Biochimie最新文献

{"title":"Optimized high-resolution respirometry defines the bioenergetic signature of Drosophila melanogaster thoracic muscle.","authors":"Felipe Berti Valer, Giulia Covolo Spegiorim, Jonatas Rodrigo Dos Santos, Ana Elisa Caleiro Seixas Azzolini, Marcus Fernandes Oliveira, Luciane Carla Alberici","doi":"10.1016/j.biochi.2026.09.007","DOIUrl":"10.1016/j.biochi.2026.09.007","url":null,"abstract":"<p><p>Drosophila melanogaster provides a robust model system for investigating mitochondrial metabolism in muscle tissue, particularly due to availability of fly strains that recapitulate genetic mitochondrial disorders, muscle atrophy, and related pathological conditions. However, most experimental approaches are adaptations of protocols originally developed for mammalian muscles, primarily rodents. Here, we systematically applied and refined variations of these mammalian-based protocols to female adult thoracic muscles of D. melanogaster, with particular emphasis on tissue permeabilization, respiratory substrates, respiratory chain inhibitors, and oxidative phosphorylation uncouplers. Our findings appointed pyruvate as the principal respiratory substrate and suggested succinate as a major anaplerotic contributor to the tricarboxylic acid cycle during NADH dehydrogenase-supported respiration. In addition, Drosophila thoracic muscle exhibits a maximal respiratory flux strongly dependent on ADP levels, with oxidative phosphorylation operating near the maximal capacity of the electron transfer system. We compared thoracic muscle with larvae fillet muscle, as well as murine and human skeletal muscle, revealing species- and tissue-specific differences in mitochondrial metabolism. Finally, we validated our refined protocol in a PINK1 RNAi Drosophila model, extending previous evidence of mitochondrial dysfunction associated with PINK1 deficiency in flies.</p>","PeriodicalId":93898,"journal":{"name":"Biochimie","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-09-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149027024","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Effects of fine particulate matter on neuroblastoma cell line: metabolic and lipid deregulations and their consequences on the onset of neurological diseases.","authors":"Candice Dulong, Ana-Maria Trunfio-Sfarghiu, Adina-Nicoleta Lazar, Nathalie Bernoud-Hubac","doi":"10.1016/j.biochi.2026.09.011","DOIUrl":"10.1016/j.biochi.2026.09.011","url":null,"abstract":"<p><p>Particulate Matter (PM) contained in atmospheric pollution can interact with pulmonary, cardiac and nervous systems. Recent studies focus on the impact of PM2.5 on the brain and on their potential role in the onset of neurodegenerative diseases such as Alzheimer's disease. Using human neuroblastoma SH-SY5Y cell line as an in vitro neurodegenerative model, multiple interconnected cellular dysfunctions induced by PM2.5 exposure have been highlighted. This includes mitochondrial damages, endoplasmic reticulum stress, oxidative stress, calcium dyshomeostasis and lipid dysregulation. These alterations activate multiple cell death pathway though apoptosis and autophagy-dependent ferroptosis. This review aims to provide an integrated overview of these mechanisms, to better understand the association between PM2.5 exposure and cell degeneration. A particular attention is given to lipid peroxidation and oxidative damage as emerging drivers of neuronal degeneration. Despite significant advances, important gaps remain regarding the translation of in vitro findings to in vivo models and particularly to human pathology. A better understanding of key molecular events is essential to identify therapeutic targets to mitigate or prevent pollution-related neurodegenerative processes.</p>","PeriodicalId":93898,"journal":{"name":"Biochimie","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-09-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149003215","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The MAPK signalling landscape of Leishmania: structural diversity, biological functions and therapeutic potential.","authors":"Priyanka Mazire, Ritika Chandekar, Amit Roy","doi":"10.1016/j.biochi.2026.09.009","DOIUrl":"10.1016/j.biochi.2026.09.009","url":null,"abstract":"<p><p>The remarkable adaptability of Leishmania throughout its complex life cycle depends on signalling networks that integrate environmental stress, developmental cues, and host-derived signals. Among these, mitogen-activated protein kinases (MAPKs) have emerged as important regulatory nodes, yet their contribution to parasite biology and their value as potential therapeutic targets remain incompletely resolved. This review examines the expanding landscape of Leishmania MAPKs, encompassing 15 homologues across pathogenic species, and synthesizes evidence linking individual kinases to parasite differentiation, intracellular survival, stress tolerance, motility, virulence, and drug response. Rather than treating the MAPK repertoire as a uniform kinase family, we critically assess the extent to which individual members have been experimentally validated and distinguish mechanistically established targets from computationally proposed candidates. Structural peculiarities, regulatory architecture, and substrate recognition offer potential opportunities for parasite-selective kinase inhibition; however, functional redundancy, incomplete kinase-substrate networks, stage-specific signalling, and conservation of MAPK pathways in the host remain substantial obstacles. We further evaluate small-molecule and natural-product inhibitors, drug-repurposing strategies, and structure-guided approaches, while considering emerging contributions from molecular dynamics, AlphaFold-based structural modelling, artificial intelligence, and nanotechnology to drug discovery. Beyond chemotherapy, evidence supporting MAPK10 as a vaccine-associated antigen broadens the translational relevance of this signalling system. Thus, the current findings position Leishmania MAPKs as a multifaceted interface between parasite signalling, adaptation, and therapeutic vulnerability. In this context, future studies should define the signalling dependencies that distinguish essential MAPKs from functionally redundant or context-dependent kinases, thereby facilitating the translation of current mechanistic insights into selective and clinically actionable therapies for leishmaniasis.</p>","PeriodicalId":93898,"journal":{"name":"Biochimie","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148984471","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Unlocking the role of fungal peptide cytotoxin candidalysin in candidiasis and non-candidiasis diseases.","authors":"Hanyu Zhu, Xue Wang, Jing Shao","doi":"10.1016/j.biochi.2026.09.004","DOIUrl":"https://doi.org/10.1016/j.biochi.2026.09.004","url":null,"abstract":"<p><p>Candida albicans is a commonly encountered opportunistic fungus that can cause both superficial discomforts and life-threatening invasive fungal infections. Ever-increasing reports have shown that C. albicans is also readily implicated in a set of metabolic and autoimmune diseases. Yeast-to-hypha transition is a hallmark of increased virulence and invasion of C. albicans, and candidalysin is the first peptide toxin secreted by C. albicans hyphae. Candidalysin contributes to fungal penetration across epithelial barrier via cytolytic activity, as well as the maintenance of commensalism and pathogenicity of Candida albicans. To specifically depict the role of candidalysin in multiple candidiasis and C. albicans associated infections, in this review, we focus on the recent advancements of candidalysin in oropharyngeal candidiasis (OPC), vulvovaginal candidiasis (VVC), disseminated candidaemia and sepsis. Meanwhile, we also pay close attention to the potential pathogenic mechanism of candidalysin in several C. albicans associated diseases including inflammatory bowel disease (IBD), asthma, Alzheimer's disease (AD) and alcohol-associated liver diseases (ALD). Based on these achievements, we propose a list of unresolved questions of interest for future investigation on biological functions and therapeutic potential of candidalysin in fungal infections caused by C. albicans. Unlocking the mode of action of candidalysin is beneficial for deep understanding of how C. albicans breaks through mucosal or blood brain barrier to colonize the niche via candidalysin and for rational design of fungal vaccine against candidalysin for clinical applications. Q1: Revision based on the comment 1 by Reviewer 1.</p>","PeriodicalId":93898,"journal":{"name":"Biochimie","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148984464","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"NFYA regulates the expression of CD44 via binding to upstream cis-element in breast cancer cells.","authors":"Sohair Salem, Shaymaa M M Yahya","doi":"10.1016/j.biochi.2026.09.010","DOIUrl":"10.1016/j.biochi.2026.09.010","url":null,"abstract":"<p><p>CRISPR/Cas9 has emerged as a powerful tool for editing of non-coding sequences, such as promoters and enhancers. Creating indels in enhancer sequences alter their function and provide valuable insights into their role in gene expression under normal and pathological conditions. This study tested the involvement of CD44-upstream cis-element in regulating CD44 expression and evaluated the potential of CRISPR/Cas9 system to manipulate the transcription factor (TF) binding site within this element. Bioinformatic tools predicted two binding sites (designated P1 and P2) for NFYA within the cis-element sequence. CRISPR/Cas9-mediated genome editing was employed to knockout NFYA gene and disrupt the NFYA-binding site at P2 in MDA-MB-231 breast cancer cells, then downstream analysis was performed using the mixed-edited cell population. Both genome editing strategies resulted in a significant reduction in CD44 gene expression at mRNA level. Notably, disruption of P2 sequence produced a greater reduction in the CD44⁺ CD24^- cell subpopulation than NFYA knockout. Both editing approaches impaired the migration capacity of MDA-MB-231 cells. Unlike NFYA knockout, altering P2 sequence induced apoptosis. ChIP assay revealed that NFYA binds to both P1 and P2 sequences, with greater enrichment observed at P2 than at P1. Site-directed mutagenesis and luciferase assay confirmed the regulatory contribution of both P1 and P2 to gene expression regulation, with stronger regulatory effect in case of P2 than in P1. Collectively, the findings demonstrated the regulation of CD44 by NFYA and highlighted the potential CRISPR/Cas9-mediated targeting of TF binding site as effective strategy for modulating CD44 expression and associated malignant phenotype.</p>","PeriodicalId":93898,"journal":{"name":"Biochimie","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148984413","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Antibacterial activity and mechanistic insights of Lucilia illustris antimicrobial peptide Cecropin A2 against Pseudomonas aeruginosa.","authors":"Liang Chen, Lei Xu, Mengyue Wang, Qilong Zhao, Chunlan Wei, Qiang Wang, Shangshang Ma, Weiping Cao","doi":"10.1016/j.biochi.2026.09.005","DOIUrl":"10.1016/j.biochi.2026.09.005","url":null,"abstract":"<p><p>Pseudomonas aeruginosa (P. aeruginosa) poses a serious public health threat due to multidrug resistance and biofilm formation. This study investigated the antibacterial mechanisms of the antimicrobial peptide, Cecropin A2 (CA2), against P. aeruginosa. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of CA2 against P. aeruginosa ATCC 9027 (PA ATCC 9027) were determined by broth microdilution. Antibacterial activity was evaluated using growth curves and time-kill assays. The mechanism was explored by assessing membrane integrity (outer/inner membrane permeability, SEM, and fluorescence microscopy), and intracellular responses (ATP, SDH activity, and ROS). Biofilm effects were assessed by crystal violet staining (biomass) and viable cell counting (biofilm-embedded bacteria). The MIC and MBC of CA2 against PA ATCC 9027 were 37.34 μM and 74.68 μM, respectively. CA2 exhibited moderate antibacterial activity against PA ATCC 9027. Scanning electron microscopy (SEM) revealed marked morphological damage after treatment. CA2 affected intracellular metabolism, potentially interacted with genomic DNA, and reduced biofilm biomass. Cecropin A2 exhibits concentration-dependent in vitro antibacterial activity against P. aeruginosa ATCC 9027, providing mechanistic insights and a theoretical basis for the development of alternative antimicrobial strategies.</p>","PeriodicalId":93898,"journal":{"name":"Biochimie","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-09-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148979050","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Advances in CRISPR-based genetic engineering of Bifidobacterium.","authors":"Xiaotong Lin, Yixin Zhang, Jiayi Cheng, Jialong Chen, Xin Liu","doi":"10.1016/j.biochi.2026.09.003","DOIUrl":"10.1016/j.biochi.2026.09.003","url":null,"abstract":"<p><p>Bifidobacterium plays a vital role in gut health, immunity, and metabolism, yet natural strains show limited native functional capabilities and genetic manipulability. Recent advances in genome editing, especially CRISPR-based systems, have provided novel approaches for engineering Bifidobacterium, though strain-specific variability and restriction-modification barriers remain substantial constraints. This review outlines the evolution from traditional homologous recombination to CRISPR/Cas technologies, highlighting applications of endogenous and exogenous systems, including CRISPRi/a and base editing. In preclinical and food biotechnology contexts, genetically engineered strains have shown potential as targeted delivery vehicles and functional probiotics. Despite these achievements, challenges such as strain variability, restriction-modification barriers, and biosafety concerns persist. Where genetic accessibility can be established, the integration of CRISPR technology with synthetic biology may enable more precise gene regulation and could support the development of next-generation engineered Bifidobacterium-based platforms.</p>","PeriodicalId":93898,"journal":{"name":"Biochimie","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148957380","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Selenium in aging and longevity.","authors":"William Gerard Pierce, John James Mackrill","doi":"10.1016/j.biochi.2026.09.002","DOIUrl":"10.1016/j.biochi.2026.09.002","url":null,"abstract":"<p><p>Selenium is an essential dietary micronutrient, exerting beneficial effects over a limited range of concentrations. Insufficient dietary intake of selenium increases the risk of type 2 diabetes, infertility, immune dysfunction, prostate cancer, cognitive decline, prostate cancer, colorectal cancer in females, skeletal muscle disorders, overall mortality and an endemic cardiomyopathy known as Keshan disease. Excessive intake of selenium is toxic, leading to a condition termed selenosis, that is associated with damage to the nails, dermatitis, alopecia, central nervous system dysfunction and death. Selenium exerts its biological effects through low-molecular weight molecules, through a small family of selenoproteins which contain the 21st amino acid selenocysteine, and via selenium-binding proteins, in which an atom of selenium is covalently bound to a cysteine residue. In addition to being regulated by body selenium status, the abundance and function of these biomolecules can be controlled by other dietary components (including copper, zinc, folic acid, resveratrol, vitamin A and α-linoleic acid) and other factors, such as exercise and caloric restriction. This review comprehensively surveys the effects of selenium-containing biomolecules on aging and aims to generate rationales underpinning the biphasic effects of this element on life-span, health-span and disease. This will provide a framework for understanding the effects of selenium on mechanisms of aging, as the starting point for development of interventions that will promote longevity and health-span.</p>","PeriodicalId":93898,"journal":{"name":"Biochimie","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-09-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148914139","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Erythrocyte trans fatty acid accumulation in elderly sarcopenic patients: biochemical evidence for oxidative stress-driven cis-trans isomerization of membrane lipids.","authors":"Amira Zarrouk, Imen Ghzaiel, Gérard Lizard, Mohammed Hammami, Sonia Hammami","doi":"10.1016/j.biochi.2026.09.001","DOIUrl":"10.1016/j.biochi.2026.09.001","url":null,"abstract":"<p><p>Free radical-mediated cis-trans isomerization of polyunsaturated fatty acids (FA) is well established in food chemistry, yet its occurrence as an endogenous biochemical process in oxidatively stressed human tissues remains poorly characterized. Sarcopenia, a geriatric syndrome defined by progressive loss of skeletal muscle mass and function, is associated with chronic oxidative stress. We hypothesized that this oxidative milieu drives in vivo cis-trans isomerization of erythrocyte membrane FA, generating trans isomers distinguishable from dietary exposure. A cross-sectional study was conducted in 160 subjects aged ≥65 years, classified as sarcopenic (n = 97) or non-sarcopenic (n = 63) by SARC-F questionnaire and handgrip strength. Erythrocyte FA profiles were determined by gas chromatography; plasma oxysterols were quantified by GC-MS. Specific trans isomers were markedly elevated in sarcopenic patients, especially C18:1 trans-11 (3.09-fold; p < 0.001) and the linoleic acid isomer C18:2 trans-9, cis-12 (2.66-fold; p < 0.001). Their cis counterparts were not significantly elevated. By contrast, two trans species that retain the cis-9 configuration, the ruminant dietary marker C18:2 cis-9, trans-11 (CLA) and the geometric isomer C18:2 cis-9, trans-12, were both paradoxically reduced (p = 0.007 and p < 0.001, respectively). Total trans FA correlated with non-enzymatic oxysterols 7-ketocholesterol (ρ = 0.49, p < 0.001),7α-hydroxycholesterol (ρ = 0.43, p = 0.003), and 7β-hydroxycholesterol (ρ = 0.49, p < 0.001), but not with enzymatically formed 24S-hydroxycholesterol. On multivariate analysis, total trans fatty acids remained independently associated with sarcopenia (OR 1.28; p = 0.018). These findings provide the first biochemical evidence that chronic oxidative stress in sarcopenic elderly patients drives endogenous cis-trans isomerization of membrane FA, challenging the notion that elevated trans FA represent a purely dietary phenomenon.</p>","PeriodicalId":93898,"journal":{"name":"Biochimie","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-09-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148914180","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cardiometabolic disease risk: the joint role of the UCP1, UCP2, UCP3, and FTO gene variants.","authors":"Anna E Pravednikova, Antonina Nikitich, Anna Andrzejczak, Anna Tomkiewicz, Lidia Karabon, Damian Skrypnik, Pawel Bogdanski, Eugene S Grebenshchikov, Boris Shulgin, Yulii V Shidlovskii","doi":"10.1016/j.biochi.2026.08.020","DOIUrl":"10.1016/j.biochi.2026.08.020","url":null,"abstract":"<p><p>Cardiometabolic diseases (CMDs) impose a significant global health burden. Although CMDs rarely occur in isolation, most genetic studies have examined individual conditions separately, overlooking shared pathogenic pathways that link them into a continuous cardiometabolic continuum. Moreover, population-specific data for Eastern and Central Europe are scarce, and combined effects of variants in key metabolism-related genes, such as UCP1-UCP3 and FTO, have not been studied within cardiometabolic continuum. In this study, we investigated individual and combinatorial effects of UCP1-UCP3, and FTO variants on risk of CMDs as a continuum in Central Russian and Polish populations. We identified novel risk-increasing combinations: minor alleles of UCP2 c.-866G > A and UCP3 c.-55C > T; and minor alleles of UCP3 c.-55C > T and FTO c.46_23525T > A with wild alleles of four UCP1 and two UCP2 variants. Protective effects were observed for two novel combinations: minor allele of FTO c.46_23525T > A with wild alleles of all studied UCPs variants; and minor allele of UCP2 c.-866G > A with wild allele of UCP2 p.Ala55Val. The FTO variant showed context-dependent effects, shifting from risk to protection depending on genetic background. Increased age, BMI, WtHR, and BF% were also significant predictors, with sex-specific differences. Predictive quality of models integrating genetic and non-genetic factors (AUC up to 0.97) further underscore the potential value of this approach. In silico analysis suggested these variants exhibit regulatory functions across various tissues, which potentially involved in CMD pathogenesis. This is the first study to demonstrate combinatorial UCP1-UCP3 and FTO effects on risk of CMDs as a continuum in Central Russian and Polish populations.</p>","PeriodicalId":93898,"journal":{"name":"Biochimie","volume":" ","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-09-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148889745","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}