Transcriptomic alterations associated with the progression of metabolic dysfunction-associated steatotic liver disease in CC042 Collaborative Cross mice.

Metabolic dysfunction-associated steatotic liver disease (MASLD), previously known as nonalcoholic fatty liver disease, has emerged as the most common chronic liver disease globally, affecting more than one-third of the world’s population. Its rate has increased by 13%, rising from 25% to 38% over the past three decades. MASLD is characterized by excessive fat accumulation in hepatocytes and, if left untreated, progresses to its more severe form, metabolic dysfunction-associated steatohepatitis (MASH), which poses a significant global health concern as one of the leading causes of cirrhosis, cirrhotic complications, and hepatocellular carcinoma. The molecular determinants of MASLD stratification are not clearly defined and require additional investigation. In this study, we used male and female CC042 mice, a Collaborative Cross mouse strain susceptible to the development of MASLD, to analyze the global transcriptomic profile in liver and to clarify the molecular mechanisms involved in the progression of the disease. Male and female CC042 mice were maintained on a high fat/high sucrose (HF/HS) diet for 20, 40, or 60 weeks, and hepatic gene expression profiles were determined by next-generation RNA sequencing. Chronic feeding of the HF/HS diet induced profound alterations in gene expression at all time points. The number of significantly differentially expressed genes (DEGs) in the livers of male mice was greater than female mice at 20 and 40 weeks and reached a maximum of 2764 genes at 40 weeks. In contrast, the number of DEGs in the livers of female mice steadily increased with the time on diet reaching a maximum of 2974 DEGs at 60 weeks after HF/HS diet initiation. The number of upregulated DEGs was higher than downregulated DEGs in both male and female mice at all time points. Functional analysis of DEGs showed that in both sexes, the most dysregulated liver toxicity-related pathways were associated with liver fibrosis, liver necrosis/cell death, inflammation, steatosis, liver damage, and liver proliferation/hyperplasia. In male mice, pathways associated with fibrosis, necrosis/cell death, inflammation, and steatosis remained constant from 20 to 40 weeks, and activation of liver hyperplasia pathway was constant from 40 and 60 weeks. In contrast, in female mice, dysregulation of genes associated with these pathways gradually increased from 20 to 60 weeks after HF/HS diet initiation. Importantly, we found that the expression of 701 and 871 genes significantly correlated with total NASH pathology scores (Pearson r > |0.8|) in the livers of male and female mice, respectively. These findings provide comprehensive insights into the dynamics of time- and sex-specific transcriptomic changes during NAFLD progression. This knowledge can be helpful for identifying promising targets for prevention and treatment, as well as for determining key potential biomarkers of NAFLD to NASH progression.