Fig. 3. Phosphorylation of protein kinase B (AKT) and extracellular signal-regulated kinase (ERK) is involved in the induction of glutamate-cysteine ligase catalytic subunit (GCLC) and glutathione synthetase (GSS) and activation of nuclear factor erythroid-2-related factor 2 (NRF2) in human HaCaT keratinocytes. (A, B) Cells were treated with LY294002 (PI3K/AKT inhibitor) or U0126 (ERK inhibitor), and following 1 h, RA was added. And (A) phospho-AKT, AKT or phospho-ERK, ERK2, and (B) GCLC, GSS protein expressions were determined by western blot analysis. Cells were transfected with 20 nM siControl and siAKT or siERK1. After 16 h of incubation, cells were treated with RA (2.5 μM) for 24 h. (C) Effect of siAKT or siERK1 on the expression of GCLC and GSS was observed via western blot using specific antibodies. (D) Cells were incubated with LY294002 or U0126, treated with RA (2.5 μM). Phospho-NRF2 and NRF2 protein expressions were detected by their specific antibodies. The TBP antibody was used as a loading control for nuclear proteins. (E) Cells were incubated with LY294002 or U0126, treated with RA (2.5 μM), and exposed to UVB for the detection of cell viability using the MTT assay. *Significantly different from UVB-treated cells (p<0.05), and **significantly different from RA plus UVB-treated cells (p<0.05).
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