Molecular Characterization of Prostate Hyperplasia in Prolactin Transgenic Mice

...................................................................................................... 1 TABLE OF CONTENTS................................................................................. 2 ORIGINAL PAPERS ....................................................................................... 4 LIST OF ABBREVIATIONS.......................................................................... 5 INTRODUCTION ............................................................................................ 6 The Prostate Gland...........................................................................................6 Prostate development ....................................................................................6 Prostate anatomy and structure in human and rodents.................................7 Prostate disorders .............................................................................................8 Benign Prostatic Hyperplasia........................................................................8 Possible theories of BPH etiology .................................................................9 Premalignant lesions of the prostate...........................................................11 Prostate carcinoma ......................................................................................11 Possible theories of prostate cancer etiology .............................................12 Prolactin...........................................................................................................13 Gene, structure, and variants.......................................................................13 Control of prolactin synthesis, secretion and regulation............................15 The prolactin receptor .................................................................................15 Prolactin signal transduction.......................................................................16 Action of prolactin in the prostate gland .....................................................16 Proliferation.................................................................................................17 Apoptosis.....................................................................................................17 Citrate production........................................................................................18 Prolactin in prostate pathophysiology .........................................................18 Prolactin in human prostate cancer and BPH.............................................18 Experimental animal data ...........................................................................19 Rodent models of prostate disease................................................................20 Transgenic prostate hyperplasia models.....................................................21 Rodent models of prostate cancer...............................................................21 Other genetically engineered mouse models with prostate phenotype .....22 Mouse models genetically engineered in the prolactin signaling pathway22 Mouse models genetically engineered in other hormones..........................23 Hormone/growth factor regulation of the prostate ....................................24 Action of androgens in the prostate............................................................24 Interactions between prolactin and androgens in the prostate gland.........25 Action of estrogens in the prostate .............................................................26 Interactions between prolactin and estrogens in the prostate gland ..........27 Action of other peptide hormones and growth factors in the prostate ......27 Functional genomics in the study of the prostate gland ............................28 AIMS OF THE THESIS ................................................................................ 29 METHODOLOGICAL CONSIDERATIONS ........................................... 30 Transgenic animals.........................................................................................30 Molecular Characterization of Prostate Hyperplasia in Prolactin Transgenic Mice 3 Approaches to gene expression analysis ......................................................32 cDNA representational difference analysis (RDA) ...................................32 Sequence analysis ........................................................................................ 34 cDNA microarray analysis .........................................................................35 Array design and Printing ........................................................................... 36 Target preparation....................................................................................... 37 Hybridization ............................................................................................... 37 Image analysis and Normalization.............................................................. 38 Data Analysis and Statistical Evaluation.................................................... 39 Experimental design .................................................................................... 40 Microarray databases.................................................................................. 41 Comparisons between cDNA RDA and cDNA microarray analyis .........42 Verification strategies .................................................................................43 cDNA microarray analysis .......................................................................... 43 Real-time RT-PCR ....................................................................................... 43 Assessment of apoptotic activity ...................................................................46 RESULTS AND TECHNICAL COMMENTS........................................... 47 Paper I..............................................................................................................47 Paper II ............................................................................................................49 Paper III...........................................................................................................53 Paper IV...........................................................................................................55 DISCUSSION .................................................................................................. 58 CONCLUSIONS ............................................................................................. 67 ACKNOWLEDGEMENTS........................................................................... 68 REFERENCES................................................................................................ 70