| Objective: By studying the differences in the expression levels of urinary exosome mi RNAs between patients with Kawasaki disease and healthy children,to select mi RNAs that specifically exist in the acute onset of Kawasaki disease,and then to evaluate the value of urinary exosome mi RNAs as potential diagnostic markers of Kawasaki disease,providing a rapid and sensitive diagnostic method for the early diagnosis of Kawasaki disease.Methods: According to the guidelines for the diagnosis and treatment of KD established by the American Heart Association(AHA)in 2017(1),30 KD patients in the acute phase of Kawasaki disease before intravenous gamma globulin(IVIG)treatment who were diagnosed at Shenzhen Children’s Hospital between June 2020 and December 2020 were included as the experimental group,and 30 healthy volunteers were recruited as the control group.Exosomes in the mixed urine of five groups of children with Kawasaki disease(n =6/group)and five groups of control mixed urine(n = 6/group)were extracted by ultra-high speed centrifugation respectively,and the extracted exosomes were identified by electron microscopy and the quality of labeled proteins;qualified exosome RNA was extracted by Trizol method;the expression profiles of mi RNAs in the exosomes of the two groups were detected by high-throughput sequencing analysis,so as to select the differentially expressed mi RNAs(differentially expressed mi RNAs)with the greatest differential expression;and DEPs were subjected to gene function annotation analysis(GO),Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis,and finally the selected mi RNAs were validated by qt-PCR.Results:(1)We selected a total of 28 differential mi RNAs in the RNAs detected in the two groups according to the Fold change > = 2 and P value < 0.05 criteria,of which 6were significantly up-regulated and 10 were significantly down-regulated.This provides a basis for the subsequent search for novel biomarkers of KD.(2)GO pathway enrichment analysis showed that differential mi RNAs were mainly involved in cell proliferation,biological regulation,metabolic processes,response to stimuli,and amino acid transport processes,and the molecular functions and cellular functions involved mainly included binding,catalytic activity,transporter activity,molecular transducer activity,and synthesis of cells,cell membranes,organelle membranes,and synaptic membranes.(3)KEGG pathway analysis indicated differential mi RNAs with cell growth and death,transport and catabolism,intracellular signaling,cardiovascular system diseases,tumors,infectious diseases,and immune diseases.(4)we could find out significantly different target mi RNAs by high-throughput sequencing analysis,and validation showed that 10 mi RNAs could be used as candidates for KD biomarkers,including mi R-26-5p,mi R-508-5p,mi R-508-3p,mi R-1271-5p,mi R-155-5p,mi R-143-3p,mi R-139-5p,mi R-217-5p,mi R-125a-3p,and mi R-193b-5p.Conclusion: By comparing the urine of KD and healthy children,10 target mi RNAs with significant differences were selected,including significantly up-regulated mi R-26-5p,mi R-508-5p,mi R-508-3p,mi R-1271-5p,mi R-155-5p,mi R-143-3p,mi R-139-5p and significantly down-regulated mi R-217-5p,mi R-125a-3p,mi R-193b-5p as differential genes,and analysis of these 10 differential mi RNAs revealed that KD was closely related to the body’s inflammatory response and damage to arterial endothelial cells at the molecular level,providing new ideas and directions for subsequent diagnosis and treatment of KD. |
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