PROJECT SUMMARY Over 160,000 abandoned mine waste sites are concentrated in the western United States and continue to degrade the environment with elevated heavy metals penetrating the surrounding sediment and groundwater. The sedimentary dust and water used by communities located near abandoned uranium mine (AUM) sites often exceeds the maximum contamination level for uranium (U, EPA max level 30 μg/L). Chronic exposure to environmental toxins and heavy metals has been linked to intestinal inflammation, increased susceptibility to pathogen-induced diseases, and higher incidences of colorectal cancer; all of which have been steadily increasing in prevalence for the past 40 years. The negative effects of heavy metals on the intestinal microbiota, barrier permeability, and inhibition of intestinal epithelial healing have been described; however, transcriptomic changes within the intestinal epithelial cells are largely unknown. Upon ingestion, uranium is poorly absorbed systemically (~1.5%) and thus distributed through the entire intestine, suggesting even low U levels are sufficient to cause intestinal damage. Thus, our goals are to characterize particulate dust (referred to as U-bearing dust, UBD) as an environmental toxicant that damages the intestinal epithelia and determine the molecular changes that develop as a response to this injury. We have previously characterized the intestinal mucus layer in human intestinal organoids (HIOs) under homeostasis and pathogen infected conditions, to characterize HIOs as an intestinal pathophysiological model. Our preliminary data of HIOs exposed to UBD found that chemical exposure negatively impacts the secretory lineage in the colon, specifically the mucus- producing goblet cells and hormone-producing enteroendocrine cells. Thus, we hypothesize that UBD has a direct effect on: 1) the mucosal surface, represented by a thinning mucus layer and goblet cell dysfunction and 2) the intestinal enteroendocrine cells in the crypt. This leads to increased susceptibility to intestinal disorders. The following Aims will address these questions in a mechanistic manner. The first aim will determine the impact of UBD on mucus layer thinning, resulting pro-inflammatory response, and subsequent goblet cell expansion. The second aim will examine the autophagy pathway in regulating changes in enteroendocrine hormone secretion and the transcription factor PROX1 as the non-canonical driver of enteroendocrine cell expansion following UBD chemical injury. There is a need to understand how heavy metals, especially those naturally occurring as particulates, induce molecular changes in the intestinal epithelia of exposed individuals to predispose them to intestinal disorders. Successful completion of this proposal will help us understand the mechanistic underpinnings of environmental-induced injury that may promote colitis or related diseases, identify potential therapeutic targets, and identify genetic variations that may contribute to susceptibility.

PROJECT NARRATIVE Abandoned uranium mines, primarily located on tribal land in the southwest United States, are not remediated and continue to degrade heavy metals into the surrounding environment, contaminating the sedimentary dust and ground water with significant concentrations of uranium. Epidemiological data suggests a direct link between inhalation and ingestion of uranium mine dusts with higher incidences of intestinal inflammation and diseases. This proposal aims to define and characterize the mechanisms underlying uranium-induced intestinal dysfunction and long-term consequences in the intestinal epithelia.