Circulating microRNAs (miRNAs) have been proposed as promising biomarkers for multiple diseases. miR-126 is reported to be associated with type 2 diabetes mellitus (T2D), diabetic nephropathy (DN) and end stage renal disease. The aim of this study was to investigate the expression of circulating miR‑126 and to assess its potential as a blood‑based biomarker for DN in T2D patients. In 52 patients with T2D without history of DN (with noromoalbuminuria), 50 patients with T2D and DN (29 with microalbuminuria and 21 with macroalbuminuria), and 50 non-diabetic healthy controls, the expression of circulating miR‑126 in peripheral whole blood was evaluated by quantitative polymerase chain reaction. The expression levels of circulating miR‑126 were significantly decreased in T2D patients and further decreased in DN patients compared with those in the controls. Multivariate logistic regression analysis confirmed the independent association of lower miR‑126 levels with T2D [adjusted odds ratio (OR), 0.797; 95% confidence interval (CI), 0.613‑0.960] and DN (adjusted OR, 0.513; 95% CI, 0.371‑0.708). miR‑126 levels were associated with the degree of albuminuria and showed significantly low expression in DN patients with microalbuminuria (adjusted OR, 0.781; 95% CI; 0.698‑0.952) and further lower expression in DN patients with macroalbuminuria (adjusted OR, 0.433; 95% CI, 0.299‑0.701), respectively compared with T2D patients with normoalbuminuria. miR‑126 levels negatively correlated with albuminuria positively with glomerular filtration rate (P<0.05), and in addition, negatively correlated with fasting glucose, glycated hemoglobin, triglyceride and LDL (P<0.05). Stepwise multiple regression analysis identified albuminuria as a significant predictor of miR‑126 (P<0.001). miR‑126 in peripheral blood yielded area under the receiver operating characteristic curves of 0.854 (95% CI, 0.779‑0.929) and 0.959 (95% CI, 0.916‑1.000) in the differentiation of DN patients from T2D patients and DN patients from non‑diabetic controls respectively. These data suggest that decreased expression of circulating miR‑126 is associated with the development of DN in T2D patients, and may be a promising blood‑based biomarker for DN risk estimation.

MicroRNAs (miRNAs) are small non-coding RNAs that function as crucial regulators of gene expression. Recently, dysregulation of miRNA expression in the blood has been demonstrated to be associated with various diseases, including type 2 diabetes mellitus (T2D), suggesting a potential for their use as biomarkers of disease prognosis. The present study examined the expression levels of T2D‑associated miR‑15a in peripheral whole blood samples from patients with T2D, pre‑diabetes individuals exhibiting impaired fasting glucose (IFG) and impaired glucose tolerance (IGT), as well as healthy control subjects, in order to investigate the potential of peripheral blood plasma miR‑15a as a biomarker for the prediction of T2D and pre‑diabetes. The present study included 24 patients with T2D, 22 IFG/IGT individuals and 24 healthy controls. The expression levels of miR‑15a were analyzed by reverse transcription-quantitative polymerase chain reaction. The results indicated that the peripheral blood miR‑15a expression levels were significantly decreased in patients with T2D and IFG/IGT individuals, compared with healthy control subjects (P<0.05). As determined by multivariate logistic regression analysis, lower miR‑15a expression was significantly associated with T2D (odds ratio [OR]; 95% confidence interval [CI]: 0.51; 0.16‑0.73, respectively; P<0.05) and pre‑diabetes (OR; 95% CI: 0.56; 0.23‑0.79, respectively; P<0.05). This association remained statistically significant following adjustment for age, body mass index and hypertension, as well as other biochemical indicators. Furthermore, a receiver operating characteristic analysis revealed that blood miR‑15a distinguished patients with T2D and IFG/IGT individuals from healthy controls (area under the curves; 95% CI: 0.864; 0.751‑0.977 and 95% CI: 0.852; 0.752‑0.953, respectively). These results demonstrated that peripheral blood miR‑15a expression levels were significantly lower in patients with T2D and IFG/IGT individuals, compared with healthy individuals. Thus, miR‑15a in peripheral whole blood may serve as a potential biomarker for T2D and pre-diabetes.

Oxidative damage to mitochondrial DNA (mtDNA) has been linked to the pathogenicity of diabetic nephropathy. We tested the hypothesis that mtDNA copy number may be increased in human mesangial cells in response to high glucose-induced reactive oxygen species (ROS) to compensate for damaged mtDNA. The effect of manganese superoxide dismutase mimetic (MnTBAP) on glucose-induced mtDNA copy number was also examined. The copy number of mtDNA was determined by real-time PCR in human mesangial cells cultured in 5 mM glucose, 25 mM glucose, and mannitol (osmotic control), as well as in cells cultured in 25 mM glucose in the presence and absence of 200 μM MnTBAP. Intracellular ROS was assessed by confocal microscopy and flow cytometry in human mesangial cells. The copy number of mtDNA was significantly increased when human mesangial cells were incubated with 25 mM glucose compared to 5 mM glucose and mannitol. In addition, 25 mM glucose rapidly generated ROS in the cells, which was not detected in 5 mM glucose. Furthermore, mtDNA copy number was significantly decreased and maintained to normal following treatment of cells with 25 mM glucose and MnTBAP compared to 25 mM glucose alone. Inclusion of MnTBAP during 25 mM glucose incubation inhibited mitochondrial superoxide in human mesangial cells. Increased mtDNA copy number in human mesangial cells by high glucose could contribute to increased mitochondrial superoxide, and prevention of mtDNA copy number could have potential in retarding the development of diabetic nephropathy.

Increased the incidence of prediabetes and type 2 diabetes (T2D) worldwide raises an urgent need to develop effective tools for early disease detection to facilitate future preventive interventions and improve patient’s care. We evaluated the suitability of diabetes-related miR-375 and miR-9 as earlier biomarkers for detecting prediabetes and T2D.TaqMan-based RT-qPCR was used to quantify the expression of miRNAs in peripheral blood of 30 prediabetes patients, 30 T2D patients and 30 non-diabetic healthy controls. Compared to controls, miR-375 and miR-9 were expressed at higher levels in prediabetes patients and progressively more enriched in T2D patients. Both miRNAs were directly associated with the presence of prediabetes and T2D independently of known risk factors to T2D and miR-375 was independently associated with the development of T2D. Both miRNAs were positively correlated with the glycemic status and other T2D risk factors. The ROC analysis indicated good diagnostic abilities for miR-375 to distinguish overall patients from control and prediabetes from T2D patients. Whereas, miR-9 showed lower values and borderline significance in discriminating the subject groups. The combination of miRNAs enhanced the predictability to discriminate patients from control. These results suggest that miR-375 and miR-9 are associated with the susceptibility to developing T2D and miR-375 alone or in combination with miR-9 could serve as biomarkers for early detection of prediabetes and T2D.

Circulating microRNAs (miRNAs) have been shown as promising biomarkers for various diseases. We investigated the predictive potential of circulating endothelium-enriched miR-126 in type 2 diabetes patients (T2D) without chronic complications and T2D patients with coronary artery diseases (CAD). The expression levels of circulating miR-126, determined by quantitative real time PCR, were decrease in peripheral blood of T2D patients and T2D with CAD compared with healthy controls. MiR-126 strongly associated with T2D and CAD, negatively correlated with LDL in CAD patients and differentiated between T2D patients, T2D patients with CAD and healthy subjects. Circulating miR-126 may serve as a biomarker for predicting patients with T2D and diabetic CAD.

The mitochondrial DNA copy number (mtDNA‑CN) is a surrogate measure of mitochondrial function and altered mtDNA‑CN reflects the oxidant‑induced cell damage. A previous study by our group demonstrated that a reduction in the renal mtDNA‑CN is implicated in the pathogenesis of diabetic nephropathy (DN), a leading cause of end‑stage renal disease in diabetic patients. In the present study, it was investigated whether the mtDNA‑CN in the peripheral blood may be utilized as a biomarker for DN in type 2 diabetes (T2D) patients. The study included 50 non‑diabetic and 100 diabetic subjects. The diabetic subjects were sub‑divided based on their albumin‑to‑creatinine ratio (ACR) into T2D patients with normoalbuminuria (n=50), DN patients with microalbuminuria (n=29) and DN patients with macroalbuminuria (n=21). The mtDNA‑CN was measured in the peripheral blood by real‑time polymerase chain reaction analysis. Patients with DN had a lower mtDNA‑CN than patients with T2D and healthy controls (P<0.05). A sub‑group analysis with stratification by the ACR indicated that a decreased mtDNA‑CN was associated with the severity and the presence of DN, as it was lower in DN patients with macroalbuminuria than in DN patients with microalbuminuria and T2D patients with normoalbuminuria (P<0.01). The area under the receiver operating characteristic curve (AUC) for mtDNA‑CN was 0.916 (sensitivity, 86% and specificity, 74%) and 0.961 (sensitivity, 96% and specificity, 88%) for differentiating DN patients from T2D patients without DN and from healthy controls, respectively. Furthermore, the AUC of mtDNA‑CN for differentiating DN patients with microalbuminuria from those with macroalbuminuria was 0.895 (sensitivity, 83% and specificity, 85%). Multivariate analysis revealed that the mtDNA‑CN was significantly associated with the occurrence and progression of DN, even after adjustment for age, mean blood pressure, glycated haemoglobin A1c and total cholesterol (P<0.05). In patients with DN, a decreased mtDNA‑CN was negatively correlated with albuminuria and conventional risk factors for DN, and was positively correlated with the estimated glomerular filtration rate. The present results therefore suggest the utilization of circulating mtDNA‑CN as a novel biomarker for the early diagnosis of DN and indicate the significance of decreased mtDNA‑CN as another independent risk factor for DN.