The expression of the structural proteins Dendrin and Neph1 in the glomerular filtration barrier in proteinuria

Background In the normal kidney, the glomerular filtration barrier successfully clears about one litre blood per minute. Damage to the filtration barrier might lead to protein leakage in the urine – proteinuria. The major ultra-structural finding in glomerular diseases with proteinuria is foot process effacement (FPE). Despite these being the most common signs of glomerular dysfunction, the underlying pathophysiological mechanisms are still not fully understood. However, a number of proteins identified in the slit diaphragm (SD) of the podocytes, including Dendrin and Neph1, are believed to be significant. Dendrin is a protein that has previously been described in mouse podocytes, associated to the actin cytoskeleton. Neph1 is a transmembrane protein that forms a complex with Nephrin in the SD. Recent studies have indicated the complex involvement in polymerization of the actin cytoskeleton and proteinuria. Aim We aimed to study the expression of Dendrin in normal human kidney and in the glomerular disease Minimal Change Nephrotic Syndrome (MCNS) with proteinuria and foot process effacement (FPE). In the second study, we wanted to investigate the subcellular localization of Neph1 in normal human kidney and the expression in Focal Segmental Glomerulosclerosis (FSGS), MCNS and in the corresponding experimental models Adriamycin nephropathy (ADR) and puromycin aminonucleoside (PAN). All characterized by substantial FPE and proteinuria. Methods The localization of Dendrin and Neph1 was first studied in normal kidney tissue and then compared to the expression in biopsy specimens of the above mentioned diseases, using light and electron microscopy. The expression was semiquantified by immuno- electron microscopy (iEM). Results Dendrin was localized solely in the podocytes close to the SD. There was no significant change in the total amount of Dendrin in MCNS compared to controls by immunofluorescence and iEM. Neph1 was also localized mainly to the SD. Double staining of Neph-1 and Nephrin showed the proteins in close connection in the SD. The total amount of Neph1 was significantly reduced in the glomerular diseases FSGS, MCNS and in ADR and PAN. The reduction of Neph1 was also seen in areas without FPE. Nephrin was reduced in MCNS and PAN but unchanged in FSGS. Conclusion In preserved slits and in areas without FPE in MCNS, the amounts of Dendrin were unchanged compared to controls. The redistribution might therefore be secondary to FPE. Neph1 co-localize with Nephrin in the SD and was reduced in FSGS, MCNS, ADR and PAN. Nephrin was however, unchanged in FSGS which could indicate a disruption of the Neph1-Nephrin complex and an involvement of Neph1 in the pathogenesis of this disease

Neph1 Is Reduced in Primary Focal Segmental Glomerulosclerosis, Minimal Change Nephrotic Syndrome, and Corresponding Experimental Animal Models of Adriamycin-Induced Nephropathy and Puromycin Aminonucleoside Nephrosis

Background/Aims: The transmembrane proteins Neph1 and nephrin form a complex in the slit diaphragm (SD) of podocytes. As recent studies indicate an involvement of this complex in the polymerization of the actin cytoskeleton and proteinuria, we wanted to study the subcellular localization of Neph1 in the normal human kidney and its expression in focal segmental glomerulosclerosis (FSGS), minimal change nephrotic syndrome (MCNS), and the corresponding experimental models of Adriamycin-induced nephropathy (ADR) and puromycin aminonucleoside nephrosis (PAN). All these disorders are characterized by substantial foot process effacement (FPE) and proteinuria. Materials and Methods: Kidney biopsies from patients with primary FSGS (perihilar type) and MCNS were compared to normal renal tissue. Mouse and rat kidney cortices from days 7 and 14 after Adriamycin injection and days 2 and 4 after puromycin aminonucleoside injection, respectively, were compared to control mouse and rat kidney. Polyclonal antibodies against Neph1 and nephrin were used for immunoelectron microscopy, and semiquantification was performed. Results: We localized Neph1 mainly to, and in close proximity to, the SD. Double staining of Neph1 and nephrin showed the proteins to be in close connection in the SD. The total amount of Neph1 in the podocytes was significantly reduced in FSGS, MCNS, ADR, and PAN. The reduction of Neph1 was also seen in areas with and without FPE. Nephrin was reduced in MCNS and PAN but unchanged in FSGS. Conclusion: With nephrin (but not Neph1) unchanged in FSGS, there might be a disruption of the complex and an involvement of Neph1 in its pathogenesis

The role of dendrin in IgA nephropathy

Background Immunoglobulin A nephropathy (IgAN) and its systemic variant IgA vasculitis (IgAV) damage the glomeruli, resulting in proteinuria, hematuria and kidney impairment. Dendrin is a podocyte-specific protein suggested to be involved in the pathogenesis of IgAN. Upon cell injury, dendrin translocates from the slit diaphragm to the nucleus, where it is suggested to induce apoptosis and cytoskeletal changes, resulting in proteinuria and accelerated disease progression in mice. Here we investigated gene and protein expression of dendrin in relation to clinical and histopathological findings to further elucidate its role in IgAN/IgAV. Methods Glomerular gene expression was measured using microarray on 30 IgAN/IgAV patients, 5 patients with membranous nephropathy (MN) and 20 deceased kidney donors. Dendrin was spatially evaluated on kidney tissue sections by immunofluorescence (IF) staining (IgAN patients, n = 4; nephrectomized kidneys, n = 3) and semi-quantified by immunogold electron microscopy (IgAN/IgAV patients, n = 21; MN, n = 5; living kidney donors, n = 6). Histopathological grading was performed according to the Oxford and Banff classifications. Clinical data were collected at the time of biopsy and follow-up. Results Dendrin mRNA levels were higher (P = .01) in IgAN patients compared with MN patients and controls and most prominently in patients with preserved kidney function and fewer chronic histopathological changes. Whereas IF staining did not differ between groups, immunoelectron microscopy revealed that a higher relative nuclear dendrin concentration in IgAN patients was associated with a slower annual progression rate and milder histopathological changes. Conclusion Dendrin messenger RNA levels and relative nuclear protein concentrations are increased and associated with a more benign phenotype and progression in IgAN/IgAV patients