Angiogenesis: In Vivo Systems, Part B

MIE445_Angiogenesis--In Vivo Systems, Part B......Page 1
SerialEditorFrontMatter ii......Page 2
Methods in Enzymology......Page 0
Copyright Page......Page 4
Contributors......Page 5
Preface......Page 9
Methods in Enzymology......Page 11
Inflammation, Angiogenesis, and Lymphangiogenesis......Page 37
Introduction......Page 38
Angiogenesis and Lymphangiogenesis in Inflammation......Page 39
Mouse Models of Inflammation......Page 41
Commonly Used Markers of Blood Vessels and of Lymphatic Vessels......Page 44
Measuring the (Lymph)angiogenic Response by Immunofluorescence......Page 46
Quantitating (Lymph)angiogenesis by Fluorescence-Activated Cell Sorting......Page 48
Measuring Vascular Permeability and Flow......Page 49
Mice......Page 50
Immunofluorescence for MECA-32 and LYVE-1 or CD31 and Podoplanin......Page 51
Immunofluorescence for Prox1 and CD31......Page 52
Ki67 Staining......Page 53
Preparation of LN Single-Cell Suspensions......Page 54
FACS Staining......Page 55
References......Page 56
Color-Coded Fluorescent Mouse Models of Cancer Cell Interactions with Blood Vessels and Lymphatics......Page 63
Introduction......Page 64
Isolation of Blood-Derived Endothelial Progenitor Cells......Page 343
Color-Coded Tumor-Host Models: GFP Nude Mouse......Page 68
Color-Coded Tumor-Host Models: Nestin-Driven-GFP Nude Mouse......Page 72
Imaging Cancer Cell Trafficking in Lymphatics: Experimental Metastasis Model......Page 73
Imaging Cancer Cell Trafficking in Lymphatics: Spontaneous Metastasis Model......Page 75
GFP Retrovirus Production......Page 77
Embryo Whole-Mount Electroporation of DNA Plasmids Expressing Fluorescent Proteins......Page 78
Double RFP and Histone H2B-GFP Gene Transduction of Cancer Cells......Page 79
Methods......Page 110
Experimental Lymphatic Metastasis Model......Page 80
Whole-Body Imaging of Tumor Vasculature......Page 81
Intravital Imaging for Deep Tumors......Page 82
Imaging with Fluorescence Microscopy......Page 83
Imaging with the Olympus IV-100 Scanning Laser Microscope System......Page 84
Summary and Conclusions......Page 85
References......Page 86
Bone Marrow-Derived Vascular Progenitors and Proangiogenic Monocytes in Tumors......Page 89
Introduction......Page 90
Culture Chamber Preparation......Page 145
Transplantation of GFP-expressing Bone Marrow to Visualize BMDC in Tumors......Page 93
Methods......Page 94
Reagents......Page 95
Methods......Page 96
Reagents......Page 97
Immunohistochemical Staining of Vasculature......Page 98
Quantification of GFP+ BMDC Subpopulations in Tumors......Page 99
Reagents......Page 100
Methods......Page 101
Analysis of Pericyte Progenitors in Tumors......Page 102
Detection of Pericyte Progenitors and Pericytes in Tumors......Page 105
Methods......Page 106
Reagents......Page 107
Methods......Page 108
Dual-Modality Imaging......Page 188
Methods......Page 109
Methods......Page 111
Reagents......Page 113
References......Page 115
Screening Phage-Display Peptide Libraries for Vascular Targeted Peptides......Page 119
Vascular Heterogeneity and its Characterization by Phage Display......Page 120
Embryo Staging and Preparation......Page 122
Normal Pregnancy......Page 123
Ligand-Directed Vectors for Vascular Targeted Gene Delivery and targeted molecular imaging......Page 124
General Recommendation......Page 126
Principle......Page 127
Phage Selection, Recovery, and Amplification......Page 128
Additional Rounds of Selection......Page 129
Principle......Page 130
Differential Centrifugation and Removal of Unbound Phage......Page 131
Additional Rounds of Selection......Page 132
Principle......Page 133
Tissue Isolation, Phage Recovery, and Phage Amplification......Page 134
Additional Rounds of Selection......Page 135
Concluding Remarks and Perspective......Page 136
References......Page 137
Avian Embryos: A Model for the Study of Primary Vascular Assembly in Warm-Blooded Animals......Page 143
Introduction......Page 144
Construction of Electroporation Chamber......Page 149
Embryonic Culture Insert/Dish Preparation......Page 151
Late Pregnancy......Page 271
Using Antibodies and Microinjection for Tagging Endothelial Cells or ECM Fibers......Page 152
Solutions......Page 153
In Ovo Electroporation of Fluorescent Protein Expressing Constructs......Page 155
Post-Incubation Fixation and Processing......Page 156
Whole-Mount Immunolabeling......Page 157
References......Page 158
Introduction......Page 161
Xenografts......Page 163
Genetic Models......Page 165
Apc+/min mice......Page 166
MEN1 mice......Page 167
RIP-Tag model......Page 168
Humanized VEGF mouse model......Page 169
Conclusions......Page 170
References......Page 171
Molecular Imaging of Tumor Vasculature......Page 177
Introduction......Page 178
Structural/Functional Imaging of Tumor Vasculature......Page 179
Origins of Coronary Vessels in the Proepicardium......Page 181
Coronary Vasculogenesis in the Subepicardium......Page 182
Molecular Imaging......Page 184
Imaging Integrin alphavbeta3 in Tumor Vasculature......Page 185
Single-Modality Imaging......Page 186
Additional Materials/Reagents......Page 357
Targeted Ultrasound......Page 190
Radionuclide-Based Imaging of VEGFR in Tumor Vasculature......Page 192
Isolation of Murine Mononuclear Cells from Bone Marrow......Page 386
PET Imaging......Page 194
Experimental Section......Page 197
DOTA Conjugation......Page 198
Cell Lines and Animal Models......Page 199
Micro-PET Imaging......Page 200
Biodistribution Studies......Page 202
Summary......Page 203
References......Page 204
Proteomic Mapping of the Vascular Endothelium In Vivo for Vascular Targeting......Page 213
Introduction......Page 214
Proteomics: An Overview......Page 216
Two-Dimensional Electrophoresis......Page 217
Liquid Chromatography......Page 218
Mass Spectrometry......Page 219
Additional Materials/Reagents......Page 222
Databases......Page 223
Quantitative Proteomics......Page 224
Label-Free Approaches......Page 225
Application of Proteomics to the Vasculature......Page 228
Isolation and Subfractionation of Plasma Membranes......Page 234
Perfusion Procedure......Page 235
Purification of Silica-Coated Luminal Endothelial Cell Plasma Membrane......Page 236
Mass Spectrometric Analysis of Endothelial Cell Membrane Proteins......Page 237
References......Page 238
Development of Coronary Vessels......Page 245
What is the PE?......Page 246
Coronary Artery Formation and Smooth Muscle Cell Recruitment......Page 248
Chorionic Villous CTBs Invade First-Trimester Decidual Explants in Vitro......Page 249
Mouse Embryos......Page 251
Isolation of Total RNA from Single PEs......Page 252
Explant Culture of the Epicardium......Page 253
Cytotrophoblast Migration and Induction of Endothelial Cell Apoptosis During Co-Culture......Page 329
Whole-Mount PECAM1 Immunostaining of Mouse Embryos......Page 256
Image Processing......Page 257
Whole Mount β-Galactosidase Staining......Page 258
Preparation of Coronary Vascular Casts......Page 259
References......Page 261
Methods for Evaluating Uteroplacental Angiogenesis and Their Application Using Animal Models......Page 265
Introduction......Page 266
Cytotrophoblast Isolation Procedure......Page 321
What we have Learned from Studies of Uteroplacental Angiogenesis Using Animal Models......Page 274
Coronary Fluorescent Microangiography: Animal Preparation and Image Acquisition (Mice)......Page 281
Conclusions......Page 285
References......Page 286
Intravital Microscopic Investigation of Leukocyte Interactions with the Blood Vessel Wall......Page 291
Transillumination Intravital Microscopy......Page 292
Measuring Leukocyte Rolling, Adhesion, and Transmigration......Page 295
Surgical Considerations......Page 298
Fluorescence Intravital Microscopy......Page 300
Lymphocyte Rolling and Adhesion in Peyer\'s Patches......Page 302
Mixed Chimeric Mice......Page 303
Spinning-Disk Confocal and Two-Photon Microscopy......Page 304
Autoperfused Flow Chamber......Page 306
Microfluidic Flow Chambers......Page 308
References......Page 310
Placental Remodeling of the Uterine Vasculature......Page 317
Introduction......Page 318
Time Considerations for Enzymatic Digestions......Page 325
Isolation and Culture of First-Trimester, Human Placental Villous Explants......Page 326
Identification of Cytotrophoblast-Modified Blood Vessels in Tissue Sections......Page 327
Co-Culture of First-Trimester CTBs with Differentiated Endometrial Endothelial Cells in a Three-Dimen......Page 330
CTB Invasion of Term Myometrial Spiral Artery Segments in Vitro......Page 331
In Vivo Models of Human Cytotrophoblast Vascular Remodeling......Page 333
Placental Xenograft of the Murine Mammary Fat Pad and Kidney Capsule......Page 334
References......Page 335
An In Vivo Experimental Model for Postnatal Vasculogenesis......Page 339
Introduction......Page 340
Materials/Reagents......Page 344
Myeloid Cells: Monocytes and Macrophages......Page 345
Isolation of Cord Blood-Derived EPCs (cbEPCs)......Page 346
Isolation of Adult Blood Derived-EPCs (abEPCs)......Page 347
Magnetic Bead Purification of Cultured CD31-Positive EPCs......Page 350
Notes......Page 351
Expansion and Characterization of Blood-derived Endothelial Progenitor Cells......Page 352
Expansion of Blood-Derived EPCs......Page 354
Notes......Page 355
Procedure......Page 356
Procedures......Page 358
Notes......Page 359
References......Page 361
Assessment of Arteriogenesis......Page 367
Introduction......Page 368
Animal Preparation and Image Acquisition (Mice)......Page 369
Micro-CT Coronary Angiography......Page 371
Complete Filling of the Coronary Arterial Vasculature......Page 372
X-ray Microangiography: Animal Preparation and Image Acquisition (Mice)......Page 373
Animal Preparation and Data Acquisition......Page 375
Electron Paramagnetic Resonance Data Acquisition......Page 376
Acknowledgments......Page 377
References......Page 378
Methods to Study Myeloid Cell Roles in Angiogenesis......Page 379
Angiogenesis......Page 380
Recruitment of Monocytes into Tumors......Page 381
Inflammation and Cancer......Page 382
Identification of Myeloid Cells in Tissues by Immunohistochemistry......Page 383
Isolation of Human and Murine Peripheral Blood Mononuclear Cells......Page 384
Fluorescence-Activated Cell Sorting Analysis......Page 387
Characterization of Integrin Activation on Myeloid Cells......Page 390
Integrin Activation as Measured by Clustering......Page 391
Adhesion of Myeloid Cells to Purified Ligands......Page 393
Adhesion of Myeloid Cells to Endothelial Cells in Vitro......Page 394
In Vitro Myeloid Migration/Invasion Assays......Page 396
In Vivo Monocyte-Trafficking Studies......Page 397
Role of Myeloid Cells in Tumor Growth Studies......Page 399
Quantification of Myeloid Cells in Experimental Tumors by FACS, qPCR, and IHC......Page 401
Bone Marrow Transplantation to Study Myeloid Cells in Tumor Growth......Page 403
References......Page 405
Author Index......Page 409
Subject Index......Page 439