Unfolded Protein Response and Cellular Stress, Part A (eBook)

Front Cover 1
Methods in Enzymology: The Unfolded Protein Response and Cellular Stress, Part A 4
Copyright 5
Contents 6
Contributors 12
Preface 18
Methods in Enzymology 20
Section 1: UPR and Cell Stress in Normal and Diseased Cells 50
Chapter 1: The Unfolded Protein Response in Mouse Cerebral Cortex 52
1. Introduction: The UPR 52
2. Transduction of the UPR Signal by PERK, IRE1, and ATF6 53
3. UPR and Apoptosis 54
4. The UPR in Health and Disease 55
5. Induction of the UPR by Sleep Deprivation 55
6. Experimental Protocols 56
7. Concluding Remarks 67
Acknowledgments 68
References 68
Chapter 2: Immunohistochemical Detection of the Unfolded Protein Response in Atherosclerotic Plaques 72
1. Endoplasmic Reticulum and the Unfolded Protein Response 73
2. Atherosclerosis 73
3. UPR in the Atherosclerotic Lesion 74
4. Mouse Models of Atherosclerosis 76
5. Methods 78
Acknowledgments 93
References 93
Chapter 3: The Role of PDI as a Survival Factor in Cardiomyocyte Ischemia 96
1. Introduction 97
2. Identification of PDI as Cardio-Protective Factor 99
3. Conclusions 111
References 111
Chapter 4: Measurement of the Increase in Endoplasmic Reticulum Stress-Related Proteins and Genes in Adipose Tissue of Obese, 116
1. Introduction 117
2. Study Subjects and Fat Biopsies 118
3. Proteomic Analysis 119
4. Western Blot Analysis 128
5. RT-PCR Analysis 129
Acknowledgments 130
References 131
Chapter 5: Measurement of the Unfolded Protein Response (UPR) in Monocytes 132
1. Introduction 133
2. Investigating UPR Activation in the Monocyte 135
Acknowledgments 142
References 142
Chapter 6: Assessment of Endurance Exercise Training in Hepatic Endoplasmic Reticulum Stress 146
1. Introduction 147
2. Assessment of an Endurance-Training Program 148
3. "Dietary-Inducible" Hepatic ER Stress 153
4. Assessment of Hepatic ER Stress Molecular Markers 154
5. Required Materials 154
Acknowledgments 155
References 155
Chapter 7: Discovery Approaches to UPR in Athero-Susceptible Endothelium In Vivo 158
1. Introduction 159
2. Procedures for the Isolation of Endothelial Cells and Preparation of RNA for Microarray Hybridization 161
3. Microarray Hybridization and Feature Extraction 165
4. Bioinformatics Analysis 166
5. Another Approach to Gene Connectivity: Weighted Gene Coexpression Network Analysis 171
6. Validation and Follow-up 173
Acknowledgments 174
References 174
Chapter 8: Assessment of Endoplasmic Reticulum Stress and the Unfolded Protein Response in Endothelial Cells 176
1. Introduction 177
2. The Acute Phase of ER Stress 178
3. The Adaptive Reaction Phase: Unfolded Protein Response in Endothelial Cells 180
4. The Final Phase: Cell Death Signaling 189
Acknowledgments 194
References 194
Chapter 9: Measuring the Impact of Cigarette Smoke on the UPR 196
1. Introduction 197
2. Preparing Samples for Analysis 198
3. Analysis of Activation of the PERK Pathway 204
4. ATF6 Pathway 207
5. IRE1 Pathway 208
6. Using LSC to Measure UPR Activation 208
Acknowledgments 210
References 211
Chapter 10: Induction and Measurement of UPR and Osmotic Stress in the Yeast Pichia pastoris 214
1. Introduction 215
2. Measuring UPR Induction in P. pastoris 218
3. Inducing and Measuring Osmotic Stress in P. pastoris 228
Acknowledgments 235
References 235
Chapter 11: Induction of ER Stress Response Leading to Programmed Cell Death in Trypanosoma brucei 238
1. Introduction 239
2. Induction of ER Stress in Trypanosomes 242
3. Measurements of ER Stress in Trypanosomes Cells 243
4. Measurements of ER Stress-Induced Cell Death 250
Acknowledgments 252
References 252
Chapter 12: Measurement of Activation of the Endoplasmic Reticulum Stress Response in Autoimmune Myositis 256
1. Introduction 257
2. Measuring Endoplasmic Reticulum Stress Response in Skeletal Muscle Tissues Using Western Blotting 258
3. Immunohistochemical Detection of ER Stress Markers in Muscle Tissues 265
4. Determination of mRNA Levels of ER Stress Targets 270
5. Concluding Remarks 272
Acknowledgments 272
References 272
Chapter 13: Activation of the Unfolded Protein Response in Human Acute Myeloid Leukemia 276
1. Introduction 277
2. Technical Aspects 281
3. Discussion 288
References 290
Chapter 14: Modulation of the Unfolded Protein Response by GRP78 in Prostate Cancer 294
1. Introduction 295
2. Methods for Monitoring UPR Modulation 297
3. Conclusions 305
References 305
Chapter 15: Endoplasmic Reticulum Stress in Brain Damage 308
1. Introduction 309
2. ER Stress and Unfolded Protein Response 310
3. Cross Talk Between ER and Mitochondria 316
4. Experimental Approaches for the Detection of ER Stress 318
Acknowledgments 320
References 321
Chapter 16: Methods and Models for Monitoring UPR-Associated Macrophage Death During Advanced Atherosclerosis 326
1. Introduction to ER Stress in Atherosclerosis 327
2. Significance of Macrophage ER Stress and Apoptosis in Advanced Atherosclerosis 328
3. Modeling Atherosclerosis-Relevant ER Stress-Induced Apoptosis 330
4. The UPR and Macrophage Death at the Murine Aortic Root 334
5. Conclusion and Future Methodological Advances 342
Acknowledgments 342
References 342
Chapter 17: Detecting Autophagy in Response to ER Stress Signals in Cancer 346
1. Introduction 347
2. Monitoring Autophagy Upon ER Stress 348
3. Monitoring Autophagy Upon ER Stress In Vivo 358
References 366
Chapter 18: The Unfolded Protein Response Induced by Salt Stress in Arabidopsis 368
1. Introduction 368
2. The Treatment of Arabidopsis Seedlings 369
3. The Analysis of Unfolded Protein Response 371
Acknowledgments 376
References 376
Chapter 19: Measurement of ER Stress Response and Inflammation in the Mouse Model of Nonalcoholic Fatty Liver Disease 378
1. Introduction 379
2. Methods to Measure ER Stress Response and Hepatic Inflammation in the Mouse Model of NAFLD 381
3. Visualizing ER Stress Response Associated with Liver Pathology in the Mouse Model of NAFLD 390
Acknowledgments 394
References 394
Author Index 398
Subject Index 414
Colour Plate 420