HMG-1/Y Regulation of NOS2 Expression in Endotoxemia

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DESCRIPTION (provided by applicant): Sepsis is a disease process representing the systemic response to severe infection and causes high morbidity and mortality. In a subset of sepsis patients, toxins such as lipopolysaccharide (LPS) are released by bacteria and activate an inflammatory response, including the release of cytokines such as interferon (IFN)-gamma. The inducible form of nitric oxide synthase (NOS2) plays an important role in endotoxemia through overproduction of nitric oxide (NO), which has been implicated in a number of pathophysiologic mechanisms of endotoxemia. The objectives of the proposal are: (a) to gain skills and knowledge in the investigation of the role of the architectural transcription factor high mobility group-I/Y (HMG-I/Y) in NOS2 expression under conditions of endotoxemia; (b) to gain experience in teaching others; and (c) to achieve the necessary expertise to lead an independent research group. This training at Brigham and Women's Hospital will incorporate the participation in several structured research activities leading to scientific independence. The laboratory is equipped with the resources required for research in cellular and molecular biology, animal physiology, and histology. The overall hypothesis of our proposed work is that HMG-I/Y facilitates the LPS/IFN-gamma synergistic induction of NOS2 and may be an important modulator of NO production in endotoxemia. In AIM 1 we will determine whether HMG-I/Y interacts with the transcription factors important for LPS/IFN-gamma induction of NOS2 using in vitro assays. In AIM 2, we will determine whether altering HMG-I/Y binding to DNA in vitro and in vivo alters NOS2 gene expression and animal survival under conditions of endotoxemia. In AIM 3, we will elucidate the role of HMG-I/Y and its effect on NOS2 expression and animal survival during endotoxemia using a transgenic mouse expressing a dominant-negative form of HMG-I/Y in the vasculature. These experiments will allow us to determine the role of HMG-I/Y in regulating NOS2 expression during endotoxemia and may reveal a novel target for improving outcomes from sepsis.