The Inflammatory Consequence of Arsenic-Mediated Urothelial Endocrine Disruption

Inorganic arsenic (iAs) is the highest-ranking toxin on the U.S. substance priority list. Chronic exposure through natural water sources is linked to a broad range of adverse health effects, including pathological changes in the urinary bladder. Due to the make up of the urothelial cell, iAs readily accumulates to elicit toxic effects. However the specific mechanisms of arsenic-induced urothelial injury are not fully characterized. Upon cell injury there is rapid influx of inflammatory mediators and increase in cellular inflammatory signaling. The glucocorticoid receptor (GR) is an anti-inflammatory transcription factor, critical o maintain the balance between pro- and anti-inflammatory processes. iAs is classified as an environmental endocrine disruptor and is known to inhibit GR transcription factor function of target genes. Angiopoietin-like protein 4 (ANGPTL4) is a target gene of GR that plays a role in wound healing in response to epithelial injury. Notably, ANGPTL4 expression is decreased following iAs exposure in human urothelial cells (HUC). Moreover, treatment of HUC with inflammatory agents increases ANGPTL4 expression, and knockdown of ANGPTL4 decreases wound closure capabilities. However, it is unknown how arsenic disrupts the GR/ANGPTL4 axis in response to urothelial injury. Thus, the hypothesis of this proposal contends that iAs decreases GR-mediated expression of ANGPTL4 and disrupts normal urothelial response to injury by decreasing wound closure and sustaining the inflammatory response. The hypothesis will be addressed by completing the following two specific aims. First, the kinetics, specificity, and mechanism of arsenic inhibition on GR transcription factor function will be explored using reporter assays, DNA-protein binding assays, and mutations at suspected iAs binding sites within the GR DNA binding domain. Second, HUC with varying combinations of GR mutations and ANGPTL4 knockdown will be subjected to functional assays to assess changes in cell motility and inflammatory response. The changes in wound healing and inflammation will be observed with and without iAs treatment. Results from these aims will provide a mechanism of arsenical endocrine disruption in the bladder and validate ANGPTL4 as a novel GR target in response to injury and inflammation. Further, aberrant physiological responses and mechanisms after urothelial injury as a result of iAs exposure will be determined. The significance of this study applies to a broad range of arsenic-induced disease in the skin, lung, liver, and kidney. There may also be improved characterization of additional endocrine disease linked to chronic iAs, such as diabetes. Because inflammation is an underlying mechanism in many chronic diseases, this proposal has the potential to translate into preventative measures and bring more awareness to the role of the environment on human health, a concept that is not yet fully appreciated. Overall, completion of this study will ensure my comprehensive research training and contribute to the development of a successful career as an independent clinician-scientist of environmental medicine and disease prevention. PUBLIC HEALTH RELEVANCE: Arsenic exposure affects select populations due to high levels in drinking water. This study explores how arsenic inhibits gene expression in response to injury of the urinary bladder. The consequences may be sustained inflammation and poor wound healing, which contributes to irreversible changes in the bladder.