRNAs differ significantly from other miRNAs across numerous properties. We observed that BmiRNAs bind with full complementarity to the mRNAs of numerous transcription factor genes. Specifically, it is demonstrated that the mRNAs of ZNF genes and other transcription factors bind to BmiRNAs with full complementarity. Nearly all these BmiRNAs exhibited binding within the 3′UTR region. We identified BmiRNAs that interact with multiple target genes, as well as target genes that interact with multiple BmiRNAs, and defined groups of BmiRNAs that bind to the mRNAs of multiple genes. Furthermore, we identified BmiRNAs interacting with the mRNAs of transcription factor genes with high free energy. These results, regarding the interaction of BmiRNAs with the mRNAs of a significant number of genes, support the hypothesis that miRNAs serve as critical regulators of human genome expression.

Abstract: U2, U6 and U5 are the components of spliceosomal ribozymes that descend from mobile self-splicing introns. The fragmentation of ancestral introns in eukaryogenesis led to the evolution of the ribozyme assembly in trans using common snRNAs and protein-chaperones. This stepwise process provides opportunities to target distinct stages of ribozyme folding. Reviewing successive spliceosomal complexes, we will discuss how to adapt them for therapy, both splicing modulation and permanent gene therapy. Adapting U2 and U6 snRNAs can overcome the limitations of U1 and U7, currently used to target early spliceosome complexes. U2 can select alternative 3’ss, such as VEGFA switch between pro- and anti-angiogenic isoforms. U6 and U2 can be used together in the pre-catalytic complex, taking advantage of the U6/U2 Helix II that can be modified to prevent intermixing with WT U6 and U2 molecules. Extending 5’ss and BP helices with adapted U6 and U2 can create specific spliceosome species for individual introns. U5 snRNA, responsible for exon recognition, is the last to pair with pre-mRNA at pre-catalytic stage, but in reverse splicing, the exon recognition loop initiates intron insertion. Completed forward splicing produces an RNA-protein complex of an intron lariat paired with U6 and U2, the active ribozyme centre and U5 snRNA associated by protein interactions – the Intron Lariat Spliceosome. ILS is homologous to the mobile Group IIA intron particle capable of invading genomic loci by reverse splicing. Group II introns are used for microbial genome engineering. Can human ILS be adapted for specific genome insertions? Group IIA introns possess a second exon-recognition loop, which adds 6bp to the interaction with the 5’exon. Engineering an additional exon-recognition loop for spliceosomal U5 snRNA is the way to bring back reverse splicing. The field of genome engineering is dominated by CRISPR/Cas derivatives, so why do we need to develop reverse splicing? It is increasingly apparent that the evolutionary fate of RNPs defines their therapeutic utility: CRISPR/Cas are prokaryotic RNPs. Targeting human RNPs is the best solution, as the therapeutic success of RNAi using RISC complex indicates. The spliceosome is a human RNP worth exploring for splicing and gene therapy.

Abstract: Drebrin (DBN1) is an F-actin side-binding protein previously implicated in myoblast differentiation, but the signaling mechanisms linking DBN1-dependent actin organization to myogenic state transitions remain unclear. Here, we combined transcriptomic prioritization with C2C12 loss-of-function analyses to address this question. DBN1 depletion reduced filamentous actin and shifted actin-associated readouts toward a more monomeric state. This was accompanied by reduced nuclear MRTFA and SRF, lower CArG-dependent SMYD1 reporter activity, and decreased expression of SRF-responsive genes. DBN1 knockdown also altered YAP1 abundance and phosphorylation, reduced EdU incorporation and cell-cycle protein expression, and increased G0/G1 accumulation. During differentiation, DBN1 depletion attenuated MyoD, MyoG, and MyHC expression and impaired myotube formation. Public skeletal-muscle datasets further showed that DBN1 was preferentially associated with a myogenic rather than an atrophy-related transcriptional program, although its regulation varied across aging and muscle-wasting contexts. Together, these findings identify DBN1-dependent actin organization as a link to MRTFA–SRF transcription during myogenic progression and associate altered YAP1 signaling with impaired precursor proliferation after DBN1 depletion.

Abstract: The mainstream approach to splicing therapy today involves synthetic antisense oligonucleotides. ASOs interfere with spliceosomal ribozyme assembly by blocking splice sites or influence protein regulators by blocking or adding their binding sites. ASOs are not compatible with human biology, which limits their efficacy and they come with a burden of chemical toxicity. Spliceosomal snRNAs adapted for a specific target can either enhance or suppress exon inclusion. Recombinant snRNAs with their pre-mRNA binding sites changed still assemble into functional snRNPs and integrate into active spliceosomes. Native to human cells, they are versatile for in vivo delivery and can be either encoded and expressed from rAAVs or delivered as RNA molecules by exosomes, benefitting from excellent biodistribution. Imported snRNAs are taken up by cytoplasmic maturation factors and transferred into the nucleus. Adapted snRNAs can be used to target different stage spliceosomes: U1 and U2 for early complexes, U2/U6 for precatalytic or U5 for catalytic complexes. U1 is the only spliceosomal snRNA thoroughly examined in pre-clinical studies (~100 mutations in 24 genes) and it is necessary to review the past 20 years of experience with U1 before moving on to other snRNAs. U1 is involved in the initial splice site selection and usually binds the 5’ss, but it also promotes 5’ss usage ‘at a distance’ if bound in the vicinity. Adapted U1 suppresses 3’ss usage, as likely does WT U1 if 3’ss CAG|GU protosplice site repeat is followed by a sequence resembling the start of the intron. However, historically the non-spliceosomal U7-OPT is often used as a scaffold molecule for exon-skipping ASOs. This chimeric U7 snRNA cannot join histone bodies, its WT destination. While both U1 and U7 show promising safety profiles in mice, WT U7 expression is 1000 times lower, than U1. U7 snRNA modulates master transcription regulators by binding histone-fold domain protein NF-Y; the effects of U7-OPT overexpression remain underexplored. U1, a splicing molecule, naturally works better for promoting exon inclusion, than U7, but years of experiments also revealed U1 and U7 limitations. New adaptations of the core spliceosomal snRNAs are necessary to overcome these limitations.

Abstract: Artificial intelligence systems deployed across medicine—drug discovery, diagnostic imaging, genomic analysis, epidemiological modeling, and clinical decision support—are trained on domain knowledge that carries both legitimate therapeutic value and dual-use potential. Prevailing AI safety practice relies on technical containment: refusal training, output classification, red-teaming, and access controls. This paper advances a bounded thesis: technical containment is a necessary but insufficient layer for managing dual-use risk in high-consequence biomedical domains, because reliable recognition of dangerous outputs requires expertise that is itself hazardous, concentrating insider risk within the very teams tasked with safety. This is not a claim that misuse is inevitable or that containment is futile; it is a claim about the limits of any single layer. The paper adopts a graded-risk model in which computational capability, tacit knowledge, materials, and infrastructure jointly determine real-world uplift, and it argues that these barriers differ substantially across domains and tools. It surveys dual-use research governance, AI safety methods, biosecurity institutions, and medical-AI regulation to locate the gaps, and proposes an operationalized, layered governance response—including explicit deployment-gating criteria for the highest-risk applications. The paper does not present novel empirical findings and identifies the empirical questions on which its practical implications depend.

Abstract: Background/Objectives: Breast cancer (BC), particularly aggressive subtypes such as triple-negative TNBC, remains a significant clinical challenge due to treatment resistance and therapeutic toxicity. This study evaluated the antitumor potential of Mexican Oregano (Lippia graveolens) infusion (MoI) and its bioactive constituent, carvacrol (Cv), in diverse BC cell lines (MCF-7, HCC-1954, and MDA-MB-231). Methods: Cellular responses were assessed by quantifying apoptosis, profiling cytokines (IL-4, IL-6, IL-10), and analyzing the gene expression of antioxidant enzymes (CAT, SOD2, GPX1) and their enzymatic activities. Results: Cv exhibited potent, subtype-independent pro-apoptotic activity, while MoI demonstrated significant efficacy with a more favorable safety profile in non-tumoral HaCaT cells. Both treatments mediated complex modulation of cytokine signaling, characterized by suppression of tumor-promoting IL-4, IL-6, and IL-10, an effect most pronounced in the highly aggressive MDA-MB-231 cell line. Transcriptional and functional analyses further revealed that these agents alter the expression and activity of antioxidant enzymes., thereby promoting pro-oxidant environments conducive to cancer cell death. Conclusions: These findings indicate that MoI and Cv exert antitumor effects through coordinated pathways of apoptosis induction, immunomodulation, and redox disruption. Consequently, MoI emerges as a promising candidate for complementary therapeutic strategies in BC management, particularly for aggressive subtypes, owing to its broad modulation of inflammatory networks and redox vulnerabilities. At the same time, Cv serves as a compelling lead compound for targeted oncological research.

Abstract: Endometrial-like tissue can establish persistent lesions outside its usual environment, but anatomical explanations for its arrival leave a second question unresolved: what permits survival during establishment? We evaluated a pre-existing selective redox-survival hypothesis using 27 historical transcriptomic comparisons across six accessions and an external donor-level analysis of proliferative eutopic endometrium from 12 endometriosis cases and 11 clinical controls. TXN reduction and ovarian ferritin/metabolic changes recurred across specified historical lesion cohorts. Historical mean survival-module shifts were null after across-test correction. The conservative expanded nondirectional sensitivity family retained eight historical but no external survival-associated sets. No external primary panel gene or population interaction met its declared threshold; limited overlapping HIF-set signals did not pass external-only correction. A later secondary test of paired peritoneal/eutopic responses found that held-out donors resembled their own cellular context more than other contexts: similarity margin 0.298, permutation P=0.0001, with 69.0% classification accuracy versus 16.7% chance. This used seven donors, six non-overlapping populations and 43 shared frozen-panel genes. Convergence varied by population and did not clearly exceed expression-matched background sets (empirical P=0.0939). The results support partial context constraint with substantial patient flexibility, while narrowing extrapolation to a common eutopic survival state. They do not establish redox-specific protection, early survival advantage, intercellular rescue or a receptor/proteoform mechanism. The central causal proposition remains unresolved.

Abstract: Specific cell tracking or targeted modulation of cellular responses are often limited by the availability, specificity and effector functions of endogenous cell-surface receptors. Synthetic receptors with customizable functionality can overcome these limitations. Based on a qualitative survey of the literature, this narrative review describes the design, the functionality and the applications of nanobody based synthetic receptor/ligand systems that employ either GFP/mCherry or the clinically approved humanized antibody Palivizumab as ligands. Depending on their molecular architecture, these systems can mediate cargo internalization for cell tracking or induce defined cellular responses through engineered interleukin, interferon, or Fas signaling pathways. We position these platforms within the broader field of synthetic receptors by discussing engineered GPCR- and Notch-based systems and briefly outline the development and clinical application of CAR T cells to illustrate the translational potential of synthetic receptor systems. Finally, we discuss the opportunities and limitations of GFP/mCherry- and Palivizumab-based systems, with particular emphasis on immunogenicity, in vivo applicability, manufacturability, scalability, and clinical translation. By linking modular receptor design to distinct functional outputs, this review highlights both the established value of GFP/mCherry- and Palivizumab-based synthetic receptor–ligand systems as experimental tools and their potential for future applications in molecular imaging and controlled signal transduction.

Abstract: In 100 and plus years of stem cell research, scientists are trying to define all the types of stem cells, their markers, differentiation abilities, and possibilities that those cells hold [1]. Induced pluripotent stem cells (iPSCS) are "human-made", induced stem cells that can be produced from different donors and cell types. They have shown to own multiple advantages compared to "natural" stem cells, such as embryonic stem cells (ESCs), adult stem cells, and fetal stem cells. iPSCs were induced by reprogramming of somatic cells, mouse embryonic fibroblasts (MEFs) in 2006, by Takahasi and Yamanaka [2]. The new era started after that, and it is still ongoing. Defining usage of different cells, the best donors, then states of cells during iPSCs reprogramming, and the markers that define them may pave the way for easier reprogramming, thus the focus of research may be on the possibilities that iPSCs hold in regenerative medicine [3], preservation of vulnerable and endangered species [4], drug discovery and testing, disease modeling [3] and cell therapy. One of the ways that can make the path easier, is tools that would collect data, analyze data and give the approximation of the solution. Artificial intelligence which applies machine learning, deep learning and other techniques, may be a valuable factor in producing models and comparing in effectiveness during the reprogramming. Human factor provides data, while artificial intelligence-based methods can bring improvement in accuracy, costs and speed of data analysis [5].

Abstract: This paper examines dihomo-γ-linolenic acid (DGLA, 20:3n-6) as a unique molecule among the polyunsaturated fatty acids (PUFAs) of the n-6 family. It has a pronounced anti-inflammatory effect and significant clinical potential. Unlike arachidonic acid (AA), from which pro-inflammatory eicosanoids are formed, DGLA is metabolized to prostaglandin E1 and 15-hydroxyeicosatrienoic acid. These compounds exhibit anti-inflammatory, vasodilating, and anti-aggregatory properties. An analysis of clinical studies indicates that low levels of DGLA are an independent predictor of all-cause mortality in patients with cardiovascular diseases. Additionally, the therapeutic use of DGLA effectively reduces the symptoms of allergic rhinitis. Particular attention is paid to genetic variability in the FADS1 gene. It modulates the rate of conversion of the precursor of DGLA (gamma-linolenic acid) to AA. This justifies the need for a personalized approach to nutritional therapy. The role of zinc as a cofactor of delta-6-desaturase is highlighted. Its deficiency causes an increase in the LA/DGLA ratio, a potential biomarker of zinc status. New therapeutic strategies are being explored, such as the target inhibition of delta-5-desaturase, which redirects DGLA metabolism toward the formation of 8-hydroxyoctanoic acid, a metabolite with potent anticancer activity. In conclusion, DGLA is a key mediator of inflammatory homeostasis. An integrated approach incorporating genetic, nutritional, and dietary factors is promising for the implementation of personalized therapeutic strategies.

Abstract: Plant pangenomics has rapidly changed the way genetic diversity is represented and studied. During the last three years, advances in long-read sequencing, chromosome-scale genome assembly, structural-variant discovery, graph genomes, haplotype-resolved assemblies and super-pangenomes have demonstrated that a single reference genome is insufficient to describe the genetic diversity of crop species. Today the current challenges in plant pangenomics is the transformation of the pangenomics based information into gleanable resources, which could be biologically interpreted and useful for plant improvement. Pangenomics has been predominantly descriptive "catalogue of variation" paradigm and transformed toward a functional and predictive pangenomics framework integrating structural variation, regulatory sequences, transcriptomics, epigenomics, phenomics and artificial intelligence. This transition is particularly important for complex and polyploid crops, where conventional reference-based approaches remain inadequate. Super-pangenomes that incorporate wild relatives provide an important opportunity to recover genetic diversity lost during domestication. To achieve, pangenomes and super-pangenomes, computational scalability, functional validation, standardization and equitable access remain major challenges. In this paper, we present computational approaches to the pangenomics and introduce computational end to end scalabale pangenomic workflow and panscape, which serves a single standalone tool for all pangenomic needs.

Abstract: Spaceflight (FLT) is associated with fluid redistribution, cardiovascular deconditioning, and altered vascular tone. We sought to determine whether FLT induces molecular alterations in human aortic smooth muscle cells (HASMCs) that could provide a mechanistic basis for altered vascular function. We analyzed NASA GeneLab/Open Science Data Repository dataset OSD-635, comprising mRNA sequencing of primary HASMCs cultured for 3 days aboard the International Space Station compared with ground controls. Up- and downregulated differentially expressed genes (DEGs), together with genes ranked by a composite significance-magnitude pi-score, overlapped with GSE225884, a transcriptomic readout of HASMCs exposed to lactacidosis. The gene intersection contained 182 genes shared in the upregulated direction and 382 shared in the downregulated direction for DEG comparisons; corresponding pi-score intersections contained 170 and 180 genes, respectively. Convergent upregulated genes included glucose/glycogen-associated genes, PYGL, PYGM, GCK, ALDOC and PGM2L1, and fatty-acid/mitochondrial metabolism genes ACADS, HADH, IDH2 and LDHD. Multiple genes involved in mitochondrial oxidative phosphorylation and maintenance were downregulated including NDUFAF8, NDUFC2, NDUFS6, COQ2, UQCC4, TIMM23, MFN2 and ACO2. Additional robustness was observed because in the FLT study 98 genes were shared between the DEG-up and pi-score-up intersections and 127 between the DEG-down and pi-score-down intersections. FLT-exposed HASMCs exhibit a lactacidosis-associated transcriptomic remodeling which may contribute to vascular dysfunction.

Abstract: Arthrogryposis–renal dysfunction–cholestasis (ARC) syndrome is a rare, usually fatal autosomal recessive multisystem disorder caused by biallelic mutations in VPS33B or VIPAS39, which encode components of a vesicular trafficking complex required for organelle biogenesis. Loss of function causes a platelet α-granule deficiency and a severe bleeding diathesis, and no curative therapy exists. Building on the efficacy of ex vivo lentiviral gene therapy in inherited haematological disorders, we evaluated haematopoietic stem cell (HSC) gene delivery in a tamoxifen-inducible Vps33b knockout mouse model (Vps33b^fl/fl-CreER^T2). Knockout mice tolerated myeloablative conditioning and supported efficient donor reconstitution after transplantation of healthy lineage-negative cells. We then transduced lineage-negative cells from knockout donors ex vivo with a third-generation lentiviral vector encoding codon-optimised human VPS33B under an elongation factor 1α short (EFS) promoter and transplanted them into conditioned knockout recipients. Transmission electron microscopy showed that platelet α-granule numbers were restored to wild-type levels after both bone marrow transplantation and ex vivo gene therapy. These findings demonstrate the feasibility of haematopoietic VPS33B gene delivery and support further preclinical evaluation of gene therapy for the bleeding phenotype of ARC syndrome.

Abstract: Rheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by persistent joint inflammation, bone erosion, and systemic involvement with genetic factors contributing significantly to disease susceptibility. Interleukin-17 (IL-17) signaling has been strongly implicated in autoimmune inflammation, making IL-17A and its receptor IL-17RC key candidates for genetic association studies. This case control study investigated IL-17A (rs2275913 G/A) and IL-17RC (rs708567 G/A) polymorphisms in 120 RA patients and 120 controls from Pakistan. Genotyping was performed using the PCR-RFLP method, and statistical analyses included allele and genotype frequency comparisons, odds ratio calculations, multiple inheritance model testing, and Hardy–Weinberg equilibrium assessment. The IL-17RC rs708567 variant showed a robust association with RA, with the A allele significantly more frequent in patients (43.3%) than controls (22.1%) and conferring an almost threefold increased risk (OR = 2.88, p = 1.42 × 10⁻⁷). Genotypic, allelic, and dominant model analyses confirmed this strong association. By contrast, the IL-17A rs2275913 variant showed no significant association, although a suggestive trend was observed under the dominant model. Hardy–Weinberg equilibrium analysis revealed deviations in the overall and control groups but equilibrium in patients, suggesting possible stratification or underlying genetic association. These findings highlight IL-17RC rs708567 as a potential genetic marker for RA susceptibility in the Pakistani population, reinforcing the importance of IL-17 signaling in RA pathogenesis and warranting further studies in larger cohorts. In conclusion, this study provides novel evidence for the role of IL-17RC in RA in Pakistan and supports its relevance as a therapeutic target. Further large-scale investigations are essential to validate these findings and advance personalized treatment strategies.