(197e) Metabolic Response of Perfused Livers to Various Oxygenation Conditions

Isolated liver perfusion systems have been used to characterize intrinsic metabolic changes in liver as a result of various perturbations, including systemic injury, hepatotoxin exposure, and warm ischemia. Most of these studies were done using hyperoxic conditions (95% O₂) but without the use of oxygen carriers in the perfusate [1-4]. Inadequate oxygenation could by itself alter gene expression levels [5] and/or promote anaerobic pathways, including glycolysis. Prior literature data do not clearly establish the impact of oxygenation, and in particular that of adding oxygen carriers to the perfusate, on the metabolic functions of the liver. Thus, the importance of using oxygen carriers in liver perfusion studies remains controversial, and the full ramifications of the impact of oxygenation have not been well characterized.

In this study, the effects of three modes of oxygen delivery to perfused livers were compared: normoxic (arterial) perfusate (21% O₂), hyperoxic perfusate (95% O₂), and hyperoxic perfusate with oxygen carriers (95% O₂ + 10% hematocrit using bovine red blood cells). In all cases, the rat liver was perfused in-situ via the portal vein at constant flow rate, 3 mL/min/g liver. The hepatic artery and the suprarenal vena cava were ligated, and the liver outflow from the hepatic vein collected through the catheter which was cannulated into the inferior vena cava via the right atrium. The concentration changes of important metabolites in the perfusate were analyzed in order to determine net extracellular fluxes which have been applied to a stoichiometric-based optimization problem incorporating flux balance and metabolic pathway analyses (elementary mode analysis). The flux distribution vectors as well as possible active elementary modes were uniquely identified by maximizing the activity of short pathways [6].

We found that perfused livers consumed oxygen at in vivo rates only when the perfusate contained RBCs. Even when using 95% O₂, in the absence of oxygen carriers, oxygen uptake was only half the in vivo rate, urea and ketone body production were significantly decreased, and metabolic pathway analysis suggests that significant anaerobic glycolysis occurred. Conversely, when RBCs were used, glucose production from lactate and glutamate, as well as pathways related to energy metabolism were upregulated. The improved physiological relevance of a perfusion system using oxygen carriers makes it a more attractive tool to investigate the effect of various perturbations on liver metabolism, including the response to toxicants and drugs, as well as disease conditions known to alter liver metabolism, such as burns and trauma leading to systemic hypermetabolism.

References

1.         Banta, S., et al., Contribution of gene expression to metabolic fluxes in hypermetabolic livers induced through burn injury and cecal ligation and puncture in rats. Biotechnology and Bioengineering, 2007. 97(1): p. 118-137.

2.         Arai, K., et al., Intrahepatic amino acid and glucose metabolism in a D-galactosamine-induced rat liver failure model. Hepatology, 2001. 34(2): p. 360-371.

3.         Lee, K., et al., Profiling of dynamic changes in hypermetabolic livers. Biotechnology and Bioengineering, 2003. 83(4): p. 400-415.

4.         Orman, M.A., et al., Metabolic Flux Determination in Perfused Livers by Mass Balance Analysis: Effect of Fasting. Biotechnology and Bioengineering, 2010. 107(5): p. 825-835.

5.         Rosmorduc, O. and C. Housset, Hypoxia: A Link between Fibrogenesis, Angiogenesis, and Carcinogenesis in Liver Disease. Seminars in Liver Disease, 2010. 30(3): p. 258-270.

6.         Orman, M.A., et al., Pathway analysis of liver metabolism under stressed condition. Journal of Theoretical Biology, 2011. 272(1): p. 131-140.