In the normal kidney, an enormous volume of blood (180 L) is filtered each day through the glomerular filtration barrier. By even the most conservative estimates, between 1 - 3 g of albumin a day make it through the filtration barrier. Podocytes are a key part of the GFB and the molecular basis of protein trafficking in podocytes is unknown. In preliminary studies, we have found that podocytes actively transcytose two major serum proteins, albumin and immunoglobulin G (IgG), and that transcytosis is the major pathway for handling these proteins. We have also found that trafficking of IgG is more complicated than that of albumin and that podocytes direct trafficking of IgG differently depending on whether IgG is coupled to antigen. In other cell types such as renal tubular cells and endothelial cells, the neonatal Fc receptor (FcRn) is required for transcytosis of albumin and IgG. In dendritic cells, FcRn is required for antigen/IgG (immune complex) trafficking to the lysosome for antigen presentation on MHC Class II. Based on our preliminary studies and the known role of FcRn in other cell types we propose a dual role for FcRn in podocytes. Specifically, we hypothesize that (1) FcRn is required for transcytosis of albumin and IgG through the podocyte and (2) FcRn is required for immune complex presentation in podocytes by directing antigen/IgG complexes to the lysosome for processing and presentation in podocyte MHC II. In this proposal, we will use primary podocytes isolated from wild type and FcRn knockout mice to determine whether FcRn is required for transcytosis of albumin and IgG. We will also determine whether the glomerular sieving coefficient for albumin and IgG is significantly less in wild type versus FcRn knockout mice and whether podocyte-specific knockout of FcRn in vivo leads to protein accumulation within the glomerulus and glomerulosclerosis. We will examine whether FcRn is required to direct antigen/IgG complexes to the lysosome for degradation and antigen presentation on podocyte MHC Class II and determine whether lack of FcRn abrogates antigen/IgG presentation in podocytes. Finally, we will determine whether podocyte-specific knockout of FcRn ameliorates disease in a mouse model of immune-mediated nephritis. We will also use intravital multiphoton microscopy, a dynamic advanced imaging technique, to examine whether podocyte-specific knockout of FcRn reduces T retention times within the glomerulus after induction of immune-mediated nephritis. The work outlined in this proposal is significant because it is the initial step in understanding exactly how serum proteins are trafficked through the podocyte. Protein accumulation in podocytes has been shown to be toxic to podocytes and correlates with podocyte loss and glomerulosclerosis. Knowledge at the molecular level of how proteins are handled by the podocyte will form the basis for the design of rational therapies to prevent protein accumulation in podocytes. An understanding of the dual role of FcRn in podocytes (both in transcytosis and antigen presentation) is also critical to the design of safe and effective therapies to treat immune-mediated glomerular diseases.
This project examines whether the neonatal Fc receptor (FcRn) is required for trafficking of the major serum proteins albumin and immunoglobulin G (IgG) through podocytes which are a critical component of the glomerular filtration barrier. This proposal also determines whether FcRn directs trafficking of IgG differently based on whether it is coupled to antigen and whether FcRn is required for immune complex presentation and disease progression in a model of immune-mediated kidney disease. Results of this work will elucidate the molecular mechanisms underlying protein trafficking in podocytes and allow for the design of rational therapies to prevent deleterious protein accumulation in podocytes and strategies to treat immune-mediated kidney diseases.
