Urinary Exosomal microRNA-451-5p Is a Potential Early Biomarker of Diabetic Nephropathy in Rats

Non-invasive renal signatures can help in serial monitoring of diabetic patients. We tested whether urinary exosomal (UE) microRNA (miR) analysis could non-invasively predict renal pathology in diabetic rats during the course of diabetes. Diabetes mellitus (DM) was induced in male Wistar rats by a single intraperitoneal injection of streptozotocin (STZ, 50 mg/kg body weight). Non-diabetic control (CTRL) rats were injected with vehicle. Insulin (INS) treatment (5U/d, s.c.) was provided to 50% of the DM rats. Urine samples were collected at weeks 3, 6, and 9 following injections and UE prepared. An increase in miR-451-5p and miR-16, observed by pilot small RNA sequencing of UE RNA, was confirmed by quantitative real-time polymerase chain reaction (qPCR) and selected for further study. Subsets of rats were euthanized after 3, 6, and 9 weeks of diabetes for renal pathology analysis, including determination of the tubulointerstitial fibrotic index (TFI) and glomerulosclerotic index (GI) scores. qPCR showed a substantial rise in miR-451-5p in UE from DM rats during the course of diabetes, with a significant rise (median fold change >1000) between 3 and 6 weeks. Moreover, UE miR-451-5p at 6 weeks predicted urine albumin at 9 weeks (r = 0.76). A delayed but significant rise was also observed for miR-16. In contrast, mean urine albumin only increased 21% between 3 and 6 weeks (non-significant rise), and renal TFI and GI were unchanged till 9 weeks. Renal expression of miR-451-5p and miR-16 (at 10 weeks) did not correlate with urine levels, and moreover, was negatively associated with indices of renal pathology (r >= -0.70, p = 0.005 for TFI and r >= -0.6, p <= 0.02 for GI). Overall, a relative elevation in renal miR-451-5p and miR-16 in diabetes appeared protective against diabetes-induced kidney fibrosis; while UE miR-451-5p may hold prognostic value as an early and sensitive non-invasive indicator of renal disease.