CXCL4 is a novel nickel‐binding protein and augments nickel allergy

Summary Background Nickel (Ni) is the most frequent metal allergen and induces a TH1‐dependent type‐IV allergy. Although Ni2+ is considered to bind to endogenous proteins, it currently remains unclear whether these Ni‐binding proteins are involved in Ni allergy in vivo. We previously reported the adjuvant effects of lipopolysaccharide (LPS) in a Ni allergy mouse model. As LPS induces a number of inflammatory mediators, we hypothesized that Ni‐binding protein(s) are also induced by LPS. Objective The objective of this study was to purify and identify Ni‐binding protein(s) from serum taken from LPS‐injected mice (referred as LPS serum) and examined the augmenting effects of these Ni‐binding protein(s) on Ni allergy in an in vivo model. Methods BALB/cA mice were sensitized with an i.p. injection of NiCl2 and LPS. Ten days after sensitization, mice were challenged with NiCl2 by an i.d. injection into ear pinnae. Ni‐binding protein(s) were purified by Ni‐affinity column chromatography and gel filtration. Results Lipopolysaccharide serum, but not serum taken from saline‐injected mice, augmented ear swelling induced by Ni‐allergic inflammation. Ni‐binding, but not non‐binding fraction, purified from LPS serum augmented Ni‐allergic inflammation. Mass spectrometry and Western blotting detected CXCL4 in the active fraction. A batch analysis with Ni‐sepharose and a surface plasmon resonance analysis revealed direct binding between CXCL4 and Ni2+. Recombinant CXCL4 augmented Ni‐allergic inflammation and exerted adjuvant effects at the sensitization phase. Conclusions These results indicate that CXCL4 is a novel Ni‐binding protein that augments Ni allergy at the elicitation and sensitization phases. This is the first study to demonstrate that the Ni‐binding protein augments Ni allergy in vivo.