Ledirac N, Delescluse C, Lesca P, Piechocki M P, Hines R N, de Sousa G, Pralavorio M, Rahmani R
Laboratoire de Pharmaco-Toxicologie Cellulaire et Moléculaire, INRA, Antibes, 06606, France.
Toxicol Appl Pharmacol. 2000 May 1;164(3):273-9. doi: 10.1006/taap.2000.8920.
Diflubenzuron (DFB) belongs to a group of compounds called benzoyphenyl ureas acting as chitin synthesis inhibitors, which also inhibit growth of B16 murine melanomas. The present study was designed to investigate the effect of this insecticide, on CYP1A1 expression and induction in human hepatoma cells HepG2. Treatment of HepG2 cells over 72 h with noncytotoxic concentrations of DFB resulted in a strong dose-dependent decrease in constitutive ethoxyresorufin-O-deethylase activity. Moreover, DFB significantly decreased CYP1A1 induction by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) after 24 h exposure, as demonstrated by ethoxyresorufin-O-deethylase (EROD) activity and Northern blot analysis. Additional studies were performed both on parental HepG2 cells and HepG2-241c.1, which were stably transfected with the chloramphenicol acetyltransferase (CAT) reporter gene, cloned under the control of the human CYP1A1 promoter (-1140 to +59). Ribonuclease protection assays (RPA) analysis clearly demonstrated an inhibition of CYP1A1 transcription in both cell lines. Surprisingly, in corresponding experiments using 3-methylcholanthrene (3-MC) as a CYP1A1 inducer, DFB was less effective. Finally, in competitive binding studies using a 9S-enriched fraction of HepG2 cytosol, DFB was capable of displacing [(3)H]-2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) from its Ah receptor binding site. Taken together, these results support the involvement of a transcriptional mechanism in the inhibition of CYP1A1 expression in HepG2 cells by DFB, possibly via an Ah receptor antagonism.
除虫脲（DFB）属于一类名为苯甲酰基苯基脲的化合物，这类化合物作为几丁质合成抑制剂，也能抑制B16小鼠黑色素瘤的生长。本研究旨在探究这种杀虫剂对人肝癌细胞HepG2中CYP1A1表达和诱导的影响。用无细胞毒性浓度的DFB处理HepG2细胞72小时以上，导致组成型乙氧异吩唑酮 - O - 脱乙基酶活性呈强烈的剂量依赖性下降。此外，如乙氧异吩唑酮 - O - 脱乙基酶（EROD）活性和Northern印迹分析所示，DFB在暴露24小时后显著降低了2,3,7,8 - 四氯二苯并 - p - 二恶英（TCDD）对CYP1A1的诱导作用。在亲本HepG2细胞和稳定转染了氯霉素乙酰转移酶（CAT）报告基因（该基因克隆在人CYP1A1启动子（-1140至+59）的控制下）的HepG2 - 241c.1细胞上都进行了额外的研究。核糖核酸酶保护分析（RPA）清楚地表明两种细胞系中CYP1A1转录均受到抑制。令人惊讶的是，在使用3 - 甲基胆蒽（3 - MC）作为CYP1A1诱导剂的相应实验中，DFB的效果较差。最后，在使用富含9S的HepG2细胞溶质部分进行的竞争性结合研究中，DFB能够从其芳烃受体结合位点取代[³H] - 2,3,7,8 - 四氯二苯并 - p - 二恶英（TCDD）。综上所述，这些结果支持转录机制参与了DFB对HepG2细胞中CYP1A1表达的抑制作用，可能是通过芳烃受体拮抗作用实现的。
