Figure 3.

The effect of 3-MC on CYP1A1 expression and EROD activity. (A) Detection of Ah receptor by RT-PCR analysis in Dami cells. RT-PCR contains a set of AhR-specific primers as shown in Table 1. A specific band of AhR is marked by the arrow (322 bp). Liver tissue cDNA was used as a reference control. (B) Western blot analysis of CYP1A1. Dami cells were cultured with 10 μM of 3-MC (+3MC) for 72 h. The control group (-3MC) received the same volume of the solvent (DMSO) without 3-MC. After 3-MC treatment, 35 μg of total protein lysates from each group was loaded in a SDS-polyacrylamide gel for Western blot analysis of CYP1A1. Immunoblotting for actin (45 kDa) was performed as an internal control. Detailed procedures for the Western blot analysis are explained in the “Materials and Methods” section. (C) Effect of 3-MC on the EROD activity. EROD assay was performed in the 3-MC-treated and control Dami cells. The intact Dami cells were incubated with 2 μg of 7-ER in a TN assay buffer for 20 min. Details regarding the EROD assay are explained in the “Materials and Methods” section. EROD activity was expressed as nanomoles per minute per milligram of protein. Statistically significant changes as compared with the DMSO treatment group are indicated as ***P < 0.001. Pairwise comparisons were performed using a Student’s paired t test.