Integrated characterization of cellular physiology underlying hepatic metabolism

Abstract

The macroscopic metabolic phenotype of a cellular system, such as insulin resistance, is the result of the integration of many hundreds or thousands of preceding cellular events, which culminates in the cell's final response to a perturbation in the environment. The data provided by DNA microarrays and multiple types of metabolic measurements can be integrated to reconstruct the actions taken by a cellular system to arrive at a particular metabolic response to a stimulus, elucidating the underlying physiology. We employed this integrated approach for the characterization of hepatic metabolism. First, we implemented a novel method for functional genomics. The metabolic response of hepatoma cells to the depletion and repletion of glutamine was characterized in time course measurements of metabolic fluxes and metabolite pool sizes. DNA microarrays characterized the expression profiles. The metabolic data were correlated with the microarray data to identify coregulated clusters of genes. This study contributed to our understanding of glutamine metabolism in hepatomas, and advanced the field of functional genomics. Next, we identified the hexosamine biosynthetic pathway (HBP) as a mechanism for hyperglycemia-induced hepatic insulin resistance.

(cont.) Glycogen deposition and glucose production data in mouse hepatocytes confirmed that HBP activity was negatively correlated with insulin sensitivity. Metabolite profiling data confirmed that prolonged incubation in hyperglycemic conditions raised the levels of hexosamine intermediates by saturating upper glycolysis. Our data, along with previous work in muscle and adipose tissue, underline the increasingly important role of the HBP in regulating insulin action and energy homeostasis. A dysfunctional HBP may contribute to the pathophysiology of Type 2 diabetes. Finally, we analyzed the control structure of the glucose production bioreaction network. We systematically perturbed the network and analyzed the effects on the fluxes. We found that gluconeogenesis was the dominant flux, and therefore regulation of gluconeogenesis determined the glucose production phenotype. G6Pase was identified as the enzyme in gluconeogenesis controlling the glucose production phenotype, whereas PEPCK played a secondary role. Our conclusions here give insight into the physiology underlying the regulation and dysregulation of hepatic glucose production with possible application to the treatment of Type 2 diabetes.