Syk inhibitor attenuates lupus in FcγRIIb-/- mice through the Inhibition of DNA extracellular traps from macrophages and neutrophils via p38MAPK-dependent pathway.

Abstract

Spleen tyrosine kinase (Syk), an important hub of immune signaling, is activated by several signalings in active lupus which could be interfered by Syk inhibitor but is still not completely evaluated in innate immune cells associated with lupus activity. Hence, a Syk inhibitor (fostamatinib; R788) was tested in vivo using Fc gamma receptor-deficient (FcγRIIb^-/-) lupus mice and in vitro (macrophages and neutrophils). After 4 weeks of oral Syk inhibitor, 40 week-old FcγRIIb^-/- mice (a full-blown lupus model) demonstrated less prominent lupus parameters (serum anti-dsDNA, proteinuria, and glomerulonephritis), systemic inflammation, as evaluated by serum TNFa, IL-6, and citrullinated histone H3 (CitH3), gut permeability defect, as indicated by serum FITC dextran assay, serum lipopolysaccharide (LPS), and serum (1 → 3)-β-D-glucan (BG), extracellular traps (ETs) and immune complex deposition in spleens and kidneys (immunofluorescent staining of CitH3 and immunoglobulin G) than FcγRIIb^-/- mice with placebo. Due to the spontaneous elevation of LPS and BG in serum, LPS plus BG (LPS + BG) was used to activate macrophages and neutrophils. After LPS + BG stimulation, FcγRIIb^-/- macrophages and neutrophils demonstrated predominant abundance of phosphorylated Syk (Western blotting), and the pro-inflammatory responses (CD86 flow cytometry analysis, supernatant cytokines, ETs immunofluorescent, and flow cytometry-based apoptosis). With RNA sequencing analysis and western blotting, the Syk-p38MAPK-dependent pathway was suggested as downregulating several inflammatory pathways in LPS + BG-activated FcγRIIb^-/- macrophages and neutrophils. Although both inhibitors against Syk and p38MAPK attenuated macrophage and neutrophil inflammatory responses against LPS + WGP, the apoptosis inhibition by p38MAPK inhibitor was not observed. These results suggested that Syk inhibitor (fostamatinib) improved the severity of lupus caused by FcγRIIb defect partly through Syk-p38MAPK anti-inflammation that inhibited both ET formation and cytokine production from innate immune cells.