Figure 2. ENO1 silencing enhances catabolic pathway adaptations.

A. Autophagy markers and SIRT-1 Western blot expression analysis in shCTRL and shENO1 CFPAC-1, MDA-MB-231 and NCI-H441 cells. Right panel: effects of the antioxidant agents NAC and TROLOX-C on LC3-II expression in CFPAC-1 cells by Western blot analysis. β-actin was used as a loading control; one representative of three independent experiments is shown B. Oxidation of palmitic acid in CFPAC-1, MDA-MB-231 and NCI-H441 cell lines transduced with shCTRL (white bars) or shENO1 (black bars). Cells were exposed to [1-¹⁴C] palmitic acid and total palmitate oxidation (sum of ¹⁴C-acid soluble metabolites and ¹⁴CO₂ production) was measured. C.–D. Analysis of phenylalanine (C) and acetoacetate (D) concentration in shCTRL and shENO1 CFPAC-1, MDA-MB-231 and NCI-H441 cell lines. E. The TCA cycle rate was evaluated measuring CO₂ emission after radiolabeling cells with [1-¹⁴C] acetylcoenzyme A in CFPAC-1, MDA-MB-231 and NCI-H441 cell lines transduced with shCTRL (white bars) or shENO1 (black bars). TCA cycle activity is expressed as pmol CO₂/min/mg protein. All graphs illustrate the mean result of three independent experiments ± SEM. *p < 0.05; **p < 0.01;***p < 0.001 relative to shCTRL.