Effect Of Liraglutide On Serum MiRNA Expressions In Patients With Newly Diagnosed Type 2 Diabetes Mellitus

Objective:(1) To explore the differences of serum miRNA expressions between patients with newly diagnosed type 2 diabetes mellitus and healthy people receiving physical examination.(2) To analyze the changes of serum miRNA-19a expression between before and after liraglutide treatment, and to analyze the correlation with insulin resistance and function changes of pancreatic β cells.Methods:Thirty patients with newly diagnosed type 2 diabetes mellitus in endocrinology department of the affiliated Jiangyin Hospital of Southeast University from January 2014 to January 2015 were enrolled as the study subjects. The levels of serum miRNAs were detected by μParafloTM miRNA microfluid chip technology. The miRNAs with significant changes of expressive abundance compared between the healthy people receiving physical examination and patients with newly diagnosed type 2 diabetes mellitus were screened out. MiR-19a was screened out as the certain miRNA with significant expression difference between the healthy people receiving physical examination and patients with newly diagnosed type 2 diabetes mellitus, strong hybridization signal and few intra-group differences. The patients were treated with liraglutide for 12 weeks, and the changes of miR-19a expression were observed. MiR-16 gene was selected as the reference gene, and the expression of miR-19a was detected by SYBR Green RT-qPCR. Each sample was detected three times at the same time, and the average value was used as the cycle threshold (Cq value). The relative expression level of miR-19a was shown as 2-ΔCq, and ΔCq=Cq miR19a-Cq miR16. The levels of body weight, waist circumference and blood pressure were detected in the 30 patients with newly diagnosed type 2 diabetes mellitus before and after 12-week liraglutide treatment. The levels of HbAlc were detected by high pressure liquid chromatography (BIO-RAD D10). The venous blood was sampled at 30 and 120 min after the patient took 75 g glucose by oral, respectively. The levels of fasting blood glucose and blood glucose at 30 and 120 min after taking glucose (G0, G30 and G120) were detected. The levels of insulin (10,130 and 1120) and C-peptide (CP0, CP30 and CP120) were detected. The data were analyzed with paired t-test and Wilcoxon signed rank test.Results:The chip detection found that there were 12 miRNAs with the most significant differential expression between the patients with newly diagnosed type 2 diabetes mellitus and healthy people receiving physical examination. There were 10 up-regulated miRNAs, including miR-4288, miR-500b-5p, miR-4442, miR-6516-3p, miR-6806-5p, miR-19a-3p,-miR-4713-5p,’miR-4291, miR-7162-5p, miR-4304, and miR-4304, and two down-regulated miRNAs, including miR-BART15 and miR-7973. According to the chip result, miR-19a with significant expression difference, strong hybridization signal and few intra-group differences was selected for the PCR experiments. The results of relative serum miR-19a expression showed abnormal distribution after the patients were treated with liraglutide for 12 weeks. The analysis by Wilcoxon signed rank test showed that the expression of serum miR-19a was significantly down-regulated after treatment, and there was statistical significance in the difference compared with that before treatment (P<0.05).The levels of body weight, HbAlC, G0, G30 and G120 after treatment were significantly lower compared with those before treatment (P<0.01). The Matsuda insulin sensitivity index (Matsuda ISI) after treatment increased compared with that before treatment (P<0.01). The ratio of the area under the curve during 0-30 min for insulin (AUCinsO-30) to the area under the curve during 0-30 min for blood glucose (AUCglu0-30) was used as the earlier phase islet secretory function index (INSR30). The area under the curve during 0-30 min for insulin (AUCinsO-30) after treatment significantly increased compared with that before treatment (P<0.01). The ratio of AUCinsO-120/AUCglu0-120 was used as the full-release islet functional index (INSR120). The area under the curve during 0-120 min for insulin (INSR120) after treatment significantly increased compared with that before treatment (P<0.01). The insulin sensitivity indices after correction were DI30= Matsuda ISI*INSR30 and DI120= Matsuda ISI*INSR120, and the levels significantly increased compared with those before treatment (P<0.01).Conclusion:The detection of serum miRNAs could be a biological indicator for effective susceptibility screen and early diagnosis of type 2 diabetes mellitus. The improvement of insulin β cell function after liraglutide treatment could be associated with the differential expressions of serum miRNAs. The expression of miR-19a was down-regulated after liraglutide treatment, suggesting that miR-19a could be a biological indicator in the treatment of diabetes mellitus with liraglutide.