A quantitative metabolomic profiling and corresponding modeling approach of hepatocytes with activated Wnt/ß-Catenin signaling

Abstract

The maintenance of liver homeostasis relies on the metabolic zonation, where pericentral (pc) and periportal (pp) zones differ in their enzyme composition, content and activity. To carry out a metabolic fingerprint along the porto-central axis of the sinusoid is of high relevance for establishing a suitable systems biology approach aiding a better mechanistic understanding of hepatocytes heterogeneity with their different cellular functions. Here, we present a metabolic modeling approach for mouse hepatocytes based on metabolite profiles in the cell and cell culture medium of of APCloxPneo mice, with an over-activated Wnt/β-Catenin signaling and control mice, with normal Wnt/β-Catenin activity. The pathway has been shown to play a key role in the functional heterogeneity of hepatocytes and fundamentally influence the metabolic zonation in adult liver. APCloxPneo mice develop an expanded zone of hepatocytes with pericentral functions, due to the partial reduction in the Adenomatous Polyposis Coli protein (APC) expression in hepatocytes. Therefore it is well suited for evaluation of the model. Hepatocytes were isolated and cultivated for up to 48 hours and medium was collected at different time points. The culture medium was analyzed with GC-MS and LC-MS methods for quantitative analyses of the compound classes' amino acids, bile acids and organic acids. A metabolic model consisting of 42 species and 67 interspecies interactions has been established to analyze the flux distribution patterns in different hepatocyte populations. The software system VANTED (Visualization and Analysis of Networks containing Experimental Data) has been used to interpret the different patterns of metabolites, occurring in various hepatocyte populations. To characterize the functional specialization of hepatocytes clustering with self-organizing maps (SOM) was performed. The flux balance analysis (FBA) was carried out to get a general idea of the metabolic capabilities of different hepatocyte populations. The model has been validated by comparison to published data of pp/pc and APCloxPneo hepatocytes. Conclusion: The model offers a valuable tool for understanding the role of hepatocyte heterogeneity in liver homeostasis under normal conditions and possibly also in diseased states. Further, it could serve as an important sub-module for a multi-scale liver model of the cellular metabolism bridging single-cell metabolism to the tissue and the whole-organ level.