Kocarek T A, Schuetz E G, Guzelian P S
Department of Medicine, Medical College of Virginia, Richmond 23298.
In Vitro Cell Dev Biol. 1993 Jan;29A(1):62-6. doi: 10.1007/BF02634372.
Because the metabolic environment can alter gene expression in cultured cells, we examined the effects of change of medium on the levels of several cytochrome P450 mRNAs in primary cultures of rat hepatocytes maintained on Matrigel. The amounts of P450 1A2, 2B1/2, or 3A1 mRNA were unaffected by changing the medium. In contrast, P450 1A1 mRNA levels were increased 1 to 2 h after media change, reached maximum levels by 6 h, and declined to baseline by 24 h. Supplementing day-old media with components of the medium revealed that only addition of amino acids resulted in 1A1 mRNA induction. From the results of direct additions and omissions, we showed that tryptophan, but not histidine, was largely responsible for the 1A1 mRNA induction. Moreover, mild photoactivation of the tryptophan resulted in a substantially increased magnitude of 1A1 mRNA induction. The time course for 1A1 mRNA induction by treatment with photoactivated tryptophan was identical to that observed after medium change. Treatment of hepatocyte cultures with beta-naphthoflavone, which is metabolized by 1A1, also resulted in a transient 1A1 mRNA induction time-course profile over a 24-h period, whereas treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin, which is relatively stable to metabolic transformation, produced sustained elevations of 1A1 mRNA, suggesting that the transient response to tryptophan also may involve metabolism of the inducer. Our results extend previous data showing that oxidized products of tryptophan induce 1A1, and suggest that the transient nature of the induction may be due to elimination of the activated tryptophan molecule.
由于代谢环境可改变培养细胞中的基因表达，我们研究了更换培养基对基质胶上原代培养大鼠肝细胞中几种细胞色素P450 mRNA水平的影响。更换培养基后，P450 1A2、2B1/2或3A1 mRNA的量未受影响。相比之下，更换培养基后1至2小时，P450 1A1 mRNA水平升高，6小时达到最高水平，24小时降至基线。用培养基成分补充一天前的培养基发现，只有添加氨基酸可诱导1A1 mRNA。从直接添加和去除成分的结果来看，我们发现色氨酸而非组氨酸是1A1 mRNA诱导的主要原因。此外，色氨酸的轻度光激活导致1A1 mRNA诱导幅度大幅增加。用光激活色氨酸处理诱导1A1 mRNA的时间进程与更换培养基后观察到的相同。用可被1A1代谢的β-萘黄酮处理肝细胞培养物，在24小时内也导致1A1 mRNA出现短暂的诱导时间进程图谱，而用对代谢转化相对稳定的2,3,7,8-四氯二苯并对二恶英处理，则使1A1 mRNA持续升高，这表明对色氨酸的短暂反应也可能涉及诱导剂的代谢。我们的结果扩展了先前的数据，即色氨酸的氧化产物可诱导1A1，并表明诱导的短暂性质可能是由于活化色氨酸分子的消除。
