Angiogenesis Assays A Critical Appraisal of Current Techniques

, by Staton, Carolyn A.; Lewis, Claire; Bicknell, Roy

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ISBN: 9780470016008 | 0470016000
Cover: Hardcover
Copyright: 2/5/2007

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Angiogenesis Assays: A Critical Appraisal of Current Techniques describes in detail the in vitro and in vivo assays currently being used to study angiogenesis, including the provision of protocols. The chapters are written by leading scientists who use these techniques in their angiogenesis research programs and who deliver here a critical appraisal of their strengths and weaknesses. Divided into three sections, the first section covers in vitro assays of endothelial cell activity; the second section details various in vivo angiogenesis assays; and the third section describes some of the biological endpoints being used in clinical trials to assess the clinical efficacy of anti-angiogenic drugs.

Foreword	p. xi
Preface	p. xv
List of contributors	p. xvii
Endothelial cell biology	p. 1
Abstract	p. 1
Introduction	p. 1
Morphology of the endothelium	p. 2
Endothelial cell adhesion and interactions	p. 4
Coagulation and haemostasis	p. 21
Transport	p. 13
Angiogenesis	p. 14
Isolation and culture of endothelial cells	p. 19
Endothelial cell heterogeneity and organ specificity	p. 21
Gene expression in endothelial cells	p. 23
Conclusions	p. 28
References	p. 28
Endothelial cell proliferation assays	p. 39
Abstract	p. 39
Introduction	p. 39
Cell proliferation	p. 41
Cell proliferation assays	p. 42
Conclusions	p. 48
References	p. 49
Endothelial cell migration assays	p. 51
Abstract	p. 51
Introduction	p. 51
Transfilter assay	p. 52
'Wound healing' assay	p. 55
Teflon fence assay	p. 57
Phagokinetic track assay	p. 58
In/ex vivo approaches for studying endothelial cell migration	p. 59
Conclusions	p. 60
References	p. 60
Tubule formation assays	p. 65
Abstract	p. 65
Introduction	p. 65
Endothelial cell sources	p. 66
Endothelial cell morphology and tubule formation	p. 67
Tubule assay matrices	p. 68
2D assay protocols	p. 69
Analysis of tubule formation in 2D assays	p. 73
Recent developments in 2D assays	p. 77
3D assays	p. 78
Analysis of 3D assays	p. 79
Co-culture assays	p. 80
Conclusions	p. 82
References	p. 82
Modelling the effects of the haemodynamic environment on endothelial cell responses relevant to angiogenesis	p. 89
Abstract	p. 89
Introduction	p. 89
Definitions	p. 91
Experimental patterns of shear exposure and relevance to angiogenesis assays	p. 93
Methods for studying responses of endothelial cells exposed to shear stress	p. 94
Readouts in flow cultures relevant to angiogenesis	p. 98
Critical considerations and conclusions	p. 99
References	p. 101
Whole or partial vessel outgrowth assays	p. 105
Abstract	p. 105
Introduction	p. 105
Rat aortic ring assay	p. 106
Mouse aorta models	p. 108
Chick aortic arch assay	p. 109
Porcine carotid artery assay	p. 110
Human explant cultures	p. 110
Recent developments	p. 112
Quantification	p. 113
Strengths and weaknesses of organ culture assays	p. 115
Applications	p. 117
Conclusions	p. 118
References	p. 118
Assaying endothelial-mural cell interactions	p. 123
Abstract	p. 123
2D models to study endothelial-mural cell interactions	p. 123
3D models to study blood vessel assembly	p. 128
Summary	p. 134
References	p. 135
Assays for membrane and intracellular signalling events	p. 139
Abstract	p. 139
Introduction	p. 139
The endothelial caveolae as the organizers of efficient spatial signal transduction mechanisms	p. 140
The prostaglandins forming system: regulation of signal transduction and angiogenesis	p. 141
Use of antisense oligo and siRNA to evaluate the function of single signalling molecules	p. 143
Knocking down and knocking out Caveolin-1 gene	p. 151
Analysing protein-protein interactions in signalling molecules involved in angiogenesis	p. 152
Final remarks	p. 155
References	p. 157
Implantation of sponges and polymers	p. 167
Abstract	p. 167
Introduction	p. 167
Materials used in implantation techniques	p. 169
Implantation technique for assessment of inflammatory processes	p. 170
Implantation technique for assessment of systemic pathological conditions	p. 170
Implantation technique for assessment of tumour angiogenesis	p. 171
Implantation technique for assessment of inflammatory angiogenesis in genetically modified mice	p. 172
Applications of cannulated sponge model for testing angiogenesis modulators	p. 172
Techniques for assessment of the vascularization in implants	p. 172
Summary of cannulated sponge assay: advantages and disadvantages	p. 178
References	p. 179
Angiogenesis assays in the chick	p. 183
Abstract	p. 183
Introduction	p. 183
Overview of the basic 'in shell' CAM assay	p. 186
Variations on the CAM assay	p. 189
Summary of the CAM assay: disadvantages and advantages	p. 199
References	p. 200
Corneal angiogenesis assay	p. 203
Abstract	p. 203
Introduction	p. 203
Anatomy and histology	p. 204
Brief history of CNV assays	p. 205
The process of corneal neovascularization	p. 206
Experimental induction of corneal neovascularization	p. 208
Discussion and conclusions	p. 220
References	p. 222
Dorsal air sac model	p. 229
Abstract	p. 229
Introduction	p. 229
Preparation of the DAS model	p. 230
Evaluation of anti-angiogenic photodynamic therapy using a DAS model	p. 231
Determination of blood volume or blood flow in the angiogenic site using the DAS model	p. 232
Quantitative or semi-quantitative analysis of angiogenesis in the DAS model	p. 234
Isolation and application of neovessel-targeting probe using DAS model	p. 236
Conclusions	p. 237
References	p. 237
Chamber assays	p. 239
Abstract	p. 239
Introduction	p. 239
In vivo imaging of angiogenesis and microcirculation	p. 240
Chamber assays	p. 241
Physiological angiogenesis	p. 246
Tumour angiogenesis	p. 248
Angiogenesis in endometriosis	p. 249
Angiogenesis in wound and bone healing	p. 250
Angiogenesis in ischaemia and hypoxia	p. 252
Angiogenesis in transplantation	p. 252
Angiogenesis in biomaterial incorporation	p. 254
Angiogenesis in tissue engineering	p. 255
Conclusions and perspectives	p. 256
References	p. 257
Tumour models: analysis of angiogenesis in vivo	p. 265
Abstract	p. 265
Introduction	p. 265
Tumour microenvironment and angiogenesis	p. 266
Tumour models	p. 267
Subcutaneous tumour models	p. 271
Orthotopic tumour models	p. 272
Transgenic mouse models of tumour angiogenesis	p. 277
Monitoring vascular permeability	p. 279
Analysis of angiogenesis in tumours	p. 280
Conclusions	p. 283
References	p. 283
Angiomouse: imageable models of angiogenesis	p. 293
Abstract	p. 293
Introduction	p. 293
Fluorescent proteins to image angiogenesis	p. 294
Dual-colour tumour host models	p. 297
ND-GFP mouse model	p. 300
Methods of angiogenesis analysis in GFP models	p. 306
Conclusions	p. 307
References	p. 308
Techniques and advances in vascular imaging in Danio rerio	p. 311
Abstract	p. 311
Introduction	p. 311
Visualizing the developing vasculature	p. 312
In situ hybridization reveals vascular specific expression	p. 318
Vascular specific fluorescent reporter lines	p. 320
Chemical mutagenesis screens reveal novel genes involved in cardiovascular development	p. 321
Conclusions	p. 323
References	p. 324
Biological and clinical implications of recruitment of stem cells into angiogenesis	p. 327
Abstract	p. 327
Introduction	p. 327
Phenotypic and functional characterization of EPCS	p. 328
Mobilization, homing and differentiation of EPCS	p. 332
Ageing	p. 335
Therapeutic applications	p. 335
Conclusions	p. 337
References	p. 337
Methods for monitoring of the anti-angiogenic activity of agents in patients: novel trial design	p. 341
Abstract	p. 341
Introduction	p. 341
Tumour micro-environmental factors	p. 343
Possible mechanisms of acquired resistance to anti-angiogenic drugs	p. 348
Standard chemotherapy versus angiogenesis inhibitors	p. 348
Monitoring clinical trials	p. 349
Tumour dormancy and tumour progression	p. 350
Tumour markers	p. 351
Microvessel density	p. 351
Surrogate endpoints	p. 352
Molecular imaging	p. 354
New insights into trial design	p. 354
Concluding remarks	p. 355
References	p. 355
An overview of current angiogenesis assays: Choice of assay, precautions in interpretation, future requirements and directions	p. 361
Abstract	p. 361
Introduction	p. 361
In vitro assays	p. 362
In vivo assays	p. 365
Clinical trials	p. 370
Future directions	p. 370
Concluding comments	p. 371
References	p. 372
Index	p. 375
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