Anti-PLA2R1 IgG-mediated podocyte PLA2R1 cleavage drives glomerular subepithelial immune complex deposition in...

In membranous nephropathy (MN), subepithelial immune deposits and glomerular basement membrane (GBM) thickening are defining lesions, yet how circulating anti-phospholipase A2 receptor 1 (PLA2R1) IgG generates GBM-anchored complexes from podocyte antigen remains unclear.

To clarify this, human podocytes stably expressing PLA2R1 and HEK293 cells expressing full-length or antibody-binding-deficient PLA2R1 (ΔN1-9) were stimulated with PLA2R1-positive MN [MN(P+)] serum or purified IgG or the metalloproteinase activator phorbol 12-myristate 13-acetate (PMA). Metalloprotease/γ-secretase dependence was tested with pharmacologic inhibitors; specificity controls included soluble ectodomain (ECD) blockade, IgG depletion, purified anti-PLA2R1 IgG, and THSD7A as unrelated antigen. Kidney biopsies underwent dual immunofluorescence and dual label immunogold electron microscopy. Recombinant ECD binding to collagen IV/laminin was assayed in vitro.

MN(P+) serum or purified anti-PLA2R1 IgG increased the PLA2R1 C-terminal fragment and soluble ECD. These effects were blocked by metalloproteinase ADAM10/17 inhibitors and by preincubating IgG/serum with soluble ECD and were complement-independent (heat-inactivation). MN(P+) IgG enhanced shedding of full-length PLA2R1 but not ΔN1-9, whereas PMA increased shedding of both, indicating epitope-dependent, antibody-triggered cleavage. MN(P+) IgG did not cleave THSD7A. In biopsies, PLA2R1-ECD was enriched within GBM deposits in MN(P+) only while PLA2R1-intracellular domain remained podocyte-restricted across groups. Quantitative electron microscopy confirmed GBM-biased ECD distribution in MN(P+). In vitro, PLA2R1-ECD bound collagen IV/laminin and selectively captured MN(P+) IgG. Early MN showed focal GBM thickening at deposit sites.

Anti-PLA2R1 IgG accelerates ADAM10/17-dependent shedding of podocyte PLA2R1 releasing ECD anchors to GBM constituents. This mechanism reconciles the serum antibody glomerular antigen match and the preferential localization of deposits.