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Photographs included in the original manuscript have been reproduced xerographically in this copy. Higher quality 6” x 9" black and white photographic prints are available for any photographs or illustrations appearing in this copy for an additional charge. Contact UMI directly to order. ProQuest Information and Learning 300 North Zeeb Road, Ann Arbor, Ml 48106-1346 USA 800-521-0600 UMI' The Immune System and Breast Carcinoma: Implications of Dietary and Other Associated Factors by Stephen Arthur Hoption Gann Dip. Bus. Admin., Camosun College, 1990 B.Sc., Universily of Victoria, 1994 A Dissertation Submitted in Partial Fulfillment o f the Requirements of the Degree of DOCTOR OF PHILOSOPHY in the Department of Biology We accept this dissertation as conforming to the required standard letten. Supervisor (Department of Biology) _____________________________________________________________________________________________________________ Dr. F.Y.M. Cho^, Supervisor ÛDepartment of Biology) ental Member (Department of Biology) ^ ^ u tsid e Member (Department of Biochemistry and Microbiology) Dr. C. van Netten, Additional Member (Department o f Health Care and Epidemiology, University of British Columbia) Dr. R. Bamie, External Examiner (Department of Laboratory Medicine, Capital Health Region) © Stephen Arthur Hoption Cann, 2001 University of Victoria All rights Reserved. This dissertation may not be reproduced in whole or in part, by photocopying or other means, without the permission o f the author. u Supervisors: Dr. J.P. van Netten and Dr. F.Y.M. Choy ABSTRACT Introduction'. A review o f animal and human studies demonstrates that the immune system is a major factor in both the enhancement and inhibition o f malignant tumour growth. Macrophages, one of the most durable and versatile immune cells, may be key to this immune duality. Macrophages have been observed in particularly high concentrations in and around breast tumours. It has been suggested that these cells generally aid tumour growth, unless activated by an acute infections, immunomodulators or other means. Study /: Using immunohistochemistry and computer-aided image analysis, macrophage concentrations in and around breast tumours were examined. Other pathological tissues were also examined for comparative purposes. Macrophage density was found to correlate positively with the Modified Bloom Richardson (MBR) grade (r = 0.41) and MBR subscore (r = 0.44), suggesting that macrophage concentrations increase as tumours become more aggressive. Similar infiltrations of macrophages were observed in lung, prostate and hyperplastic thyroid tissues; although in these latter tissues, macrophages were generally confined to the tumour periphery. Study II'. Iodine has been shown to play many roles in normal human physiology. In addition to its incorporation into thyroid hormones, iodine also has antibiotic and anti­ tumour properties. Epidemiological studies of iodine in breast cancer have not been conducted. In this pilot case-control study, whole blood levels of 10 trace elements (Br, Cr, Fe, I, Mb, Mg, Mn, Se, V, and Zn) and their association with breast cancer was investigated. Other general, medical and dietary characteristics were examined as well. In comparison with iodine levels in Japan, iodine levels in the population under study were considerably lower, with a mean of 28.4 jj.g/1 and a range of 19-35 p.g/1. In the univariate logistic regression analysis, a number of significant associations with breast cancer were observed. A high education status (OR = 0.31) and high iron status (OR = 0.15) were associated with reduced risks, whereas previous hysterectomy or ovariectomy was associated with an increased risk of breast cancer (OR = 3.64). In the adjusted multivariate analysis, a high iron status remained associated with a reduced risk (OR = O.OI) and a history of breast pain with an increased risk (OR =11.25). U1 Conclusion: Understanding the duality o f immune function, not only provides insight into cancer progression, but offers two primary avenues for treatment. First, one may down- regulate immune reparative activities, which aid tumour growth. This may be accomplished by using immunosuppressants. A second approach is to take advantage of the large population o f tumour-associated immune cells, particularly macrophages, and stimulate these cells into their defensive activities. A wide variety of infectious agents may be used to stimulate this response. Finally, iodine is one immunomodulator that may be used to enhance immune activity for treatment, or alternatively, prevent tumour growth through long-term intake; unfortunately, blood iodine levels noted in this study would be too low to afford protection. Dr. J^. van Netten, Supervisor (Department of Biology) >r. F.Y.M. dhoy, Supèr irvisor (Department o f Biology) ental Member (Department of Biology) isiu d^^fember (Department of Biochemistry and Microbiology) Dr. C. van Netten, Additional Member (Q^>artlSent of Health Care and Epidemiology, University of British Columbia) . R. Baillie, External Examiner (DepartmeniDr. R. Baillie, External Examiner (DepartmeniDr. (Department of Laboratory Medicine, Capital Health Region) IV TABLE OF CONTENTS ABSTRACT .................................................................................................................. ü TABLE OF CONTENTS............................................................................................. iv LIST OF TABLES ........................................................................................................ vi LIST OF FIGURES.............................................................................................................. vii ABBREVIATIONS U S E D ........................................................................................ x ACKNOWLEDGEMENTS.............................................................................................. xü GENERAL INTRODUCTION .................................................................................. 1 CHAPTER I Immunohistochemical and computed-aided image analysis of macrophage content in breast carcinoma Introduction ................................................................................................................... 31 M e t h o d s ........................................................................................................................ 51 R e s u lts ..............................................................................................................................55 D iscussion ........................................................................................................................ 70 CHAPTER 2 A case-control study of iodine, selenium and other associatedfactors in patients with newly diagnosed breast carcinoma Introduction ...................................................................................................................75 M e t h o d s ........................................................................................................................ 86 R e su lts............................................................................................................................. 94 D iscussion ...................................................................................................................... 110 OVERVIEW ...................................................................................................................... 135 LITERATURE CITED.......................................................................................................142 APPENDIX I Breast Cancer Grading and Staging ................................................... 185 APPENDIX n Ethical Approval — Macrophage Study .............................................. 187 APPENDIX in Informed Consent - Case Subjects ................................................... 188 APPENDIX IV Informed Consent - Control Subjects ..............................................192 APPENDIX V Questionnaire — Case S u b jects ........................................................ 196 APPENDIX VI Questionnaire - Control S u b jects ...................................................203 APPENDIX Vn Ethical Approval I - Case-Control Study .........................................209 APPENDIX Vm Ethical Approval II - Case-Control Study .........................................210 VI LIST OF TABLES Table 1 Staging and grading o f breast carcinomas. 6 Table 2 Tumour-derived chemokines involved in the regulation of 36 tumour-infiltrating macrophages. Table 3 Factors