Glucokinase Inactivation Reduces Lysophosphatidylcholine Accumulation to Alleviate High‐Fat Diet–Induced Hepatic Steatosis

ABSTRACT Partial inactivation of glucokinase (GCK) is typically characterized by mild hyperglycemia and a favorable lipid profile compared to type 2 diabetes. Previous studies have shown that GCK activity influences serum lipid profiles in a diet‐dependent manner; however, its role in hepatic lipid metabolism in the context of metabolic dysfunction‐associated steatotic liver disease (MASLD) remains unclear. To address this, we utilized a newly established heterozygous GCK mutation knock‐in mouse model (GCK Mut ) fed either a normal diet (ND) or a high‐fat diet (HFD). Under ND conditions, GCK Mut mice developed mild hyperglycemia without overt hepatic injury but displayed reduced hepatic glycogen storage, likely due to decreased energy flux. Metabolomic analyses further revealed substantial reprogramming of hepatic amino acid and lipid metabolism in GCK Mut mice. Notably, levels of lysophosphatidylcholines (LPCs)—bioactive metabolites implicated in lipotoxicity and the pathogenesis of MASLD—were significantly reduced, as confirmed by ELISA. Under HFD conditions, GCK inactivation markedly attenuated hepatic lipid accumulation, as demonstrated by biochemical quantification and histological analysis. This protective effect was associated with downregulation of genes involved in de novo lipogenesis and fatty acid uptake, as revealed by transcriptomic analyses of primary hepatocytes. Moreover, both the expression of phospholipase A2 (PLA2) and its product LPC were significantly reduced in GCK Mut mice, whereas pharmacologic activation of GCK increased hepatic LPC accumulation. These findings suggest that partial GCK inactivation reprograms hepatic metabolism and mitigates lipid‐induced hepatic stress, highlighting reduced hepatic GCK activity as a potential therapeutic strategy for early intervention in MASLD.