Unveiling the role of age in MASLD development and biological process activation: Implications for translational research.

Age is a risk factor for severity and progression of Metabolic Dysfunction-Associated Steatotic Liver Disease (MASLD), yet its specific contributions to liver injury and molecular alterations remain unclear. Current animal models often overlook this variable. This study investigates how aging influences MASLD progression in a murine model, focusing on histological, biochemical, and transcriptomic differences. Young (2-month-old) and middle-aged (10-month-old) C57BL/6N male and female mice were fed chow (CD) or high-fat diet (HFD) for 20 weeks. Metabolic and biochemical parameters were assessed, and liver tissue analyzed by histology and RNA sequencing. Differentially expressed genes were identified with Limma, followed by gene ontolgy enrichment. Translational relevance was evaluated using the MASLD Human Proximity Score (MHPS). Middle-aged males developed insulin resistance and more severe hepatic injury than young males after HFD. Histological and biochemical assessments confirmed advanced disease in middle-aged mice, with extensive lipid accumulation, hepatocellular hypertrophy, inflammation and fibrosis. Females showed milder damage. Diet was the main determinant of transcriptomic changes in males: young HFD mice activated tissue remodeling pathways, whereas middle-aged mice showed inflammatory activation and reduced energy metabolism. In females, transcriptomic responses were attenuated, with metabolic and stress-related changes restricted mostly to young HFD animals. MHPS ranked the middle-aged male model highest in similarity to human MASLD. Age is a critical determinant of hepatic responses to dietary stress, with sex-specific modulation. The middle-aged male model closely mirrors human MASLD, providing a more relevant preclinical platform to improve patient stratification and therapeutic discovery.