*2.1. Fucoxanthin Increases Expression of GCLC and GSS at the mRNA and Protein Levels*

HaCaT cells were treated with various concentrations of fucoxanthin for 12 h. The mRNA levels of GCLC and GSS in fucoxanthin-treated cells increased, demonstrating that 20 ΐM of fucoxanthin caused the maximal induction both in GCLC and GSS expression (Figure 1A). Next, HaCaT cells were treated with 20 ΐM fucoxanthin for various amounts of time. The mRNA levels of GCLC and GSS were highest at 12 h of treatment (Figure 1B). Furthermore, when HaCaT cells were treated with various concentrations of fucoxanthin for 12 h, the protein levels of GCLC and GSS increased, exhibiting that 20 ΐM of fucoxanthin induced maximal in GCLC and GSS expression (Figure 1C). Treatment with 

20 ΐM of fucoxanthin for 12 h markedly increased the protein levels of GCLC and GSS (Figure 1D). These results indicate that fucoxanthin treatment increases expression of GCLC and GSS at both the mRNA and protein levels. 

**Figure 1.** Effects of fucoxanthin treatment on the mRNA and protein expression of glutamate-cysteine ligase catalytic subunit (GCLC) and glutathione synthetase (GSS) in HaCaT cells. Cells were incubated with various concentrations of fucoxanthin (0–20 ΐM) for various amounts of time (0–24 h). The mRNA levels of GCLC and GSS were detected by reverse transcription-PCR (RT-PCR) following treatment ( **A**) with various concentrations of fucoxanthin for 12 h; and ( **B**) with 20 ΐM fucoxanthin for various amounts of time. The protein levels of GCLC and GSS were detected by Western blotting following treatment ( **C**) with various concentrations of fucoxanthin for 12 h; and ( **D**) with 20 ΐM fucoxanthin for various amounts of time. **\*** and **#** indicates significantly different from control of GCLC and GSS, respectively ( *p* < 0.05). 
