Efficient biomarkers for diabetic nephropathy (DN) have not been established. Using enzyme-linked immunosorbent assay, we found previously that urinary levels of full-length megalin (C-megalin), a multiligand endocytic receptor in proximal tubules, was positively correlated with DN progression in type 2 diabetes mellitus (T2DM). Here, we found that urinary extracellular vesicle (UEV) excretion and C-megalin content in UEVs or in their exosomal fraction increased along with the progression of the albuminuric stages in T2DM patients. Cultured immortalized rat proximal tubule cells (IRPTCs) treated with fatty acid-free bovine serum albumin (BSA) or advanced glycation end product-modified BSA (AGE-BSA), megalin’s endocytic ligands, increased EV excretion and their C-megalin content. C-megalin excretion from IRPTCs via EVs was significantly blocked by an exosome-specific inhibitor, GW4869, indicating that this excretion is mainly exocytosis-mediated. AGE-BSA treatment of IRPTCs caused apparent lysosomal dysfunction, which stimulated multivesicular body formation, resulting in increased exosomal C-megalin excretion. In a high-fat diet-induced, megalin-mediated kidney injury model in mice, urinary C-megalin excretion also increased via UEVs. Collectively, exocytosis-mediated urinary C-megalin excretion is associated with the development and progression of DN in T2DM, particularly due to megalin-mediated lysosomal dysfunction in proximal tubules, and hence it could be a candidate biomarker linked with DN pathogenesis.
This article contains Supplementary Data online at http://diabetes.diabetesjournals.org/lookup/suppl/doi:10.2337/db16-1031/-/DC1.
- Received August 24, 2016.
- Accepted February 14, 2017.
- © 2017 by the American Diabetes Association.