(300f) In Vivo Metabolic Fluxes In Rat Livers: Effect of Burn Injury

Available animal models provide a limited understanding of the complex, integrated effects of burn injury systemically and on the liver; in vivo studies commonly lack a thorough analysis of metabolic pathways, whereas ex vivo studies such as isolated perfusion carry an increased potential risk of experimental artifacts. The purpose of this study is to develop an in vivo animal model for the comprehensive investigation of the effects of burn injury. The data is presented both as concentrations of metabolites and their fluxes to enable the separation of systemic changes from changes brought about by the liver. A comprehensive representation of the metabolic pathways implicated in in vivo normal vs. burn liver can help identify differences in intracellular pathways for metabolic intervention using therapeutic targets to reduce complications associated with thermal burn injury.

Three groups of Sprague Dawley rats received a sham, 20% or 40% TBSA (Total Body Surface Area) burn and on the fourth day post-injury, following an 18-24 hour fast, flow rates of the portal vein, hepatic artery and suprahepatic vena cava were measured per rat followed by blood sampling of the suprahepatic vena cava, portal vein and hepatic artery. The liver was then excised and weighed for the purposes of normalizing all the data obtained. Blood samples were processed for blood gases and multiple metabolites as well as amino acids were measured with HPLC.

The results of this study verify the use of a 20 % and 40% TBSA burn in 6 week old Sprague Dawley rats as a clinically relevant in vivo animal model to study the effects of this kind of trauma. The data shows significant metabolic alterations as a consequence of burn injury and clearly demonstrates the role the liver plays in facilitating hypermetabolism. Metabolic Flux analysis (MFA) shows an increase in pyruvate synthesis, intracellular glutamate synthesis from histidine and increased tyrosine and phenylalanine uptake and metabolism for the 20 % burn injury conditions as compared to the sham control condition. For the 40 % burn condition, a significant increase in urea cycle, gluconeogenic, TCA cycle, acetyl-CoA synthesis, oxygen uptake and mitochondrial metabolism is observed as compared to the sham condition.

In addition to comparison of in vivo conditions, we performed a comparison of in vitro perfusion vs. in vivo conditions for the available data that revealed striking differences. In vitro perfusion data shows an increase in urea cycle fluxes, glutamine to glutamate conversion, asparagine to aspartate conversion, triacylglycerol conversion to beta-hydroxybutyrate and acetoacetate as compared to the in vivo perfusion data. For the 20 % burn condition, in vitro burn perfusion shows an increase in few hepatic fluxes viz. glycine, histidine, glutamine, arginine, threonine uptake with decrease in alanine, phenylalanine, asparagine and lactate uptake rate. Intracellularly, there is an increase in glutamine and histidine to glutamate metabolism and acetyl-CoA to â-hydroxybutyrate conversion. Overall, a similarity in PPP, TCA cycle and gluconeogenic fluxes between in vivo and in vitro burn injury models might imply application of appropriate metabolic intervention previously applied in the context of therapeutic efficacy of antioxidants in vitro to revert the hepatic hypermetabolic state post-burn injury in vivo.