Exploratory Study On Regulatory Network Of CircRNA-miRNA-mRNA In Type 2 Diabetes Mellitus

Objective:The incidence rate of type 2 diabetes(T2DM)is high,but its pathogenesis is not clear.This study used high-throughput sequencing technology and bioinformatics analysis to explore the role of differentially expressed micro RNA(mi RNA),circular RNA(circ RNA),m RNA and non coding RNA(noncoding RNA)co expression regulatory network in T2 DM,and provided new ideas for exploring the occurrence,diagnosis and treatment of T2 DM at the transcriptome level.Method:Three newly diagnosed T2 DM patients and three healthy people in the Second Affiliated Hospital of Jilin University from August 2017 to June 2018 were selected as the case group and the control group;six subjects were male,aged from 40 to 60 years old.The peripheral blood was collected,and the whole blood RNA was extracted and sequenced.PE150 sequencing strategy was used to sequence circ RNA and m RNA,SE50 sequencing strategy was used to sequence mi RNA.The differential expression of circ RNA,mi RNA and m RNA was analyzed by DEseq2 based on negative binomial distribution.The corrected P<0.05 was selected as the RNA screening criteria for differential expression.The ggplot2 package in R was used to draw volcano map and hierarchical clustering heat map.According to the relationship among circ RNA,mi RNA and m RNA,competitive endogenous RNA coexpression regulatory network was constructed and presented by Cytoscape software.Gene ontology(GO)and kyoto encyclopedia of genes and genomes(KEGG)function enrichment analysis were performed by KOBAS3.0 website,and signal pathway bubble diagram was drawn by ggplot2.Protein protein interaction(PPI)of m RNA in regulatory network was analyzed by using string online database.Result:1.According to the adjusted P<0.05,65 differentially expressed circ RNAs(18up-regulated and 47 down regulated),202 differentially expressed mi RNAs(107up-regulated and 95 down regulated),and 199 differentially expressed m RNAs(71up-regulated and 128 down regulated)were screened.2.GO and KEGG enrichment analysis of differentially expressed circrna.There were 1947 items and 1422 biological process(BP)items in GO enrichment,there were 233 cell component(CC)items,and 292 molecular function(MF)items.KEGG enrichment analysis showed that the main enrichment pathways were ubiquitin mediated proteolysis,m RNA monitoring pathway,hematopoietic cell lineage,endocytosis,long-term depression,proteins in cancer,progesterone mediated oocyte maturation,HIF-1 signaling pathway and African trypanosomiasis.3.According to the relationship between the differentially expressed RNAs,construct the regulatory network of circrna mi RNA m RNA coexpression.According to the degree >10,betweenness centrality > 0.1 and closeness centrality > 0.2,nine key RNAs were screened out: five mi RNAs including mi R-1343-3p,mi R-6881-3p,mi R-1301-3p,mi R-130b-5p and mi R-15b-5p were the key nodes in the regulatory network;four circrnas including circ₀029408?circ₀009015?circ₀000230 and circ₀002722.4.In the functional enrichment analysis of non coding RNA coexpression regulatory network,there were 3563 items in go enrichment analysis,including 2656 items of BP,379 items of CC and 527 items of MF.KEGG enrichment analysis showed that the main enrichment pathways were metabolic pathway,m TOR signaling pathway,biosynthesis of other types of o-sugars,other glycine,AMPK signaling pathway,FC?RI signaling pathway,FC?R-mediated phagocytosis,MAPK signaling pathway and Notch signaling pathway.5.The protein interaction analysis of m RNA in circrna mi RNA m RNA co expression network showed that GAPDH,UBE2 s,PTK2B and MAP2K3 were the key proteins in PPI network.Conclusion Conclusion:1.Circ RNA in differential expression analysis₀029408,circ₀009015,circ₀000230,circ₀002722,mi R-1343-3p,mi R-6881-3p,mi R-1301-3p,mi R-130b-5p and mi R-15b-5p were the main regulators of type 2 diabetes.2.Notch signaling pathway,m TOR signaling pathway,MAPK signaling pathway,FC?R-mediated phagocytosis and HIF-1 signaling pathway are the main signaling pathways related to T2 DM.3.GAPDH,UBE2S?PTK2B?MAP2K3 may be closely related to insulin secretion.