Loss of filtration function in diabetic glomeruli is associated with ultrastructural changes in glomerular endothelial cell fenestrations

Background: The study of glomerular endothelial cell (GEnC) fenestrations including key regulatory factors is neglected despite their loss in diabetic nephropathy, a disease associated with decreased filtration function, being previously described. Methods: We comprehensively characterised GEnC fenestral and renal filtration functional changes including measurement of glomerular ultrafiltration coefficient and glomerular filtration rate (GFR) in diabetic mice and humans. We further evaluated Eps homology domain protein-3 (Ehd3) as a potential regulator of GEnC fenestrations. Results: This study identified loss of GEnC fenestration density which was associated with decreased renal filtration function in diabetic nephropathy. We also identified increased GEnC fenestration width, an ultrastructural change that may develop to maintain filtration surface area. GEnC fenestration width was negatively associated with renal filtration function considered a result of development of diaphragms in widening fenestrations providing resistance to filtration. The increased presence of diaphragmed fenestrations in diabetes was supported by increased PLVAP1 expression. We identified decreased glomerular Ehd3 expression in diabetes and demonstrated its association with GEnC fenestration measurements suggesting its role in regulating fenestrations. We further demonstrated reduced fenestration formation in vitro in an Ehd3 knockdown cell line. Ehd3 was positively associated with filtration function suggesting loss of glomerular Ehd3 expression in disease may contribute to declining glomerular filtration function through aberrant GEnC fenestration regulation. Conclusions: This is the first study to demonstrate the critical role of GEnC fenestrations in renal filtration function and identify a key regulator, Ehd3, that may serve as a therapeutic target to retore filtration function in disease.