Identification of novel potential molecular targets associated with pediatric septic shock by integrated bioinformatics analysis and validation of in vitro septic shock model

Identifies hub genes associated with pediatric septic shock

Abstract

Background/Aim: Sepsis is a major cause of morbidity, mortality, and healthcare utilization among children all over the world. Sepsis, characterized as life-threatening organ failure, results from a dysregulated host response to infection. When combined with critically low blood pressure, it causes septic shock, resulting in high mortality  rates. The aim of this study was to perform a bioinformatic analysis of gene expression profiles to predict  septic shock risk.

Methods: Four datasets related to pediatric septic shock were retrieved from the Gene Expression Omnibus (GEO) database for a total of 240 patients and 83 controls. GEO2R tools based on R were used to find differentially expressed genes (DEGs). The Database for Annotation, Visualization and Integrated Discovery (DAVID) was used to examine the functional enrichment of DEGs. STRING was used to create a protein–protein interaction (PPI) network. After separately analyzing the four datasets, commonly affected genes were removed using the Venny program. Finally, human umbilical vein endothelial cells (HUVECs) were stimulated with supernatants of lipopolysaccharide (LPS)-stimulated RAW267.4 macrophage cells and expression of selected genes was confirmed by real-time reverse-transcriptase polymerase chain reaction (qRT-PCR) and used to construct an in vitro septic shock model.

Results: Seven-hundred seventy-one common differentially expressed genes in the four  groups were found. Of these, 433 genes showed increased expression, while 338 had reduced expression. In the DAVID analysis results, DEGs up-regulated according to gene ontology results were enriched in the regulation of innate and adaptive immune responses, complement receptor-mediated signaling, and cytokine secretion processes. Down-regulated DEGs were  significantly enriched in the regulation of immune response, T-cell activation, antigen processing, and presentation and integral component of plasma membrane processes. According to The Search Tool for the Retrieval of Interacting Genes/Proteins (STRING), Cystoscape Molecular Complex Detection (MCODE), nine down-regulated genes in the center of the PPI network, ZAP70, ITK, LAT, PRKCQ, LCK, IL2RB, FYN, CD8A, CD247 and four up-regulated genes, MMP9, TIMP1, LCN2, HGF, were associated with septic shock. Expressions of FYN and MMP9 genes in the in vitro septic shock model were consistent with the bioinformatic results.

Conclusion: Comparative bioinformatics analysis of data from four different septic shock studies was performed. As a result, molecular processes and important signal networks and 13 genes that we think will play a role in the development and risk prediction of septic shock are proposed.

Methods: Four datasets related to Pediatric septic shock were retrieved from the Gene Expression Omnibus (GEO) database for a total of 240 patients and 83 controls. GEO2R tools based on R were used to find differentially expressed genes (DEGs). DAVID was used to examine the functional enrichment of DEGs. STRING was used to create a protein-protein interaction (PPI) network. After separately analyzing the four datasets, commonly affected genes were removed using the Venny program. Finally, HUVECs were stimulated with supernatants of LPS-stimulated RAW267.4 macrophage cells and expression of selected genes was confirmed by qRT-PCR, constructing an in vitro septic shock model.

Results: There were 771 common differentially expressed genes in the 4 groups. Of these, 433 genes showed increased expression, while 338 had reducing expression. In the DAVID analysis results, DEGs upregulated by gene ontology were enriched in the regulation of innate and adaptive immune responses, complement receptor-mediated signaling, and cytokine secretion processes. Downregulated DEGs are significantly enriched in the regulation of immune response, T cell activation, antigen processing, and presentation and integral component of plasma membrane processes. According to STRING, cystoscape MCODE, and cytohubba analysis, 9 downregulated genes in the center of the PPI network, ZAP70, ITK, LAT, PRKCQ, LCK, IL2RB, FYN, CD8A, CD247, and 4 upregulated genes, MMP9, TIMP1, LCN2, HGF, were associated with septic shock. Expressions of FYN and MMP9 genes in the in vitro septic shock model were consistent with bioinformatic results.

Conclusion: Important signaling networks and 13 genes potentially indicating molecular processes for the incidence, development, and risk prediction in septic shock were found using bioinformatic analysis of gene expression profiles.