Fig. 4.

Inhibitory effects of rapamycin (RP) and mycophenolic acid (MPA) on tumor necrosis factor (TNF)-α-induced intercellular adhesion molecule 1 (ICAM-1) and inducible nitric oxide synthase (iNOS) expression in HaCaT cells. (A) HaCaT cells were pre-incubated for 24 h, and then stimulated with TNF-α (20 ng/ml) in the presence of RP, MPA, or a combination of the two drugs for 3 h (for RNA) or 12 h (for protein). ICAM-1 and iNOS expression were determined after treatment with 100 nM RP, 100 nM MPA, and different combinations of the two drugs (20 nM RP and 80 nM MPA, 50 nM RP and 50 nM MPA, and 80 nM RP and 20 nM MPA) by reverse transcription-polymerase chain reaction (RT-PCR) and western blot. (B) HaCaT cells were pretreated with an extracellular signal-related kinases (ERK) inhibitor (PD98059), p38 inhibitor (SB203580), or c-Jun N-terminal kinases (JNK) inhibitor (SP600125) for 1 h, and then stimulated with TNF-α for 12 h. The band densities are expressed as percentages of the band density of the TNF-α-only group. ^**p<0.01 compared to treatment with TNF-α alone. The experiment was repeated independently at least three times. The densitometric analysis of the western blot was standardized using glyceraldehyde 3-phosphate dehydrogenase (GAPDH) as an internal control.