Differences in Toxic Response Induced by Three Variants of the Diarrheic Shellfish Poisoning Phycotoxins in Human Intestinal Epithelial Caco-2 Cells
Abstract
:1. Introduction
2. Results
2.1. Cytotoxicity
2.2. Transcriptomic Profile
2.3. OA-, DTX-1- and DTX-2-Induced Gene Expression
2.4. Cell Cycle Arrest and MAP Kinase Pathway Quantified by High Content Analysis
3. Discussion
4. Materials and Methods
4.1. Chemicals
4.2. Cell Culture and Toxin Exposure
4.3. Cytotoxicity Evaluation by Neutral Red Uptake Assay
4.4. Microarray Experiments and Data Analysis
4.5. RT-qPCR
4.6. HCA of Multiparametric Toxicity Endpoints
4.7. Statistical Analysis
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
References
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Gene | Control | OA1 3.8 nM | OA 27.7 nM | OA 55.3 nM | DTX-1 3.5 nM | DTX-1 6.9 nM | DTX-1 13.8 nM | DTX-2 13.9 nM | DTX-2 27.8 nM | DTX-2 55.6 nM |
---|---|---|---|---|---|---|---|---|---|---|
ccnd1 | 0.18 ± 0.01 | 0.25 ± 0.03 | 0.35 ± 0.01 b | 0.31 ± 0.01 | 0.19 ± 0.01 | 0.24 ± 0.01 | 0.27 ± 0.01 | 0.20 ± 0.01 | 0.26 ± 0.01 | 0.26 ± 0.01 |
ccnd2 | 0.28 ± 0.02 | 0.32 ± 0.01 | 0.38 ± 0.08 | 0.09 ± 0.01 b | 0.26 ± 0.04 | 0.29 ± 0.03 | 0.26 ± 0.02 | 0.30 ± 0.02 | 0.27 ± 0.02 | 0.15 ± 0.05 |
rela | 0.99 ± 0.17 | 0.96 ± 0.05 | 0.96 ± 0.16 | 2.60 ± 0.26 c | 0.83 ± 0.13 | 0.86 ± 0.09 | 1.28 ± 0.17 | 0.86 ± 0.11 | 1.02 ± 0.13 | 1.44 ± 0.25 |
nras | 0.48 ± 0.05 | 0.48 ± 0.02 | 0.54 ± 0.07 | 0.86 ± 0.05 c | 0.39 ± 0.04 | 0.45 ± 0.03 | 0.51 ± 0.05 | 0.45 ± 0.04 | 0.44 ± 0.04 | 0.60 ± 0.08 |
map2k1 | 0.64 ± 0.12 | 0.46 ± 0.03 | 0.54 ± 0.12 | 0.93 ± 0.20 | 0.69 ± 0.07 | 0.59 ± 0.04 | 0.54 ± 0.02 | 0.48 ± 0.06 | 0.63 ± 0.09 | 1.13 ± 0.23 a |
mapk8 | 0.39 ± 0.02 | 0.34 ± 0.04 | 0.35 ± 0.04 | 0.52 ± 0.03 | 0.39 ± 0.03 | 0.41 ± 0.03 | 0.38 ± 0.05 | 0.33 ± 0.03 | 0.40 ± 0.05 | 0.48 ± 0.01 |
cdkn1b | 0.34 ± 0.04 | 0.30 ± 0.03 | 0.32 ± 0.05 | 0.55 ± 0.07 a | 0.38 ± 0.02 | 0.38 ± 0.04 | 0.44 ± 0.01 | 0.31 ± 0.03 | 0.42 ± 0.08 | 0.60 ± 0.06 b |
hras | 0.38 ± 0.03 | 0.47 ± 0.04 | 0.53 ± 0.04 | 0.62 ± 0.03 b | 0.39 ± 0.06 | 0.41 ± 0.05 | 0.52 ± 0.03 | 0.37 ± 0.02 | 0.46 ± 0.05 | 0.52 ± 0.05 |
rfc1 | 0.79 ± 0.16 | 0.66 ± 0.03 | 0.48 ± 0.06 a | 0.33 ± 0.01 c | 0.68 ± 0.05 | 0.58 ± 0.05 | 0.45 ± 0.03 b | 0.55 ± 0.06 | 0.45 ± 0.02 b | 0.45 ± 0.04 b |
rfc4 | 0.63 ± 0.05 | 0.53 ± 0.04 | 0.46 ± 0.05 | 0.20 ± 0.02 c | 0.56 ± 0.05 | 0.59 ± 0.07 | 0.35 ± 0.03 b | 0.51 ± 0.04 | 0.44 ± 0.05 | 0.37 ± 0.07 b |
rpa1 | 0.58 ± 0.05 | 0.53 ± 0.07 | 0.50 ± 0.02 | 0.23 ± 0.01 c | 0.49 ± 0.04 | 0.54 ± 0.02 | 0.41 ± 0.01 a | 0.52 ± 0.06 | 0.43 ± 0.04 | 0.38 ± 0.05 a |
rpa3 | 0.50 ± 0.03 | 0.49 ± 0.03 | 0.46 ± 0.05 | 0.25 ± 0.03 b | 0.55 ± 0.04 | 0.51 ± 0.02 | 0.41 ± 0.01 | 0.62 ± 0.05 | 0.53 ± 0.09 | 0.38 ± 0.06 |
OA | Control | 1.7 nM | 3.5 nM | 6.9 nM | 13.8 nM | 27.7 nM | 55.3 nM | 110.6 nM | 221.3 nM | 442.5 nM |
Cell count | 100 ± 0.0 | 94.2 ± 5.1 | 97.8 ± 6.0 | 86.6 ± 4.2 a | 73.2 ± 8.4 a | 48.8 ± 11.2 c | 22.8 ± 6.9 c | 16.3 ± 3.5 c | 11.2 ± 4.0 c | 6.8 ± 1.6 c |
Phospho-H3 | 2.0 ± 0.0 | 2.2 ± 0.2 | 2.2 ± 0.5 | 2.2 ± 0.5 | 3.2 ± 0.9 | 8.5 ± 1.8 | 13.2 ± 3.1 | 12.0 ± 3.6 | 14.5 ± 5.5 | 19.0 ± 6.3 b |
EdU | 5.7 ± 0.2 | 14.0 ± 1.2 b | 13.2 ± 2.7 a | 13.7 ± 2.2 b | 9.7 ± 2.4 | 7.2 ± 1.8 | 0.2 ± 0.2 | 0.5 ± 0.3 | 0.5 ± 0.5 | 0.0 ± 0.0 |
Phospho-MEK2 | 5.0 ± 0.0 | 5.2 ± 1.1 | 5.5 ± 1.6 | 5.0 ± 1.4 | 8.0 ± 2.7 | 6.0 ± 2.0 | 12.5 ± 3.3 | 20.0 ± 4.6 | 29.0 ± 8.3 b | 31.2 ± 9.2 b |
DTX1 | Control | 1.7 nM | 3.5 nM | 6.9 nM | 13.8 nM | 27.7 nM | 55.3 nM | 110.6 nM | 221.3 nM | 442.5 nM |
Cell count | 100 ± 0.0 | 106.2 ± 3.2 | 90.9 ± 5.9 | 75.5 ± 7.5 a | 46.5 ± 9.9 c | 23.0 ± 6.6 c | 19.2 ± 5.8 c | 21.1 ± 5.2 c | 15.4 ± 5.4 c | 7.8 ± 2.5 c |
Phospho-H3 | 2.0 ± 0.0 | 2.7 ± 0.3 | 3.8 ± 0.9 | 4.8 ± 1.6 | 8.3 ± 2.4 | 9.4 ± 4.2 | 8.6 ± 2.7 | 13.1 ± 6.5 | 21.5 ± 13.5 | 16.3 ± 9.2 |
EdU | 5.7 ± 0.2 | 9.2 ± 0.7 | 5.0 ± 2.7 | 9.4 ± 6.2 | 2.0 ± 1.4 | 0.2 ± 0.1 | 0.1 ± 0.1 | 0.2 ± 0.1 | 0.2 ± 0.2 | 0.0 ± 0.0 |
Phospho-MEK2 | 5.0 ± 0.0 | 9.8 ± 2.4 | 5.7 ± 1.2 | 9.0 ± 2.9 | 13.9 ± 4.6 | 16.8 ± 2.7 | 13.2 ± 4.1 | 18.3 ± 3.9 | 22.1 ± 10.7 | 42.4 ± 14.0 b |
DTX2 | Control | 1.7 nM | 3.5 nM | 7 nM | 13.9 nM | 27.8 nM | 55.6 nM | 111.3 nM | 222.5 nM | 445 nM |
Cell count | 100 ± 0.0 | 88.9 ± 7.1 | 84.3 ± 6.8 | 81.4 ± 5.0 b | 66.3 ± 9.1 b | 42.3 ± 8.9 c | 24.2 ± 6.9 c | 18.2 ± 4.7 c | 22.7 ± 6.4 c | 16.5 ± 4.2 c |
Phospho-H3 | 2.0 ± 0.0 | 3.0 ± 0.4 | 2.5 ± 0.1 | 2.5 ± 0.5 | 6.1 ± 3.1 | 7.8 ± 3.1 | 10.0 ± 3.3 | 13.0 ± 2.9 | 22.1 ± 10.4 a | 15.3 ± 7.2 |
EdU | 5.7 ± 0.2 | 4.1 ± 2.0 | 2.7 ± 1.5 | 9.1 ± 5.1 | 7.7 ± 5.9 | 0.7 ± 0.6 | 4.5 ± 4.1 | 3.5 ± 3.1 | 0.1 ± 0.1 | 0.1 ± 0.1 |
Phospho-MEK2 | 5.0 ± 0.0 | 4.3 ± 0.8 | 4.6 ± 1.3 | 5.2 ± 0.4 | 5.8 ± 2.0 | 7.7 ± 2.1 | 12.3 ± 3.9 | 14.1 ± 3.1 | 13.8 ± 4.3 | 19.2 ± 5.3 a |
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Huguet, A.; Drapeau, O.; Rousselet, F.; Quenault, H.; Fessard, V. Differences in Toxic Response Induced by Three Variants of the Diarrheic Shellfish Poisoning Phycotoxins in Human Intestinal Epithelial Caco-2 Cells. Toxins 2020, 12, 783. https://doi.org/10.3390/toxins12120783
Huguet A, Drapeau O, Rousselet F, Quenault H, Fessard V. Differences in Toxic Response Induced by Three Variants of the Diarrheic Shellfish Poisoning Phycotoxins in Human Intestinal Epithelial Caco-2 Cells. Toxins. 2020; 12(12):783. https://doi.org/10.3390/toxins12120783
Chicago/Turabian StyleHuguet, Antoine, Olivia Drapeau, Fanny Rousselet, Hélène Quenault, and Valérie Fessard. 2020. "Differences in Toxic Response Induced by Three Variants of the Diarrheic Shellfish Poisoning Phycotoxins in Human Intestinal Epithelial Caco-2 Cells" Toxins 12, no. 12: 783. https://doi.org/10.3390/toxins12120783