Regulation Of Macrophage Polarization By IL-1R2 In Patients With Sepsis

Objective : To screen potential core targets by using sepsis peripheral blood sequencing and bioinformatics technology,and explore the potential function of core gene IL1-R2 in sepsis macrophages.Methods:From January 2019 to December 2020,15 patients admitted to the ICU of Southwest Medical University’s Affiliated Hospital were collected using the SEPSIS 3.0 standard.Normal volunteers were taken as the control group(n=10).RNA-seq treatment of peripheral blood was conducted within24 hours of admission;IDEP online analysis software,written in R,was utilized to ensure data quality control and differential gene screening.(P<0.01;log FC ≥ 2);The differential genes were analyzed for gene function enrichment,and then the ligand-receptor interaction related genes were selected for protein interaction analysis using STRING to screen potential core targets;Collect public data sets and use meta strategy to verify the expression trend of core genes in sepsis group;Use the data from the public data set GSE65682 to explore the prognostic relevance of potential core genes;Then,five PBMC samples(NC=2;SIRS=1;SEPSIS=2)were collected for cell line location analysis of core genes using 10 X single cell sequencing;Furthermore,LPS(100ng/ml)was used to stimulate THP1 cells to build a sepsis model,and RT-q PCR was used to verify the expression of core genes.A cell sepsis model was constructed by knocking out IL1-R2 by si RNA.Flow cytometry was employed to explore the influence of IL1-R2 on THP1 cell polarization,while RT-q PCR was utilized to observe the expression of IL-1B and TNF.Finally,sequencing and bioinformatics analysis of cells in both the knockout and control groups were conducted.we explored the effect of IL1-R2 knockdown on downstream genes.Results: In comparison to the normal group,913 DEGs were present in the peripheral blood cells of those suffering from sepsis;Of which 673 were increased;240 decreased.These up-regulated differential genes are mainly concentrated in: neutrophil activation participates in immune response,bone marrow leukocyte activation,leukocyte degranulation,etc;The down-regulated genes are enriched in cell component morphogenesis,neuron projection morphogenesis,axonogenesis,etc In this study,the PPI interaction network was analyzed,and multiple genes such as IL1-R2,HGF,MMP9,IL10,TGFA,LCN2,EPHA2,SDC1,ERBB3,MIC1 were selected to be located in the core region,and such genes are related to cell communication and signal transduction and are located in the extracellular structure.Through the survival curve,it was found that the genes in the core region of IL1-R2 were negatively correlated with the survival rate of patients with sepsis.Multiple public data sets show that the target gene IL1-R2 is highly expressed in the peripheral blood of patients with sepsis.Single cell sequencing analysis showed that they were mainly expressed in monocytes.The high expression of IL-1R2 gene in sepsis was verified by the detection of THP1 by PCR in vitro.Finally,through si RNA knockout,it was found that the expression of TNF increased in knockdown group,but there was no difference in IL-1B;M1 polarization increased.After knocking down IL1-R2,3718 downstream genes changed,including 2038 up-regulated and 1680 down-regulated.IL-1R2 is mainly involved in cytokine-mediated signal pathway,positive regulation of cell adhesion,positive regulation of cytokine products and other functions.Conclusion: Compared with the normal group,IL1-R2 gene is mainly located in human peripheral blood mononuclear cell line;The expression was increased in sepsis group and correlated with the prognosis of patients;IL-1R2 is involved in the regulation of macrophage polarization,and may be a potential research and treatment target.