The objective of the studies described herein are directed at developing strategies for enhancing muscle mass through understanding the biochemical basis for the impairment in protein synthesis that characterizes the metabolic response to sepsis. Sustained skeletal muscle wasting contributes to the morbidity and mortality associated with sepsis. We established that sepsis, but not sterile inflammation, induces specific maladjustments in at least two cell signaling pathways culminating in the inhibition of protein synthesis by limiting the process of mRNA translation initiation in skeletal muscle. One includes the mammalian target of rapamycin (mTOR), a serine/threonine kinase that controls phosphorylation of two other protein factors S6K1 and 4E-BP1 that regulate mRNA translation and formation of an active elF4E-elF4G complex and elF4G phosphorylation. The second is the phosphorylation of elF2Be through activation of glycogen synthase kinase-3p (GSKSfi). The following specific aims use transgenic, pharmacologic and/or nutritional approaches to validate the relative contribution and potential therapeutic relevance of these signaling pathways in the restraint in protein synthesis, and hence skeletal muscle protein, to a septic insult.
Specific Aim 1 will investigate the contribution of lowered mTOR activity to the decrease in skeletal muscle protein synthesis and the ability of leucine (or norleucine) to reverse that inhibition of mTOR and elF4G phopshorylation during sepsis. We will also determine if stimulating mTOR activity with nutrients reverses the sepsis-induced inhibition of skeletal muscle protein synthesis and the role of pharacolgical inhibition and genetic ablation of mTOR in nutrient stimulated protein synthesis. Access to the BCAT2 knockout mouse will allow us to establish whether leucine or a leucine metabolite is responsible for leucine's activation of mTOR signaling.
Specific Aim 2 will investigate the impact of reversing the septic-induced increase in phosphorylation and decrease in cellular content of elF2Be using genetically-dependent anti-inflammatory therapy, pharmacological inhibition of GSK-3(3 and over expression of non-phosphorylatable elF2Be on muscle protein synthesis during sepsis. We will test the hypothesis that treatment of muscles from septic rats with the GSK3P inhibitors LiCI, SB216763 and SB415286 or expression of active, non-phosphorylatable elF2Be will abrogate the sepsis-induced inhibition of protein synthesis in skeletal muscle. ? ? ?
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