Peptidylarginine Deiminase 4 Promotes the Renal Infiltration of Neutrophils and Exacerbates the TLR7 Agonist-Induced Lupus Mice

Abstract

Peptidylarginine deiminase 4 (PAD4), encoded by PADI4, plays critical roles in the immune system; however, its contribution to the pathogenesis of lupus nephritis remains controversial. The pathological roles of PAD4 were investigated in lupus model mice. An imiquimod (IMQ)-induced lupus model was analyzed in wild-type (WT) and Padi4-knockout (KO) mice. Proteinuria, serum anti-double stranded DNA (anti-dsDNA) antibody, and renal infiltrated cells were evaluated. Neutrophil migration and adhesion were assessed using adoptive transfer and adhesion assay. PAD4-regulated pathways were identified by RNA-sequencing of Padi4 KO neutrophils. Padi4 KO mice exhibited significant improvements in proteinuria progression compared with WT mice, whereas, serum anti-dsDNA antibody and immune complex deposition in the glomeruli showed no difference between both mice strains. Padi4 KO mice showed decreased neutrophil infiltration in the kidneys. Adoptively transferred Padi4 KO neutrophils showed decreased migration to the kidneys of IMQ-treated WT mice, and adhesion to ICAM-1 was impaired in Padi4 KO neutrophils. Padi4 KO neutrophils exhibited reduced upregulation of p38 mitogen-activated protein kinase (MAPK) pathways. Toll-like receptor 7 (TLR7)-primed Padi4 KO neutrophils demonstrated reduced phosphorylation of p38 MAPK and lower expression of JNK-associated leucine zipper protein (JLP), a p38 MAPK scaffold protein. Neutrophils from heterozygous Jlp KO mice showed impaired adhesion to ICAM-1 and decreased migration to the kidneys of IMQ-treated WT mice. These results indicated a pivotal role of PAD4-p38 MAPK pathway in renal neutrophil infiltration in TLR7 agonist-induced lupus nephritis, and the importance of neutrophil-mediated kidney inflammation. Inhibition of the PAD4-p38 MAPK pathway may help in formulating a novel therapeutic strategy against lupus nephritis.

PAD4 controls chemoattractant production and neutrophil trafficking in malaria

“…Normal or slightly elevated CXCR2 expression levels were confirmed in bone marrow‐isolated neutrophils (Figures S3(C)–S3(F)). This is in agreement with studies showing normal in vitro migration capacity of PAD4−/− neutrophils 27 …”

“…In summary, PAD4 promotes malaria immunopathology by enhancing immune cell trafficking without affecting parasite sequestration patterns. This is consistent with findings in a lupus model, where renal neutrophil infiltration and pathology were also reduced in PAD4−/− mice 27 …”

“…To test if PAD4 controls chemokine production in human cells, we treated monocyte‐derived macrophages ( n = 5–6 donors) with GSK484, a PAD4 pharmacologic inhibitor. As with mouse macrophages, PAD4 inhibition significantly reduced CXCL1 (Figure 4(A)) and CXCL2 (Figure 4(B)) secretion in TLR7‐stimulated human macrophages (Figure 4(A)), consistent with a reported role for PAD4 in TLR7 responses in lupus 4,27 . We obtained similar results with human monocytes (Figure S4(D)).…”

“…In summary, PAD4 promotes malaria immunopathology by enhancing immune cell trafficking without affecting parasite sequestration patterns. This is consistent with findings in a lupus model, where renal neutrophil infiltration and pathology were also reduced in PAD4−/− mice 27. To understand the mechanistic basis of the observed reduction in neutrophil accumulation in the liver, we analyzed abundance ofF I G U R E 2 PAD4 promotes trafficking of neutrophils into infected livers (A) Representative immunofluorescence image of neutrophils (green) in livers (DAPI, blue) at peak parasitemia; (B) quantification of (A), n = 3 noninfected and 5 infected mice; (C) flow cytometric enumeration of neutrophils in blood, n = 9-10 animals, 3 independent experiments.…”

“…The reduction in immunopathology in PAD4−/− mice is likely to be caused by a combination of reduced granulopoiesis, impaired chemokine production as well as previously described neutrophilintrinsic defects. The latter include dampened p38 MAPK signaling in response to TLR7 ligands 27 as well as altered immunostimulatory properties of NETs. 10 Several mechanisms could explain the effect of PAD4 inhibition on chemokine production.…”