Conceptual Background and Bioenergetic/Mitochondrial Aspects of Oncometabolism

1st Edition, Volume 542 - May 23, 2014
Editors: Lorenzo Galluzzi, Guido Kroemer
Language: English
Hardback ISBN:
9 7 8 - 0 - 1 2 - 4 1 6 6 1 8 - 9
eBook ISBN:
9 7 8 - 0 - 1 2 - 4 1 6 6 5 7 - 8

Volume 542 of Methods in Enzymology continues the legacy of this premier serial with quality chapters authored by leaders in the field. This new volume covers research method… Read more

Conceptual Background and Bioenergetic/Mitochondrial Aspects of Oncometabolism

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Volume 542 of Methods in Enzymology continues the legacy of this premier serial with quality chapters authored by leaders in the field. This new volume covers research methods providing a theoretical overview on metabolic alterations of cancer cells and a series of protocols that can be employed to study oncometabolism, in vitro, ex vivo and in vivo. Malignant cells exhibit metabolic changes when compared to their normal counterparts, owing to both genetic and epigenetic alterations. Although such a metabolic rewiring has recently been indicated as "yet another" general hallmark of cancer, accumulating evidence suggests that the metabolic alterations of each neoplasm rather represent a molecular signature that intimately accompanies, and hence cannot be severed from, all facets of malignant transformation.

Preface
- Acknowledgments
Chapter One: The Metabolic Alterations of Cancer Cells
- Abstract
- 1 Introduction
- 2 Aerobic Glycolysis: A Hallmark of Cancer Cell Metabolism
- 3 Mitochondrial Metabolism in Cancer Cells
- 4 Lipid Metabolism in Cancer Cells
- 5 Future Perspectives: Altered Metabolism Shapes Tumor Evolution
- 6 Conclusions
Chapter Two: Autophagy and Cancer Metabolism
- Abstract
- 1 Introduction
- 2 Overview of the Autophagy Machinery
- 3 Metabolic Stimuli Regulating Autophagy
- 4 Autophagy and Tumor Suppression
- 5 Tumor-Promoting Functions of Autophagy
- 6 Conclusion
- Acknowledgments
Chapter Three: Regulation of Cancer Metabolism by Oncogenes and Tumor Suppressors
- Abstract
- 1 Introduction
- 2 HIF-1: Regulator of Hypoxic Responses and Cancer Metabolism
- 3 The PI3K–AKT–PTEN Pathway Regulates Metabolism
- 4 mTOR Controls Anabolism and It Is Inhibited By AMPK Upon Metabolic Stress
- 5 c-Myc Promotes Aerobic Anabolism
- 6 Ras Stimulates Glycolysis and the PPP
- 7 NF-kappaB Regulates Inflammation and Proliferation But Also Metabolism
- 8 Retinoblastoma: Suppressing Tumorogenesis and Anabolism
- 9 p53 Regulates Multiple Metabolic Pathways
- 10 Conclusions
- Acknowledgments
Chapter Four: Cross Talk Between Cell Death Regulation and Metabolism
- Abstract
- 1 Introduction
- 2 Signaling to Programmed Cell Death
- 3 Bioenergetic Regulation of Programmed Cell Death
- 4 Redox Signaling and ROS: Critical Regulators of Programmed Cell Death
- 5 Destructive ROS During the Execution of Cell Death
- 6 Conclusions
- Acknowledgments
Chapter Five: Techniques to Monitor Glycolysis
- Abstract
- 1 Introduction
- 2 Measuring Glucose Uptake and Lactate Production
- 3 Measuring the Activity of Rate-Limiting Glycolytic Enzymes
- 4 Metabolite Measurements and Glucose Tracing
- 5 Summary
- Acknowledgments
Chapter Six: Measurement of Enolase Activity in Cell Lysates
- Abstract
- 1 Introduction
- 2 Assay of Enolase Activity
- 3 DEAE-Cellulose Chromatography
Chapter Seven: Extracellular Flux Analysis to Monitor Glycolytic Rates and Mitochondrial Oxygen Consumption
- Abstract
- 1 Introduction
- 2 Techniques to Evaluate Immune Cell Bioenergetics
- 3 Bioenergetic Profiling of Immune Cells Using the XF Extracellular Flux Analyzer
- 4 Summary
Chapter Eight: Conventional Techniques to Monitor Mitochondrial Oxygen Consumption
- Abstract
- 1 Introduction
- 2 Principles of Cell Respiration Measurements
- 3 Conventional Measurement of Oxygen Consumption
Chapter Nine: Use of Safranin for the Assessment of Mitochondrial Membrane Potential by High-Resolution Respirometry and Fluorometry
- Abstract
- 1 Introduction
- 2 Preparation of Mitochondrial Thom
- 3 HRR and Determination of mtMP by Safranin
- 4 Limitations of Safranin as mtMP Indicator
- 5 Respirometry with Homogenate
- 6 Optimization of Conditions for Use of Safranin
- 7 An Application Example
- 8 Conclusions
Chapter Ten: Kinetic Analysis of Local Oxygenation and Respiratory Responses of Mammalian Cells Using Intracellular Oxygen-Sensitive Probes and Time-Resolved Fluorometry
- Abstract
- 1 Introduction
- 2 Experimental Design and Protocols
- Acknowledgments
Chapter Eleven: Cell-Based Measurements of Mitochondrial Function in Human Subjects
- Abstract
- 1 Introduction
- 2 Blood Lymphocyte Protocol
- 3 Skeletal Muscle Myoblast Protocol
- 4 Data Analysis
- Acknowledgments
Chapter Twelve: Use of Chemical Probes to Detect Mitochondrial ROS by Flow Cytometry and Spectrofluorometry
- Abstract
- 1 Introduction
- 2 Mitochondrial ROS Production
- 3 Methods to Detect ROS Production
- 4 Concluding Remarks
- Acknowledgments
Chapter Thirteen: Methods to Monitor ROS Production by Fluorescence Microscopy and Fluorometry
- Abstract
- 1 Introduction
- 2 Measurement of Reactive Oxygen Species Production in Intact Adherent Cells Using Fluorescent Probes
- 3 Quantification of Protein Level with the Use of the SRB Assay
- 4 Measurement of Mitochondrial ROS Production in Permeabilized Cells with the Amplex Red Fluorescent Probe
- 5 Measurement of ROS by Fluorescence Microscopy
- Acknowledgments
Chapter Fourteen: Genetically Encoded Redox Sensors
- Abstract
- 1 Introduction
- 2 Endogenously Encoded Redox Sensors
- 3 Engineered Genetically Encoded Redox Sensors
- 4 Oxidative Stress in Cancer and Potential Applications of Redox Sensors in Oncology Research
- 5 Limitations and Future of Genetically Encoded Redox Sensors
- 6 Summary
Chapter Fifteen: Use of Genetically Encoded Sensors to Monitor Cytosolic ATP/ADP Ratio in Living Cells
- Abstract
- 1 Introduction
- 2 Experimental Components and Considerations
- 3 Typical Experiment: [ATP/ADP]_cyt in Pancreatic β-Cells
- 4 Summary
- Acknowledgments
Chapter Sixteen: Methods to Monitor and Compare Mitochondrial and Glycolytic ATP Production
- Abstract
- 1 Introduction
- 2 Monitoring Intracellular ATP Using Luciferase-Based Techniques
- 3 Monitoring Intracellular ATP Using Alternative Enzymatic Methods
- 4 Summary
- Acknowledgments
Chapter Seventeen: Measurement of ADP–ATP Exchange in Relation to Mitochondrial Transmembrane Potential and Oxygen Consumption
- Abstract
- 1 Introduction
- 2 Methodology
- 3 Conclusions and Comments
- Acknowledgments
Chapter Eighteen: Real-Time Assessment of the Metabolic Profile of Living Cells with Genetically Encoded NADH Sensors
- Abstract
- 1 Introduction
- 2 Experimental Components and Considerations
- 3 Initial Characterization Experiments
- 4 Steady-State and Kinetics Experiments
- 5 Data Handling/Processing
- 6 Summary
- Acknowledgments
Chapter Nineteen: ¹³C Isotope-Assisted Methods for Quantifying Glutamine Metabolism in Cancer Cells
- Abstract
- 1 Introduction
- 2 Methods
- 3 Applications
- 4 Summary
- Acknowledgments
Chapter Twenty: Measurement of Fatty Acid Oxidation Rates in Animal Tissues and Cell Lines
- Abstract
- 1 Introduction
- 2 Protocol Overview
- 3 Equipment
- 4 Materials
- 5 Protocol
- 6 Data Handling and Calculations
- Acknowledgments
Chapter Twenty-One: Methods to Assess Lipid Accumulation in Cancer Cells
- Abstract
- 1 Introduction
- 2 Materials
- 3 Methods
- 4 Notes
- Acknowledgments
Chapter Twenty-Two: Analysis of Hypoxia-Induced Metabolic Reprogramming
- Abstract
- 1 Introduction
- 2 Exposure of Cells to Hypoxic Culture Conditions
- 3 Alterations in Glucose Uptake and Metabolism
- 4 Induction of Mitochondria-Selective Autophagy
- 5 Maintenance of Intracellular pH
- Acknowledgments
Author Index
Subject Index

Lorenzo Galluzzi

Lorenzo Galluzzi is Assistant Professor of Cell Biology in Radiation Oncology at the Department of Radiation Oncology of the Weill Cornell Medical College, Honorary Assistant Professor Adjunct with the Department of Dermatology of the Yale School of Medicine, Honorary Associate Professor with the Faculty of Medicine of the University of Paris, and Faculty Member with the Graduate School of Biomedical Sciences and Biotechnology of the University of Ferrara, the Graduate School of Pharmacological Sciences of the University of Padova, and the Graduate School of Network Oncology and Precision Medicine of the University of Rome “La Sapienza”. Moreover, he is Associate Director of the European Academy for Tumor Immunology and Founding Member of the European Research Institute for Integrated Cellular Pathology. Galluzzi is best known for major experimental and conceptual contributions to the fields of cell death, autophagy, tumor metabolism and tumor immunology. He has published over 450 articles in international peer-reviewed journals and is the Editor-in-Chief of four journals: OncoImmunology (which he co-founded in 2011), International Review of Cell and Molecular Biology, Methods in Cell biology, and Molecular and Cellular Oncology (which he co-founded in 2013). Additionally, he serves as Founding Editor for Microbial Cell and Cell Stress, and Associate Editor for Cell Death and Disease, Pharmacological Research and iScience.

Affiliations and expertise

Assistant Professor of Cell Biology in Radiation Oncology, Department of Radiation Oncology, Weill Cornell Medical College, NY, USA

Guido Kroemer

Guido Kroemer got his M.D. in 1985 from the University of Innsbruck, Austria, and his Ph.D. in molecular biology in 1992 from the Autonomous University of Madrid, Spain. He is currently Professor at the Faculty of Medicine of the University of Paris Descartes/Paris V, Director of the INSERM research team ‘Apoptosis, Cancer and Immunity’, Director of the Metabolomics and Cell Biology platforms of the Gustave Roussy Cancer Campus, and Practitioner at the Hôpital Européen George Pompidou (Paris, France). He is also the Director of the Paris Alliance of Cancer Research Institutes (PACRI) and the Labex 'Immuno-Oncology'. Dr. Kroemer is best known for the discoveries that mitochondrial membrane permeabilization constitutes a decisive step in regulated cell death; that autophagy is a cytoprotective mechanism with lifespan-extending effects; and that anticancer therapies are successful only if they stimulate tumour-targeting immune responses. He is currently the most-cited cell biologist in Europe (relative to the period 2007-2013), and he has received the Descartes Prize of the European Union, the Carus Medal of the Leopoldina, the Dautrebande Prize of the Belgian Royal Academy of Medicine, the Léopold Griffuel Prize of the French Association for Cancer Research, the Mitjavile prize of the French National Academy of Medicine and a European Research Council Advanced Investigator Award.

Affiliations and expertise

INSERM Cordeliers Research Cancer Paris; Hopital Europeen Georges Pompidou; Universite Paris Descartes, France

Life Sciences

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