Sphingolipid metabolites such as ceramides and sphingoid bases are important modulators of cell survival, cell proliferation, angiogenesis, migration and vascular integrity. Our in vitro and in vivo data indicate that sphingosine 1-phosphate (SIP), a naturally occurring bioactive lipid, is an essential angiogenic factor that regulates vascular endothelial cell (EC) barrier permeability, a crifical and defining feature of ALL SIP acts extracellularly through its G-protein coupled S1P1-5 receptors and there is evidence that supports an intracellular role of SIP in calcium release and cell proliferation.SIP mediated cellular responses are regulated by its synthesis, catalyzed by sphingosine kinases (SphKs), and degradation mediated by SIP phosphatases, and SI P lyase (SI PL). In the course of our studies, we observed that alteration of SphKs and SI PL expression in animals or human lung microvascular endothelial cells (jHLMVECs) modulated LPSnduced inflammatory responses and barrier function suggesting a key role for these enzymes in maintenance of barrier function and integrity. Project 1 will test the hypothesis that """"""""Modulation of intracellular S1P by SphKs and S1 PL of the endothelium regulates LPS-induced lung inflammation and barrier dysfunction"""""""". SA#1 will define molecular mechanisms of SphKI and 2 and SI PL regulation of LPSinduced inflammatory responses in animal and HLMVEC culture models.. We will utilize a combination of molecular, biochemical and lipidomics approach to evaluate and quantify intracellular SIP accumulation and determine its role in modulation IL-6 release and barrier dysfunction in the endothelium. SA#2 will characterize ALI associated single nucleotide polymorphisms (SNPs) in SphKs and SI PL and conduct SNPspecific association studies. SA#3 will determine the effectiveness of SphK activator(s) and S1PL inhibitor(s) as potential therapeutic agents in murine model of LPS-induced lung injury. Together, this Project will greatly advance studies proposed in other Projects, clarify the key roles of SphKs and SI PL in ALI pathobiology and facilitate the development of novel therapeutic agents of sphingolipid metabolism in ameliorating sepsisinduced inflammation and lung injury.
Acute and sub-acute inflammatory lung injuries are common but devastating disorders resulting from insults such as sepsis, and radiation therapy for thoracic malignancies with a mortality rate of 30-50%! Specific. therapies currently do not exist for the treatment of ALI. The propsed studies will define key sphingolipid metabolizing enzymes as novel targets of therapy against sepsis-induced lung injury.
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