Food-grade TiO2 is trapped by intestinal mucus in vitro but does not impair mucin O-glycosylation and short-chain fatty acid synthesis in vivo: implications for gut barrier protection

Abstract

BackgroundTitanium dioxide (TiO2) particles are commonly used as a food additive (E171 in the EU) for its whitening and opacifying properties. However, the risk of gut barrier disruption is an increasing concern because of the presence of a nano-sized fraction. Food-grade E171 may interact with mucus, a gut barrier protagonist still poorly explored in food nanotoxicology. To test this hypothesis, a comprehensive approach was performed to evaluate in vitro and in vivo interactions between TiO2 and intestinal mucus, by comparing food-grade E171 with NM-105 (Aeroxyde P25) OECD reference nanomaterial.ResultsWe tested E171-trapping properties of mucus in vitro using HT29-MTX intestinal epithelial cells. Time-lapse confocal laser scanning microscopy was performed without labeling to avoid modification of the particle surface. Near-UV irradiation of E171 TiO2 particles at 364 nm resulted in fluorescence emission in the visible range, with a maximum at 510 nm. The penetration of E171 TiO2 into the mucoid area of HT29-MTX cells was visualized in situ. One hour after exposure, TiO2 particles accumulated inside “patchy” regions 20 µm above the substratum. The structure of mucus produced by HT29-MTX cells was characterized by MUC5AC immunofluorescence staining. The mucus layer was thin and organized into regular “islands” located approximately 20 µm above the substratum. The region-specific trapping of food-grade TiO2 particles was attributed to this mucus patchy structure. We compared TiO2-mediated effects in vivo in rats after acute or sub-chronic oral daily administration of food-grade E171 and NM-105 at relevant exposure levels for humans. Cecal short-chain fatty acid profiles and gut mucin O-glycosylation patterns remained unchanged, irrespective of treatment.ConclusionsFood-grade TiO2 is trapped by intestinal mucus in vitro but does not affect mucin O-glycosylation and short-chain fatty acid synthesis in vivo, suggesting the absence of a mucus barrier impairment under “healthy gut” conditions.Electronic supplementary materialThe online version of this article (10.1186/s12951-018-0379-5) contains supplementary material, which is available to authorized users.

Jejunal villus absorption and paracellular tight junction permeability are major routes for early intestinal uptake of food-grade TiO2 particles: an in vivo and ex vivo study in mice

“…The E171 sample was obtained from a French commercial supplier of food coloring. The TiO 2 particle suspensions (> 95% anatase, [6]) were prepared in milliQ water following the generic Nanogenotox dispersion protocol [44], and the physicochemical properties of the particles in the E171 batch have been characterized in previous studies [6,7,45].…”

The food additive E171 and titanium dioxide nanoparticles indirectly alter the homeostasis of human intestinal epithelial cells in vitro

“…On top of HT29-MTX cells, TiO 2 particles would be trapped in mucus islands. 28 Conversely, on top of Caco-2 cells, which are not protected by mucus, agglomerates of TiO 2 can cause damage to epithelial cells, including disruption of the brush border, because of the mass of TiO 2 that settles down on the microvilli, 63 but also because of the movement of TiO 2 agglomerates in the vicinity of the microvilli, as previously demonstrated. 4 This bias could be avoided by using inverted cultures or microgravity reactors.…”

“…13 However, Talbot et al recently showed that the sub-chronic oral exposure of rats to E171 had little or no impact on the quantity and quality of intestinal mucus, and concluded that the protective function of mucus remained intact. 28 Therefore, this suggests that the reduced number of goblet cells reported by Urrutia-Ortega et al would have a limited physiological consequence for healthy persons.…”

“…57 Likewise, TiO 2 particles may trigger this stimulated secretion, as already shown in bronchial cells, where it was triggered by changes in cytosolic calcium concentration. 27 Since mucus secreted by HT29-MTX has recently been shown to trap TiO 2 particles, including E171, 28 increased mucus secretion would increase TiO 2 entrapment in the vicinity of cells. In similar experimental conditions, we previously showed that Ti intracellular accumulation is not significantly different in Caco-2 cells and in Caco-2/HT29-MTX cells.…”

“…27 However, a recent study rather showed that the sub-chronic oral exposure of rats to E171 had little or no impact on the quantity and quality of intestinal mucus. 28 Overall, the impact of TiO 2 on intestinal microbiota is limited. Waller et al demonstrated a modification of the microbial composition and biochemical responses of the microbiota after exposure to food-grade TiO 2 particles in an in vitro model human colon reactor.…”