Study On The Evaluating Severity And Injury Mechanism Of Acute Paraquat Poisoning Rats Based On GC-MS Metabolomics

Objective Screening of plasma metabolite biomarkers for the evaluation of the severity of acute paraquat(PQ)poisoning in rats based on GC-MS metabolomics technology,providing a basis for the early diagnosis and disease severity of PQ poisoning,The mechanism of acute paraquat poisoning could be further discussed.Methods 24 SD rats were divided into the normal saline control group,the low-dose intoxicati6 on group(40mg/Kg),the medium-dose intoxication group(80mg/Kg)and the high-dose intoxication group(120mg/Kg)according to the random number table method.Every group has 6SD rats.Observed the general conditions of rats in 24 hours after gastrointestinal perfusion,then collected abdominal aorta blood of rats in24 hours.GC-MS was used to detect endogenous small molecule metabolites in plasma,and metabolic profiles and random forest analysis were performed to screen potential biomarkers.Liver and kidney function of rats were detected.The lung injury degree of paraquat poisoning rats was observed by HE staining.Results(1)The rats in each group were observed,soon after intragastric administration,in 40 mg/Kg PQ poisoning group,infected rats showed poisoning symptoms such as less movement,less food,etc;In 80 mg/Kg PQ poisoning group,the infected symptoms were worsened,and showed shortness of breath,were unresponsive after stimulation;As to the 120mg/Kg PQ poisoning group,infected rats were slow in reacting,keep curling up under stimulation,no drinking and eating activities basically,also appear labored breathing,foam-like liquid overflow from nose and other symptoms,even worse,one died;Normal saline control group rats were diet normally,vital signs were stable,show no discomfort symptoms.(2)S-N-K(S)method was used for pairwise comparison in each group,At the level of alpha =0.05,the contents of ALT,BUN and CR in plasma of paraquat intoxicated rats in control ?40 mg/Kg ?80 mg/Kg and 120mg/Kg dose groups were significantly different,and there was a dose-dependent relationship.kruskal-wallis H test,a non-parametric statistical method,was adopted for this group of data,at the level of=0.05,AST elevation was considered to be different in each group.(3)The lung injury degree of paraquat poisoning rats were observed by Hematoxylin-eosin(HE)staining.Result showed: In the blank control group,the alveolar structure was clear and complete,no exudation of alveolar cavity and lung interstitium,and no inflammatory cell infiltration;In 40 mg/Kg PQ poisoning group,the alveolar structure obviously changed.partial alveolar collapse was observed,alveolar cavity and interstitium exuded a little.In the 80mg/Kg and 120mg/Kg group,the normal structure of alveoli basically disappeared,alveolar occlusion collapsed,and inflammatory cells in the interstitial lung infiltrated.(4)In combination with standard substance and NIST05 mass spectrometry library,the endogenous metabolites in rat paraquat poisoning serum were qualitatively determined.A total of 30 metabolites were identified,including 14 metabolites with statistically significant difference from the control group,including amino acids,organic acids,sugars and fatty acids.Combining random forest algorithm to analyze the contribution of differential metabolites to metabolic profile changes in poisoned rats,it was found that d-Erythrotetrofuranose,propionic acid,L-alanine,D-fructose,glutamine,and phenylacetic acid caused paraquat key differential metabolites in the changes in serum metabolic profile of poisoned rats.The metabolic pathways of metabolites that play an important role in the classification of the four groups of rats are mainly alanine,aspartic acid and glutamic acid metabolism.(5)Using canonical correlation analysis to analyze metabolite characteristics and clinical features,correlation coefficient R is 0.92,metabolites L-valine,L-phenylalanine,tyrosine,inositol and biochemistry Indicator BUN contributed the most to the correlation of the two groups of features.Conclusion(1)Compared with the control group,the toxic group showed obvious lung injury when various symptoms appeared,and the glucose metabolism,amino acid metabolism,lipid metabolism and energy metabolism were changed.(2)Compared with the control group,the content of L-alanine,L-valine,glutamine and L-tyrosine decreased in the poisoning group,the content of benzeneacetic acid,L-serine,9'12-Octadecadienoic acid and trans-9-Octadecenoic acid increased,and they were related to the toxic dose.(3)The metabolic pathways of metabolites that play an important role in the classification of 4 groups of rats are mainly alanine,aspartic acid and glutamic acid metabolic pathways.By affecting ATP metabolism and nitrogen balance,it can cause organ injury.(4)Metabolite and liver and kidney function indicators correlation coefficient R is 0.92,metabolites L-valine,L-phenylalanine,tyrosine,inositol and indicator BUN contributed the most to the correlation of the two groups of features.