Competing Endogenous RNA Expression Profile Research In BV2 Microglial Cells Exposed To Lipopolysaccharide

Sepsis-associated encephalopathy is one of the most common complications of sepsis,which seriously increases the cost of medical care and impairs the quality of life of patients.Microglia activation plays a key role in the neuroinflammatory progression of sepsis-associated encephalopathy.A better understanding of new molecules that regulate microglia activation and their potential functions will greatly facilitate the exploration of new therapies.Recent studies have shown that lncRNA is related to sepsis-associated encephalopathy,but whether another popular non-coding RNA,circRNA,is related to sepsis-associated encephalopathy remains unclear.Their potential function in microglia activation and the mechanism is also unclear.At the same time,studies point out that lncRNA,circRNA,and mRNA can affect the occurrence and development of diseases through the ceRNA mechanism.Therefore,we expand the study of non-coding RNA expression profiles of sepsis-associated encephalopathy in order to have a deeper understanding of the pathophysiological mechanism of sepsis-associated encephalopathy.Objective:To dentify the differentially expressed lncRNA,circRNA,and mRNA that lipopolysaccharide induces early activation of BV2 microglia,and understand its diagnostic and therapeutic value as well as potential functions and mechanisms of ceRNA in sepsis-associated encephalopathy.Methods:Culturing BV2 microglia in vitro,and using LPS to stimulate BV2 microglia to establish an in vitro model of sepsis-associated encephalopathy.The cultured microglia were divided into two groups,CN group and LPS group.The cells in the CN group were cultured in vitro as the control group,and the cells in the LPS group were stimulated with LPS 1 μg/ml as the experimental group.The TRIzol method was used to extract total RNA from each group cells.The extracted total RNA samples of 3 in each group were sent to relevant companies for ceRNA chip detection and bioinformatics analysis.Including:(1)Using a super-differential photometer to perform quality control on total RNA samples in accordance with Agilent Bioanalyzer quality inspection standards;(2)Using ceRNA chip scanning to identify differential expression of lncRNA,circRNA,and mRNA in the LPS group relative to the CN group(2)Compared with the CN group,use the DAVID bioinformatics database to perform GO enrichment analysis and KEGG pathway enrichment analysis on the differentially expressed mRNAs in the LPS group;(3)Construct a ceRNA network based on the correlation analysis between differentially expressed RNAs.Results:(1)The quality of all RNA samples meets the experimental requirements of the chip manufacturer,and subsequent experiments can be carried out;(2)Compared with the CN group,the LPS group differentially expressed lncRNA,circRNA,mRNA hierarchical clustering diagram,scatter diagram,volcano Figure hints: 2488 lncRNAs,263 circRNAs and 2765 mRNAs were differentially expressed,of which 1744 and 744 lncRNAs,140 and 123 circRNAs,1135 and 630 mRNAs were up-regulated and down-regulated,respectively.(3)Compared with the CN group,the bar graph of differentially expressed mRNA GO enrichment analysis in the LPS group indicated that the top 3 molecular function GO items were "protein binding","cytokine activation",and "RNA polymerase II activation" Subsequence specific DNA binding ";the top 3 cell components GO entries are " cell membrane "," plasma membrane outside ",and" cytoplasm ";the top 3 biological process GO entries are " inflammatory response " and" immune system " Process","immune response".The GO bubble chart indicates that "protein binding" is the GO item of molecular function with the most differential genes and the highest degree of significance;"cell membrane" is the GO item of the cell component with the most differential genes and the highest degree of significance;"Inflammation response" means that there are more differential genes,Go items for biological processes with a higher degree of significance.(4)Compared with the CN group,the differentially expressed mRNA KEGG pathway enrichment analysis of the LPS group indicated that the first three pathways with the highest significance were the "TNF signaling pathway","NOD-like receptor signaling pathway",and "IL-17 signal path".The KEGG bubble chart prompts: "TNF signaling pathway" is a pathway with more genes and a higher degree of significance.(5)In the top 100 ceRNA networks of lncRNA-mi RNA-mRNA,the lncRNAs with strong interaction ability are NONMMUT144861.1,NONMMUT043540.2,ENSMUST00000203956.1;mi RNAs with strong interaction ability include mmu-mi R-762,mmu-mi R-149-3p,mmu-mi R-709;mRNAs with strong interaction ability include Rab11fip1,Birc3,Abhd15.Among the top 100 ceRNA networks of circRNA-mi RNA-mRNA,circRNAs with strong interaction ability include MMU＿CIRCpedia＿27949,mmu＿circ＿0008161,MMU＿CIRCpedia＿509;mi RNAs with strong interaction ability include mmu-mi R-7069-5p,mmu-mi R-7081-5p,mmu-miR-7686-5p;mRNAs with strong interaction ability include Helz2,Dcbld2,and Fam129 a.Conclusion:(1)The mechanisms of lncRNA,circRNA,mRNA and ceRNA may be involved in the occurrence and progression of sepsis-associated encephalopathy.(2)A large number of new candidate lncRNAs,circRNAs and mRNAs involved in the early activation of microglia induced by lipopolysaccharide are revealed.(3)Research may provide relevant information for the development of promising biomarkers and therapeutic targets for sepsis-associated encephalopathy in the body in the future.