Acute lung injury (ALI) is frequently associated with sepsis or severe blood loss, conditions in which there is increased production of reactive oxygen intermediates (ROI). ALI is characterized by the accumulation of large numbers of neutrophils in the lungs that show activation of PI3-K/Akt kinases and the transcriptional factor NF-kappaB. Data from the present project period show that ROI, particularly superoxide and hydrogen peroxide, modulate activation of PI3-K, Akt, and NF-kappaB in neutrophils and participate in the development of ALI after hemorrhage or exposure to TLR2 and TLR4 ligands. In addition, we found that PI3-K and Akt participate in the development of LPS induced ALI and also enhance nuclear translocation of NF-kappaB and phosphorylation of the p65 subunit of NF-kappaB in neutrophils stimulated through TLR2 or TLR4. Our hypothesis for work proposed in this competing continuation application is that ROI through interactions with PI3-K/Akt and related intracellular pathways modulate NF-kappaB dependent transcriptional activity and play a central role in neutrophil activation as well as in the development of sepsis or hemorrhage induced ALI.
The specific aims of this project are: 1) To characterize the distinct mechanisms by which hydrogen peroxide and superoxide modulate NF-kappaB activity in neutrophils and affect NF-kappaB activation induced through TLR2 or TLR4; 2) To characterize the mechanisms by which PI3-K and associated kinases and phosphatases (SHIP, PTEN, and PI3-Kgamma) modulate NF-kappaB activity in neutrophils stimulated by hydrogen peroxide, superoxide, TLR2, or TLR4; and 3) To examine how interactions between hydrogen peroxide, superoxide, and PI3-K/Akt modulate the development of TLR2, TLR4, or hemorrhage induced ALI. Our goal is to characterize mechanisms involving ROI, PI3-K/Akt, and related intracellular events that modulate NF-kappaB dependent transcription in neutrophils after in vitro or in vivo exposure to ligands interacting with TLR2 or TLR4, and that contribute to the development of sepsis or hemorrhage induced ALI. These studies should provide insights into fundamental mechanisms that modulate neutrophil function and contribute to the development of ALI. The proposed experiments are likely to suggest new therapeutic approaches to improve outcome from important clinical conditions, such as sepsis and hemorrhage induced acute lung injury, in which neutrophil dependent pathways play major roles. ? ? ?
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