Radiation-Induced Senescence Reprograms Secretory and Metabolic Pathways in Colon Cancer HCT-116 Cells
Abstract
:1. Introduction
2. Results
2.1. HCT-116 Cells Exhibit Cellular Senescence Phenotype Following Ionizing Radiation
2.2. IR Dose and Time Dependent Senescence Responses by HCT-116 Cells
2.3. Detection of Senescence-Associated Proteins, P53 and p21 by Immunoblot Analysis
2.4. Secretion of Cytokines and Chemokines by Senescent HCT-116 (SCC) Cell Cultures
2.5. Secretion of Extracellular Matrix Proteins, MMPs and TIMPs, and Angiogenic and Growth Factors by Senescent HCT-116 (SCC) Cell Cultures
2.6. Analysis of Metabolites in Control and Senescent HCT-116 Cells (SCC)
2.7. Adenylate, Guanylate Energy Charge and Creatine Levels in Senescent HCT-116 Cells (SCC)
2.8. Redox System Metabolites, NAD, NADP and Glutathione in Senescent HCT-116 Cells (SCC)
2.9. Aerobic Glycolysis with Accumulation of Pyruvate and Lactate in Senescent HCT-116 Cells (SCC)
2.10. TCA (Citric Acid) Cycle Metabolites in Senescent HCT-116 Cells
2.11. Pentose Phosphate Pathway Metabolites in Senescent HCT-116 Cells (SCC)
2.12. Levels of Amino Acids Decreased Significantly in Senescent HCT-116 Cells (SCC)
2.13. Polyamine Metabolites Are Differentially Regulated in Senescent HCT-116 Cells (SCC)
3. Discussion
4. Material & Methods
4.1. Cell Cultures
4.2. Radiation Treatments
4.3. Senescence-Associated Beta Galactosidase (SA-β Gal) Detection
4.4. Protein Extraction and Quantitation
4.5. Western Immunoblot Analysis of Proteins
4.6. Radiation Treatments and Collection of Culture Supernatant Samples
4.7. Multiplex Luminescence Analysis of Secreted Proteins in Culture Supernatants
4.8. Preparation of Proliferating (Control) and Senescent Cells for Metabolite Extraction
4.9. Analysis of Intracellular Metabolites by CE-TOFMS and CE-QqQMS
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
CRC | Colorectal cancer |
ECM | Extracellular matrix CRC |
HCT-116 | Human colon cancer cells |
SCC | Senescent cancer cells |
SCCVII | Mouse squamous cell carcinoma cells |
IR | Ionizing radiation |
cGy | cGray units, radiation intensity |
SA- β-Gal | Senescence-associated β-galactosidase |
NAD | Nicotinamide adenine dinucleotide |
NADH | Nicotinamide adenine dinucleotide hydride |
TCA | Tricarboxylic acid cycle (citric acid cycle) |
PPP | Pentose phosphate pathway |
Aldolase | Fructose, 1, 6 biphosphate aldolase |
PRPP | 5-Phospho-D ribose 1-pyrophosphoric acid |
VEGF | Vascular endothelial growth factor |
MCT | Monocarboxylate transporter |
SASP | Senescence-associated secretory phenotype |
TGF | Transforming growth factor |
PDGF | Platelet derived growth factor |
IL | Interleukin |
MMPs | Matrix metalloproteases |
TIMPs | Tissue inhibitors of MMPs |
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Nagineni, C.N.; Naz, S.; Choudhuri, R.; Chandramouli, G.V.R.; Krishna, M.C.; Brender, J.R.; Cook, J.A.; Mitchell, J.B. Radiation-Induced Senescence Reprograms Secretory and Metabolic Pathways in Colon Cancer HCT-116 Cells. Int. J. Mol. Sci. 2021, 22, 4835. https://doi.org/10.3390/ijms22094835
Nagineni CN, Naz S, Choudhuri R, Chandramouli GVR, Krishna MC, Brender JR, Cook JA, Mitchell JB. Radiation-Induced Senescence Reprograms Secretory and Metabolic Pathways in Colon Cancer HCT-116 Cells. International Journal of Molecular Sciences. 2021; 22(9):4835. https://doi.org/10.3390/ijms22094835
Chicago/Turabian StyleNagineni, Chandrasekharam N., Sarwat Naz, Rajani Choudhuri, Gadisetti V. R. Chandramouli, Murali C. Krishna, Jeffrey R. Brender, John A. Cook, and James B. Mitchell. 2021. "Radiation-Induced Senescence Reprograms Secretory and Metabolic Pathways in Colon Cancer HCT-116 Cells" International Journal of Molecular Sciences 22, no. 9: 4835. https://doi.org/10.3390/ijms22094835