Abstract
Nonalcoholic fatty liver disease (NAFLD) is associated with metabolic syndromes spanning a wide spectrum of diseases, from simple steatosis to the more complex nonalcoholic steatohepatitis. To identify the deregulation that occurs in metabolic processes at the molecular level that give rise to these various NAFLD phenotypes, algorithms such as pathway enrichment analysis (PEA) can be used. These analyses require the use of predefined pathway maps, which are composed of reactions describing metabolic processes/subsystems. Unfortunately, the annotation of the metabolic subsystems can differ depending on the pathway database used, making these approaches subject to biases associated with different pathway annotations, and these methods cannot capture the balancing of cofactors and byproducts through the complex nature and interactions of genome-scale metabolic networks (GEMs). Here, we introduce a framework entitled Minimum Network Enrichment Analysis (MiNEA) that is applied to GEMs to generate all possible alternative minimal networks (MiNs), which are possible and feasible networks composed of all the reactions pertaining to various metabolic subsystems that can synthesize a target metabolite. We applied MiNEA to investigate deregulated MiNs and to identify key regulators in different NAFLD phenotypes, such as a fatty liver and liver inflammation, in both humans and mice by integrating condition-specific transcriptomics data from liver samples. We identified key deregulations in the synthesis of cholesteryl esters, cholesterol, and hexadecanoate in both humans and mice, and we found that key regulators of the hydrogen peroxide synthesis network were regulated differently in humans and mice. We further identified which MiNs demonstrate the general and specific characteristics of the different NAFLD phenotypes. MiNEA is applicable to any GEM and to any desired target metabolite, making MiNEA flexible enough to study condition-specific metabolism for any given disease or organism.