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. 2013 Sep 17;170(3):489–505. doi: 10.1111/bph.12111

Figure 3.

Figure 3

URB597 and AEA induce HO-1 transcripts, independent of CB receptors. (A, B) AM251 or AM630 cannot block URB597 and AEA from activating nqo1-ARE-Luc reporter in MCF-7 cells. MCF-7 cells co-transfected with 0.5 μg of nqo1-ARE-Luc reporter and 0.15 μg of pCMV-β-galactosidase plasmids were pretreated with 1.0 μM of AM251 or AM630 for 1 h, followed by 5 and 10 μM of URB597 or 2.5 μM of AEA treatment for 16 h. Luciferase activity assay and data analysis were performed as in Figure 1C. The data represent the mean ± SD of at least three independent experiments performed in triplicate. (C, D) AM251 and AM630 fail to block URB597 and AEA from inducing HO-1 mRNA expression. MCF-7 cells were pretreated with vehicle (DMSO), 1 μM of AM251, AM630, or 5 mM of NAC for 1 h, followed by URB597 (5 μM) and AEA (2.5 μM) treatment for 6 h. Semi-quantitative RT-PCR for HO-1 mRNA expression was performed as in Figure 1A. Furthermore, 5 mM of NAC blocks AEA from inducing HO-1 transcripts. All figures are representative of three independent experiments. (E) Capsazepine cannot block AEA from activating nqo1-ARE-Luc reporter. MCF-7 cells co-expressed with 0.5 μg of ARE reporter and 0.15 μg of pCMV-β-galactosidase were pretreated with 1.0 and 2.0 μM of capsazepine for 1 h, followed by 2.5 μM of AEA treatment for 16 h. Luciferase activity assay and data analysis were performed as in Figure 1C. The data represent the mean ± SD of at least three independent experiments performed in triplicate. Caps.: capsazepine *P < 0.05 as compared with the control untreated cells; **P < 0.001 as compared with the control untreated cells.