Relationships between p53 status, apoptosis and induction of micronuclei in different human and mouse cell lines in vitro: Implications for improving existing assays

Whitwell, James; Smith, Robert; Jenner, Karen; Lyon, Heather; Wood, D.J.; Clements, Julie; Aschcroft-Hawley, Kelly; Gollapudi, B. Bhaskar; Kirkland, David; Lorge, Elisabeth; Pfuhler, Stefan; Tanir, Jennifer Y.; Thybaud, Véronique

doi:10.1016/j.mrgentox.2015.05.011

Cited by 43 publications

(27 citation statements)

References 33 publications

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“…However, these assays are time‐consuming and therefore not suitable for high throughput screening. What is more, they are performed with cell lines derived from mice (L5178Y) or hamster (CHO, AS52 and V79), which are mainly p53 defective and of little relevance for humans as well as being devoid of biotransformation properties (Whitwell et al, ; Rees et al, ). For this reason, we chose the HepG2 cells, the same cell model we used for the γH2AX assay, to perform the in vitro PIG‐A gene mutation assay originally developed with the TK6 cell line (Krüger et al, ; ; Nicklas et al, ; Rees et al, ) and more recently on the L5178Y cell line (Bemis et al, ; David et al, ; Wang et al, ).…”

Section: Discussionmentioning

confidence: 99%

Genotoxicity and mutagenicity assessment of food contaminant mixtures present in the French diet

Kopp

Vignard

Mirey

et al. 2018

Environ and Mol Mutagen

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Through diet, people are exposed simultaneously to a variety of contaminants (e.g. heavy metals, mycotoxins, pesticides) that could have combined adverse effects on human health. A previous study identified six main mixtures of food contaminants to which French adult consumers are exposed. These complex mixtures are comprised of 11 to 19 chemicals that have numerous toxic properties. In the present study, we investigated the genotoxic effects of these food contaminants, as single molecules and in mixtures that reflect their occurrence in the French diet, using the γH2AX assay in two human cell lines (HepG2, LS-174 T). Results of detailed analysis of the 49 individual contaminants (including 21 tested in this study) demonstrated a positive genotoxic response to 14 contaminants in HepG2 and 12 in LS-174 T cells. Next, our results indicated that two mixtures out of six triggered significant γH2AX induction after 24 hr of treatment, at concentrations for which individual compounds did not induce any DNA damage, suggesting more than additive interactions between chemicals. γH2AX positive mixtures were then tested for mutagenicity with the innovative in vitro PIG-A assay in HepG2 cells coupled with the soft agar colony formation assay. The two γH2AX positive mixtures led to a significant increase in the frequency of PIG-A GPI-deficient cells and in the number of colonies formed in soft agar. In conclusion, our study demonstrates that two mixtures of contaminants present in the French diet induce genotoxicity and mutagenicity, and that the combined effects of single molecules present in these mixtures are likely not additive, highlighting potential problems for hazard assessment of mixtures. Environ. Mol. Mutagen. 59:742-754, 2018. © 2018 Wiley Periodicals, Inc.

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Section: Discussionmentioning

confidence: 99%

Genotoxicity and mutagenicity assessment of food contaminant mixtures present in the French diet

Kopp

Vignard

Mirey

et al. 2018

Environ and Mol Mutagen

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“…Therefore the advantages offered by the use of human cell lines or primary nonhuman cells are often overshadowed by incongruous gene expression making them nonreliable or irrelevant. 9,16,[31][32][33] For instance, considerably higher expression of P450 and UGTIA1 enzymes and considerably lower expression of uptake transporters were documented for corneal tissues reconstructed from immortalized human corneal epithelial cells, thus limiting their utility for drug absorption studies. 34 Recent success in the isolation and expansion of primary human corneal epithelial cells has allowed for a reliable and reproducible source of primary human corneal epithelial cells for the development of novel organotypic corneal tissue models.…”

mentioning

confidence: 99%

New Human Organotypic Corneal Tissue Model for Ophthalmic Drug Delivery Studies

Kaluzhny

Kinuthia

Truong

et al. 2018

Invest. Ophthalmol. Vis. Sci.

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PURPOSE. The purpose of the current work was to develop a physiologically relevant, in vitro human three-dimensional (3D) corneal epithelial tissue model for use in ophthalmic drug development.METHODS. Normal human corneal epithelial cells were cultured at the air-liquid interface to produce the 3D corneal tissue model. Corneal barrier was determined by measuring transepithelial electrical resistance (TEER). Quantitative PCR arrays were utilized to investigate expression of 84 phase I/II metabolizing enzymes and 84 drug transporter genes. Permeability was evaluated using model compounds with a wide range of hydrophobicity, molecular weight, and excipients. Finally, different formulations of latanoprost and bimatoprost were administered and drug absorption and tissue viability and integrity were investigated.RESULTS. Histologic assessment and TEER of the corneal tissue model revealed tissue structure, thickness, and barrier formation (1000 6 146 XÁcm 2 ) comparable to native human corneal epithelium. The 3D corneal tissue expressed tight junctions, mucins, and key corneal epithelial detoxification enzymes. Drug-metabolizing enzyme and transporter gene expression in 3D corneal tissue and excised human corneal epithelium were highly correlated (r 2 ¼ 0.87). Coefficients of permeation for model drugs in the tissue model and excised rabbit corneas also showed a high correlation (r 2 ¼ 0.94). As expected, latanoprost and bimatoprost free acids had much lower permeability (Papp ¼ 1.2 3 10 À6 and 1.9 3 10 À6 ) than the corresponding prodrugs (Papp ¼ 2.5 3 10 À5 and 5.6 3 10 À5 ), respectively. The presence of 0.02% benzalkonium chloride in ophthalmic formulations significantly affected tissue barrier and viability.CONCLUSIONS. The newly developed 3D corneal tissue model appears to be very useful for evaluation of corneal drug permeability and safety during ophthalmic drug development.

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“…A number of studies are available on in vitro micronucleus (MN) assay in different rodent (V79, CHL, CHO and L5178Y) and human cell lines, p53 competent and p53 mutated, in the presence and absence of cytochalasin B (Fowler et al., ,b; Whitwell et al., ). Ethyl acrylate was tested at different concentrations up to 55% of cytotoxicity for 3 h followed by recovery and for 24 h without recovery.…”

Section: Procedures Of the Safety Assessmentmentioning

confidence: 99%

“…In contrast, small but statistically significant increases in MN frequency were observed at the highest two concentrations analysed following treatment of L5178Y cells. Significant increases in caspase activity (greater than threefold over the vehicle control) associated with high level of toxicity were observed in TK6 cells after treatment with ethyl acrylate, suggesting apoptosis as a mechanism of induction of micronuclei frequencies (Fowler et al., ,b; Whitwell et al., ).…”

Section: Procedures Of the Safety Assessmentmentioning

confidence: 99%

Safety of ethyl acrylate to be used as flavouring

Silano¹,

Bolognesi²,

Castle³

et al. 2017

EFS2

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JECFA (2006) and EFSA (2010). Moreover, new data on the use levels of ethyl acrylate as flavouring substance have been provided. For use as flavouring substance, the chronic dietary exposure estimated using the added portions exposure technique (APET), is calculated to be 3,545 lg/person per day for a 60-kg adult and 2,233 lg/person per day for a 15-kg 3-year-old child. Exposure from food contact materials may be up to 6,000 lg/person per day. The Panel considered that based on the available data, which covers all relevant genetic endpoints (i.e. gene mutations, structural and numerical chromosomal aberrations) there is no concern with respect to genotoxicity of ethyl acrylate. The Panel evaluated the available carcinogenicity studies conducted in rats and mice and agreed with the NTP evaluation (1998) concluding that the forestomach squamous cell papilloma and carcinoma observed in rodents were not relevant to humans. Additionally, there was no evidence of systemic toxicity in short-term and subchronic toxicity studies. Therefore, the Panel concluded that there is no safety concern for the use of ethyl acrylate as a flavouring substance, under the intended conditions of use.

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Relationships between p53 status, apoptosis and induction of micronuclei in different human and mouse cell lines in vitro: Implications for improving existing assays

Cited by 43 publications

References 33 publications

Genotoxicity and mutagenicity assessment of food contaminant mixtures present in the French diet

Genotoxicity and mutagenicity assessment of food contaminant mixtures present in the French diet

New Human Organotypic Corneal Tissue Model for Ophthalmic Drug Delivery Studies

Safety of ethyl acrylate to be used as flavouring

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