Comparing the Effects of Black and Navy Beans on Vascular Properties in Spontaneously Hypertensive Rats by Tara B. Loader A Thesis submitted to the Faculty of Graduate Studies of The University of Manitoba In partial fulfillment of the requirements of the degree MASTER OF SCIENCE Department of Human Nutritional Sciences University of Manitoba Winnipeg, Manitoba, Canada R3T 2N2 Copyright © 2017 by Tara Loader Abstract Pulses such as dried beans may be useful in the treatment of hypertension. Vascular protection by beans may relate to their phenolic compounds, which are associated with darker-coloured seeds. Thus, black beans were compared to navy beans for their effects on hypertension and vascular remodelling in spontaneously hypertensive rats (SHR). Assessments of blood pressure, pulse wave velocity (PWV), and vessel morphology were conducted to ascertain the vascular effects of the beans. Results showed that neither black beans nor navy beans influenced blood pressure or PWV in SHR. However, vascular remodelling was mitigated by black beans but not by navy beans. This was indicated by the absence of a significant difference in the ratio of aortic elastin content to vessel wall area in black bean-fed SHR compared to normotensive Wistar Kyoto (WKY) rats. The consumption of black beans could therefore be a therapeutic dietary strategy for combating vascular remodelling in hypertension. ii Acknowledgements Writing this thesis and completing this graduate program has been quite a journey. My decision to take on the role of a Master’s student was not an easy one as my life was quite comfortable and stress free prior to this time. I didn’t fully know what I would be getting into, but I knew it would be a challenge and that for me, it was a “now or never” type of event. I thank my parents, Carol and Greg Loader, as well as my partner, Max Porozny for being supportive of my decision to return to school and accomplish this personal mission. In January 2015 I hopped up the steps of the St. Boniface Research Centre ready to conquer my first day. Having never worked in a laboratory before, I was uncertain how I’d perform. There was definitely a learning curve, but I had excellent teachers. Thank you to Leslie Tworek for being the first to teach me how to conduct protein extractions, Western blotting, and cell culture experiments. As well, thank you to all past and present lab members for your aid and inspiration. I began involvement with the beans and lentils study a few months into 2015. My kindred study partner, Jaime Clark, and I would spend the next few months practically joined at the hip. Hours were spent boiling beans and lentils, feeding and tending to animal experiments, and complaining about the entire process. If it weren’t for Jaime acting as my partner in crime along the way, my current mental state may be in question. Thus, a big thank you to Jaime, not only for her hard work with this study but also for the fun times and great laughs along the way. #spillthebeans iii Of course, thank you to all who were involved in the study. This includes the R.O. Burrell Laboratory and their amazing staff (for animal care and pulse wave velocity analysis), Dennis Labossiere (for freeze-drying the pulses), and Li Ren (for conducting serum and urine analysis). I’d also like to express gratitude to the organizations that provided financial support for the study and my stipends, namely NSERC, Growing Forward 2, Manitoba Pulse and Soybean Growers, Graduate Enhancement of Tri-Council Stipends, and Research Manitoba. In addition, I’d like to sincerely thank Dr. Grant Pierce and Dr. Michel Aliani for agreeing to be on my committee, reading and offering valuable input on my thesis, and challenging me during my oral examination. Finally, this section would not be complete without two tremendous and overwhelming thank yous to my advisors, Dr. Carla Taylor and Dr. Peter Zahradka. Together, you have provided me with so much support and encouragement, and have enabled me to become the “scientist” that I am today. The two of you spent countless hours helping me with this thesis and for that I will be forever grateful. Almost twenty-seven months after starting this degree, I sit here writing this final portion of my thesis. It is amazing to reflect upon my time in the program, and contemplate the events that occurred and the wealth of knowledge I obtained. I hope to never stop exploring and I look forward to my next adventure. “If your dreams do not scare you, they are not big enough.” – Ellen Johnson Sirleaf iv Table of Contents Abstract ..................................................................................................................... ii Acknowledgements ................................................................................................... iii Table of Contents ....................................................................................................... v List of Tables ............................................................................................................ vii List of Abbreviations .................................................................................................. ix Literature Review ....................................................................................................... 1 Introduction .............................................................................................................................................. 1 Hypertension ............................................................................................................................................ 3 Determinants of Blood Pressure ....................................................................................................... 6 Cardiovascular Physiology ..................................................................................................................... 6 Endothelial Dysfunction ....................................................................................................................... 10 Vascular Remodelling and Arterial Stiffness ............................................................................... 13 Dietary Influences ................................................................................................................................... 17 Beans and Lentils ................................................................................................................................. 20 Definition of Pulses ................................................................................................................................. 20 Consumption in Canada ....................................................................................................................... 21 Production in Canada ............................................................................................................................ 22 Nutritional Content ................................................................................................................................ 24 Phenolic Compounds .............................................................................................................................. 26 Vascular Benefits ..................................................................................................................................... 30 The Spontaneously Hypertensive Rat (SHR) ............................................................................ 56 Knowledge Gaps ....................................................................................................... 60 Study Rationale ........................................................................................................ 62 Hypothesis ............................................................................................................... 64 Objectives ................................................................................................................ 65 Materials and Methods ............................................................................................ 66 Experimental Animals and Diets ................................................................................................... 66 Assessment of Disease Risk ............................................................................................................. 71 Body Weight .............................................................................................................................................. 71 Body Composition ................................................................................................................................... 72 Serum Biochemistry ............................................................................................................................... 72 Urine Biochemistry ................................................................................................................................. 72 Assessments of Arterial Stiffness and Vascular Function .................................................... 73 Blood Pressure .......................................................................................................................................... 73 v Pulse Wave Velocity (PWV)................................................................................................................. 73 Aorta Collection and Morphology .................................................................................................... 74 Western