The Core-Clock Gene NR1D1 Impacts Cell Motility In Vitro and Invasiveness in a Zebrafish Xenograft Colon Cancer Model
Abstract
:1. Introduction
2. Results
2.1. The Knockdown of Core-Clock Genes Affects the Oscillatory Phenotype of HCT116 Cells In Vitro
2.2. The Downregulation of Core-Clock Genes Impacts Cell Proliferation, Apoptosis and Migration in CRC Cell Lines
2.3. Downregulation of BMAL1 and NR1D1 Impairs Tumorigenesis In Vivo
3. Discussion
4. Materials and Methods
4.1. Cell Culture
4.2. Lentivirus Production
4.3. Transduction with Lentiviral Vectors
4.4. Bioluminescence Measurements
4.5. Animal Care and Handling
4.6. Cell Labelling
4.7. Zebrafish Xenograft Microinjection
4.8. Micrometastasis Assay
4.9. Immunofluorescence
4.10. Imaging Processing and Analysis
4.11. RNA Extraction, cDNA Synthesis (Reverse Transcription) and Quantitative Real-Time PCR (qPCR)
4.12. Cell Cycle Assay
4.13. Proliferation, Apoptosis, and Migration Measurements
4.13.1. Proliferation Assays
4.13.2. Apoptosis Assays
4.13.3. Migration Assays
4.13.4. 3D Spheroid Assay
4.14. Rhythmicity Analysis of Time-Course Data
4.14.1. Harmonic Regression Using R
4.14.2. Cosinor Analysis
4.15. Statistical Analysis
4.16. Mathematical Modelling
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Basti, A.; Fior, R.; Yalҫin, M.; Póvoa, V.; Astaburuaga, R.; Li, Y.; Naderi, J.; Godinho Ferreira, M.; Relógio, A. The Core-Clock Gene NR1D1 Impacts Cell Motility In Vitro and Invasiveness in a Zebrafish Xenograft Colon Cancer Model. Cancers 2020, 12, 853. https://doi.org/10.3390/cancers12040853
Basti A, Fior R, Yalҫin M, Póvoa V, Astaburuaga R, Li Y, Naderi J, Godinho Ferreira M, Relógio A. The Core-Clock Gene NR1D1 Impacts Cell Motility In Vitro and Invasiveness in a Zebrafish Xenograft Colon Cancer Model. Cancers. 2020; 12(4):853. https://doi.org/10.3390/cancers12040853
Chicago/Turabian StyleBasti, Alireza, Rita Fior, Müge Yalҫin, Vanda Póvoa, Rosario Astaburuaga, Yin Li, Julian Naderi, Miguel Godinho Ferreira, and Angela Relógio. 2020. "The Core-Clock Gene NR1D1 Impacts Cell Motility In Vitro and Invasiveness in a Zebrafish Xenograft Colon Cancer Model" Cancers 12, no. 4: 853. https://doi.org/10.3390/cancers12040853