Bars R G, Bell D R, Elcombe C R
Zeneca Central Toxicology Laboratory, Macclesfield, Cheshire, U.K.
Biochem Pharmacol. 1993 May 25;45(10):2045-53. doi: 10.1016/0006-2952(93)90015-o.
We have analysed the induction of microsomal and peroxisomal proteins and their RNAs after treatment of hepatocytes with the peroxisome proliferator, clofibric acid, in vitro and in vivo. After treatment of hepatocytes with 1 mM clofibric acid for 4 days, P450 4A1 RNA is induced 500-fold, and acyl-CoA oxidase and P450 2B1 280-fold, relative to control cultures. These RNAs are detectably induced after administration of 25 microM clofibric acid, and show a similar induction response with increasing doses of clofibric acid. Western blot analysis of the P450 4A and bifunctional enzyme (BFE) proteins showed that both were induced in parallel with increasing doses of clofibric acid, over a range of 25 microM-1 mM. The distribution of the induced proteins was examined by immunocytochemistry. Increasing doses of clofibric acid led to an increase in the average intensity of staining for both proteins throughout the hepatocyte population. There was, however, a graded variation between hepatocytes in the intensity of staining, both for P450 4A and BFE proteins. The heterogeneity in response of the hepatocyte population in vitro may be related to differential sensitivity of hepatocytes to induction in vivo. Therefore, rats were dosed with 0, 50 or 300 mg/kg of clofibric acid for 4 days by gavage, and the livers were examined by immunocytochemistry. After 50 mg/kg of clofibric acid, both P450 4A and BFE were induced mainly in zones 3 and 2 of the liver acinus. However, after 300 mg/kg of clofibric acid, staining for both proteins was strong and homogenous throughout the liver acinus. Thus, hepatocytes from zones 3 and 2 of the acinus are differentially responsive to induction by clofibric acid.
我们已经分析了在体外和体内用过氧化物酶体增殖剂氯贝酸处理肝细胞后，微粒体和过氧化物酶体蛋白及其RNA的诱导情况。用1 mM氯贝酸处理肝细胞4天后，相对于对照培养物，P450 4A1 RNA被诱导了500倍，酰基辅酶A氧化酶和P450 2B1被诱导了280倍。在给予25 microM氯贝酸后，这些RNA可被检测到诱导，并且随着氯贝酸剂量的增加呈现出相似的诱导反应。对P450 4A和双功能酶（BFE）蛋白的蛋白质印迹分析表明，在25 microM - 1 mM的范围内，两者均随着氯贝酸剂量的增加而平行诱导。通过免疫细胞化学检查诱导蛋白的分布。氯贝酸剂量的增加导致整个肝细胞群体中两种蛋白染色的平均强度增加。然而，对于P450 4A和BFE蛋白，肝细胞之间的染色强度存在分级变化。体外肝细胞群体反应的异质性可能与肝细胞在体内对诱导的不同敏感性有关。因此，通过灌胃给大鼠分别给予0、50或300 mg/kg的氯贝酸4天，并通过免疫细胞化学检查肝脏。给予50 mg/kg氯贝酸后，P450 4A和BFE主要在肝腺泡的3区和2区被诱导。然而，给予300 mg/kg氯贝酸后，整个肝腺泡中两种蛋白的染色都很强且均匀。因此，腺泡3区和2区的肝细胞对氯贝酸诱导的反应不同。
