Extended Data Fig. 3: Age is a major risk factor of MASLD development.

(A) Re-analysis of the Duke MASLD cohort (GSE213623) revealed that the chronological age positively correlates with MASLD histological markers including hepatocyte ballooning, portal inflammation and fibrosis in MASLD patients (control n = 69; MASLD patients n = 299; ballooning score 0, n = 32, score 1, n = 118, score 2, n = 149; portal_inf score 0, n = 146, score 1, n = 142; fibrosis F0F1, n = 97; F2, n = 107; F3F4, n = 95). **p < 0.01; ***p < 0.001; ****p < 0.0001. (B, C) GSEA using KEGG revealed that genes related to longevity and its associated mechanisms (for example nicotinamide; FoxO signaling) are depleted (B), while genes related to programmed cell death (C) are highly enriched in the transcriptomics of MASLD patients. Red arrows point to the pathways of interest. (D) AHGS enrichment increases with chronological age in MASLD patients but not control subjects. (E) AHGS enrichment score with chronological age in RNA-seq data of 226 normal liver samples from GTEx v8 database (https://gtexportal.org/home/). p-values were calculated using Wilcoxon Rank Sum test. Boxplot shows the upper quantile (75%), median (50%) and lower quantile (25%) of overall data distribution (A, D, E). p values in A, D, E were calculated using Wilcoxon Rank Sum test; p values in GSEA plots in B and C were calculated using permutation test, then adjusted for multiple comparison testing using the Benjamini-Hochberg method.