FEBS Open Bio最新文献

Heavy alcohol drinking is known to increase the risk of bacterial pneumonia. However, the link between alcohol levels and risk of infection remains underexplored. Recently, we found that alcohol induced α2-6sialo mucin O-glycans in human tracheobronchial epithelial cells, which mediated the killing of U937 macrophages. By extending this study, we focus here on whether altered glycans induced by alcohol in human airway epithelial cells can promote adhesion of Klebsiella pneumoniae (Kp) and Streptococcus pneumoniae (Sp). We have found that exposure of human tracheal epithelial cells to alcohol also induces high mannose N-glycans terminated with α3mannose and increases adhesion of Kp, which is inhibited by αmethylmannoside or aldehyde dehydrogenase 2 activator 1. Further, the α2-6sialo mucin O-glycans induced by alcohol in human tracheal epithelial cells also enhance the adhesion of Sp, which is inhibited by ovine submaxillary mucin or aldehyde dehydrogenase 2 activator 1. We conclude that alcohol induces altered glycans in human airway epithelial cells, which increase the risk of bacterial pneumonia by compromising immune function and promoting the adhesion of Kp and Sp.

A variety of dual reporter HIV-1 derivatives have been developed that enable detection of infected cells independently from transcription initiated from the 5' LTR promoter. These reporters enable isolation of cells that are latently infected with HIV-1 provirus. We have previously described several dual reporter derivatives, including Red Green HIV-1 (RGH), which expresses mCherry from a constitutive internal promoter and GFP from the 5' LTR. A limitation of most dual HIV-1 reporter derivatives, including RGH, is that the Nef ORF is often disrupted to accommodate insertion of the internal reporter. Consequently, the potential role of Nef for establishment of latency using these reporters has not been clarified. To address this issue, we created three different RGH derivatives (RGHI), which express Nef from IRES elements downstream of the mCherry internal promoter. We found that each of these derivatives produced Nef protein at higher levels than wild-type LAI virus and that Nef expressed from each of the IRES elements was localized to the cell membrane. All of the Nef-expressing RGHI reporter viruses formed latent and productive infections and could be reactivated from latency at similar levels as the parental RGH derivative. These results indicate that expression of Nef does not affect the capability of HIV-1 to form latent infections. Furthermore, we propose that the RGHI derivatives described here represent novel tools to examine the role of Nef for HIV-1 replication.

Primary fibroblasts are widely used in a variety of experimental and therapeutic studies. Patient-derived skin biopsies are an accessible way to generate dermal fibroblasts for wound and burn therapeutics and can be easily reprogrammed to induced pluripotent stem cells (iPSCs). Despite the increasing use and interest in skin biopsies, there is limited information regarding the culturing potential of a single biopsy and the effects of extended culture on fibroblast formation and reprogramming potential. To better understand the potential of long-term skin biopsy culture, we cultured biopsy samples for 6–16 months, resulting in 6–16 generations of explant reculturing and then analyzed subsequent generations of fibroblasts. Our results showed that fibroblast morphology and physiology are maintained over time, but although older generations remained proliferative, they did so at a decreased rate. Gene expression analyses uncovered transcriptional changes with long-term skin culture, but deep DNA sequencing did not reveal any large deletions or amplifications. Spontaneous DNA mutations in fibroblast generations appeared to be random and not enriched for any specific signaling pathways. Importantly, fibroblasts generated after 16 months and over 16 generations in explant culture retained competency for reprogramming into induced pluripotent stem cells. Taken together, our results support long-term culture of skin biopsies to generate large numbers of primary fibroblasts. These cells maintain their identity and integrity, enabling the study of fibroblast maintenance as well as rare human disorders.

Genome stability and faithful DNA replication are essential for cell viability. Numerous interlinked pathways in DNA damage recognition, repair and maintenance of physiologically competent nucleotide pools contribute to providing a solid framework to uphold DNA integrity. The enzyme family of dUTPases is involved in balancing the appropriate nucleotide pools by removing dUTP from the cellular milieu and providing dUMP for thymidylate de novo biosynthesis. In the present study, we show that dUTPase is essential for normal development in zebrafish. We also found that the fish dut gene from different genomes contains several single-nucleotide variations (SNPs). This observation prompted structural and functional investigations of the SNP variants at the protein level. Results indicated that none of the mutation sites of the variants are within the active site. Still, one of the variants showed drastically lower protein stability and catalytic efficiency as compared to the other two dUTPase variants, underlining the importance of detailed characterization of SNPs even at sites distant from the active site. In conclusion, we demonstrate the importance of dUTPase function in zebrafish development and unveil the role of several point mutations on protein structure and function.

Senescence is a complex cellular state characterised by irreversible growth arrest and metabolic reprogramming. In neurons, senescence has been mainly observed in the context of ageing and age-related neurodegeneration. Lipid metabolism plays a critical role in cellular homeostasis, with emerging evidence suggesting that alterations in lipid species, including fatty acids, cholesterol, sphingolipids and phospholipids, fundamentally drive or contribute to the senescent phenotype in both neuronal and non-neuronal cells in the brain. Namely, changes in lipid species levels result in the accumulation of lipid droplets (LDs), leading to dysregulation of membrane dynamics, and in turn to the production of bioactive lipid mediators, which collectively shape the senescence-associated secretory phenotype (SASP) in the brain. In this review, we describe the cell type-specific patterns of lipid dysregulation in neurons, astrocytes, microglia and other glial cells during senescence, highlighting the role of key lipid species and their association with senescence markers and phenotypes. Furthermore, we discuss the bidirectional relationship between lipid metabolism and mitochondrial dysfunction in cellular senescence. We also examine the molecular mechanisms through which lipid metabolic pathways can orchestrate neural senescence and their contribution to ageing and age-related neurodegenerative disorders, such as Alzheimer's disease and Parkinson's disease. Finally, we review emerging therapeutic strategies targeting lipid metabolic pathways to modulate neural senescence and potentially ameliorate age-associated brain pathology.

Gemcitabine/Cisplatin (Gem/Cis) chemotherapy is a standard treatment for muscle-invasive bladder cancer (MIBC) but yields suboptimal response rates. The contribution of tumor-stromal crosstalk and macrophage recruitment to chemoresistance remains poorly understood. This study investigated these mechanisms using a functional ex vivo bladder cancer tissue slice model combined with n = 64 spatial transcriptomics. Spatial analysis revealed transcriptomic changes involving the immunomodulating gene SPP1 that has been also recently presented as a putative predictive biomarker for neoadjuvant chemotherapy in bladder cancer. Moreover, Non-Responders exhibited upregulation of chemokines including CXCL1 and CXCL8 and enrichment of immunoregulatory M2 macrophages in tumor regions, suggesting active macrophage recruitment from the stroma. On the contrary, Responders showed upregulation of complement components, proinflammatory macrophage subsets and signals associated with cytotoxic lymphocyte recruitment. Tissue slices corresponding cell cultures confirmed overexpression of immunomodulating markers including checkpoints PD-L1 and PD-L2 in Non-Responder cancer cells upon Gem/Cis treatment. Using TCGA bladder cancer data, the transcriptomic gene set was further validated revealing a prognostic signature associated with patients' outcome. These findings uncover a novel mechanism of chemotherapy resistance in bladder cancer driven by tumor-stromal interactions and macrophage recruitment and suggest that targeting macrophage infiltration may improve chemotherapy response in bladder cancer.

A growing demand for sustainable materials across various industries has sparked an increasing interest in bio-based polymers as eco-friendly alternatives to conventional fossil-based polymers. Sourced from renewable materials, bio-based polymers offer significant advantages, such as biocompatibility, the ability to modify their functional properties for specific applications and, increasingly sought after, the capability for biodegradation. This review article provides an overview of bio-based polymer sources, discussing their unique functional properties, environmental impact and potential for end-of-life options, such as composting and anaerobic digestion. It highlights the importance of ensuring human health and environmental hazard assessment, by incorporating principles like a Safe and Sustainable by Design (SSbD) approach and assessing the product's life cycle (LCA). The dual role of the anaerobic digestion of biodegradable polymers and its potential for methane generation is reviewed, emphasising its contribution to reducing environmental impact and renewable energy production through waste management. Lastly, possibilities of applications in different industries and future market trends are reviewed. By integrating current knowledge, this review highlights the potential of bio-based polymers in advancing sustainability across various sectors, while addressing key existing challenges and future opportunities in their development, production, and application across various sectors, while addressing key existing challenges and future opportunities in their development, production and application.

Plastic waste from fossil-derived polymers remains a major environmental challenge, driving interest in biopolymers and enzyme-enabled end-of-life strategies. This review synthesizes current understanding of how polymer structure and thermal state govern enzymatic degradability, with emphasis on semicrystalline architectures and state-dependent accessibility. Within the Keller-Flory two-phase framework, crystalline lamellae embedded in an amorphous matrix dictate water/enzyme diffusion, chain mobility, and hydrolysis kinetics. Enzymatic attack preferentially initiates in amorphous regions, producing characteristic biphasic behavior as amorphous domains erode faster than crystalline regions, leading to crystallinity enrichment and subsequent slowing of degradation. Thermal transitions further modulate this balance: near or above T_g, segmental mobility and free volume rise, accelerating hydrolysis if enzymes remain stable; above T_m, chain mobility is maximal, but enzyme stability typically limits feasibility. Processing and architecture also strongly influence outcomes: annealing increases crystallinity and slows mass loss, quenching suppresses crystallization and hastens degradation, random copolymerization disrupts packing and lowers T_m, while block copolymers often degrade selectively by domain. Recent advances expand the operational window toward rubbery or near-molten states for low-melting aliphatic polyesters (e.g., PCL, PLGA, PEG-b-PLA), leveraging thermophilic/engineered hydrolases (cutinases, PETases, lipases, carboxylesterases) with demonstrated stability at 60-90 °C. Emerging strategies-including enzyme thermostabilization, AI-guided design, disulfide grafting, smart encapsulation, and in-situ enzyme embedding-enable self-degradation of materials and accelerate inside-out depolymerization under mild triggers. Integrating thermal analysis with polymer morphology and enzyme engineering offers a path to programmable, circular end-of-life for biopolymers, translating fundamental structure-property-reactivity relationships into practical enzymatic recycling and reduced environmental impact.

Herbal and dietary supplements (HDS) are popular among consumers seeking a 'natural' approach for improving their health; however, at present, there is a lack of evidence to support the claims of efficacy and safety for most of these products. Herbal weight loss supplements (WLS) are a group of HDS that are frequently implicated in cases of toxicity; however, the causative substances often remain unknown due to the complex chemical nature of such supplements. This study aimed to analyse the in vitro safety (in human liver carcinoma (HepG2) cells and colon carcinoma (Caco-2) cells) of 12 active compounds commonly found in WLS, first with safety screening using the MTT cytotoxicity assay, followed by metabolic profiling with ¹H NMR spectroscopy. Of the phytochemicals evaluated, epigallocatechin-3,0-gallate (EGCG) was the only compound that caused a significant reduction in the viability of both cell lines (25.3% in HepG2 cells and 18.5% in Caco-2 cells), and this decrease was potentiated by CYP450 induction with rifampicin. Subsequent ¹H NMR analysis showed changes in key metabolites such as amines, amino acids, carboxylic acids, and glucose that were indicative of protein degradation and disrupted energy and lipid metabolism. While the remaining 11 active compounds analysed did not demonstrate significant toxicity in isolation, these require further assessment to determine their safety when used in combination with other phytochemicals. Given that the majority of WLS contain multiple herbal ingredients, each with a complex chemical composition, it is important to understand the role of interactions in adverse events.

Fluorescence recovery after photobleaching (FRAP) is a quantitative technique to study the dynamics of fluorescently tagged proteins in living cells. Current FRAP workflows are limited in throughput because of the requirement for human interaction. Here, we present RoboMic, a fully automated confocal microscopy platform for high-throughput imaging assays such as FRAP. We demonstrate its capabilities using two complementary approaches: sequential FRAP (sFRAP) and a novel parallel FRAP (pFRAP). The latter enables simultaneous photobleaching and monitoring of multiple cells within one imaging cycle, increasing throughput by approximately five- to 10-fold while maintaining spatiotemporal resolution. The protocol consists of microscope control software for automated, AI-based selection and segmentation of cell nuclei, sub-nuclear ROI definition, photobleaching, and time-lapse imaging. As proof of concept, we examined the nuclear dynamics of the androgen receptor and the cohesin complex under diverse conditions, demonstrating that RoboMic generates robust and reproducible data. In a single session, the platform yields hundreds of FRAP measurements, thereby increasing statistical power and scalability for large-scale studies of protein mobility. While we focus here on FRAP, RoboMic can be readily applied to a wide range of quantitative functional imaging assays.