Myeloid IPMK promotes the resolution of serum transfer-induced arthritis in mice

Abstract

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by widespread joint inflammation, which leads to joint damage, disability, and mortality. Among the several types of immune cells, myeloid cells such as macrophages are critical for controlling the pathogenesis of RA. Inositol phosphates are water-soluble signaling molecules, which are synthesized by a series of enzymes including inositol phosphate kinases. Previous studies revealed actions of inositol phosphates and their metabolic enzymes in the modulation of inflammation such as Toll-like receptor-triggered innate immunity. However, the physiological roles of inositol polyphosphate (IP) metabolism in the regulation of RA remain largely uncharacterized. Therefore, our study sought to determine the role of inositol polyphosphate multikinase (IPMK), a key enzyme for IP metabolism and various cellular signaling control mechanisms, in mediating RA. Using myeloid cell-specific IPMK knockout (KO) mice, arthritis was induced via intraperitoneal K/BxN serum injection, after which disease severity was evaluated. Both wild-type and IPMK KO mice developed similar RA phenotypes; however, conditional deletion of IPMK in myeloid cells led to elevated arthritis scores during the resolution phase, suggesting that IPMK deficiency in myeloid cells impairs the resolution of inflammation. Bone marrow-derived IPMK KO macrophages exhibited no apparent defects in immunoglobulin Fc receptor (FcR) activation, osteoclast differentiation, or resolvin signaling. Taken together, our findings suggest that myeloid IPMK is a key determinant of RA resolution.

Keywords: IPMK; Rheumatoid arthritis; inflammation; inositol polyphosphate; resolution.

Figure 1.

Myeloid IPMK inhibits the severity of inflammatory arthritis. (A) and (B) K/BxN serum-transfer arthritis was induced in wild-type (Ipmk^WT) (n = 14) or myeloid-specific IPMK KO (Ipmk^ΔLysM) mice (n = 10) by intraperitoneal injection of K/BxN serum on day 0 and 2. Arthritis score (A) and paw thickness (B) were assessed. Data are shown as mean ± standard error of the mean (SEM). *p < 0.05, **p < 0.01.

Figure 2.

Myeloid IPMK reduces synovitis, pannus formation, and joint erosion. (A) Representative images of H&E staining of ankle joints from Ipmk^ΔLysM or Ipmk^WT mice in K/BxN serum-transfer arthritis (original magnification X20). (B) Histologic scores of synovitis, pannus formation, and bone erosion of ankle joints. Data are shown as mean ± SEM. *p < 0.05, **p < 0.01.

Figure 3.

Macrophage IPMK does not impair FcR signaling. mRNA expression of the proinflammatory cytokines Tnf-α (A), Il-1β (B), and Il-6 (C) was quantified by RT-qPCR in Ipmk^ΔLysM and Ipmk^WT BMDMs treated with HAGG (25 μg/ml) on designated time point. Data are shown as means ± SEM (n = 3). *p < 0.05, **p < 0.01. (D) Phosphorylation of NF-κB and JNK was analyzed from Ipmk^ΔLysM and Ipmk^WT BMDM lysates treated with HAGG (25 μg/ml) by immunoblotting. Images shown are representative of experiments performed twice.

Figure 4.

Myeloid IPMK deletion does not alter osteoclast differentiation. mRNA expression of the markers for osteoclast differentiation Trap, Ctsk, H⁺-ATPase was quantified by RT-qPCR in Ipmk^ΔLysM and Ipmk^WT BMDMs after 5 days treatment with M-CSF (50 ng/ml) and RANKL (100 ng/ml). Data are shown as means ± SEM (n = 3). *p < 0.05, **p < 0.01, ***p < 0.001.

Figure 5.

Macrophage IPMK is dispensable for resolvin D1 signaling. (A) Phosphorylation of Akt and ERK was analyzed by immunoblotting. Lysates of Ipmk^ΔLysM and Ipmk^WT BMDMs were prepared by resolvin D1 (50 nM) treatment for designated timepoint. (B) All blots are representative of four independent experiments. Densitometric quantitation results were normalized to ethanol-treated controls. Data are shown as means ± SEM (n = 4). *p < 0.05.