Kim H, Reddy S, Novak R F
Institute of Chemical Toxicology, Wayne State University, Detroit, MI 48201-2654, USA.
Drug Metab Dispos. 1995 Aug;23(8):818-24.
The effects of 3-methylcholanthrene (3-MC) and pyridine on rat renal cytochrome P450 (CYP) 1A1 and 1A2 mRNA expression have been examined by Northern-blot analysis and reverse transcriptase-polymerase chain reaction (RT-PCR), followed by Southern-blot analysis of the PCR products. Northern-blot hybridization and RT-PCR analyses of kidney poly(A)+ RNA revealed that 3-MC treatment produced a time-dependent increase in the renal CYP1A1 and 1A2 mRNA levels, with CYP1A1 and 1A2 mRNA levels maximally increased at 24 and 18 hr, respectively, after treatment. These data were confirmed via RT-PCR analysis using a subsaturating level of cDNA template and by Southern-blot analysis of the PCR products. This approach served as the foundation for examining the effects of pyridine on CYP1A1 and 1A2 expression in renal tissue. RT-PCR analysis of renal CYP1A1 and 1A2 poly(A)+ RNA levels after treatment with pyridine (200 mg/kg/day for 3 consecutive days) revealed that CYP1A1 mRNA levels were maximally elevated approximately 10-fold after pyridine treatment for 2 consecutive days, whereas CYP1A2 mRNA levels were maximally elevated approximately 3-fold at 24 hr after treatment. The mRNA levels of glyceraldehyde 3-phosphate dehydrogenase, which served as an internal control, remained constant after 3-MC or pyridine treatment. These results show that expression of CYP1A1 and 1A2 mRNAs is enhanced in renal tissue after exposure to 3-MC or pyridine, and that constitutive expression of CYP1A1 seems to be greater than that of CYP1A2 in renal tissue.
通过Northern印迹分析和逆转录聚合酶链反应（RT-PCR），随后对PCR产物进行Southern印迹分析，研究了3-甲基胆蒽（3-MC）和吡啶对大鼠肾细胞色素P450（CYP）1A1和1A2 mRNA表达的影响。对肾多聚腺苷酸（poly(A)+）RNA的Northern印迹杂交和RT-PCR分析显示，3-MC处理使肾CYP1A1和1A2 mRNA水平呈时间依赖性增加，处理后CYP1A1和1A2 mRNA水平分别在24小时和18小时达到最大值。这些数据通过使用亚饱和水平的cDNA模板的RT-PCR分析以及PCR产物的Southern印迹分析得到证实。该方法为研究吡啶对肾组织中CYP1A1和CYP1A2表达的影响奠定了基础。对用吡啶（200 mg/kg/天，连续3天）处理后的肾CYP1A1和1A2 poly(A)+ RNA水平进行RT-PCR分析显示，连续2天用吡啶处理后，CYP1A1 mRNA水平最大升高约10倍，而处理后24小时CYP1A2 mRNA水平最大升高约3倍。作为内对照的甘油醛-3-磷酸脱氢酶的mRNA水平在3-MC或吡啶处理后保持恒定。这些结果表明，暴露于3-MC或吡啶后，肾组织中CYP1A1和1A2 mRNA的表达增强，并且肾组织中CYP1A1的组成型表达似乎大于CYP1A2。
