Reference gene panel for urinary exosome-based molecular diagnostics in patients with kidney disease

BACKGROUND Kidney disease is a severe complication of diabetes that often leads to end-stage renal disease. Early diagnosis is crucial for prevention or delay. However, the current diagnostic methods, with their limitations in detecting the disease in its early stages, underscore the urgency and importance of finding new solutions. miRNAs encapsulated inside urinary exosomes (UEs) have potential as early biomarkers for kidney diseases. The need for reference miRNAs for accurate interpretation currently limits their translational potential. AIM To identify consistently expressing reference miRNAs from UEs of controls and patients with type 2 diabetesmellitus (T2DM) and biopsy-confirmed kidney diseases. METHODS miRNA profiling was performed on UEs from 31 human urine samples using a rigorous and unbiased method. The UEs were isolated from urine samples collected from healthy individuals (n = 6), patients with T2DM (n = 13), and T2DM patients who also had kidney diseases (including diabetic nephropathy, n = 5; membranous nephropathy, n = 5; and IgA nephropathy, n = 2) through differential ultracentrifugation. After characterizing the UEs, miRNA expression profiling using microarray technology was conducted. RESULTS Microarray data analysis identified 14 miRNAs that were consistently expressed in UEs from 31 human samples, representing various kidney conditions: diabetic controls, diabetic nephropathy, membrane nephropathy, IgA nephropathy, and healthy controls. Through in silico analysis, we determined that 10 of these miRNAs had significant potential to serve as reference genes in UEs. CONCLUSION We identified uniformly expressing UE miRNAs that could serve as reference genes kidney disease biomarkers.