| ObjectiveIn this study,circBank,starBase and circIteractome and other bioinformatics databases were conducted to predict the miRNA interacting with hsa＿circ＿0071106 which obtained by the gene chip and population verification by our research group early,and verified its differential expression between type 2 diabetes and controls by qRT-PCR.Subsequently,the lncRNA interacting with the related miRNA was predicted by starBase and Diana-lncRNABase bioinformatics,and the lncRNA related to type 2 diabetes was selected in combination with the literature,and its expression in type 2 diabetes and controls was verified by qRT-PCR.The ROC curve was used to explore the diagnostic value of related hsa＿circ＿0071106,miRNA and lncRNA alone and in combination,and the interaction between them on diabetes was analyzed by the mediation effect.Finally,this study provides scientific basis for the study of the mechanism on type 2 diabetes,through the enrichment of miRNA target genes and their possible functional pathways predicting that affect diabetes.MethodsThis study used an individual matched case-control study design.In2019,based on health screening,patients diagosed with type 2 diabetes in5 community populations in Huadu District,Guangzhou were selected as cases.The inclusion criteria were fasting blood glucose≥7.0 mmol/L and/or an oral glucose tolerance test for 2 hours≥11.1mmol/L and/or diagnosed as type 2 diabetes by a hospital county level above.The inclusion criteria of control were FBG<7.0mmol/L and an oral glucose tolerance test for 2 hours<11.1mmol/L according to the standard of the same sex and the age is no more than 5 years,from the same residential area and no history of diabetes.An epidemiological questionnaire investigation was conducted on the subjects to obtain information on general demographic data,smoking,drinking,past history,etc.Blood pressure was measured and fasting peripheral venous blood was drew.The miRNA and lncRNA that interact with hsa＿circ＿0071106 were selected based on the results of bioinformatics prediction and previous literature,and the expression of miRNA,lncRNA and circRNA in type 2diabetes was checked by RT-q PCR.ROC curve was conducted to evaluate the diagnostic value of miRNA,lncRNA and circRNA alone and in combination.Multivariate conditional logistic regression was applied to estimate the effect of miRNA,lncRNA and circRNA factor on the risk of type 2 diabetes.The mediation effect was used to analyze the interaction relationship among hsa＿circ＿0071106,target miRNA and target lncRNA on diabetes.GO enrichment analysis and KEGG pathway enrichment analysis are used to predict the possible functional pathways of miRNA target genes that affect diabetes.Finally,correlation analysis was used to explore the correlation between hsa＿circ＿0071106,target miRNA,target lncRNA and clinical indicators of type 2 diabetes.Results1.In this study,circBank,starBase and circIteractome were used to predict the miRNA interacting with hsa＿circ＿0071106 online,and Cytoscape software were used to draw a network diagram to describe their relationship.The miRNA predicted by the three databases were hsa-miR-1206,hsa-miR-607,hsa-miR-450b-3p,hsa-miR-556-5p,hsa-miR-409-3p,hsa-miR-3690 and hsa-miR-29a-5p,etc.2.A total 101 cases and 101 controls were included.The relative expressions of hsa-miR-29a-5p,hsa-miR-607,hsa-miR-409-3p and hsa-miR-3690 in the peripheral blood of type 2 diabetes cases and controls were detected by qRT-PCR.And the expressions of hsa-miR-29a-5p,hsa-miR-3690 and hsa-miR-607 in the diabetes group were higher than those in the control group significantly.The multivariate conditional logistic regression was used to analyze,and the age,gender,smoking,drinking,TG,TC and LDL-C were controlled at the same time.The results showed that hsa-miR-29a-5p was a risk factor affecting type 2diabetes(OR=10.28,95%CI:1.95～54.10,P=0.018).The risk of type 2diabetes with high expression of hsa-miR-607 was 5.17 times that of low expression(95%CI:1.48～18.09,P=0.012).And the risk of type 2diabetes with high expression of hsa-miR-3690 was 26.88 times that of low expression(95%CI:2.65～272.87,P=0.030).3.In this study,lncRNA interacting with hsa-miR-29a-5p,hsa-miR-607 and hsa-miR-3690 were predicted by starBase and Diana-lncRNABase online.And through the intersection of the Venn diagram,it was found that there were 27 lncRNA interacting with hsa-miR-29a-5p,hsa-miR-607 and hsa-miR-3690,including lncRNA TUG1,lncRNA MALTA1 and lncRNA MEG3.4.In this study,the relative expressions of lncRNA TUG1,lncRNA MALTA1 and lncRNA MEG3 in the peripheral blood of type 2 diabetes cases and controls were tested by qRT-PCR.And the expression of lncRNA TUG1 and lncRNA MEG3 in diabetes group was higher than in controls.The multivariate conditional logistic regression model was conducted with the age,gender,smoking,drinking,TG,TC and LDL-C being controlled at the same time.The results showed that lncRNA MEG3 was a risk factor affecting type 2 diabetes(OR=5.75,95%CI:1.55～21.36,P=0.014).The risk of type 2 diabetes with high expression of lncRNA TUG1 was 5.98 times higher than that of low expression(95%CI:1.62～22.10,P=0.014).5.In this study,the ROC curve was used to evaluate the diagnostic value of hsa-miR-29a-5p,hsa-miR-607,hsa-miR-3690,lncRNA MEG3,lncRNA TUG1 and hsa＿circ＿0071106 in type 2 diabetes.And the area under the curve for the diagnosis of the expression level of lncRNA TUG1 was 0.642(95%CI:0.549～0.735),with a sensitivity of 63.4%and a specificity of 97.0%.The area under the ROC curve of the combined effect of hsa-miR-607,lncRNA TUG1 and hsa＿circ＿0071106 was 0.763(95%CI:0.682～0.8441),which yielded a sensitivity of 74.3%and a specificity of 97.0%.6.This study used mediation analysis and found that the effects of hsa＿circ＿0071106 and hsa-miR-607 on type 2 diabetes was mainly attributable to the pure mediation effect(excess relative risk=0.34,95%CI:-0.03～0.66,P=0.031).The effect on type 2 diabetes between lncRNA TUG1 and hsa-miR-607 was mainly attributable to the pure mediating effect(excess relative risk=0.32,95%CI:0.03～0.61,P=0.030).The influence of hsa＿circ＿0071106 and lncRNA TUG1 on type2 diabetes was mainly due to pure mediating effect(excess relative risk=0.32,95%CI:0.04～0.60,P=0.026).The effect of lncRNA MEG3and hsa-miR-29a-5p on type 2 diabetes was mainly due to pure mediating effect(excess relative risk=0.36,95%CI:0.04～0.68,P=0.025).The influence of lncRNA MEG3 and hsa-miR-3690 on type 2 diabetes was mainly due to the pure mediating effect(excess relative risk=0.28,95%CI:0.01～0.55,P=0.046).7.In this study,online predictions of bioinformatics target genes for hsa-miR-29a-5p,hsa-miR-607 and hsa-miR-3690 were carried out using databases such as starBase,miRwalk,miRDB and Targetscan.And the intersection was found by the Venn diagram.There were 122 target genes of hsa-miR-29a-5p predicted by the four databases,352 target genes of hsa-miR-607 and 116 target genes of hsa-miR-3690.GO functional enrichment analysis found that hsa-miR-29a-5p,hsa-miR-607 and hsa-miR-3690 may regulate the DNA template transcription and RNA polymerase II promoter transcription and other biological processes on protein and ATP binding elements in the cytoplasm or nucleus.KEGG pathway enrichment analysis revealed that it may participate in the insulin resistance,the Fox O signaling pathways,AMPK signaling pathways and other signaling pathways.8.This study used correlation analysis and found that hsa-miR-3690was positively correlated with SBP(r_s=0.19,P=0.008).And hsa-miR-29a-5p was positively correlated with LDL-C(r_s=0.17,P=0.037).Hsa-miR-607 was positively correlated with LDL-C(r_s=0.27,P<0.001),and negatively correlated with HDL-C(r_s=-0.17,P=0.040).ConclusionsThis study found that hsa-miR-29a-5p,hsa-miR-607,hsa-miR-3690,lncRNA MEG3,lncRNA TUG1 and hsa＿circ＿0071106 were associated with the risk of type 2 diabetes.LncRNA TUG1 and hsa＿circ＿0071106regulate type 2 diabetes through the mediation of hsa-miR-607.And hsa＿circ＿0071106 regulates type 2 diabetes through the mediation of lncRNA TUG1.LncRNA MEG3 affects type 2 diabetes through the mediation of hsa-miR-29a-5p and hsa-miR-3690.They may participate in complex regulation process of type 2 diabetes in cytoplasm or nucleus with protein binding,ATP binding and other elements through DNA template transcription,RNA polymerase II promoter transcription regulation and other biological processes.And participate in insulin resistance,Fox O signaling pathway,AMPK signaling pathway and other signaling pathways.The combination of lncRNA TUG1,hsa-miR-607and hsa＿circ＿0071106 is more valuable in type 2 diabetes diagnosing. |
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