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Abstract: PO1976

Knockout of the Neonatal Fc Receptor (FcRn) Alters Lysosomal Function in Podocytes

Session Information

  • Podocyte Biology
    October 22, 2020 | Location: On-Demand
    Abstract Time: 10:00 AM - 12:00 PM

Category: Glomerular Diseases

  • 1204 Podocyte Biology

Authors

  • Blaine, Judith, University of Colorado Denver School of Medicine, Aurora, Colorado, United States
  • Ng, Madeline D., University of Colorado Denver School of Medicine, Aurora, Colorado, United States
  • Tonsawan, Pantipa, Khon Kaen University, Khon Kaen, Thailand
  • Lewis, Linda, University of Colorado Denver School of Medicine, Aurora, Colorado, United States
  • Dylewski, James F., University of Colorado Denver School of Medicine, Aurora, Colorado, United States
Background

FcRn is a trafficking protein that diverts monomeric IgG from the lysosome but sorts multimeric IgG and immune complexes (ICs) to the lysosome for processing. FcRn is required in dendritic cells to traffic ICs to the lysosome for proteolytic processing and presentation on MHC II. Podocytes express FcRn and previous work had proposed that podocytes can act as antigen presenting cells. Here we show that cultured podocytes are weak antigen presenting cells (APCs) and that knockout (KO) of FcRn in podocytes does not alter podocyte response to an immune stimulus but does result in significant downregulation of lysosomal function.

Methods

Cultured wild type (WT) and FcRn KO podocytes were treated with interferon gamma (IFNg) to simulate proinflammatory conditions. MHC II and costimulatory marker expression was assessed by flow cytometry. Antigen presentation was evaluated by examining T cell response when WT or FcRn KO podocytes were treated with ICs and used as APCs. Lysosomal size and cellular location in WT and FcRn KO podocytes were examined using confocal microscopy. WT or FcRn KO podocytes were treated with ICs and colocalization of lysosomes and ICs was quantitated. RNA-seq was used to examine lysosomal pathways.

Results

Both WT and FcRn KO podocytes upregulated MHC II after treatment with IFNg but there was no difference in expression levels between WT and KO. There was no change in CD80 expression between WT and KO after treatment with IFNg. CD86 and ICOSL expression levels in WT and FcRn KO were minimal at baseline and after treatment with IFNg. When used as APCs, WT podocytes induced a very modest amount of IL-2 production by T cells ( a marker of T cell activation) whereas KO podocytes induced none. After treatment with ICs, lysosomes in WT podocytes were significantly larger and were also clustered around the nucleus, indicating lysosomal activation. In contrast, after IC treatment lysosomes in the KO were smaller and more peripherally located. Treatment with ICs also resulted in significantly greater colocalization between lysosomes and ICs in WT versus FcRn KO podocytes, demonstrating that ICs were not directed to the lysosome in the KO. RNA-seq showed significant downregulation of lysosomal pathways in KO podocytes compared to WT after treatment with ICs.

Conclusion

FcRn KO in podocytes alters lysosomal trafficking and function.

Funding

  • NIDDK Support