Annas A, Granberg A L, Brittebo E B
Department of Pharmaceutical Biosciences, Biomedical Centre, Uppsala University, SE-751 24, Sweden.
Toxicol Appl Pharmacol. 2000 Nov 15;169(1):94-101. doi: 10.1006/taap.2000.9054.
In the present study, 7-ethoxyresorufin O-deethylase (EROD), 7, 12-dimethylbenz[a]anthracene (DMBA)-hydroxylase, and covalent binding of (3)H-labeled 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole ((3)H-Trp-P-1) and (3)H-DMBA were examined in human umbilical vein endothelial cells (HUVEC) and human umbilical artery endothelial cells (HUAEC) exposed to the aryl hydrocarbon (Ah) receptor agonist beta-naphthoflavone (BNF) or vehicle only. The results revealed a marked induction of enzymatic activity in BNF-treated HUVEC compared with vehicle-treated cells, whereas no similar response was observed in BNF-treated HUAEC. EROD, DMBA hydroxylase, and covalent binding of (3)H-Trp-P-1 and (3)H-DMBA in BNF-treated HUVEC were reduced in the presence of the CYP1A inhibitor ellipticine. Addition of other CYP1A inhibitors alpha-naphthoflavone, miconazole, 1-ethynylpyrene, 1-(1-propynyl)pyrene), or the CYP1A substrate ethoxyresorufin to the incubation buffer of BNF-treated HUVEC reduced covalent binding of (3)H-Trp-P-1 by 93-98%. Western blot analysis confirmed an induction of CYP1A1 in BNF-treated HUVEC, but not in BNF-treated HUAEC. CYP1A1 was, however, detected in both vehicle- and BNF-treated HUAEC. The results showed that BNF exposure induced CYP1A1 and metabolic activation of xenobiotics in HUVEC, whereas the catalytic activity remained low in BNF-treated HUAEC. Our results suggest that endothelial lining of human veins may be a target for adverse effects of xenobiotics activated into reactive metabolites by Ah receptor-regulated enzymes. Several studies have detected CYP1A1 in endothelial linings, whereas expression of CYP1A2 and CYP1B1 seems to be negligible at this site. This suggests that the metabolic activation and covalent binding of (3)H-Trp-P-1 and (3)H-DMBA in HUVEC are most likely mediated by CYP1A1.
在本研究中，检测了人脐静脉内皮细胞（HUVEC）和人脐动脉内皮细胞（HUAEC）中7-乙氧基试卤灵O-脱乙基酶（EROD）、7,12-二甲基苯并[a]蒽（DMBA）羟化酶，以及（3）H标记的3-氨基-1,4-二甲基-5H-吡啶并[4,3-b]吲哚（（3）H-Trp-P-1）和（3）H-DMBA的共价结合情况，这些细胞分别暴露于芳烃（Ah）受体激动剂β-萘黄酮（BNF）或仅溶剂中。结果显示，与溶剂处理的细胞相比，BNF处理的HUVEC中酶活性有显著诱导，而BNF处理的HUAEC中未观察到类似反应。在CYP1A抑制剂玫瑰树碱存在的情况下，BNF处理的HUVEC中EROD、DMBA羟化酶以及（3）H-Trp-P-1和（3）H-DMBA的共价结合减少。向BNF处理的HUVEC的孵育缓冲液中添加其他CYP1A抑制剂α-萘黄酮、咪康唑、1-乙炔基芘、1-（1-丙炔基）芘或CYP1A底物乙氧基试卤灵，可使（3）H-Trp-P-1的共价结合减少93%-98%。蛋白质印迹分析证实，BNF处理的HUVEC中CYP1A1有诱导，但BNF处理的HUAEC中没有。然而，在溶剂处理和BNF处理的HUAEC中均检测到CYP1A1。结果表明，BNF暴露可诱导HUVEC中CYP1A1和外源性物质的代谢活化，而BNF处理的HUAEC中催化活性仍然较低。我们的结果表明，人静脉的内皮衬里可能是被Ah受体调节酶激活为反应性代谢物的外源性物质产生不良反应的靶点。多项研究在内皮衬里中检测到CYP1A1，而CYP1A2和CYP1B1在此部位的表达似乎可以忽略不计。这表明HUVEC中（3）H-Trp-P-1和（3）H-DMBA的代谢活化和共价结合很可能由CYP1A1介导。
