Spink David C, Katz Barbara H, Hussain Mirza M, Pentecost Brian T, Cao Zhimin, Spink Barbara C
Wadsworth Center, New York State Department of Health, Albany, NY 12201-0509, USA.
Carcinogenesis. 2003 Dec;24(12):1941-50. doi: 10.1093/carcin/bgg162. Epub 2003 Sep 11.
Cytochrome P450 (CYP)1A1 and CYP1B1, which are under the regulatory control of the aryl hydrocarbon (Ah) receptor (AhR), catalyze the metabolic activation of numerous procarcinogens and the hydroxylation of 17beta-estradiol (E2) at the C-2 and C-4 positions, respectively. There is evidence of cross-talk between estrogen receptor alpha (ERalpha)- and AhR-mediated signaling in breast and endometrial cells. To further examine these interactions, we investigated the short- and long-term effects of E2 exposure on Ah responsiveness in MCF-7 human breast cancer cells. Short-term exposure to 1 nM E2 elevated the ratio of the 4- to 2-hydroxylation pathways of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced E2 metabolism and the ratio of the induced CYP1B1 to CYP1A1 mRNA levels, as determined by real-time PCR. Cells maintained long-term (9-12 months) in low-E2 medium progressively lost Ah responsiveness, as indicated by diminished rates of TCDD-induced E2 metabolism and ethoxyresorufin O-deethylase activity, and the reduced expression of the CYP1A1 and CYP1B1 mRNAs and proteins levels. These E2-deprived cells showed elevated levels of ERalpha mRNA, depressed levels of AhR mRNA, and unchanged levels of the AhR nuclear translocator mRNA. Transient transfection studies using a CYP1B1-promoter-luciferase reporter construct showed that reduced CYP1B1 promoter activity in E2-deprived cells could be restored by co-transfection with an AhR expression construct, indicating that AhR expression was limiting in these cells. The reduced Ah responsiveness of E2-deprived cells was reversed by culture for four passages in medium supplemented with 1 nM E2; ERalpha and AhR mRNAs returned to near-normal levels and the inducibility of the CYP1A1 and CYP1B1 mRNAs, proteins, and E2 metabolic activities by TCDD was restored. These studies indicate that the continued presence of estrogen is required to maintain high levels of AhR expression and inducibility of the procarcinogen-bioactivating enzymes, CYP1A1 and CYP1B1, in MCF-7 cells.
细胞色素P450（CYP）1A1和CYP1B1受芳烃（Ah）受体（AhR）的调控，分别催化多种前致癌物的代谢活化以及17β-雌二醇（E2）在C-2和C-4位的羟基化。有证据表明，雌激素受体α（ERα）和AhR介导的信号在乳腺和子宫内膜细胞中存在相互作用。为了进一步研究这些相互作用，我们调查了E2暴露对MCF-7人乳腺癌细胞中Ah反应性的短期和长期影响。短期暴露于1 nM E2可提高2,3,7,8-四氯二苯并对二恶英（TCDD）诱导的E2代谢中4-羟基化途径与2-羟基化途径的比率，以及通过实时PCR测定的诱导型CYP1B1与CYP1A1 mRNA水平的比率。长期（9-12个月）维持在低E2培养基中的细胞逐渐丧失Ah反应性，这表现为TCDD诱导的E2代谢速率和乙氧基异吩恶唑酮O-脱乙基酶活性降低，以及CYP1A1和CYP1B1 mRNA和蛋白质水平的表达降低。这些E2缺乏的细胞显示ERα mRNA水平升高，AhR mRNA水平降低，而AhR核转运蛋白mRNA水平不变。使用CYP1B1启动子-荧光素酶报告基因构建体进行的瞬时转染研究表明，E2缺乏细胞中降低的CYP1B1启动子活性可通过与AhR表达构建体共转染来恢复，这表明AhR表达在这些细胞中是受限的。E2缺乏细胞降低的Ah反应性通过在补充有1 nM E2的培养基中传代培养四代而逆转；ERα和AhR mRNA恢复到接近正常水平，并且TCDD对CYP1A1和CYP1B1 mRNA、蛋白质以及E2代谢活性的诱导能力得以恢复。这些研究表明，雌激素的持续存在是维持MCF-7细胞中高水平AhR表达以及前致癌物生物活化酶CYP1A1和CYP1B1诱导能力所必需的。
