Syk/Src Pathway-Targeted Inhibition of Skin Inflammatory Responses by Carnosic Acid
Figure 5
Effect of CA on the mRNA expression of proinflammatory genes, the activation of transcription factors, and upstream signaling cascades for NF-κB activation. (a) The mRNA levels (left panel) of genes encoding iNOS, TNF-α, and COX-2 were determined by semiquantitative PCR. (b) HEK293 cells cotransfected with NF-κB-Luc, IFN-β-promoter-Luc, or AP-1-Luc plasmid constructs (1 μg/mL each) and β-gal (as a transfection control) were treated with CA (0 to 20 μg/mL) in the presence or absence of PMA (100 nM) or by cotransfection with the adapter molecule, TRIF. Luciferase activity was measured using a luminometer. (b) Levels of NF-κB family proteins, p50 and p65, in the nuclear fraction were determined by immunoblotting analyses using antibodies against total protein. (d) and (e) Phosphoprotein or total protein levels of IκBα, IKK, Akt, PDK1, p85, Src, Syk, and β-actin from cell lysates were determined by immunoblotting analyses using phosphospecific or total protein antibodies. Relative intensity was calculated by densitometric scanning. and compared to the control.
