Pro-inflammatory adjuvant properties of pigment-grade titanium dioxide particles are augmented by a genotype that potentiates interleukin 1β processing

Background: Pigment-grade titanium dioxide (TiO2) particles are an additive to some foods (E171 on ingredients lists), toothpastes, and pharma-/nutraceuticals and are absorbed, to some extent, in the human intestinal tract. TiO2 can act as a modest adjuvant in the secretion of the pro-inflammatory cytokine interleukin 1 beta (IL-1 beta) when triggered by common intestinal bacterial fragments, such as lipopolysaccharide (LPS) and/or peptidoglycan. Given the variance in human genotypes, which includes variance in genes related to IL-1 beta secretion, we investigated whether TiO2 particles might, in fact, be more potent pro-inflammatory adjuvants in cells that are genetically susceptible to IL-1 beta-related inflammation. Methods: We studied bone marrow-derived macrophages from mice with a mutation in the nucleotide-binding oligomerisation domain-containing 2 gene (Nod2(m/m)), which exhibit heightened secretion of IL-1 beta in response to the peptidoglycan fragment muramyl dipeptide (MDP). To ensure relevance to human exposure, TiO2 was food-grade anatase (119 +/- 45 nm mean diameter +/- standard deviation). We used a short 'pulse and chase' format: pulsing with LPS and chasing with TiO2 +/-MDP or peptidoglycan. Results: IL-1 beta secretion was not stimulated in LPS-pulsed bone marrow-derived macrophages, or by chasing with MDP, and only very modestly so by chasing with peptidoglycan. In all cases, however, IL-1 beta secretion was augmented by chasing with TiO2 in a dose-dependent fashion (5-100 mu g/mL). When co-administered with MDP or peptidoglycan, IL-1 beta secretion was further enhanced for the Nod2(m/m) genotype. Tumour necrosis factor a was triggered by LPS priming, and more so for the Nod2(m/m) genotype. This was enhanced by chasing with TiO2, MDP, or peptidoglycan, but there was no additive effect between the bacterial fragments and TiO2. Conclusion: Here, the doses of TiO2 that augmented bacterial fragment-induced IL-1 beta secretion were relatively high. In vivo, however, selected intestinal cells appear to be loaded with TiO2, so such high concentrations may be 'exposure-relevant' for localised regions of the intestine where both TiO2 and bacterial fragment uptake occurs. Moreover, this effect is enhanced in cells from Nod2(m/m) mice indicating that genotype can dictate inflammatory signalling in response to (nano) particle exposure. In vivo studies are now merited.