Project Summary/Abstract Alcohol-related traumatic and burn injuries remain a considerable health and economic burden to the American society. Studies have shown that patients who are intoxicated at the time of injury are more susceptible to infection and exhibit significantly higher morbidity and mortality compared to burn patients who are not intoxicated at the time of injury. Similar to clinical findings, studies performed in experimental conditions also suggest that ethanol intoxication at the time of burn injury decreases host resistance to infectious complications. Yet, the mechanism by which ethanol enhances post burn pathogenesis remains largely unclear. Gut barrier dysfunction is frequently associated with ethanol exposure and major injury. We have shown that the ethanol intoxication combined with moderate burn injury causes intestinal tissue damage, leakiness, and a significant increase in bacterial translocation within 24 hours after injury. We further found that ethanol combined with burn injury increases bacterial load (primarily Gram-negative bacteria) in the gut. Such a shift in gut microbiota may perturb bacteria/host interactions to produce a microenvironment conducive for neutrophil accumulation and activation, leading to tissue damage and gut leakiness following ethanol and burn injury. Our hypothesis is that accumulation of intestinal Gram-negative (i.e. enterobactericeae) bacteria following ethanol and burn injury perturbs gut microbiota-epithelial interactions, which become exacerbated by altered microRNA homeostasis, thus, culminating in gut inflammation and barrier disruption. The hypothesis will be tested in 3 AIMS in a well-established mouse model of ethanol intoxication and burn injury. Studies in Aim1 are to delineate the mechanism by which ethanol and burn induced changes in gut bacteria influence gut barrier integrity following injury. We will use TLR-2, TLR-4, Nod-1 and/or Nod-2 deficient mice to identify the key PRR by which intestine epithelial cells detect Gram-negative bacteria and/or their products. Aim 2 will determine whether restoration of microRNA biogenesis or miR-150 in intestine epithelial cells following alcohol and burn injury reduces gut inflammation and prevents barrier disruption. Furthremore studies in Aim3 will determine whether treatment of animals with probiotics re-establishes gut microbiota and gut barrier following burn injury. The findings from these studies will reveal a novel role for gut microbiota in altered gut barrier function following ethanol intoxication and burn injury and in turn may help in developing new therapeutic strategies for patients suffering from a combined insult of ethanol intoxication and burn injury.