Investigation of toxicity of a panel of food-grade nanomaterials on an in vitro triculture model of the mucus-secreting intestinal epithelium

I.S. Sohal, G. Deloid, K. O’Fallon, Ph. Demokritou, D. Bello
University of Massachusetts-Lowell,
United States

Keywords: food grade, nanomaterial, toxicity, triculture, epithelium


Background: Engineered nanomaterials (ENMs) have been increasingly used in food products to improve appearance, color, texture, for nutritional fortification, and in food preservation. Existing data on their safety assessment is limited and rather inconclusive. Several methodological aspects of the existing in vitro ingested, food-grade ENM (iENM) toxicity studies – (i) use of surrogate non-food grade nanomaterials; (ii) inadequate characterization of nanomaterials in dry powder form and in the test media; (iii) limited data on ENM biokinetics and dissolution in test media; (iv) lack of standardized dispersion protocols and in vitro dosimetry considerations; and (v) limited consideration on the use of realistic doses, could account for the disparity among the studies. In addition, most in vitro studies employ Caco-2 monocultures as a test model of the intestinal epithelium, but the intestinal mucosa is a more complex system. Objective: The current study investigates toxicity of representative iENMs in an in vitro triculture model representative of a mucus-secreting intestinal epithelium as well as a C2BBe1 monoculture, by carefully considering the aforementioned study design elements. Methods: Certified food-grade iENMs (titania, silica, zinc oxide and iron oxide) were acquired and extensively characterized in the dry state and all test media. Nanoparticle dispersions were prepared using a standard dispersion protocol [1]; dissolution kinetics of test nanomaterials was also examined in test cell culture media. A realistic working dose range was calculated based on reported estimates of human daily intake for respective iENMs, in conjunction with in vitro dosimetry estimates [2]. Results: We will present the effects of iENM on trans-epithelial electrical resistance (TEER), modifications in the brush border epithelium morphology and tight junction expression of the monoculture and triculture systems, mitochondrial activity, membrane damage, inflammatory cytokines/chemokines production, oxidative stress markers, and extracellular vesicle production. We will further compare and contrast the findings from the triculture system with the monoculture, as well as between various iENM.