Fig. 2

Effect of CA treatment on the mRNA levels of specific genes or on the expression of the protein they encode in HepG2 cells. Except for the control, in which cells were not exposed to any chemical, cells were incubated with CA at different concentrations (5, 10, and 20 μM) — or not at all — for 24 h before being incubated with 0.3 mM t-BHP for 2 h. Total RNA was extracted using the Trizol reagent and an equivalent amount of RNA was converted into cDNA using the reverse transcriptase kit implementing manufacturer’s instructions. RT-PCR and qRT-PCR experiments were ultimately performed to analyze the amount of a HO-1, c glutamate-cysteine ligase (GCL) catalytic unit (GCLC), and e GCL modifier subunit (GCLM) mRNA in HepG2 cells. Proteins (10 μg) were separated by 10% SDS-PAGE and electro-transferred to a polyvinylidene difluoride (PVDF) membrane. Immunoblotting was performed using monoclonal or polyclonal antibodies against b heme oxygenase-1 (HO-1), d GCLC, and f GCLM. Values are expressed as mean ± standard deviation (n = 3). Different letters indicate signification differences at p < 0.05 by Tukey’s studentized range tests