In Vitro Genotoxicity Evaluation of PAHs in Mixtures Using Experimental Design
Abstract
:1. Introduction
2. Materials and Methods
2.1. Chemicals and Mixture Preparation
2.2. Cellular Culture
2.3. Cytokinesis-Block Micronucleus Assay
2.4. Cytokinesis-Block Micronucleus Assay
2.5. Experimental Factors and Domain of Interest
2.6. Determination of Genotoxic Equivalent Factors
3. Results and Discussion
3.1. Cytostasis
3.2. DNA Damage–Comet Assay
3.3. Chromosomal Damage–CBMN Assay
3.4. Determination of Genotoxic Equivalent Factors
4. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Variable | Factors | Principal Effects | Comet Assay | Micronucleus Assay | ||
---|---|---|---|---|---|---|
Xi | bi | Level − | Level + | Level − | Level + | |
X1 | Naphthalene | b1 | 1 | 10 | 1 | 10 |
X2 | Phenanthrene | b2 | 1 | 10 | 1 | 10 |
X3 | Pyrene | b3 | 1 | 10 | 1 | 10 |
X4 | Benzo(b)fluoranthene | b4 | 0.1 | 1 | 0.2 | 2 |
X5 | Benzo(a)pyrene | b5 | 0.0025 | 1 | 0.0025 | 2 |
X6 | Dibenz(a,h)anthracene | b6 | 0.1 | 1 | 0.2 | 2 |
Experiment | Nap | Phe | Pyr | BbF | BaP | dBahA | TEQ | %Increase in Tail DNA |
---|---|---|---|---|---|---|---|---|
1_1 | 1 | 1 | 1 | 0.1 | 0.0025 | 0.1 | 0.12 | 75.41 |
1_2 | 1 | 1 | 1 | 0.1 | 0.0025 | 0.1 | 0.12 | 55.16 |
1_3 | 1 | 1 | 1 | 0.1 | 0.0025 | 0.1 | 0.12 | 84.94 |
2 | 1 | 10 | 10 | 1 | 1 | 1 | 2.12 | 172.36 |
3 | 10 | 1 | 10 | 1 | 1 | 1 | 2.12 | 122.65 |
4 | 10 | 10 | 1 | 1 | 1 | 1 | 2.12 | 244.26 |
5 | 10 | 10 | 10 | 0.1 | 1 | 1 | 2.04 | 141.01 |
6 | 10 | 10 | 10 | 1 | 0.0025 | 1 | 1.13 | 109.41 |
7 | 10 | 10 | 10 | 1 | 1 | 0.1 | 1.23 | 118.05 |
8 | 10 | 10 | 1 | 0.1 | 0.0025 | 0.1 | 0.13 | 104.17 |
9 | 10 | 1 | 10 | 0.1 | 0.0025 | 0.1 | 0.13 | 117.40 |
10 | 10 | 1 | 1 | 1 | 0.0025 | 0.1 | 0.21 | 61.90 |
11 | 10 | 1 | 1 | 0.1 | 1 | 0.1 | 1.12 | 87.87 |
12 | 10 | 1 | 1 | 0.1 | 0.0025 | 1 | 1.02 | 60.52 |
13 | 1 | 10 | 10 | 0.1 | 0.0025 | 0.1 | 0.13 | 71.06 |
14 | 1 | 10 | 1 | 1 | 0.0025 | 0.1 | 0.21 | 73.10 |
15 | 1 | 10 | 1 | 0.1 | 1 | 0.1 | 1.12 | 92.09 |
16 | 1 | 10 | 1 | 0.1 | 0.0025 | 1 | 1.02 | 82.46 |
17 | 1 | 1 | 10 | 1 | 0.0025 | 0.1 | 0.21 | 60.65 |
18 | 1 | 1 | 10 | 0.1 | 1 | 0.1 | 1.12 | 77.90 |
19 | 1 | 1 | 10 | 0.1 | 0.0025 | 1 | 1.02 | 71.00 |
20 | 1 | 1 | 1 | 1 | 1 | 0.1 | 1.20 | 150.34 |
21 | 1 | 1 | 1 | 1 | 0.0025 | 1 | 1.11 | 78.98 |
22 | 1 | 1 | 1 | 0.1 | 1 | 1 | 2.01 | 80.99 |
Experiment | Nap | Phe | Pyr | BbF | BaP | dBahA | TEQ | %Cytostasis | χ2 |
---|---|---|---|---|---|---|---|---|---|
1_1 | 1 | 1 | 1 | 0.2 | 0.0025 | 0.2 | 0.23 | 6.67 | 4.27 |
1_2 | 1 | 1 | 1 | 0.2 | 0.0025 | 0.2 | 0.23 | −0.63 | 6.08 |
1_3 | 1 | 1 | 1 | 0.2 | 0.0025 | 0.2 | 0.23 | 2.20 | 4.76 |
2 | 1 | 10 | 10 | 2 | 2 | 2 | 4.22 | 45.11 | 39.69 |
3 | 10 | 1 | 10 | 2 | 2 | 2 | 4.22 | 34.92 | 50.78 |
4 | 10 | 10 | 1 | 2 | 2 | 2 | 4.22 | 31.68 | 56.01 |
5 | 10 | 10 | 10 | 0.2 | 2 | 2 | 4.05 | 27.60 | 5.01 |
6 | 10 | 10 | 10 | 2 | 0.0025 | 2 | 2.23 | 4.31 | 0.57 |
7 | 10 | 10 | 10 | 2 | 2 | 0.2 | 2.43 | 40.96 | 14.44 |
8 | 10 | 10 | 1 | 0.2 | 0.0025 | 0.2 | 0.24 | 9.96 | 8.58 |
9 | 10 | 1 | 10 | 0.2 | 0.0025 | 0.2 | 0.24 | 13.99 | 6.27 |
10 | 10 | 1 | 1 | 2 | 0.0025 | 0.2 | 0.41 | 12.59 | 1.26 |
11 | 10 | 1 | 1 | 0.2 | 2 | 0.2 | 2.23 | 26.18 | 0.62 |
12 | 10 | 1 | 1 | 0.2 | 0.0025 | 2 | 2.03 | 18.31 | 2.35 |
13 | 1 | 10 | 10 | 0.2 | 0.0025 | 0.2 | 0.24 | 14.99 | 8.49 |
14 | 1 | 10 | 1 | 2 | 0.0025 | 0.2 | 0.41 | 11.45 | 18.33 |
15 | 1 | 10 | 1 | 0.2 | 2 | 0.2 | 2.23 | 38.46 | 35.05 |
16 | 1 | 10 | 1 | 0.2 | 0.0025 | 2 | 2.03 | 10.74 | 29.60 |
17 | 1 | 1 | 10 | 2 | 0.0025 | 0.2 | 0.41 | 14.30 | 1.31 |
18 | 1 | 1 | 10 | 0.2 | 2 | 0.2 | 2.23 | 34.99 | 15.53 |
19 | 1 | 1 | 10 | 0.2 | 0.0025 | 2 | 2.03 | 11.30 | 30.72 |
20 | 1 | 1 | 1 | 2 | 2 | 0.2 | 2.4 | 43.09 | 26.86 |
21 | 1 | 1 | 1 | 2 | 0.0025 | 2 | 2.21 | 16.79 | 36.53 |
22 | 1 | 1 | 1 | 0.2 | 2 | 2 | 4.02 | 39.14 | 64.79 |
Single PAH | PAH in Mixtures | ||||
---|---|---|---|---|---|
PAH | TEF | GEF (γ H2AX) | GEF (CBMN Assay) | GEF (Comet-CBMN Assays) | Mean GEF for PAH Mixtures |
from [22] | from [31] | from Present Study | |||
Nap | 0.001 | 0 | 0.10 | 0.06–0 | 0.03 |
Phe | 0.001 | 0 | 0.01 | 0.08–0.03 | 0.05 |
Pyr | 0.001 | 0 | 0.01 | 0.02–0 | 0.01 |
BbF | 0.1 | [0.69–1.46] | 0.22 | 0.74–0.48 | 0.61 |
dBahA | 1 | [0.5–2.77] | 1.13 | 0.61–1.24 | 0.93 |
BaP | 1 | 1.00 | 1.00 | 1.00 | 1.00 |
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Castel, R.; Tassistro, V.; Claeys-Bruno, M.; Malleret, L.; Orsière, T. In Vitro Genotoxicity Evaluation of PAHs in Mixtures Using Experimental Design. Toxics 2023, 11, 470. https://doi.org/10.3390/toxics11050470
Castel R, Tassistro V, Claeys-Bruno M, Malleret L, Orsière T. In Vitro Genotoxicity Evaluation of PAHs in Mixtures Using Experimental Design. Toxics. 2023; 11(5):470. https://doi.org/10.3390/toxics11050470
Chicago/Turabian StyleCastel, Rebecca, Virginie Tassistro, Magalie Claeys-Bruno, Laure Malleret, and Thierry Orsière. 2023. "In Vitro Genotoxicity Evaluation of PAHs in Mixtures Using Experimental Design" Toxics 11, no. 5: 470. https://doi.org/10.3390/toxics11050470