Infectious complications, especially pneumonia, are six times more common in burn patients who have been consuming alcohol prior to injury, than patients with burn alone. In the studies outlined below, we will use a murine model of ethanol exposure and burn injury to determine the mechanisms by which ethanol consumption prior to burn injury renders mice with decreased pulmonary bacterial clearance and increased mortality. We propose that ethanol affects the phagocytic function of circulating neutrophils, and since neutrophils are the first innate immune cells to response to bacterial infection, pulmonary bacterial clearance is compromised in the burned patient with prior ethanol exposure.
In aim 1, we will determine if mice exposed to ethanol prior to burn injury are more susceptible to a bacterial challenge given intratracheally after the burn by examining 1) the survival, 2) the degree of pulmonary bacterial clearance, and 3) septicemia in mice from all groups.
Aim 2 will focus on the neutrophil infiltration in the lungs after ethanol and burn injury followed by pulmonary infection. First, we will quantitate neutrophil infiltration in the lung by using three methods; 1) hematoxylin & eosin stained lung sections, 2) immunohistochemistry, using an anti- Gr-1 antibody (neutrophil marker) in combination with MOMA-2 (macrophage marker) in order to exclude the monocyte population expressing both markers GR-1 and MOMA-2. 3). ln order to determine the composition of cells in the alveolar space, we will perform bronchoalveolar lavage, label the cells with specific antibodies, and analyze the samples by flow cytomerty. If we determine, that there are differences in neutrophil infiltration into the lungs of infected mice exposed to ethanol and burn injury when compared to burn alone, we will assess cytokine and chemokine production in the lungs required for recruitment after injury.
In aim 3 of the proposal, we will examine the effects of ethanol and burn injury on neutrophil activation and pahgocytosis. To determine neutrophil activation, we will study the superoxide and elastase production by neutrophils. The phagocytic ability of neutrophils will be determined by using an in vitro phagocytosis assay. The studies proposed in this application are critical for the understanding of how ethanol exposure prior to burn injury modulates the inflammatory response and contributes to decreased pulmonary bacterial clearance. Understanding the mechanisms leading to increased morbidity and mortality after such injury, may lead to the development of therapeutic targets that can potentially help patient that have been onsuming alcohol prior to burn injury overcome an infectious challenge. ? ? ?
| Karavitis, John; Murdoch, Eva L; Deburghgraeve, Cory et al. (2012) Ethanol suppresses phagosomal adhesion maturation, Rac activation, and subsequent actin polymerization during FcýýR-mediated phagocytosis. Cell Immunol 274:61-71 |
| Murdoch, Eva L; Karavitis, John; Deburghgraeve, Cory et al. (2011) Prolonged chemokine expression and excessive neutrophil infiltration in the lungs of burn-injured mice exposed to ethanol and pulmonary infection. Shock 35:403-10 |
