Explore the vast range of titles available.

Background/Aims: It is still unclear what happens in the glomerulus when proteinuria starts. Using puromycin aminonucleoside nephrosis (PAN) rats, we studied early ultrastructural and permeability changes in relation to the expression of the podocyte-associated molecules nephrin, α-actinin, dendrin, and plekhh2, the last two of which were only recently discovered in podocytes. Methods: Using immune stainings, semiquantitative measurement was performed under the electron microscope. Permeability was assessed using isolated kidney perfusion with tracers. Possible effects of ACE inhibition were tested. Results: By day 2, some patchy foot process effacement, but no proteinuria, appeared. The amount of nephrin was reduced in both diseased and normal areas. The other proteins showed few changes, which were limited to diseased areas. By day 4, foot process effacement was complete and proteinuria appeared in parallel with signs of size barrier damage. Nephrin decreased further, while dendrin and plekhh2 also decreased but α-actinin remained unchanged. ACE inhibition had no significant protective effect. Conclusions: PAN glomeruli already showed significant pathology by day 4, despite relatively mild proteinuria. This was preceded by altered nephrin expression, supporting its pivotal role in podocyte morphology. The novel proteins dendrin and plekhh2 were both reduced, suggesting roles in PAN, whereas α-actinin was unchanged. Copyright © 2010 S. Karger AG, Basel [PUBLICATION ABSTRACT]

Pleckstrin homology domain–containing, family H (with MyTH4 domain), member 2 (Plekhh2) is a 1491-residue intracellular protein highly enriched in renal glomerular podocytes for which no function has been ascribed. Analysis of renal biopsies from patients with focal segmental glomerulosclerosis revealed a significant reduction in total podocyte Plekhh2 expression compared to controls. Sequence analysis indicated a putative α-helical coiled-coil segment as the only recognizable domain within the N-terminal half of the polypeptide, while the C-terminal half contains two PH, a MyTH4, and a FERM domain. We identified a phosphatidylinositol-3-phosphate consensus-binding site in the PH1 domain required for Plekhh2 localization to peripheral regions of cell lamellipodia. The N-terminal half of Plekkh2 is not necessary for lamellipodial targeting but mediates self-association. Yeast two-hybrid screening showed that Plekhh2 directly interacts through its FERM domain with the focal adhesion protein Hic-5 and actin. Plekhh2 and Hic-5 coprecipitated and colocalized at the soles of podocyte foot processes in situ and Hic-5 partially relocated from focal adhesions to lamellipodia in Plekhh2-expressing podocytes. In addition, Plekhh2 stabilizes the cortical actin cytoskeleton by attenuating actin depolymerization. Our findings suggest a structural and functional role for Plekhh2 in the podocyte foot processes.

Background: The podocytes are injured in many acquired glomerular diseases. This may present as proteinuria or morphological changes such as foot process effacement (FPE) or loss of podocytes in the urine. The exact mechanisms are still mainly unknown. However, a number of proteins identified in the slit diaphragm (SD) of the podocytes, including Dendrin and Neph1, are believed to be of significance in the podocytes’ response to injury and in the resulting pathophysiological development. Dendrin was primarily found to be associated to the actin cytoskeleton in mouse podocytes. Neph1 is a transmembrane protein that together with Nephrin forms a complex in the SD, involved in polymerization of the actin cytoskeleton and proteinuria. Plekhh2 is an uncharacterized protein that we localized to the podocyte cytoplasm.Aim: The principal aim was to study the expression and role of novel structural glomerular proteins in acquired human glomerular disease. In the first study, focusing on the expression of Dendrin in normal human kidney and in the glomerular disease Minimal Change Nephrotic Syndrome (MCNS), light and immune electron microscopy (iEM) was used. In the second study, the subcellular localization of the uncharacterized protein Plekhh2 in normal human kidney and Focal Segmental Glomerulosclerosis (FSGS) was investigated by immunofluorescence (IFL) and iEM. Neph1 and Nephrin were studied in FSGS, MCNS and in the corresponding experimental models Adriamycin nephropathy (ADR) and puromycin aminonucleoside nephrosis (PAN). Lastly, we returned to Dendrin, and studied the expression in the podocyte nuclei in IgA Nephropathy (IgAN) and Membranous Nephropathy (MN) with iEM and analyzed the gene expression with microarray.Results: Dendrin was localized to the glomerular slit diaphragm (SD). There was no significant change in the amount of Dendrin in MCNS compared to controls by IFL and iEM. In areas of FPE Dendrin was redistributed from the SD to the podocyte cytoplasm. Plekhh2 was localized mainly to the podocyte cytoplasm. The expression was reduced in FSGS. Double staining of Neph-1 and Nephrin showed the proteins in close connection in the SD. Neph1 was significantly reduced in FSGS, MCNS, ADR and PAN. The reduction of Neph1 was also seen in areas without FPE. Nephrin was unchanged in FSGS, but reduced in MCNS and PAN. Dendrin was increased in the podocyte nuclei in IgAN, with a corresponding two fold increase of the gene expression. Both protein and gene expression of Dendrin was unchanged in MN.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, per se. However, the increase of nuclear Dendrin in IgAN suggests an upregulated apoptotic pathway and a possible role for Dendrin in the pathogenesis of this disease. Plekhh2 was reduced and relocated from the plasma membrane to centrally in the foot processes in FSGS. Neph1 was reduced in FSGS, MCNS, ADR and PAN, in contrast to Nephrin which was unchanged in FSGS. This could indicate a disruption of the Neph1-Nephrin complex and an involvement of Neph1 in the pathogenesis of this disease.

BackgroundIn 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.AimWe 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.MethodsThe 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 immunoelectron microscopy (iEM).ResultsDendrin 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.ConclusionIn 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.