Impact of copper oxide nanomaterials on differentiated and undifferentiated Caco-2 intestinal epithelial cells; assessment of cytotoxicity, barrier integrity, cytokine production and nanomaterial penetration

Abstract

BackgroundCopper oxide nanomaterials (CuO NMs) are exploited in a diverse array of products including antimicrobials, inks, cosmetics, textiles and food contact materials. There is therefore a need to assess the toxicity of CuO NMs to the gastrointestinal (GI) tract since exposure could occur via direct oral ingestion, mucocillary clearance (following inhalation) or hand to mouth contact.MethodsUndifferentiated Caco-2 intestinal cells were exposed to CuO NMs (10 nm) at concentrations ranging from 0.37 to 78.13 μg/cm2 Cu (equivalent to 1.95 to 250 μg/ml) and cell viability assessed 24 h post exposure using the alamar blue assay. The benchmark dose (BMD 20), determined using PROAST software, was identified as 4.44 μg/cm2 for CuO NMs, and 4.25 μg/cm2 for copper sulphate (CuSO4), which informed the selection of concentrations for further studies. The differentiation status of cells and the impact of CuO NMs and CuSO4 on the integrity of the differentiated Caco-2 cell monolayer were assessed by measurement of trans-epithelial electrical resistance (TEER), staining for Zonula occludens-1 (ZO-1) and imaging of cell morphology using scanning electron microscopy (SEM). The impact of CuO NMs and CuSO4 on the viability of differentiated cells was performed via assessment of cell number (DAPI staining), and visualisation of cell morphology (light microscopy). Interleukin-8 (IL-8) production by undifferentiated and differentiated Caco-2 cells following exposure to CuO NMs and CuSO4 was determined using an ELISA. The copper concentration in the cell lysate, apical and basolateral compartments were measured with Inductive Coupled Plasma Optical Emission Spectrometry (ICP-OES) and used to calculate the apparent permeability coefficient (Papp); a measure of barrier permeability to CuO NMs. For all experiments, CuSO4 was used as an ionic control.ResultsCuO NMs and CuSO4 caused a concentration dependent decrease in cell viability in undifferentiated cells. CuO NMs and CuSO4 translocated across the differentiated Caco-2 cell monolayer. CuO NM mediated IL-8 production was over 2-fold higher in undifferentiated cells. A reduction in cell viability in differentiated cells was not responsible for the lower level of cytokine production observed. Both CuO NMs and CuSO4 decreased TEER values to a similar extent, and caused tight junction dysfunction (ZO-1 staining), suggesting that barrier integrity was disrupted.ConclusionsCuO NMs and CuSO4 stimulated IL-8 production by Caco-2 cells, decreased barrier integrity and thereby increased the Papp and translocation of Cu. There was no significant enhancement in potency of the CuO NMs compared to CuSO4. Differentiated Caco-2 cells were identified as a powerful model to assess the impacts of ingested NMs on the GI tract.Electronic supplementary materialThe online version of this article (doi:10.1186/s12989-017-0211-7) contains supplementary material, which is available to authorized users.

Toxicological evaluation of metal oxide nanoparticles and mixed exposures at low doses using zebra fish and THP1 cell line

“…IL8 leads to the recruitment of neutrophils to the site of damage and, therefore, implying that CuO nanomaterials lead to cellular damage in cells. In previous studies, the level of IL8 was found to increase twofolds when Caco 2 cells were treated with CuO NP . No significant effect on IL8 was observed in case of cerium oxide NP and the mixture of these NP.…”

The Nano–Intestine Interaction: Understanding the Location‐Oriented Effects of Engineered Nanomaterials in the Intestine

“…[58,59] Apart from silver nanomaterials, CuO and ZnO nanomaterials also exhibited similar oxidative stress effects that induced cytotoxicity and damage to the ultrastructure of the intestine (Figure 5b). [60,61] Recently, Fe 2 O 3 NMs have been shown to disrupt the integ rity of intestinal barrier through affecting the expression of gammacatenin, Zona occludens1, and other integrity associated proteins in a sizedependent manner, in the order of 17 nm > 53 and 100 nm. [62] Consistent with this finding, another in vivo study has shown intestinal absorption of par ticles to be size dependent, with smaller particles (polystyrene microspheres and colloidal gold) being absorbed to a greater degree than larger ones.…”

“…For example, CuO NMs showed totally different behavior on the morphology of both differentiated and undifferentiated monolayer models of Caco2 cells (Figure 5c). [60] In spite of their adverse effects, NMs also show some benefits on designing test platform for nanotoxicity. .…”

Copper Oxide Nanoparticles Stimulate Cytotoxicity and Apoptosis in Glial Cancer Cell Line

“…5 CuO NPs are able to generate oxidative stress and significantly decrease cell viability in tumor cells. [16][17][18] Glioma is a type of brain and central nervous system tumors, which starts in supportive glial cells that surround nerve cells and help them function. 19 Cell division is a normal process to build new cells and tissues but it is stopped whenever there is no longer need for more cells or is a threat for the organisms to be destroyed by apoptotic cell death.…”