Genetic Inactivation of Peroxiredoxin-I Impairs the Growth of Human Pancreatic Cancer Cells
Abstract
:1. Introduction
2. Materials and Methods
2.1. Mice and Ethical Approval
2.2. Cell Lines, Small Interfering RNA (siRNA) Transfections and Pharmacological Treatments
2.3. Cell Counting and Crystal Violet Staining
2.4. Cell Cycle Analysis
2.5. Reactive Oxygen Species (ROS) Measurements
2.6. Western Blot
2.7. Immunofluorescence and Immunohistochemistry
2.8. Detection of 8-Oxoguanine (8-oxoG) Adducts
2.9. Statistical Analysis
3. Results and Discussion
3.1. PRX-I Expression Is Induced during Pancreatic Carcinogenesis
3.2. Genetic Ablation of PRX-I Induces a Prooxidant Response Leading to Cell Cycle Blockade and Reduced Growth of Human PDAC Cells
3.3. Genetic Ablation of PRX-I Alters the Autophagic Flux in PDAC Cells
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
8-oxoG | 8-oxoguanine |
ACC | Acetyl-CoA carboxylase |
AMPK | AMP-activated protein kinase |
ATG | Autophagy-related gene |
Cdc25C | M-phase inducer phosphatase 3 |
CHK2 | Checkpoint kinase 2 |
ERK/MAPK | Extracellular signal-regulated kinases/mitogen-activated protein kinases |
HRAS | Harvey rat sarcoma viral oncogene homolog protein |
HSC70 | Heat-shock cognate 70 |
JAK/STAT3 | Janus kinase/signal transducer and activator of transcription 3 |
Kras | Kirsten rat sarcoma gene |
KRAS | Kirsten rat sarcoma protein |
LAMP1 | Lysosomal-associated membrane protein 1 |
LC3 | Microtubule-associated protein 1A/1B-light chain |
NRF2 | Nuclear factor erythroid 2-related factor 2 gene |
NRF2 | Nuclear factor erythroid 2-related factor 2 protein |
pH3 | Phospho-histone 3 |
PRDX1 | Human Peroxiredoxin-1 gene |
PRX-I | Peroxiredoxin-I protein |
PRX-II | Peroxiredoxin-II protein |
ROS | Reactive oxygen species |
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Antibody | Reference | Dilution | Conditions |
---|---|---|---|
PRX-I | NBP1-82558, BioTechne Minneapolis, MN, USA | 1/1000 | 5% Milk, overnight, 4 °C |
PRX-II | Sc-515428, SCB, Heidelberg, Germany | 1/500 | 5% Milk, overnight, 4 °C |
P-STAT3Y705 | 9145S, CST, Leiden, The Netherlands | 1/1000 | 5% BSA, overnight, 4 °C |
STAT3 | 12640S, CST, Leiden, The Netherlands | 1/1000 | 5% BSA, overnight, 4 °C |
P-CHK2T68 | 2197S, CST, Leiden, The Netherlands | 1/1000 | 5% BSA, overnight, 4 °C |
CHK2 | 2662S, CST, Leiden, The Netherlands | 1/1000 | 5% BSA, overnight, 4 °C |
P-Cdc25CS216 | 4901S, CST, Leiden, The Netherlands | 1/1000 | 5% BSA, overnight, 4 °C |
p27Kip1 | 3686S, CST, Leiden, The Netherlands | 1/1000 | 5% BSA, overnight, 4 °C |
LC3 | 12741S, CST, Leiden, The Netherlands | 1/1000 | 5% BSA, overnight, 4 °C |
ATG5-ATG12 | 4180T, CST, Leiden, The Netherlands | 1/1000 | 5% BSA, overnight, 4 °C |
ATG7 | 8558T, CST, Leiden, The Netherlands | 1/1000 | 5% BSA, overnight, 4 °C |
ATG16L1 | 8089T, CST, Leiden, The Netherlands | 1/1000 | 5% BSA, overnight, 4 °C |
p62 | Sc-28359, SCB, Heidelberg, Germany | 1/500 | 5% Milk, overnight, 4 °C |
HSC70 | Sc-7298, SCB, Heidelberg, Germany | 1/5000 | 5% Milk, overnight, 4 °C |
β-Actin | A5441-100UL, SA, Overijse, Belgium | 1/5000 | 5% Milk, overnight, 4 °C |
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Dahou, H.; Minati, M.-A.; Jacquemin, P.; Assi, M. Genetic Inactivation of Peroxiredoxin-I Impairs the Growth of Human Pancreatic Cancer Cells. Antioxidants 2021, 10, 570. https://doi.org/10.3390/antiox10040570
Dahou H, Minati M-A, Jacquemin P, Assi M. Genetic Inactivation of Peroxiredoxin-I Impairs the Growth of Human Pancreatic Cancer Cells. Antioxidants. 2021; 10(4):570. https://doi.org/10.3390/antiox10040570
Chicago/Turabian StyleDahou, Hajar, Marie-Albane Minati, Patrick Jacquemin, and Mohamad Assi. 2021. "Genetic Inactivation of Peroxiredoxin-I Impairs the Growth of Human Pancreatic Cancer Cells" Antioxidants 10, no. 4: 570. https://doi.org/10.3390/antiox10040570