Fig. 2

Association of transcriptomic data with serum ferritin. a Permutation test for the goodness-of-fit (R²Y) and goodness of prediction (Q²Y) obtained from the O-PLS model between serum ferritin and hepatic transcriptome in a subsample of the discovery and replication cohorts from Italy and Spain (n = 86). b Significant transcripts associated with serum ferritin after further validation of the O-PLS significant variables by pSC adjusting for age, sex, BMI and country. c Pathways significantly associated with serum ferritin based on mapping associated transcripts by over-representation analysis with hypergeometric test. d Permutation tests for the O-PLS model between serum ferritin and SLCs (n = 86). e Significant SLCs associated with serum ferritin after further validation of the O-PLS results by pSC adjusting for age, sex, BMI, and country. f O2-PLS scores for the joint variation between microbial families and genes associated with serum ferritin. A model with 2 predictive components, and 1 orthogonal component for the genes and bacterial families blocks, was constructed based on 7-fold cross-validation. g O2-PLS joint loadings plots, where pcorr represents the correlation-scaled loadings from the gene block and qcorr represents the correlation-scaled loadings from the bacterial families block. h Heatmap displaying z-scores of the ferritin-associated transcripts for each subject. Clustering was based on Euclidean distances and Ward linkage. Genes associated with liver fat accumulation from O-PLS modelling are highlighted in bold, whereas those associated with bacterial families from O2-PLS modelling are highlighted in colour boxes. i Heatmap for the pSC adjusted by age, BMI, sex, and country between ferritin-associated plasma and j urine metabolites with ferritin-associated transcripts (n = 86). k Significant (p < 0.05) pSC adjusted for age, BMI and country, between ferritin-associated families and transcripts (n = 56). Only significant associations (p < 0.05) are displayed. Significant associations after a pFDR correction (pFDR < 0.05) are highlighted with a black box. l–n Expression of upregulated (GSK3B, PDE7A, SBNO2) and o–s downregulated genes (GYS2, SEC24B, SOCS2, MTUS1 and SLC51A) in human primary hepatocytes after treatment with iron and palmitic acid. Data are mean ± SEM. Comparisons by one-way ANOVA. *p < 0.05, **p < 0.01, ***p < 0.001 compared to control group based on t test. ^#p < 0.05, ^##p < 0.01, ^###p < 0.001 compared to PA group based on t test. Ctrl, control group; PA, palmitic acid; Fe48h, pre-treatment iron 50 μM for 48h; Fe72h, pre-treatment iron 50 μM for 72h; Fe48h + PA, pre-treatment iron 50 μM for 48h + palmitic acid 200 μM for 24 h; Fe72h + PA, pre-treatment iron 50 μM for 72 h + palmitic acid 200 μM for 24 h