VASCULAR DISEASE: MOLECULAR BIOLOGY AND GENE THERAPY PROTOCOLS

By RICHARD LEE

CONTENTS PART I GENETICS IN VASCULAR DISEASE DETECTION OF MUTATIONS AND DNA POLYMORPHISMS IN GENES INVOLVED IN CARDIOVASCULAR DISEASES BY POLYMERASE CHAIN REACTION–SINGLE-STRAND CONFORMATION POLYMORPHISM ANALYSIS ANALYSIS OF GENETIC VARIANTS IN CARDIOVASCULAR RISK GENES BY HETERODUPLEX ANALYSIS RADIATION HYBRID (RH) MAPPING OF HUMAN SMOOTH MUSCLE-RESTRICTED GENES PART II ISOLATION OF GENES EXPRESSED IN VASCULAR TISSUE EFFICIENT EXTRACTION OF RNA FROM VASCULAR TISSUE PREPARATION OF CDNA LIBRARIES FROM VASCULAR CELLS SCREENING CDNA LIBRARIES USING PARTIAL PROBES TO ISOLATE FULL-LENGTH CDNAS FROM VASCULAR CELLS CLONING FULL-LENGTH CDNAS FROM VASCULAR TISSUES AND CELLS BY RAPID AMPLIFICATION OF CDNA ENDS (RACE) AND RT-PCR USE OF CDNA REPRESENTATIONAL DIFFERENCE ANALYSIS TO IDENTIFY DISEASE-SPECIFIC GENES IN HUMAN ATHEROSCLEROTIC PLAQUES THE USE OF DIFFERENTIAL MRNA DISPLAY (DDRT-PCR) TO IDENTIFY GENES DIFFERENTIALLY EXPRESSED IN NORMAL AND DISEASED VASCULAR CELLS IDENTIFICATION OF NOVEL PROTEIN KINASES IN VASCULAR CELLS PART III METHODS FOR MAPPING TRANSCRIPTIONAL START SITES AND MEASUREMENT OF PROMOTER ACTIVITY IN VASCULAR CELLS PRIMER EXTENSION ANALYSIS TO MAP TRANSCRIPTION START SITES OF VASCULAR GENES USE OF LIPOSOME-MEDIATED DNA TRANSFECTION TO DETERMINE PROMOTER ACTIVITY IN SMOOTH MUSCLE CELLS NUCLEAR RUN-ON ASSAY TO STUDY GENE TRANSCRIPTION IN VASCULAR CELLS ELECTROMOBILITY SHIFT ANALYSIS (EMSA) APPLIED TO THE STUDY OF NF-KAPPA B BINDING INTERACTIONS IN VASCULAR SMOOTH MUSCLE CELLS PART IV MOLECULAR ANALYSIS OF MRNA EXPRESSION IN VASCULAR CELLS MEASUREMENT OF GENE EXPRESSION IN THE VASCULAR WALL BY REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION (RT-PCR) ANALYSIS NORTHERN BLOT ANALYSIS TO QUANTIFY GENE EXPRESSION IN VASCULAR DISEASES LOCALIZATION OF GENE EXPRESSION IN HUMAN ATHEROSCLEROTIC LESIONS BY IN SITU HYBRIDIZATION RNASE PROTECTION ASSAYS FOR QUANTIFICATION OF GENE EXPRESSION IN VASCULAR TISSUE PART V MOLECULAR METHODS TO STUDY APOPTOSIS AND PHENOTYPIC CHANGES IN VASCULAR CELLS AND TISSUE DETECTION OF APOPTOSIS IN ATHEROSCLEROSIS AND RESTENOSIS BY TERMINAL DUTP NICK END LABELING (TUNEL) IN VITRO DETECTION OF APOPTOSIS IN ISOLATED VASCULAR CELLS MOLECULAR ASSESSMENT OF THE PHENOTYPIC CHANGES ASSOCIATED WITH SMOOTH MUSCLE CELLS USING TWO-DIMENSION ELECTROPHORESIS AND MICROSEQUENCE ANALYSIS PART VI IN VITRO METHODS TO EXPRESS FOREIGN GENES IN VASCULAR CELLS SIMPLE METHODS FOR PREPARING RECOMBINANT ADENOVIRUSES FOR HIGH-EFFICIENCY TRANSDUCTION OF VASCULAR CELLS GENERATION OF RECOMBINANT ADENO-ASSOCIATED VIRUSES FOR DELIVERY OF GENES INTO VASCULAR CELLS HEMAGGLUTINATING VIRUS OF JAPAN LIPOSOME-MEDIATED GENE DELIVERY TO VASCULAR CELLS HIGH-EFFICIENCY AND LOW-TOXICITY ADENOVIRUS-ASSISTED ENDOTHELIAL TRANSFECTION GENE TRANSFER IN VASCULAR CELLS USING AN ENGINEERED NA-H EXCHANGER (NHE1) AS A SELECTABLE MARKER USE OF RETROVIRUSES TO EXPRESS EXOGENOUS GENES IN VASCULAR SMOOTH MUSCLE CELLS EMBRYONAL STEM (ES) CELL–DERIVED MACROPHAGES: A CELLULAR SYSTEM THAT FACILITATES THE GENETIC DISSECTION OF MACROPHAGE FUNCTION DELIVERY OF RECOMBINANT ADENOVIRUSES TO HUMAN SAPHENOUS VEIN PART VII IN VIVO VASCULAR GENE TRANSFER PROTOCOLS AN IN VIVO ANGIOGENESIS ASSAY TO STUDY POSITIVE AND NEGATIVE REGULATORS OF NEOVASCULARIZATION EFFICIENT LIPOSOME-MEDIATED GENE TRANSFER TO RABBIT CAROTID ARTERIES IN VIVO GENE DELIVERY TO RABBIT ARTERIES USING THE COLLAR MODEL DELIVERY OF ANTISENSE OLIGONUCLEOTIDES TO THE VASCULAR WALL LOCAL GENE DELIVERY OF RECOMBINANT ADENOVIRUSES TO THE RAT CAROTID ARTERY IN VIVO