White R D, Hahn M E, Lockhart W L, Stegeman J J
Biology Department, Woods Hole Oceanographic Institution, Massachusetts 02543.
Toxicol Appl Pharmacol. 1994 May;126(1):45-57. doi: 10.1006/taap.1994.1089.
Understanding the effects of environmental contaminants on cetaceans and other marine mammals will require information on the biochemistry of xenobiotic metabolism in these species. We characterized the hepatic microsomal cytochrome P450 system in beluga whales (Delphinapterus leucas) from the Canadian Arctic. The content of native P450 averaged 0.203 and 0.319 nmol/mg microsomal protein, cytochrome b5 content averaged 0.199 and 0.236 nmol/mg, and rates of NADPH-cytochrome c reductase were 79 and 76 nmol/min/mg, for females and males respectively. Ethoxyresorufin O-deethylase (EROD), pentoxyresorufin O-depentylase (PROD), and benzo[a]pyrene (BP) hydroxylase (AHH) activities were significantly greater in males than in females, and were highly correlated with one another (r2 between 0.853 and 0.912). HPLC analysis of in vitro BP metabolites revealed benzo-ring (7,8- and 9,10-) dihydrodiols, consistent with activation of this compound, as well as 4,5-dihydrodiol,3-OH-, 7-OH-, and 9-OH-BP and 1,6- and 3,6-quinones. Estradiol 2-hydroxylase activity did not differ between sexes, and rates did not correlate with those of the other activities. Antibodies against scup P450B (an apparent teleost CYP2B) and rat CYP2B1 did not recognize proteins in beluga liver microsomes, but there was a protein detected by antibodies to PB-inducible rabbit CYP2B4. Antibodies to ethanol and ketone-inducible rat CYP2E1 reacted with two proteins in beluga liver microsomes. Antibodies specific to hydrocarbon-inducible CYP1A1 and/or CYP1A2 forms showed a single protein band, apparently more closely related to CYP1A1. The content of CYP1A was fivefold greater in male than in female beluga. CYP1A content was highly correlated with EROD, PROD, and AHH activities, suggesting that this P450 form is a primary catalyst for these reactions in beluga. CYP1A content and activity were highly correlated with the concentrations in blubber of non-ortho and mono-ortho PCB congeners, compounds that induce CYP1A in other mammals. These results indicate that a CYP1A is a catalyst for the metabolism of aromatic hydrocarbon pollutants in the beluga whale, and strongly suggest that this protein is induced in these organisms by environmental contaminants, including PCBs. The results support the measurement of CYP1A expression as a biomarker of exposure to inducers in marine mammals. The full functional and evolutionary relationships of beluga CYP1A and of beluga proteins immunologically related to other P450 forms are uncertain.
要了解环境污染物对鲸类和其他海洋哺乳动物的影响，需要掌握这些物种中异源物质代谢的生物化学信息。我们对来自加拿大北极地区的白鲸（白鲸属）肝脏微粒体细胞色素P450系统进行了表征。雌性和雄性白鲸的天然P450含量平均分别为0.203和0.319 nmol/mg微粒体蛋白，细胞色素b5含量平均分别为0.199和0.236 nmol/mg，NADPH-细胞色素c还原酶的速率分别为79和76 nmol/min/mg。雄性白鲸的乙氧基异吩恶唑酮O-脱乙基酶（EROD）、戊氧基异吩恶唑酮O-脱戊基酶（PROD）和苯并[a]芘（BP）羟化酶（AHH）活性显著高于雌性，且彼此高度相关（r2在0.853至0.912之间）。体外BP代谢产物的HPLC分析显示存在苯环（7,8-和9,10-）二氢二醇，这与该化合物的活化一致，以及4,5-二氢二醇、3-OH-、7-OH-和9-OH-BP以及1,6-和3,6-醌。雌二醇2-羟化酶活性在两性之间没有差异，其速率与其他活性的速率也不相关。针对鲷鱼P450B（一种明显的硬骨鱼CYP2B）和大鼠CYP2B1的抗体未识别出白鲸肝脏微粒体中的蛋白质，但用PB诱导的兔CYP2B4抗体检测到一种蛋白质。针对乙醇和酮诱导的大鼠CYP2E1的抗体与白鲸肝脏微粒体中的两种蛋白质发生反应。对烃诱导的CYP1A1和/或CYP1A2形式具有特异性的抗体显示出一条单一的蛋白带，显然与CYP1A1关系更密切。雄性白鲸的CYP1A含量比雌性高五倍。CYP1A含量与EROD、PROD和AHH活性高度相关，这表明这种P450形式是白鲸中这些反应的主要催化剂。CYP1A含量和活性与非邻位和单邻位多氯联苯同系物在鲸脂中的浓度高度相关，这些化合物在其他哺乳动物中可诱导CYP1A。这些结果表明，CYP1A是白鲸体内芳香烃污染物代谢的催化剂，并强烈表明这种蛋白质在这些生物中是由包括多氯联苯在内的环境污染物诱导产生的。这些结果支持将CYP1A表达的测量作为海洋哺乳动物接触诱导剂的生物标志物。白鲸CYP1A以及与其他P450形式免疫相关的白鲸蛋白质的完整功能和进化关系尚不确定。
