Schuetz E G, Wrighton S A, Barwick J L, Guzelian P S
J Biol Chem. 1984 Feb 10;259(3):1999-2006.
We administered a series of steroid hormones to primary nonproliferating cultures of adult rat hepatocytes and found that dexamethasone and other glucocorticoids but not sex steroid hormones, mineralocorticoids, or derivatives of pregnenolone other than pregnenolone 16 alpha-carbonitrile (PCN) stimulated de novo synthesis of an immunoreactive protein, indistinguishable from the form of cytochrome P-450 (P450PCN) induced by PCN in rat liver. No difference were discerned among purified liver cytochromes from rats treated with dexamethasone, PCN or dexamethasone plus PCN, among proteolytic digests of these proteins, or among the immunoprecipitated cytochromes prepared from cultured hepatocytes treated with these steroids as judged by electrophoresis on polyacrylamide gels containing sodium dodecyl sulfate followed by immunoblot analysis. Of the steroids tested, dexamethasone proved to be the most efficacious inducer increasing the rate of synthesis of P450PCN from 0.05% of total cellular protein synthesis in incubated control cultures (measured as incorporation of [3H]leucine into immunoprecipitable P450PCN) to as much as 9.4% in cultures incubated for 5 days in medium containing dexamethasone (10(-5) M). As with traditional glucocorticoid-responsive liver functions, induction of immunoreactive P450PCN was dependent on the concentration of dexamethasone (10(-8) to 10(-5) M) and was promptly reversed upon withdrawal of the steroid. However, during the 24-h interval between 24 to 48 h of culture age the hepatocytes were refractory to either induction or de-induction of immunoreactive P450PCN even though continuous exposure of the cells to dexamethasone (including this interval) was mandatory for maximal induction of P450PCN at 120 h in culture. Unlike cultured rat hepatocytes, HTC hepatoma cultures failed to exhibit dexamethasone-responsive expression of immunoreactive P450PCN. We conclude that glucocorticoids and PCN constitute a specific "class" of synthetic and endogenous inducers of a single form of cytochrome P-450.
我们对成年大鼠肝细胞的原代非增殖培养物施用了一系列类固醇激素，发现地塞米松和其他糖皮质激素可刺激一种免疫反应性蛋白的从头合成，而性类固醇激素、盐皮质激素或孕烯醇酮的衍生物（除孕烯醇酮16α-腈（PCN）外）则不能。这种免疫反应性蛋白与PCN在大鼠肝脏中诱导产生的细胞色素P-450（P450PCN）形式无法区分。在用含地塞米松、PCN或地塞米松加PCN处理的大鼠的纯化肝细胞色素之间、这些蛋白质的蛋白水解消化产物之间，或在用这些类固醇处理的培养肝细胞制备的免疫沉淀细胞色素之间，通过在含十二烷基硫酸钠的聚丙烯酰胺凝胶上电泳，随后进行免疫印迹分析，未发现差异。在所测试的类固醇中，地塞米松被证明是最有效的诱导剂，可使P450PCN的合成速率从孵育对照培养物中总细胞蛋白合成的0.05%（以[3H]亮氨酸掺入免疫沉淀的P450PCN来衡量）增加到在含地塞米松（10(-5)M）的培养基中孵育5天的培养物中的9.4%。与传统的糖皮质激素反应性肝功能一样，免疫反应性P450PCN的诱导取决于地塞米松的浓度（10(-8)至10(-5)M），并且在撤除类固醇后迅速逆转。然而，在培养24至48小时的24小时间隔内，肝细胞对免疫反应性P450PCN的诱导或去诱导均无反应，尽管细胞持续暴露于地塞米松（包括此间隔）对于在培养120小时时最大程度诱导P450PCN是必需的。与培养的大鼠肝细胞不同，HTC肝癌培养物未表现出免疫反应性P450PCN的地塞米松反应性表达。我们得出结论，糖皮质激素和PCN构成了单一形式细胞色素P-450的合成和内源性诱导剂的特定“类别”。
