Quantitative Proteomics And Bioinformatics Analysis Of Human Coronary Artery Endothelial Cell Injury Induced By Kawasaki Disease

Objective:Coronary artery injury is one of the most important complications of Kawasaki disease,which seriously endangers the cardiovascular health of children and adolescents.In recent years,more and more studies have confirmed that in Kawasaki disease,coronary endothelial cell damage such as coronary endothelial cell apoptosis,proliferation disturbance,decreased migration ability,decreased angiogenesis ability and so on,will increase the risk of secondary coronary artery damage in Kawasaki disease.Therefore,this study intends to establish a model of serum-induced coronary artery endothelial cell injury in children with Kawasaki disease,and screen the protein markers of coronary artery endothelial cell injury in(KD)of Kawasaki disease by isobaric tags for relative and absolute quantitation(i TRAQ)proteomics,so as to provide theoretical basis and experimental scheme for further exploring the molecular regulation mechanism of coronary artery endothelial cell injury in Kawasaki disease.Methods:(1)A total of 20 children with KD(11 males and 9 females)in Shenzhen Children's Hospital and 20 healthy children(10 males and 10 females)in the outpatient clinic of the Department of Child Health were included in the study.(1)A total of 20 children with KD(11 males and 9 females)and 20 healthy children(10males and 10 females)were included in the study.The diagnostic criteria of KD refer to the 2017 Kawasaki Disease guidelines of AHA in the United States.The experiment was divided into two groups: the(HCAECs)of human coronary artery endothelial cells incubated with serum of children was KD group,and the HCAECs incubated with serum of healthy children was group N.(2)The HCAECs incubated in KD group and N group was cleaved,the total protein was extracted,and the protein samples were qualitatively annotated and quantitatively detected by i TRAQ protein.Then the biological function of the protein data detected by i TRAQ was annotated by bioinformatics scheme,gene ontology(GO)analysis,KEGG analysis,PPI network analysis,and finally the expression of protein markers was verified by Western blot method.Results:(1)The main results were as follows:(1)among the children with KD,there were 11 males and 9 females,with an average age of 1.8±0.6.in the healthy control group,there were 10 males and 10 females,with an average age of 2.0±0.5.there was no significant difference in gender and average age between the two groups(P >0.05).(2)518 differentially expressed proteins were identified by proteomics(P <0.05).GO analysis showed that the differential proteins were significantly enriched in cell process,cell localization,immune system process,cell proliferation,cell aggregation,biological adhesion,metabolic process,regeneration,biological regulation and other biological processes.It was significantly enriched in cellular components such as cells,organelles and cellular parts,and in molecular functions such as binding,catalytic activity and molecular function regulators(P < 0.05 and FDR < 0.01).The KEGG results showed that the differential proteins were significantly enriched in ribosome,PI3K-Akt signal pathway,endoplasmic reticulum protein processing,transcriptional disorders in cancer,calcium signal pathway,NF-k B signal pathway,B cell receptor signal pathway and so on(P < 0.05 and FDR < 0.01).According to the relationship density of PPI network,the protein markers of PWP2,MCM4,MCM7,SLD5,HDAC2,MCM6,MCM5,MCM3 and MCM2 were selected as protein markers of coronary artery endothelial cell injury in KD.(3)Western blot verified the expression level of protein markers HDAC2,PWP2 and MCM2.The results showed that the expression of three protein markers decreased in KD coronary artery endothelial cells,which was consistent with the expression pattern of proteomics.Conclusion:This study suggests that the serum of KD can affect the protein expression pattern of HCAECs and mobilize the signal pathways related to cardiovascular system.PWP2,MCM4,MCM2,SLD5,HDAC2,MCM6,MCM7,MCM5 and MCM3 can be used as candidate protein markers for coronary artery endothelial cell injury in KD,which provides molecular targets for the pathogenesis and treatment of coronary artery injury in Kawasaki disease,and provides a new theoretical basis for accurate diagnosis and treatment of KD.