Project description:A total of 35 samples of renal tissues (15 DN, 10 DM, and 10 CON) were selected randomly from archived deidentified formalin-fixed paraffin-embedded DN biopsy tissue specimens. Normal control kidney tissues were obtained from kidneys that had been refused for use in transplantation due to vascular anomalies or from preimplant biopsy samples. Three random samples from each group were used for lncRNA microarrays
Project description:Current diagnostic methods for diabetic nephropathy (DN) lack precision, especially in early stages and monitoring progression. This study aims to find potential biomarkers for DN progression and evaluate their accuracy. Using serum samples from healthy controls (NC), diabetic patients (DM), early-medium stage DN (DN-EM), and late-stage DN (DN-L), researchers employed quantitative proteomics and Mfuzz clustering analysis revealed 15 proteins showing increased expression during DN progression, hinting at their biomarker potential. Combining Mfuzz clustering with weighted gene co-expression network analysis (WGCNA) highlighted five candidates (HMGB1, CD44, FBLN1, PTPRG, and ADAMTSL4). HMGB1 emerged as a promising biomarker, closely correlated with renal function changes. Experimental validation supported HMGB1’s upregulation under high glucose conditions, reinforcing its potential as an early detection biomarker for DN. This research advances DN understanding and identifies five potential biomarkers, notably HMGB1, as a promising early monitoring target. These findings set the stage for future clinical diagnostic applications in DN.
Project description:Diabetic mellitus (DM) is a disease that affects glucose homeostasis and has cause of complications, such as diabetic nephropathy (DN). For diagnose of the early DN, microalbuminuria is one of the mostly used biomarker currently. However, more early diagnostic biomarkers are desired rather than microalbuminuria.
Project description:Urinary proteome was analyzed and quantified by Tandem Mass Tag (TMT) labeling followed by bioinformatics analysis to study DN pathophysiology and to identify biomarkers of clinical outcome. We included type 2 diabetic normotensive non-obese males with and without incipient DN (microalbuminuria). Sample collection included blood and urine at baseline (control and DN-basal) and, in DN patients, after 3 months of losartan treatment (DN-treated). Urinary proteome analysis identified 166 differentially abundant proteins between controls and DN patients, 27 comparing DN-treated and DN-basal patients, and 182 between DN-treated patients and controls. Mathematical modeling analysis identified 80 key proteins involved in DN pathophysiology, 15 in losartan effect and 7 of these 95 are essential in both processes. VCAM-1 and neprilysin are the only ones that are differentially expressed in the urinary proteome. We observed an increase of VCAM-1 urine levels in DN-basal patients compared to diabetic controls and an increase of urinary neprilysin in DN-treated patients with persistent albuminuria, the latter confirmed by ELISA . Our results point to neprilysin and VCAM-1 as potential candidates in DN pathology and treatment.
Project description:Compared with plasma sEV (Con-sEV) from control rats, plasma sEV (DM-sEV) from 8-week diabetic rats significantly induced cardiomyocyte apoptosis as evidenced by increased percentage of apoptotic cells and activity of pro-apoptotic protein caspase 3. The proapoptotic effect of DM-sEV was blunted by RNase but not proteinase K, suggesting that DM-sEV exerted the cardiotoxic effects mainly through their contained RNAs. Increasing evidence suggests that miRNAs are the most important molecules by which sEV regulate recipient cell function. To identify the sEV-containing specific miRNAs responsible for the effects, Con-sEV and DM-sEV were subjected to miRNA sequencing.
Project description:Epithelial and stromal/mesenchymal limbal stem cells contribute to corneal homeostasis and cell renewal. Extracellular vesicles (EVs), including exosomes (Exos), can be paracrine mediators of intercellular communication. Previously, we described cargos and regulatory roles of limbal stromal cell (LSC)-derived Exos in non-diabetic (N) and diabetic (DM) limbal epithelial cells (LEC). Presently, we quantify the miRNA and proteome profiles of human LEC-derived Exos and their regulatory roles in N- and DM-LSC. We revealed some miRNA and protein differences in DM vs. N-LEC-derived Exos' cargos including proteins involved in Exo biogenesis and packaging that may affect Exo production and ultimately cellular crosstalk and corneal function. Treatment by N-Exos, but not by DM-Exos enhanced wound healing in cultured N-LSC and increased proliferation rate in N and DM LSCs vs. corresponding untreated (control) cells. N-Exos treated LSC reduced keratocyte markers ALDH3A1 and lumican, and increased MSC markers CD73, CD90 and CD105 vs. control LSC. These being opposite to the changes quantified in wounded LSCs. Overall, N-LEC Exos have a more pronounced effect on LSC wound healing, proliferation, and stem cell marker expression than DM-LEC Exos. This suggests that regulatory miRNA and protein cargo differences in DM- vs. N-LEC-derived Exos could contribute to the disease state.
Project description:Diabetic Nephropathy (DN) is a chronic complication of diabetes and the primary cause of end stage renal disease. DN can be differentially diagnosed only through histological investigation. Therefore, there is need for molecular biomarkers, such as miRNAs, to discriminate among different histological lesions in diabetics. Aim of this study was to identify a pattern of differentially expressed miRNAs in kidney biopsies of DN patients and to assess their potential as differential diagnostic biomarkers. Using microarray, we assayed miRNA expression in kidney tissue from 8 DN patients, 6 diabetic patients with membranous nephropathy and 4 patients with normal histology. Nine miRNAs were differentially expressed among the three groups of patients, thus underlying their potential role as markers of specific histological damage. In silico we identified 2 miRNAs (i.e., miR-27b-3p and miR-1228-3p) showing an interaction with UBE2v1, an ubiquitin-conjugating E2 enzyme variant that mediates the formation of lysine 63-linked ubiquitin chains, a mechanism we showed as involved in DN kidney fibrosis. Both miRNAs were confirmed to be down-regulated in DN biopsies, using qPCR and in situ hybridization, and in urines of DN patients. Interestingly, the urinary levels of both miRNAs were also able to discriminate among different degrees of renal fibrosis. Finally, we showed that the combined urinary expression of both miRNAs was also able to discriminate DN patients from other glomerulonephritides, both in the presence and absence of T2D. We identified two regulatory miRNAs potentially useful as diagnostic biomarkers of tubular-interstitial fibrosis in diabetic patients with DN.
Project description:Despite recent advances, diabetic nephropathy (DN) remains a major public health concern. The precise underlying molecular mechanisms of DN remain elusive. Accumulating recent evidence suggests that mitochondrial integrity and lipid metabolism in podocytes significantly contribute to the pathogenesis of DN. However, the interplay between these two key metabolic regulators of DN is not fully understood. This study examines the role of ChREBP (carbohydrate-response element-binding protein), a master regulator of lipogenesis, on mitochondrial morphology and progression of DN. Our findings suggest that diabetic mice with podocyte-specific deletion of ChREBP are protected against mitochondrial fragmentation and progression of DN. Using liquid chromatography coupled with mass spectrometry, we identified the central role of ChREBP-induced plasmalogen phospholipids in linking mitochondrial lipidomes with mitochondrial dynamics in DN.
Project description:To decipher the effect of endothelial specific inhibition of Brg1 on histone marker H3K4me3 in the zebrafish genome during zebrafish heart regeneration, we achieved endothelium-specific over-expression of DN-xBrg1 by using the compound zebrafish line consisting of Tg(ubi:loxp-DsRed-STOP-loxp-DN-xBrg1; kdrl:CreER) (defined as DN), while we used Tg(ubi:loxp-DsRed-STOP-loxp-DN-xBrg1) as control (Ctrl) in the presence of 4-HT starting at 3 days before ventricular resection. The whole ventricles from CtrlK and DNK hearts at 7 dpa were subjected to ChIP-seq analysis using H3K4me3 antibody.