Age is a critical determinant of hepatic responses to dietary stress, with sex-specific modulation, and the middle-aged male model closely mirrors human MASLD, providing a more relevant preclinical platform to improve patient stratification and therapeutic discovery.
Key Findings
Results
Middle-aged male mice developed insulin resistance and more severe hepatic injury than young males after high-fat diet feeding.
Young (2-month-old) and middle-aged (10-month-old) C57BL/6N male and female mice were fed chow diet (CD) or high-fat diet (HFD) for 20 weeks.
Middle-aged males showed insulin resistance as a metabolic consequence of HFD not observed to the same degree in younger animals.
Histological and biochemical assessments confirmed advanced disease in middle-aged mice, including extensive lipid accumulation, hepatocellular hypertrophy, inflammation, and fibrosis.
Female mice showed milder hepatic damage compared to males regardless of age group.
Results
Diet was the main determinant of transcriptomic changes in male mice, with age determining the nature of pathway activation.
Differentially expressed genes were identified using the Limma statistical framework followed by gene ontology enrichment analysis.
Young HFD male mice activated tissue remodeling pathways in response to dietary stress.
Middle-aged HFD male mice showed inflammatory pathway activation and reduced energy metabolism transcriptomic signatures.
The age-specific transcriptomic responses suggest distinct disease mechanisms operating at different life stages.
Results
Transcriptomic responses in female mice were attenuated compared to males, with metabolic and stress-related changes restricted mostly to young HFD females.
Female mice of both age groups showed milder histological and biochemical damage than their male counterparts.
Transcriptomic changes in females were less pronounced than in males across diet and age conditions.
Metabolic and stress-related transcriptomic changes in females were largely confined to young HFD animals.
These findings indicate sex-specific modulation of hepatic responses to dietary stress.
Results
The MASLD Human Proximity Score (MHPS) ranked the middle-aged male HFD model highest in similarity to human MASLD.
Translational relevance was evaluated using the MASLD Human Proximity Score (MHPS), a metric for comparing animal model transcriptomics to human disease.
Among all groups tested, the middle-aged male HFD model received the highest MHPS ranking.
This indicates that middle-aged male mice on HFD most closely mirror the molecular landscape of human MASLD.
The result supports the use of middle-aged male mice as a more relevant preclinical platform for therapeutic discovery and patient stratification research.
Background
Current animal models of MASLD often overlook age as a variable, despite age being a recognized risk factor for disease severity and progression.
The study identifies a gap in the preclinical literature where age is frequently not controlled or reported as a study variable.
Age is established as a risk factor for severity and progression of Metabolic Dysfunction-Associated Steatotic Liver Disease (MASLD).
The specific contributions of aging to liver injury and molecular alterations had remained unclear prior to this investigation.
The authors argue this oversight limits translational relevance of existing preclinical MASLD research.
Methods
Both male and female C57BL/6N mice of two age groups were included, enabling sex- and age-stratified analyses of MASLD development.
The study used a 2×2×2 factorial design: two sexes (male and female), two ages (2-month-old young and 10-month-old middle-aged), and two diets (chow diet and HFD).
HFD was administered for 20 weeks in all diet groups.
Metabolic and biochemical parameters were assessed alongside histological and RNA sequencing analyses of liver tissue.
The inclusion of both sexes allowed for identification of sex-specific modulation of hepatic responses.
What This Means
This research suggests that the age of mice used in laboratory studies of fatty liver disease (formally called Metabolic Dysfunction-Associated Steatotic Liver Disease, or MASLD) has a major impact on how closely those animal models reflect the human disease. Researchers fed young (2-month-old) and middle-aged (10-month-old) mice a high-fat diet for 20 weeks and measured changes in metabolism, liver tissue damage, and gene activity. They found that middle-aged male mice developed more severe liver disease — including fat accumulation, inflammation, and scarring — compared to younger males, and that the specific biological processes activated in the liver depended strongly on both the age of the animal and its sex. Female mice showed much milder disease overall.
When the researchers compared the gene activity patterns from each mouse group to patterns seen in human MASLD patients, middle-aged male mice on a high-fat diet were the closest match to human disease. This was measured using a tool called the MASLD Human Proximity Score. Younger male mice activated pathways related to tissue remodeling, while middle-aged males showed more inflammation and reduced energy processing — patterns more consistent with what is seen in human patients.
This research suggests that many previous animal studies of fatty liver disease may have used mice that were too young to accurately model how the disease works in adult humans. By showing that middle-aged male mice more faithfully replicate human MASLD biology, this work implies that future preclinical drug testing and disease research should take animal age — and sex — into account. Doing so could improve the chances that treatments developed in the lab will actually work in human patients.
Lameroli Mauriz L, Scelza-Figueredo A, Orellano M, Martínez Ruiz B, Bayo J, Fiore E, et al.. (2026). Unveiling the role of age in MASLD development and biological process activation: Implications for translational research.. Journal of physiology and biochemistry. https://doi.org/10.1007/s13105-026-01217-8