Na+-K+-ATPase Enables Pathological B Cell Survival and Leads to Glomerular Basement Membrane (GBM) Thickening in Lupus Nephritis

Background: The kidney in lupus nephritis (LN) is characterized by a significant immune cell infiltrate and glomerular pathology. How lymphocytes lead to the characteristic damage of glomerular structures in LN is poorly understood. We recently published that intrarenal B lymphocytes upregulate Na+-K+-ATPase (NKA) in LN and that pharmacologic or genetic blockade of NKA results in fewer intrarenal B cells and reduced albuminuria, dependent on the genotype of the immune cell and not the host. As changes in kidney B cells were the only notable difference between control and NKA-inhibited mice, we hypothesize that kidney B cells affect glomerular basement membrane (GBM) structure and that mice with fewer kidney B cells would have preserved GBM and improved albuminuria. Methods: We compared lupus-prone MRLlpr and MRLlpr.Fxyd2-/- mice—which lack a gamma NKA subunit and have fewer intrarenal B cells—to define glomerular pathology. Podocyte and GBM pathology were investigated using immunofluorescence (IF), electron microscopy (EM) and real-time PCR. Results: We found that MRLlpr mice have proteinuria and thickened GBM on EM as compared to non-lupus B6 mice (277nm and 162nm, respectively, p<0.0001). Both proteinuria and GBM thickness were significantly improved in MRLlpr.Fxyd2-/- mice as compared to MRLlpr controls (200nm and 277nm, respectively, p<0.0001). We found immune complex deposits in both genotypes, with no notable differences on EM. IF showed homogeneously thickened collagen IV staining in MRLlpr as compared to MRLlpr.Fxyd2-/- mice (p<0.0001) while whole-kidney gene expression studies confirmed that the increased expression of glomerular collagen IV genes in MRLlpr was reduced in the MRLlpr.Fxyd2-/- mice. We are currently investigating collagen IV expression in contributing cell types (i.e. podocytes and endothelial cells) as well as looking at metalloproteinases to see if collagen breakdown is affected. Additionally, we are characterizing lupus kidney cytokine profiles to test the hypothesis that B cell-derived cytokines are promoting abnormal collagen deposition in LN. Conclusion: We show that lupus-prone MRLlpr mice have a significantly thickened GBM when compared to non-lupus and MRLlpr.Fxyd2-/- animals, likely due to abnormal collagen IV deposition. We are currently investigating whether these findings are due to differences in intrarenal B cell numbers and cytokine levels. Funding: NIDDK Support, Private Foundation Support