17α-Ethynylestradiol and Levonorgestrel Exposure of Rainbow Trout RTL-W1 Cells at 18 °C and 21 °C Mainly Reveals Thermal Tolerance, Absence of Estrogenic Effects, and Progestin-Induced Upregulation of Detoxification Genes
Abstract
:1. Introduction
2. Materials and Methods
2.1. RTL-W1 Cell Culture
2.2. Exposure to EDCs
2.3. RNA Extraction and cDNA Synthesis
2.4. Gene Expression Analysis
2.5. Statistical Analysis
3. Results
3.1. Cell Density and Viability
3.2. Gene Expression
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Gene * | Primer (5′-3′) | Protocol | E (%) | Reference |
---|---|---|---|---|
CYP1A | F: GATGTCAGTGGCAGCTTTGA R: TCCTGGTCATCATGGCTGTA | 95 °C—3 min (95 °C—20 s; 60.0 °C—20 s; 72 °C—20 s) 40× 95 °C—1 min | 97.7 | [60] |
CYP3A27 | F: GACGGTGGAGATCAACG R: GAGGATCTCGACCATGG | 95 °C—3 min (95 °C—20 s; 60.0 °C—20 s; 72 °C—20 s) 40× 95 °C—1 min | 99.0 | [60] |
HSP70b | F: AGGCCCAACCATTGAAGAGA R: GCAATGTCCAGCAATGCAATA | 95 °C—3 min (95 °C—10 s; 60.0 °C—30 s; 72 °C—30 s) 40× 95 °C—1 min | 92.2 | [61] |
UGT | F: ATAAGGACCGTCCCATCGAG R: ATCCAGTTGAGGTCGTGAGC | 94 °C—3 min (94 °C—20 s; 60.0 °C—20 s; 72 °C—20 s) 40× 94 °C—1 min | 94.9 | [60] |
GST | F: AGCTGCTCCCAGCTGATCC R: CAAACCACGGCCACATCATGTAATC | 94 °C—3 min (94 °C—20 s; 60.0 °C—20 s; 72 °C—20 s) 40× 94 °C—1 min | 93.9 | [62] |
CAT | F: CACTGATGAGGGCAACTGGG R: CTTGAAGTGGAACTTGCAG | 95 °C—3 min (95 °C—10 s; 58.0 °C—30 s; 72 °C—30 s) 40× 95 °C—30 s | 104.5 | [63] |
MRP2 | F: CCATTCTGTTCGCTGTCTCA R: CTCGTAGCAGGGTCTGGAAG | 94 °C—3 min (94 °C—20 s; 60.0 °C—20 s; 72 °C—20 s) 40× 94 °C—1 min | 100.2 | [60] |
ERα | F: GACATGCTCCTGGCCACTGT R: TGGCTTTGAGGCACACAAAC | 95 °C—4 min (95 °C—1 min; 61.6 °C—1 min) 40× 95 °C—1 min | 95.6 | [64] |
VtgA | F: AACGGTGCTGAATGTCCATAG R: ATTGAGATCCTTGCTCTTGGTC | 95 °C—3 min (95 °C—10 s; 62.9 °C—1 min) 40× 95 °C—30 s | 102.5 | [64] |
FAS | F: ACCGCCAAGCTCAGTGTGC R: CAGGCCCCAAAGGAGTAGC | 95 °C—3 min (95 °C—20 s; 60.0 °C—20 s; 72 °C—20 s) 40× 95 °C—1 min | 95.0 | [65] |
FABP1 | F: GTCCGTCACCAACTCCTTC R: GCGTCTCAACCATCTCTCC | 94 °C—3 min (94 °C—20 s; 57.0 °C—20 s; 72 °C—20 s) 40× 95 °C—1 min | 102.5 | [66] |
FATP1 | F: AGGAGAGAACGTCTCCACCA R: CGCATCACAGTCAAATGTCC | 95 °C—3 min (95 °C—20 s; 60.0 °C—20 s; 72 °C—20 s) 40× 95 °C—1 min | 92.9 | [67] |
β-act | F: TCTGGCATCACACCTTCTAC R: TTCTCCCTGTTGGCTTTGG | 94 °C—3 min (94 °C—20 s; 55.0 °C—20 s; 72 °C—20 s) 40× 94 °C—1 min | 99.7 | [66] |
ef1α | F: TGCCACACTGCTCACATC R: TCTCCAGACTTCAGGAACTTG | 94 °C—3 min (94 °C—20 s; 55.0 °C—20 s; 72 °C—20 s) 40× 94 °C—1 min | 97.4 | [68] |
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Vilaça, M.; Lopes, C.; Seabra, R.; Rocha, E. 17α-Ethynylestradiol and Levonorgestrel Exposure of Rainbow Trout RTL-W1 Cells at 18 °C and 21 °C Mainly Reveals Thermal Tolerance, Absence of Estrogenic Effects, and Progestin-Induced Upregulation of Detoxification Genes. Genes 2024, 15, 1189. https://doi.org/10.3390/genes15091189
Vilaça M, Lopes C, Seabra R, Rocha E. 17α-Ethynylestradiol and Levonorgestrel Exposure of Rainbow Trout RTL-W1 Cells at 18 °C and 21 °C Mainly Reveals Thermal Tolerance, Absence of Estrogenic Effects, and Progestin-Induced Upregulation of Detoxification Genes. Genes. 2024; 15(9):1189. https://doi.org/10.3390/genes15091189
Chicago/Turabian StyleVilaça, Margarida, Célia Lopes, Rosária Seabra, and Eduardo Rocha. 2024. "17α-Ethynylestradiol and Levonorgestrel Exposure of Rainbow Trout RTL-W1 Cells at 18 °C and 21 °C Mainly Reveals Thermal Tolerance, Absence of Estrogenic Effects, and Progestin-Induced Upregulation of Detoxification Genes" Genes 15, no. 9: 1189. https://doi.org/10.3390/genes15091189