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. 2015 May 28;6(23):19792–19806. doi: 10.18632/oncotarget.4075

Figure 2. Effect of HO-1 on the stability of ICAM-1 mRNA and TTP expression.

Figure 2

A. HT-29 cells were treated with or without 20 μM hemin for 8 h. The cells were then stimulated with 10 ng/ml TPA for 2 h followed by the addition of 5 μg/ml actinomycin (Act D) for up to 150 min. ICAM-1 mRNA was measured using real time PCR. The expression levels were normalized to ICAM-1 mRNA levels. The normalized level of ICAM-1 mRNA at time 0 was set at 100%, and all other normalized mRNA levels were graphed relative to that value. Each point represents the mean ± SD from three independent experiments. B. Hemin-induced expression of TTP. HT-29 cells were pretreated with 10 and 20 μM hemin for 1 h followed by stimulation with 10 ng/ml TPA for an additional 16 h, TTP mRNA was quantified by real time PCR (upper panel). Values represent TTP mRNA normalized to GAPDH mRNA. Immunoreactive TTP was measured by western blot (lower panel). C. The effects of hemin on TTP protein expression in Caco-2 cells. Caco-2 cells were pretreated with 10 and 20 μM hemin for 1 h followed by stimulation with 10 ng/ml TPA and TTP prtoein was analyzed by western blot. D. HT-29 cells were treated with 5 μM CoPPIX for 1 h followed by stimulation with 10 ng/ml TPA and TTP prtoein was analyzed by western blot. E. HT-29 cells were pretreated with 20 μM ZnPP for 30 min, followed by 20 μM hemin for 1 h and further stimulated with 10 ng/ml TPA. The cells were harvested for western blot analysis and measurement of HO activity. A representative immunoblot out of three independent experiments is shown. Results were repeated in at least three independent experiments. The data are expressed as mean ± SD. *P < 0.05, **P < 0.01 and ***P < 0.001.