Establishment of Rat Karoshi Model and Model-based Proteomic and Metabolomic Analyses of Energy Metabolism and Oxidative Stress
Objective Karoshi,death from overwork,is a serious problem with unclear identification standards and mechanisms.This study aims to establish a karoshi rat model by integrating weight-bearing swimming and sleep deprivation.This model will enable us to investigate the adverse effects of acute physical and mental fatigue on cardiac functions and explore the response mechanisms to overwork using integrated omics approaches,specifically metabonomics and proteomics.Methods The experimental design involved healthy male sprague-dawley(SD)rats subjected to weight-bearing swimming and sleep deprivation for 7 d.The rats were monitored for changes in physiological function indexes,including electrocardiogram and respiration.Protein digestion,iTRAQ labeling,and quantitative data analyses were performed to determine differentially expressed proteins(DEPs).Additionally,GC-MS analysis was conducted to identify differential metabolites.The integration analysis of differential metabolites and proteins shared by the fatigue group and the overwork group was performed to construct a relevant metabolic pathway network and integrate the proteomics and metabolomics data.Statistical analysis was carried out using one-way ANOVA and Duncan's multiple range t-tests.Results The rats subjected to weight-bearing swimming and sleep deprivation showed various physical and behavioral changes associated with fatigue,including hair disorder,decreased muscle tension,reduced food intake,and weight loss.Analysis of cardiac functions revealed cardiac hypertrophy and heart failure in the fatigue and karoshi groups,as evidenced by changes in heart color,myocardial fiber structure,heart rate,respiratory rate,and cardiac ultrasound measurements.Metabolomics analysis using GC-MS identified several differential metabolites in response to overwork,including amino acids involved in various metabolic pathways.Proteomic analysis using iTRAQ technology identified DEPs in the fatigue and karoshi groups,with a subset of DEPs shared by both groups.The GO analysis revealed that the up-regulated DEPs were primarily associated with mitochondria and peroxisomes in the cellular component category.The Reactome analysis further highlighted the enrichment of DEPs in the transfer of ferriheme from methemoglobin to hemopexin pathway.Integration analysis of the DEPs and differential metabolites revealed the activation of autophagy,increased mitochondrial oxidative phosphorylation,enhanced branched-chain amino acid degradation,and altered peroxisomal β-oxidation.These findings suggested complex metabolic adaptations to meet the increased energy demands during overwork while also dealing with oxidative stress.Furthermore,the reprogramming of energy metabolism was observed,with upregulation of fatty acid β-oxidation enzymes and glycolysis-related enzymes in the fatigue group,indicating a shift towards glucose metabolism.In contrast,the karoshi group showed a decreased dependence on fatty acids as an energy source and increased utilization of glucose.The model proposed in this study highlights the interconnected metabolic changes involving mitochondria,peroxisomes,and lysosomes in response to overwork.The findings contribute to our understanding of the mechanisms involved in overwork-related pathologies and provide a basis for further research in the field of karoshi.Conclusion Overall,metabolic reprogramming might provide sufficient energy to the heart,alleviate oxidative stress and damage to cardiac cells in response to excessive exertion and fatigue.Our findings provide an insight into response mechanism to overwork death and lay a foundation for further research on overwork death.
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维普
