Type 2 Diabetes mellitus: mRNA and miRNA profiling
ABSTRACT: This SuperSeries is composed of the following subset Series: GSE21321: Blood microRNA profiles and upregulation of hsa-miR-144 in males with type 2 diabetes mellitus. GSE26167: MicroRNA 144 impairs insulin signaling by inhibiting the expression of insulin receptor substrate 1 in Type 2 Diabetes mellitus Refer to individual Series
Project description:This SuperSeries is composed of the following subset Series: GSE21321: Blood microRNA profiles and upregulation of hsa-miR-144 in males with type 2 diabetes mellitus. GSE26167: MicroRNA 144 impairs insulin signaling by inhibiting the expression of insulin receptor substrate 1 in Type 2 Diabetes mellitus Refer to individual Series
Project description:Dysregulation in expression of microRNAs (miRNAs) in various tissues has been linked to a wide spectrum of diseases, including Type 2 Diabetes mellitus (T2D). In this study, we compared the expression profiles of miRNAs in blood samples from Impaired Fasting Glucose (IFG) and T2D male patients with tissues from T2D rat models. Healthy adult males with no past history of T2D (n=158) and with desirable cholesterol and blood pressure profiles were enrolled in this study. They were then classified according to fasting glucose levels to have T2D, IFG or as healthy controls (CTL), for comparison of miRNA expression profiles. Employing miRNA microarray, we identified ‘signature miRNAs’ in peripheral blood samples that distinguished IFG and T2D. Eight selected miRNAs were further validated using stem-loop real-time RT-PCR. miR-144 expression was found to be dysregulated in Type 2 Diabetes, wherein its expression was significantly higher than in healthy controls. Insulin receptor substrate 1 (IRS1) has been predicted to be a potential target of miR-144. Consistent with this observation, IRS1 mRNA and protein levels, verified by quantitative real-time PCR and western blotting respectively, were found to be down-regulated. Using luciferase assay, we further demonstrated that miR-144 directly targets IRS1 and showed its effects on protein expression via immunocytochemistry. From this cross-sectional study in humans, we have identified signature miRNAs which could explain the pathogenesis of T2D. Whether miRNAs like miR-144 could be potential therapeutic targets for management of T2D will need to be explored by further mechanistic and functional studies. miRNA profiling of tissues from T2D rat models. Total RNA (plus miRNAs) was isolated using a modification of the RiboPure™-Blood kit from Ambion (Austin,TX) according to the manufacturer’s protocol. The concentration of total RNA and integrity were determined by using Nano-Drop ND-1000 Spectrophotometry (NanoDrop Tech, Rockland, Del) and gel electrophoresis respectively.
Project description:Over 40 % of microRNAs are located in introns of coding genes, and many intronic microRNAs are co-regulated with their host genes. In such cases of co-regulation, the products of host genes and their intronic microRNAs can cooperate to coordinately regulate biologically important pathways. Therefore, we screened intronic microRNAs dysregulated in liver of obese mouse models to identify previously uncharacterized coding host genes that may contribute to the pathogenesis of obesity-associated insulin resistance and type 2 diabetes mellitus. Our approach identified that expression of both Ectodysplasin A (Eda), the causal gene of X-linked hypohidrotic ectodermal dysplasia (XLHED; MIM 305100) and its intronic microRNA, miR-676, was strongly increased in liver of obese mouse models. Moreover, hepatic EDA expression is increased in obese human subjects, reduced upon weight loss, and its hepatic expression correlates with systemic insulin resistance. Eda expression in murine liver is controlled via PPARg activation, increases in circulation and promotes JNK activation and inhibitory serine phosphorylation of IRS1 in skeletal muscle. Consistently, bi-directional modulation of hepatic Eda expression in mouse models affects systemic glucose metabolism with alterations of muscle insulin signaling, revealing a novel role of EDA as an obesity-associated hepatokine, which impairs insulin sensitivity in skeletal muscle.
Project description:We analyzed the associations between whole blood microRNA profiles and the indices of glucose metabolism and impaired fasting glucose and examined whether the discovered microRNAs correlate with the expression of their mRNA targets. MicroRNA and gene expression profiling were performed for the Young Finns Study participants (n = 871). Glucose, insulin, and glycated hemoglobin (HbA1c) levels were measured, the insulin resistance index (HOMA2-IR) was calculated, and the glycemic status (normoglycemic [n = 534]/impaired fasting glucose [IFG] [n = 252]/type 2 diabetes [T2D] [n = 24]) determined. Levels of hsa-miR-144-5p, -122-5p, -148a-3p, -589-5p, and hsa-let-7a-5p associated with glycemic status. hsa-miR-144-5p and -148a-3p associated with glucose levels, while hsa-miR-144-5p, -122-5p, -184, and -339-3p associated with insulin levels and HOMA2-IR, and hsa-miR-148a-3p, -15b-3p, -93-3p, -146b-5p, -221-3p, -18a-3p, -642a-5p, and -181-2-3p associated with HbA1c levels. The targets of hsa-miR-146b-5p that correlated with its levels were enriched in inflammatory pathways, and the targets of hsa-miR-221-3p were enriched in insulin signaling and T2D pathways. These pathways showed indications of co-regulation by HbA1c-associated miRNAs. There were significant differences in the microRNA profiles associated with glucose, insulin, or HOMA-IR compared to those associated with HbA1c. The HbA1c-associated miRNAs also correlated with the expression of target mRNAs in pathways important to the development of T2D.
Project description:Results Platelets in non-diabetic patients demonstrated miRNA expression profiles comparable to previously published data. The miRNA expression profiles of platelets in diabetics were similar. Statistical analysis unveiled only three miRNAs (miR-377-5p, miR-628-3p, miR-3137) with high reselection probabilities in resampling techniques, corresponding to signatures with only modest discriminatory performance. Functional annotation of predicted targets for these miRNAs pointed towards an influence of diabetes mellitus on mRNA processing. Conclusions/interpretation We did not find any major differences in platelet miRNA profiles between diabetics and non-diabetics. Minor differences pertained to miRNAs associated with mRNA processing. Thus, previously described differences in plasma miRNAs between diabetic and nondiabetic patients cannot be explained by plain changes in the platelet miRNA profile. Platelet miRNA profiles were assessed in clinically stable diabetic and nondiabetic patients (each n=30). Platelet miRNA was isolated from leucocyte-depleted platelet-rich plasma, and miRNA profiling was performed using LNA micro-array technology (miRBase 18.0, containing 1,917 human miRNAs). Effects of diabetes mellitus were explored by univariate statistical tests for each miRNA, adjusted for potential confounders, and by developing a multivariable signature, which was evaluated by resampling techniques. Platelet phenotype was assessed by light transmission aggregometry and impedance aggregometry.
Project description:Pterygium is a relatively common human ocular surface fibroproliferative disease that affects vision. Endogenously produced microRNA (miRNA) regulates gene expression in various ocular surface diseases and possibly pterygium. We aimed to investigate the role of miRNA in pterygium. Paired human pterygium and conjunctival tissues were obtained from patients diagnosed with primary pterygium. miRNA microarray profiling identified statistically significant miRNA changes which were matched to reciprocal significant changes in their target transcripts. We employed quantitative real-time polymerase chain reaction and found that hsa-miR-766 was up-regulated (2.57-fold) whilst hsa-miR-215 was down-regulated (0.49-fold) in pterygium compared to conjunctival control. Localization of miRNA was performed using in-situ hybridization. Transcript levels of predicted hsa-miR-766 targets, nuclear receptor subfamily 4, group A, member 1 and epidermal growth factor-containing fibulin-like extracellular matrix protein 1, were down-regulated in pterygium compared to conjunctiva by 0.53- and 0.64-fold, respectively. Collagens type 3, alpha 1 and type 4, alpha 2, both targets of hsa-miR-215, were up-regulated in pterygium by 3.01- and 3.11-fold, respectively. These changes were confirmed in the protein levels using immunofluorescent staining. Derangement of hsa-miR-766 and hsa-miR-215 may cause dysregulation of matrix rearrangement, cell proliferation and adhesion proteins, resulting in pterygium formation. Targeting miRNA may be a possible therapeutic approach in this disease. 3 pterygium samples and 3 matched conjuctiva samples from patients diagnosed with primary pterygium. A pool of all 6 samples was used as the common reference.
Project description:BACKGROUND: Regardless the regulatory function of microRNAs (miRNA), their differential expression pattern has been used to define miRNA signatures and to disclose disease biomarkers. To address the question of whether patients presenting the different types of diabetes mellitus could be distinguished on the basis of their miRNA and mRNA expression profiling, we obtained peripheral blood mononuclear cell (PBMC) RNAs from 7 type 1 (T1D), 7 type 2 (T2D), and 6 gestational diabetes (GDM) patients, which were hybridized to Agilent miRNA and mRNA microarrays. Data quantification and quality control were obtained using the Feature Extraction software, and data distribution was normalized using quantile function implemented in the Aroma light package. Differentially expressed miRNAs/mRNAs were identified using Rank products, comparing T1DxGDM, T2DxGDM and T1DxT2D. Hierarchical clustering was performed using the average linkage criterion with Pearson uncentered distance as metrics. RESULTS: The use of the same microarrays platform permitted the identification of sets of shared or specific miRNAs/mRNA interaction for each type of diabetes. Nine miRNAs (hsa-miR-126, hsa-miR-1307, hsa-miR-142-3p, hsa-miR-142-5p, hsa-miR-144, hsa-miR-199a-5p, hsa-miR-27a, hsa-miR-29b, and hsa-miR-342-3p) were shared among T1D, T2D and GDM, and additional specific miRNAs were identified for T1D (20 miRNAs), T2D (14) and GDM (19) patients. ROC curves allowed the identification of specific and relevant (greater AUC values) miRNAs for each type of diabetes, including: i) hsa-miR-1274a, hsa-miR-1274b and hsa-let-7f for T1D; ii) hsa-miR-222, hsa-miR-30e and hsa-miR-140-3p for T2D, and iii) hsa-miR-181a and hsa-miR-1268 for GDM. Many of these miRNAs targeted mRNAs associated with diabetes pathogenesis. CONCLUSIONS: These results indicate that PBMC can be used as reporter cells to characterize the miRNA expression profiling disclosed by the different diabetes mellitus manifestations. Shared miRNAs may characterize diabetes as a metabolic and inflammatory disorder, whereas specific miRNAs may represent biological markers for each type of diabetes, deserving further attention.
Project description:BACKGROUND: Dysregulation of microRNA (miRNA) expression in various tissues and body fluids has been demonstrated to be associated with several diseases, including Type 2 Diabetes mellitus (T2D). Here, we compare miRNA expression profiles in different tissues (pancreas, liver, adipose and skeletal muscle) as well as in blood samples from T2D rat model and highlight the potential of circulating miRNAs as biomarkers of T2D. In parallel, we have examined the expression profiles of miRNAs in blood samples from Impaired Fasting Glucose (IFG) and T2D male patients. METHODOLOGY/PRINCIPAL FINDINGS: Employing miRNA microarray and stem-loop real-time RT-PCR, we identify four novel miRNAs, miR-144, miR-146a, miR-150 and miR-182 in addition to four previously reported diabetes-related miRNAs, miR-192, miR-29a, miR-30d and miR-320a, as potential signature miRNAs that distinguished IFG and T2D. Of these microRNAs, miR-144 that promotes erythropoiesis has been found to be highly up-regulated. Increased circulating level of miR-144 has been found to correlate with down-regulation of its predicted target, insulin receptor substrate 1 (IRS1) at both mRNA and protein levels. We could also experimentally demonstrate that IRS1 is indeed the target of miR-144. CONCLUSION: We demonstrate that peripheral blood microRNAs can be developed as unique biomarkers that are reflective and predictive of metabolic health and disorder. We have also identified signature miRNAs which could possibly explain the pathogenesis of T2D and the significance of miR-144 in insulin signaling.
Project description:microRNA profiling of mouse small intestinal crypt cells comparing control untreated with cells treated with insulin growth factor-1 (IGF-1). IGF-1 stimulated cell proliferation, as observed in Brdu incoporation assay. Two condition experiment. Control vs IGF-1 treatment. Biological replicates: 3 control, 3 treated. Independently grown and harvested. One replicate per array
Project description:BACKGROUND The aim of this study was to compare the transcriptome between impaired fasting glucose (IFG) and type 2 diabetes mellitus (T2DM), and further research their molecular mechanisms. MATERIAL AND METHODS The original microarray GSE21321, including miRNA and mRNA expression profiles, was downloaded from the GEO database. Data preprocessing was processed by limma package, and differentially expressed genes (DGs) and miRNA (DMs) were screened. Then, the regulatory relationships among miRNA, TF, and genes were screened and the regulatory network was constructed. Finally, DAVID was used for KEGG enrichment analysis. RESULTS There were 11 upregulated IFG-related DMs and five upregulated T2DM-related DMs. Three of the DMs overlapped. In addition, there were eight downregulated IFG-related DMs and two downregulated T2DM-related DMs. Only one downregulated DM overlapped. Similarly, there were 264 upregulated IFG-related DGs and 331 upregulated T2DM-related DGs; and 196 overlapping genes were obtained. In addition, there were 400 downregulated IFG-related DMs and 568 downregulated T2DM-related DMs. A total of 326 downregulated DMs were overlapped. The overlapped DGs were enriched in various pathways, including hematopoietic cell lineage, Fc gamma R-mediated phagocytosis, and MAPK signaling pathway. TAF1 (upregulated gene) and MAFK (downregulated gene) were hub nodes both in IFG- and T2DM-related miRNA-TF-gene regulatory network. In addition, miRNAs, including hsa-miR-29a, hsa-miR-192, and hsa-miR-144, were upregulated hub nodes in the two regulatory networks. CONCLUSIONS Genes including TAF1 and MAFK, and miRNAs including hsa-miR-29a, hsa-miR-192, and hsa-miR-144 might be potential target genes and important miRNAs for IFG and T2DM.