Figure 5.

Effect of ferrocene-1H-1,2,3-triazole hybrids on the protein expression of cPLA₂ and COX-2 in RMCs. (a,c) Western blot assay to detect COX-2 and cPLA₂ expression in cell lysate. (b) The COX-2 mRNA level was determined by real-time PCR. The sequence of addition of TNF-α reagent and compounds was as follows. In panels (a,b), RMCs were treated with ferrocene-1H-1,2,3-triazole hybrids (12.5 μg/mL) for 2 h and then exposed to TNF-α for another 24 h. In panel (c), RMCs were treated without or with F, X1, X2, X3, X4, or X5 at 12.5 μg/mL for 24 h. COX-2 and cPLA₂ expression were measured by comparison with that of GAPDH (internal control). In panel (d), the enzyme activity assay kits to detect of endogenous cPLA₂ and COX-2 activity in RMC cell lysate (6 μg) stimulated without or with ferrocene-1H-1,2,3-triazole hybrids or AACOCF3 (AA) in the presence of 10 ng/mL TNF-α. Data were obtained from at least three independent experiments and are expressed as mean ± SEM. One-way analysis of variance (ANOVA) followed by Bonferroni’s multiple-comparisons test was used to identify significant differences between multiple groups. Asterisks indicate a significant difference compared with the control group, ** p < 0.01. Hashes indicate a significant difference compared with the TNF-α-treated group, ^## p < 0.01; ^# p < 0.05.