Objective: To investigate the differential expression of circRNAs in human blood, as a diagnostic marker for type 2 diabetes mellitus (T2DM) and to explore the mechanism of action of circRNA on miRNA and its target genes.
Methods: Circular RNA microarrays were performed to identify T2DM-related circRNAs. qPCR assays were performed to detect circRNA expression pattern in clinical samples. TargetScan and miRBase program was used to predict circRNA/miRNA interaction. Dual luciferase reporter gene assay to detect the mechanism of action of circRNA on miRNA.A total of 1,439 up-regulated expressed circRNAs were found in microarray analysis. Nine of the carcRNAs were selected by qPCR in pre-research.Blood collected from normal people(NC, N=20), prediabetes(pre-DM, n=20) and T2DM(DM, N=20) for qPCR test.
Results: The results showed that the difference expression of hsa-circ-000094 among the nine circRNAs were the most obvious in the three groups, the area under the maximum curve was found by ROC curve analysis, Spre-DM =0.8025 (95% CI, 0.6655-0.9395, p=0.001); ST2DM=0.77(95% CI,0.624-0.916, p=0.003). In order to verify the clinical diagnostic ability of hsa-circ-000094, the sample were expanded to 60 in each three groups. The results showed that the expression of hsa-circ-000094 in the three groups were different, the difference and ROC curve analysis were statistically significant, Spre-DM =0.6733 (95% CI, 0.5757-0.7710, P<0.001);ST2DM=0.7231 (95% CI,0.6327-0.8134, P<0.001). The results of Dual luciferase reporter gene assay showed that the fluorescence activity was reduced by about 40% after transfection of hsa-circ-000094 WT and hsa-miR-370-5p mimic into cells. It suggests that hsa-circ-000094 acted as miR-370 sponge to inhabit miR-370 activity.
Conclusions: CircRNAs are involved in T2DM pathogenesis, and thus serve as a potential biomarkers of T2DM diagnosis.
C. Zhu: None. L. Wei: None. Y. Xiao: None.
National Natural Science Foundation of China (81500615)