Fig. 5. Copper modulates the HepG2 cell secretome.

a Volcano plot of proteins identified by secretome analysis in the medium of HepG2 cells cultured for 48 h in the presence of 50 nM selenite and with 0 or 100 µM CuSO₄. Two-sided t-test-based statistics were applied on normalized and logarithmized protein ratios to extract the significantly regulated proteins. b Extracellular APOE of HepG2 cells co-cultured with Cu and Se for indicated time points. APOE was analyzed using dot blot and normalized to Ponceau (P) staining (n = 3). c Intracellular APOE in HepG2 cells treated with 0 or 100 µM CuSO₄ without (-Se) or with 50 nM selenite for 72 h measured by Western Blot normalized to Ponceau staining (n = 4). An siRNA-mediated knockdown of APOE was generated and intracellular APOE (d) and extracellular SELENOP (e) were analyzed after 72 h treatment with selenite and/or Cu by Western Blot normalized to Ponceau staining (n = 4). Data are depicted as mean ± SD. Biological replicates are indicated by individual dots. Statistical analyses were based on two-way ANOVA with Bonferroni’s post-test (b–e). Adjustments for multiple comparisons were made for data provided in (a). *p < 0.05; **p < 0.01; ***p < 0.001 vs. -Cu; ⁺⁺p < 0.01; ⁺⁺⁺p < 0.001 vs. siControl. Source data are provided as a source Data file.