The Expression And Biological Significance Of Circulating LncRNA In Patients With Diabetic Kidney Disease

Objective By analyzing the differential expression of lncRNA in peripheral blood of patients with diabetic kidney disease（DKD）and normal controls,it will bring theoretical foundations for in-depth exploration of the pathogenesis and treatment targets of diabetic kidney disease.Methods Randomly collect 3 peripheral blood samples of DKD patients and normal controls（NC）from Taizhou people’s Hospital from June to July 2020 for second-generation high-throughput sequencing.Combining the sequencing results,first log CPM The（log₂Counts Per Million）column is sorted in descending order,and the gene ID with the larger log CPM value is selected first（to prevent amplification failure）,and then the pvalue is less than 0.05,and the absolute value of Fold Change（|FC|）≥1.5 to screen out differentially expressed lncRNAs.According to|FC|,and select the 8differentially expressed lncRNAs with the highest differential multiples,namely:ENSG00000212939,ENSG00000233968,ENSG0000025717,ENSG00000228203,ENSG00000223534,ENSG00000265218,ENSG00000236438,ENSG00000216863.From June to November 2020,peripheral blood samples of 20 DKD patients,20diabetes mellitus（DM）patients,and normal controls were randomly selected,using Quantitative Real-time Polymerase Chain Reaction（q RT-PCR）verifies the above 8lncRNAs.Go analysis（Gene Ontology analysis,gene ontology analysis）,KEGG（Kyoto Encyclopedia of Genes and Genomes）pathway analysis and analysis of target mRNA genes with differential expression of lncRNA screened by the second-generation high-throughput sequencing through bioinformatics software analysis.The interaction analysis between lncRNA and target mRNA gene further clarifies the function of differentially expressed lncRNA in organisms by regulating mRNA.Results 1.In the peripheral blood samples of DKD patients,with|FC|≥1.5,P<0.05as the standard,there were 77 differentially expressed lncRNAs,of which 24 were up-regulated lncRNAs and 53 were down-regulated lncRNAs;there were 305 mRNAs are differentially expressed,210 of which up-regulate mRNAs and 95 down-regulate mRNAs.2.The results of GO enrichment analysis show that from the biological process level,the differentially expressed lncRNAs are mainly related to the response to nutritional level,celluar carbohydrate biosynthetic process,negative regulation of cytoskeleton organization,regulation of synaptic plasticity and positive regulation of lipid metabolism process,etc.From the cell component level,the differentially expressed lncRNAs are mainly related to extracellular matrix,hemoglobin complex,organelles,and synapses,etc.From the molecular function level,the differentially expressed lncRNAs are mainly related to ion gated channel activity,gated channel activity,ligand-gated cation channel activity and hormone binding,etc.3.KEGG pathway analysis shows that some enrichment pathways related to differentially expressed lncRNA include metabolic pathways,lysosome,chemokine signaling pathway,insulin signaling pathway,c AMP signaling pathway,TNF signaling pathway,Ras signaling system,PPAR signal pathway and MAPK signal pathway,etc.4.The 8differentially expressed lncRNAs were screened for verification by q RT-PCR.2^-△△Ctwas calculated based on the CT value,and statistical analysis showed that among the 8genes verified,the gene ENSG00000265218 is down-regulated in DKD patients.The q RT-PCR result of this gene is consistent with the trend of the second-generation high-throughput sequencing results,verifying the reliability of the sequencing results.Conclusion 1.Compared with normal controls,there are significantly differentially expressed lncRNAs in the peripheral blood of DKD patients.2.The second-generation high-throughput sequencing technology is used to detect genes differentially expressed in peripheral blood of DKD patients and normal controls,and screen out lncRNAs that are differentially expressed significantly up or down in DKD patients.3.The analysis of biological information shows that the interaction and biological functions of these differentially expressed lncRNAs and mRNAs may be related to the occurrence and development of diabetic nephropathy,and provide experimental evidence and new targets for its early diagnosis and clinical diagnosis and treatment.4.The expression of lncRNA ENSG00000265218 is down-regulated in diabetic patients,and it is further down-regulated in DKD patients,which is promising as a new marker of diabetes and diabetic kidney disease.