Diabetic nephropathy (DN) is the leading cause of chronic kidney disease. Animal models are essential tools for designing new strategies to prevent DN. C57Bl/6 (B6) mice are widely used for transgenic mouse models, but are relatively resistant to DN. This study aims to identify the most effective method to induce DN in a type 1 (T1D) and a type 2 diabetes (T2D) model in B6 mice. For T1D-induced DN, mice were fed a control diet, and randomised to streptozotocin (STZ) alone, STZ+unilateral nephrectomy (UNx), or vehicle/sham. For T2D-induced DN, mice were fed a western (high fat) diet, and randomised to either STZ alone, STZ+UNx, UNx alone, or vehicle/sham. Mice subjected to a control diet with STZ +UNx developed albuminuria, glomerular lesions, thickening of the glomerular basement membrane, and tubular injury. Mice on control diet and STZ developed only mild renal lesions. Furthermore, kidneys from mice on a western diet were hardly affected by diabetes, UNx or the combination. We conclude that STZ combined with UNx is the most effective model to induce T1D-induced DN in B6 mice. In our hands, combining western diet and STZ treatment with or without UNx did not result in a T2D-induced DN model in B6 mice.

Background Ischaemia-reperfusion (IR) injury is an important determinant of delayed graft function (DGF) affecting allograft function. Mitochondrial DNA (mtDNA) is released upon cell death and platelet activation into the extracellular environment and has been suggested to be a biomarker in several diseases. Whether extracellular mtDNA accumulates in plasma and/or urine upon renal IR and predisposes DGF is unknown. Methods C57BL/6J wild-type mice were subjected to renal IR. In addition, an observational case–control study was set up enrolling 43 patients who underwent kidney transplantation. One day post-IR in mice and a few days following renal transplantation in human, blood and urine were collected. Patients were stratified into DGF and non-DGF groups. Results mtDNA-encoded genes accumulate in urine and plasma in both mice subjected to renal IR injury and in humans following renal transplantation. In human renal transplant recipients, cold ischaemia time and renal function correlate with urinary mtDNA levels. Urinary mtDNA levels but not urinary nuclear DNA levels were significantly higher in the DGF group compared with the non-DGF group. Multiple receiver operating characteristic curves revealed significant diagnostic performance for mtDNA-encoded genes cytochrome c oxidase III (COXIII); nicotinamide adenine dinucleotide hydrogen subunit 1 (NADH-deh); mitochondrially encoded, mitochondrially encoded nicotinamide adenine dinucleotide dehydrogenase 2 (MT-ND2) with an area under the curve of, respectively, 0.71 [P = 0.03; 95% confidence interval (CI) 0.54–0.89], 0.75 (P = 0.01; 95% CI 0.58–0.91) and 0.74 (P = 0.02; 95% CI 0.58–0.89). Conclusions These data suggest that renal ischaemia time determines the level of mtDNA accumulation in urine, which associates with renal allograft function and the diagnosis of DGF following renal transplantation.

Background Diabetic nephropathy (DN) is a major complication of diabetes and the main cause of end-stage renal disease. Extracellular vesicles (EVs) are small cell-derived vesicles that can alter disease progression by microRNA (miRNA) transfer. Methods In this study, we aimed to characterize the cellular origin and miRNA content of EVs in plasma samples of type 2 diabetes patients at various stages of DN. Type 2 diabetes patients were classified in three groups: normoalbuminuria, microalbuminuria and macroalbuminuria. The concentration and cellular origin of plasma EVs were measured by flow cytometry. A total of 752 EV miRNAs were profiled in 18 subjects and differentially expressed miRNAs were validated. Results Diabetic patients with microalbuminuria and/or macroalbuminuria showed elevated concentrations of total EVs and EVs from endothelial cells, platelets, leucocytes and erythrocytes compared with diabetic controls. miR-99a-5p was upregulated in macroalbuminuric patients compared with normoalbuminuric and microalbuminuric patients. Transfection of miR-99a-5p in cultured human podocytes downregulated mammalian target of rapamycin (mTOR) protein expression and downregulated the podocyte injury marker vimentin. Conclusions Type 2 diabetes patients with microalbuminuria and macroalbuminuria display differential EV profiles. miR-99a-5p expression is elevated in EVs from macroalbuminuria and mTOR is its validated mRNA target.