Metabolomic Characterization And Post-mortem Diagnostic Biomarkers In Rats With Sudden Death From Anaphylaxis

Objective:To explore the changes of endogenous small molecule metabolites in the plasma of rats with sudden death from anaphylaxis,to identify the specific post-mortem diagnostic biomarkers of sudden death from anaphylaxis and differential diagnostic biomarkers from sudden coronary death,to explore the metabolic pathways and the mechanisms related to sudden death from anaphylaxis,and to provide accurate and reliable diagnostic indicators and basis for the forensic identification of sudden death from anaphylaxis.Methods:Forty-eight male SD rats were randomly divided into four groups,and four rat models were prepared:(1)sudden death from anaphylaxis rats were prepared by ovalbumin(OVA)injection;(2)coronary atherosclerosis rats were prepared by high-fat emulsion gavage combined with vitamin D3 intraperitoneal injection,and then sudden coronary death was induced by surgical ligation of the anterior descending branch of the left coronary artery;(3)coronary atherosclerosis rats were injected by OVA and died of anaphylaxis;(4)control rats were not treated.Immediately after death,2 m L of plasma was collected from each group of rats,and the metabolites in the plasma were detected by UHPLC-Q Exactive Orbitrap-MS.Compound Discoverer 2.0 software was used to pre-process the raw data,and the obtained data matrix was normalized and imported into SIMCA-P 13.0 software for multivariate statistical analysis to identify differential metabolites in the plasma of each group,and to search for specific biomarkers of sudden death from anaphylaxis.The pathway enrichment analysis was performed by Metabo Analyst 3.0 to investigate the metabolic pathways and mechanisms of sudden death from anaphylaxis.Finally,the diagnostic efficacy of the identified differential metabolites for sudden death from anaphylaxis was assessed by ROC curve analysis.Results:1.Thirty-seven specific metabolites of sudden death from anaphylaxis,including betaine,hippuric acid,3-indolyl sulfate,and arachidonic acid were identified by metabolomics techniques.In sudden death from anaphylaxis,D-glutamine and D-glutamate metabolism,alanine,aspartate and glutamate metabolism and arginine biosynthesis played key roles.2.Nineteen specific metabolites of sudden coronary death,including phytosphingosine,docosahexaenoic acid,linoleic acid and L-tyrosine were identified,and linoleic acid metabolism and biosynthesis of phenylalanine,tyrosine and tryptophan played important roles in sudden coronary death.3.In the plasma of rats in the coronary atherosclerosis with anaphylaxis,a total of 59 specific metabolites were identified,of which 22 belonged to the specific metabolites of sudden death from anaphylaxis and 14 belonged to the specific metabolites of sudden coronary death,suggesting that two pathological processes occurred simultaneously in this group of rats.There may be an interaction and modulation between anaphylaxis and coronary artery lesions,and the mechanism of their interaction needs to be further explored.4.The AUC value was 0.997 for the combined diagnosis of sudden death from anaphylaxis using five metabolites: imidazoleacetic acid,D-synuclein,taurine goodeoxycholic acid,taurine and dodecanoic acid,and 0.94 for the combined diagnosis of sudden coronary death using three metabolites: triethanolamine,glycyrrhetinic acid and phytosphingosine,indicating the high diagnostic accuracy of the screened metabolite combinations.Conclusion:This study explored the plasma metabolite profiles of rats with sudden death from anaphylaxis,sudden coronary death and coronary atherosclerosis with anaphylaxis from the perspective of metabolomics,and identified 37 specific metabolites,including betaine,hippuric acid,3-indolyl sulfate and arachidonic acid,which are expected to be used for postmortem diagnosis of sudden death from anaphylaxis.We also explored a series of metabolic changes that occur in vivo during sudden death from anaphylaxis,including D-glutamine and D-glutamate metabolism,alanine,aspartate and glutamate metabolism and arginine biosynthesis,which can provide a scientific basis for future forensic identification of sudden death from anaphylaxis and related mechanism studies.