Tracewell W, Desjardins J, Iversen P
Department of Pharmacology, University of Nebraska Medical Center, Omaha 68198-6260, USA.
Toxicol Appl Pharmacol. 1995 Dec;135(2):179-84. doi: 10.1006/taap.1995.1221.
CYP1A1 gene expression is regulated by known cis- and transacting elements controlling inhibition and induction of CYP1A1 transcription. The influence of a double-stranded phosphorothioate oligonucleotide (dsODN) with sequence identical to the CYP1A1 negative regulatory element (NRE) was examined in Sprague-Dawley rats. Two strategies were employed: (i) two single-stranded complementary 25-mer ODNs that form a double-stranded ODN (ODN1) and (ii) a 54-base, self-complementary ODN which forms a dsODN hairpin (ODN2). A dsODN hairpin with scrambled NRE sequence was evaluated as a control (ODN3). Zoxazolamine paralysis times, an in vivo marker of CYP1A1 activity, were reduced from 184 +/- 18 min in saline-treated rats to 103 +/- 12.5 min 24 hr after a single 1.7-mg ODN1 iv injection. Liver microsomal EROD, an in vitro marker of CYP1A1/2 activity, was increased from 210 +/- 10 pmol in saline-treated animals to 703 +/- 73 and 623 +/- 89 pmol resorufin/mg protein/min after iv ODN1 and iv ODN2, respectively. ODN1's activity did not change PNP hydroxylation and PROD, markers of CYPs 2E1 and 2B1/2. ODN2 did not significantly change PNP but did significantly alter PROD. The ODN3 did not cause any significant changes in any assay measured. The ODN1-induced responses in ZX paralysis and EROD were observed post-iv injection, but not following ip injection of ODN1. Western blot analysis of ODN1- and HPO-treated rat liver microsomes also revealed increased in CYP1A1 protein. These data indicate double-stranded ODNs mimic the cis-acting NRE in vivo inducing CYP1A1 in the absence of other xenobiotics.
CYP1A1基因表达受已知的顺式和反式作用元件调控，这些元件控制着CYP1A1转录的抑制和诱导。在斯普拉格-道利大鼠中研究了一种与CYP1A1负调控元件（NRE）序列相同的双链硫代磷酸酯寡核苷酸（dsODN）的影响。采用了两种策略：（i）两条形成双链ODN的单链互补25聚体ODN（ODN1）和（ii）一条形成dsODN发夹结构的54个碱基的自互补ODN（ODN2）。将具有随机NRE序列的dsODN发夹作为对照进行评估（ODN3）。唑沙宗麻痹时间是CYP1A1活性的体内标志物，在单次静脉注射1.7 mg ODN1后24小时，经盐水处理的大鼠的唑沙宗麻痹时间从184±18分钟降至103±12.5分钟。肝脏微粒体EROD是CYP1A1/2活性的体外标志物，经静脉注射ODN1和静脉注射ODN2后，经盐水处理动物的肝脏微粒体EROD分别从210±10 pmol增加至703±73和623±89 pmol间苯二酚/毫克蛋白/分钟。ODN1的活性未改变PNP羟化作用以及CYPs 2E1和2B1/2的标志物PROD。ODN2未显著改变PNP，但确实显著改变了PROD。ODN3在任何所测试验中均未引起任何显著变化。静脉注射ODN1后观察到ODN1诱导的唑沙宗麻痹和EROD反应，但腹腔注射ODN1后未观察到。对经ODN1和HPO处理的大鼠肝脏微粒体进行的蛋白质免疫印迹分析也显示CYP1A1蛋白增加。这些数据表明双链ODN在体内模拟顺式作用NRE，在不存在其他异生素的情况下诱导CYP1A1。
