Epigenetic Influence of Diet on Bladder Physiology - PROJECT SUMMARY The high incidence and prevalence of Lower Urinary Tract Dysfunction (LUTD), a common factor in many bladder disorders in African populations (about 66%, 20% and 6% of the male population show mild, moderate and severe symptoms respectively), has been related to environmental and lifestyle factors such as diet. We have previously reported in both normal and diseased bladders, that diet alters bladder morphology, specifically cell hyperplasia and hypertrophy, causing changes in function and influencing key calcium-signalling mechanisms responsible for detrusor contractility. Understanding of the mechanisms responsible for LUTD and other bladder disorders is still very limited; epigenetics may play a role. Thus, the long-term goal is to determine the epigenetic effects of diet on bladder physiology and pathophysiology, while also attaining the level of training and expertise required for me to become an independent researcher. The overall objectives in this application are to (i) characterize the influence of diet on specific histone modifications and gene expression changes in the bladder and (ii) relate any observed alterations in histones and gene expression to phenotypic and functional changes in the bladder. The central hypothesis is that epigenetic modifications contribute to the molecular changes underlying alterations in bladder morphology and function with different diets. The rationale for this project is that identification of the epigenetic effects of diet in the bladder will provide a foundation, and supportive preliminary data, for subsequent studies on the roles played by diet in the normal bladder and in bladder dysfunction. Diet, being a modifiable factor with great public health impact, could provide an insight into the mechanisms by which LUTD occur. This project also offers me the opportunity to establish my independence as an epigeneticist. The central hypothesis will be tested by pursuing three specific aims: 1) Identify changes in bladder morphology and function after consumption of different diets (high fat, high carbohydrate, and high protein diets); 2) Identify bladder transcriptome changes after dietary treatment using RNAseq; and 3) Identify effects of the diets on histone acyl marks for proliferative status (H3S10P, H3S28P), transcriptional activation (H3K9Ac, H3K27Ac) and DNA replication (H4K5Ac, H4K12Ac) in the bladder of rats from each dietary group after feeding. Under the first aim, excised whole rat bladder tissue will be used to identify changes in bladder weight, detrusor contractility, and bladder histology (haematoxylin and Eosin stain, and immunohistochemistry) to determine morphological and functional changes as a result of each diet. For the second aim, RNA sequencing will be done to identify global changes in transcriptome abundance in each dietary group. To achieve the third aim, protein separation techniques (SDS-PAGE) and western immunoblotting will be employed to identify changes in specific histone targets. The proposed research is innovative, because it focuses on a new direction for research into bladder physiology and pathophysiology, investigating diet, a common modifiable public health factor in all patients and a common denominator in the aetiologies of several non-communicable diseases, and how its effects on epigenetic modifications, influencing transcription and expression of genes could result in LUTD. The proposed research is significant because it is expected to advance scientific knowledge on the mechanisms of bladder function, and provide an initial standpoint and data upon which subsequent studies on the epigenetic roles played by diet in bladder dysfunction will build. It will also provide data to support new intervention strategies for the management of LUTD and other non-communicable dieases.
