Cellular Effects of Heavy Metals (eBook)

Preface 5
Contents 7
Contributors 9
Abbreviations 12
Part I Introduction 14
Chapter 1 15
Heavy Metals, Trace Elements and Their Cellular Effects 15
Introduction 15
Why Another Book on Heavy Metals? 15
Brief Review of Chapters 16
Definition of Heavy Metals 18
Trace Metal Elements 21
Cellular Effects of Heavy Metals 23
Non-essential Harmful Heavy Metals 24
Cellular Toxicity of Heavy Metals 25
Detoxification of Heavy Metals 26
Detection of Cellular Toxicity of Heavy Metals 26
Replacing In Vivo Animal Studies with In Vitro Systems 26
Bacterial, Fungal and Mammalian In Vitro Systems 27
Mammalian Cell Cultures 27
Permeability Changes Caused by Heavy Metals 28
Oxidative Damages Caused by Heavy Metals 29
Lipid Peroxidation 29
Oxidative DNA Damage 29
Estimation of Toxic Effects of Heavy Metals 30
Tumorigenic Potential of Heavy Metals 30
Metabolic Parameters 30
Cytoskeletal and Nucleoskelatal Changes 31
Chromosomal and Chromatin Changes Induced by Heavy Metals 31
Detection of Apoptotic and Necrogenic Chromatin Changes 32
Detection and Determination of Heavy Metals in Cells 32
Spectroscopy, Spectrometry 32
Atomic Absorption Spectrophotometry 32
Inductively Coupled Plasma Atomic Emission Spectroscopy (ICP-AES) 33
X-ray Fluorescence 33
Backscatter Electron (BSE) Imaging and Energy Dispersive Spectroscopy 33
Amperometric Detection of Unithiol Complexes of Heavy Metals 33
Isotope Techniques 33
Isotope Dilution Mass Spectrometric Method (IDMS) 33
Tritium-Labelled Chelates 34
Whole-Cell-Sensing Systems 34
Protein Based Bisosensors 34
Construction of Metal Detection Circuits in E. coli 34
Luminescence-Based Whole-Cell-Sensing Systems using Genetically Engineered Bacteria 34
Whole-Cell Heavy Metal Detecting Yeast System using Cadmium-Inducible Gene Promoter 35
Heavy Metal Toxicity Detected by Cardiac Cell-Based Biosensor 35
Antibody-Based Sensors for Heavy Metal Ions 36
Porphyrin Test 36
Disposable Cuvette Test for the Enzymatic Determination of Heavy Metals 36
References 36
Part II Heavy Metal Toxicity in Microbes 41
Chapter 2 42
Toxic Metal/Metalloid Tolerance in Fungi—A Biotechnology-Oriented Approach 42
Introduction 43
First Line of Defense: Extracellular Chelation and Binding to Cell Wall Constituents 44
Second Line of Defense: Transport, Intracellular Chelation and Compartmentalization 45
Third Line of Defense: The Antioxidative Defense System 50
Screening for Future Targets to Engineer Heavy Metal Tolerant Fungi 53
References 57
Chapter 3 70
Interference of Chromium with Cellular Functions 70
Introduction 70
Chromium and Environment 70
Extracellular Reduction of Chromate 72
Metal Ion Uptake by Yeasts and Fungi 73
Biosorption of Chromium 73
Bioaccumulation of Chromium 74
Cellular Interactions of Chromium 75
Interactions of Chromium with Plasma Membrane 75
ROS Formation During Intracellular Cr(VI) Metabolism 77
Mechanisms of Chromium Sensitivity and Resistance 78
Interactions of Chromium with Biomolecules 81
Mechanism of Chromium Toxicity 81
Risk Assessment in Human Exposure to Cr(VI) 82
Inhalation 83
Dermal Absorption 84
Oral Intake 84
Kinetics and Metabolism 85
Excretion 86
Nutritional Practices and Assessment of Risk Involved in Human Exposure to Chromium(III) 86
Prevention and Repair of Chromium-Induced Damage 88
Conclusions 89
References 90
Chapter 4 98
Saccharomyces Cerevisiae as a Model Organism for Elucidating Arsenic Tolerance Mechanisms 98
Introduction 98
Impact of Arsenic on Yeast Cells 99
Arsenic Uptake Routes in Yeast 100
Arsenate Uptake 100
Arsenite Uptake 101
Arsenic Detoxification Systems in Yeast 103
ACR Gene Cluster—A Major Determinant of Arsenic Tolerance in Yeast 103
As(III)-responsive Transcription Factor Yap8p/Acr1p 104
Arsenate Reductase Acr2p 106
Arsenite Permease Acr3p 106
Role of Fps1p in As(III) Efflux 107
Vacuolar Sequestration of Metalloids 108
Glutathione Biosynthesis and the Role of Met4p 109
Oxidative Stress Defence and Yap1p 110
Hog1p and Cell Cycle Regulation in Response to As(III) Exposure 112
Other Detoxification and Tolerance Systems 112
Global Analysis of Tolerance Factors in Yeast 112
Proteasomal Degradation of Damaged Proteins 113
TOR- and PKA-Pathway: Regulation of General Stress Responses and Ribosomal Proteins 113
Yeast as a Model System for Elucidating the Molecular Biology of Arsenic Toxicity and Tolerance 114
Aquaporins, Metalloid Transport and Human Health 114
ACR-Proteins in Plants 115
Conclusions 115
References 115
Part III Heavy Metal Induced Toxicity 
124 
Chapter 5 125
Heavy Metal Toxicity in an Insect Cell Line (Methyl-HgCl, HgCl2, CdCl2 and CuSO4) 125
Introduction 125
Materials 126
Cell Culture 126
Metal Exposure 127
Viability and Proliferation Assays 127
Light and Electron Microscopy 127
Atomic Absorption and Fluorescence Spectrometry 128
Biochemical Assays 128
Methods 128
Viability Assays 128
Growth Assays 129
Light Microscopy and Cytoskeleton Staining 129
Electron Microscopy 130
Autometallography 130
Atomic Absorption and Fluorescence Spectrometry 131
Biochemical Assays 131
Results and Discussion 132
Viability Tests 132
Proliferation Assays 134
General Cell Morphology (Light Microscopy) 136
Ultrastructural Effects (Electron Microscopy) 138
Metal Uptake 142
Mitochondrial Impairment and Anaerobic Metabolism in Cd-Treated Cells 145
Cadmium-Induced Molecular Defense Mechanisms 146
Conclusions 148
References 150
Part IV Genotoxic Effects of Heavy Metals 155
Chapter 6 156
Cellular Changes in Mammalian Cells Induced by Cadmium 156
Introduction 156
Oxydative DNA Damage Caused by Heavy Metals 156
Methods 158
Chemicals 158
Solutions 158
Cell Growth 158
Heavy Metal Treatment 159
Cell Cycle Synchronization 159
Flow Cytometry 159
Reversible Permeabilization of Cells 160
DNA Synthesis in Reversibly Permeabilized Cells 160
DNA Isolation 160
Random Oligonucleotide-Primed Synthesis (ROPS) Assay 161
Analysis of 8-hydroxy-2'-deoxyguanosine 161
Isolation of Nuclei 161
Spreads of Nuclear Structures 161
Visualization of Chromatin Structures 161
Results 162
Cellular Effects of Cadmium 162
Effect of Cd on Replicative and Repair Synthesis 162
DNA Strand Breaks and Oxidative DNA Damage Generated by Cd 162
Chromatin Changes Induced by Cd 163
Growth Inhibition by Cd in Murine PreB Cells 164
Chromatin Changes Induced by Cd in Murine PreB Cells 166
Discussion 169
References 170
Chapter 7 172
Chromatin Toxicity of Ni(II) Ions in K562 Erythroleukemia Cells 172
Introduction 172
Materials and Methods 174
Chemicals and Reagents 174
Cell Growth 174
Treatment with Nickel Chloride 
174 
Reversible Permeabilization of Cells 175
Isolation of Nuclei 175
Spreads of Nuclear Structures 176
Visualization of Large Scale Chromatin Structures 176
Time-Lapse Photography 176
Results 176
Cellular Toxicity of NiCl2 176
Chromatin Structures of Normal Untreated Cells 177
Density Changes in Chromatin Structures at Low (0.2 and 0.5 µM) Concentrations of Ni(II) 177
Apoptotic Chromatin Changes at Elevated (1–5 µM) Concentrations of Nickel Chloride 178
Chromatin Changes at Higher (10 µM) Nickel Chloride Concentration 181
Necrotic Chromatin Changes at High (50 µM) Nickel Chloride Concentration 181
Cellular Motion After 100 µM NiCl2 Treatment 181
Discussion 185
Conclusions 186
References 187
Chapter 8 188
Genotoxic Chromatin Changes in Schizosaccharomyces Pombe Induced by Hexavalent chromium (CrVI) Ions 188
Introduction 188
Materials and Methods 190
Materials and Solutions 190
Cell Growth 190
Toxicity of Cr (VI) on S. Pombe 191
Preparation of Protoplasts 191
Isolation and Visualization of Large Scale Chromatin Structures 191
Results 191
Visualization of Interphase Chromatin Structures of S. Pombe 191
Cellular Toxicity of Cr(VI) 192
Apoptotic Chromatin Changes at Low Cr(VI) Concentration (10–50 µM) 194
Necrotic Chromatin Changes at Higher Cr(VI) Concentration 194
Discussion 197
Conclusions 199
References 200
Chapter 9 203
Chromatin Changes upon Silver Nitrate Treatment in Human Keratinocyte HaCaT and K562 Erythroleukemia Cells 203
Introduction 203
Materials and Methods 205
Chemicals and Reagents 205
Cell Culture and Silver Nitrate Exposure 205
Isolation and Visualization of Large Scale Chromatin Structures 205
Time-Lapse Photography 206
Changes in Chromatin Structure upon Silver Nitrate Exposure 206
Results 206
Cell Viability After AgNO3 Treatment 206
Time-Lapse Analysis of Cell Death 208
Chromatin Structures in Control HaCaT Cells and After Subtoxic (0.5 µM) Concentration of Silver Nitrate Treatment 209
Chromatin Changes at Low (5–10 µM) Concentrations of Silver Nitrate in Nuclei of HaCaT Cells 213
Chromatin Changes at Elevated (15–20 µM) Concentrations of Silver Nitrate in Nuclei of HaCaT Cells 213
Chromatin Changes at Higher (30–50 µM) Concentrations of Silver Nitrate in Nuclei of HaCaT Cells 213
Chromatin Structures of Normal Untreated K562 Cells 213
Chromatin Changes at Low (0.5–5 µM) Concentrations of Silver Nitrate in Nuclei of K562 Cells 215
Shrinkage and Expansion of Nuclear Structures of K562 Cells at Elevated (10–50 µM) Ag+ Concentrations 215
Discussion 215
Biological Effects of Metallic Silver 215
Toxic Effects of Silver Ions 217
Cellular Effect of Silver Nitrate on Eukaryotic Cells 217
Conclusions 223
References 224
Part V Chemical Carcinogenesis Induced by Heavy Metals 226
Chapter 10 227
Heavy Metal-Induced Carcinogenicity: Depleted Uranium and Heavy-Metal Tungsten Alloy 227
Introduction 227
Depleted Uranium (DU) 228
Heavy-Metal Tungsten Alloy 229
Routes of Exposure 230
In Vitro Studies 231
Depleted Uranium 231
Heavy-Metal Tungsten Alloy 233
In Vivo Studies 235
Depleted Uranium 235
Heavy-Metal Tungsten Alloy 236
Human Exposures 237
Depleted Uranium 237
Heavy-Metal Tungsten Alloy 238
Conclusions 238
References 239
Chapter 11 243
Role of Oxidative Damage in Metal-Induced Carcinogenesis 243
Introduction 243
Basic Redox Biochemistry of Carcinogenic Metals 245
Oxidative DNA Damage 248
DNA Base Damage 248
Cross-Linking 250
Strand Scission 251
Depurination 252
Oxidative Protein Damage 255
Discussion 256
Conclusion 258
References 259
Part VI Cellular Responses to Heavy Metal Exposure 266
Chapter 12 267
Non-native Proteins as Newly-Identified Targets of Heavy Metals and Metalloids 267
Introduction 267
Principles of Protein Folding 268
Interaction of Heavy Metals with Functional Groups of Proteins 269
Interference of Heavy Metals with the Refolding of Chemically Denatured Proteins 270
Mechanism of Folding Inhibition by Heavy Metal Ions 274
Interference of As(III) Species with Oxidative Refolding of Disulfide Bond-Containing Proteins 274
Possible Sequels of Protein Folding Inhibition in Cells 275
Conclusions 276
References 277
Chapter 13 279
Cellular Mechanisms to Respond to Cadmium Exposure: Ubiquitin Ligases 279
Introduction 279
Ubiquitin System 279
E3 Ubiquitin Ligases 281
Cullin-Ring Ligases (CRLs) 281
The SCF-Complex 281
Cadmium and Ubiquitin Ligases 282
Cellular Response to Cadmium Exposure 282
Saccharomyces Cerevisiae Transcription Factor Met4 284
Cadmium Exposure Leads to the Disassembly of SCFMet30 286
The Schizosaccharomyces Pombe Transcription Factor Zip1 286
SCFPof1 is Responsible for the Ubiquitination of Zip1 287
Mammalian Transcription Factor Nrf2 287
KEAP1-CUL3 Ubiquitin Ligase is Responsible for the Ubiquitination of Nrf2 288
Conclusions 289
References 290
Chapter 14 294
Metals Induced Disruption of Ubiquitin Proteasome System, Activation of Stress Signaling and Apoptosis 294
Introduction 295
The Ubiquitin Proteasome System (UPS) 295
UPS and Neurodegenerative Disease 297
Environmental Metals Exposure and Neurodegenerative Disease 298
Results and Discussion 300
MeHg, Cd2+, and As3+ Induced Alteration of the Proteasome Activity 301
MeHg, Cd2+, and As3+ Induced Accumulation of HMW-polyUb 302
MeHg, Cd2+, and As3+ Induced Activation of MAPK Signaling 302
Integrative Genomic Gene Expression Analysis and Pathway Mapping 305
Interruptions of UPS Pathway 306
Conclusions 308
References 309
Part VII Biomarkers 315
Chapter 15 316
Blood Lead Level (BLL, B-Pb) in Human and Animal Populations: B-Pb as a Biological Marker to Environmental Lead Exposure 316
Introduction 316
Aims 317
Methods 317
Biomonitoring and Biomarkers: Human and Animal Approach 318
Toxicokinetics of Lead 319
Effects of Lead on Red Blood Cells 321
Biomarkers of Lead Exposure 323
Blood Lead Concentration 323
Blood Lead Levels’ Reference Values 324
Alternative Biomarkers 326
Lead in Plasma/Serum 326
Animal Populations 327
Biomonitoring in Pets 328
Conclusions 328
References 329
Part VIII Removal of Heavy Metals 332
Chapter 16 333
Removal of Heavy Metal Sulfides and Toxic Contaminants from Water 333
Introduction 333
Methods 334
Chemicals and Reagents 334
Precipitation and Removal of Heavy Metals 335
Determination of Heavy Metal Content 335
Fish 335
Guppy Ecotoxicity Test 336
Treatment with Carbogen Gas and Air Flow 336
Results 336
Analogy Between the Chemistry of Removing Cyanide and Heavy Metals 336
Removal of Hg2+, Ni2+ and Pb2+ as their Sulfides 338
Ecotoxicity Test of Heavy Metal Ions 339
Removal of Sodium Sulfide from Water 340
pH Changes during Carbogen Treatment 341
Survival of Fish upon Removal of Heavy Metal and Sodium Sulfide 342
Discussion 342
Conclusions 344
References 345
Index 347