Evidence That Cytochrome b5 and Cytochrome b5 Reductase Can Act as Sole Electron Donors to the Hepatic Cytochrome P450 Systems
Evidence That Cytochrome b5 and Cytochrome b5 Reductase Can Act as Sole Electron Donors to the Hepatic Cytochrome P450 Systems
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DOI：
10.1124/mol.112.084616
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发表时间：
2013-06-01
影响因子：
3.6
通讯作者：
Wolf, C. Roland
中科院分区：
文献类型：
--
作者：
Henderson, Colin J.;McLaughlin, Lesley A.;Wolf, C. Roland
我们先前描述了遗传模型的开发，以研究肝细胞色素P450（P450）系统的体内功能，通过肝细胞色素P450氧化还原酶[POR; HRN（肝还原酶无效）线]或细胞色素b5 [HBN（肝细胞色素B（5）无效）线]的缺失。然而，HRN小鼠仍然表现出低水平的单加氧酶活性，尽管没有可检测的还原酶蛋白。为了研究这是否是因为细胞色素B（5）和细胞色素B（5）还原酶可以作为P450系统的唯一电子供体，我们将HRN与HBN小鼠杂交以产生缺乏两种电子供体（HBRN）的肝表达的品系。HBRN小鼠表现出HRN系表型特征的恶化：肝肿大、肝脂肪变性和某些P450表达增加。此外，相对于HRN模型，体外药物代谢活性进一步降低，在某些情况下降低至不可检测的水平。体内药代动力学研究表明，咪达唑仑半衰期、C-max和浓度-时间曲线下面积增加，清除率降低，在HBRN线中的程度大于HBN或HRN模型。使用低于细胞色素B（5）还原酶表观Km但远高于POR表观Km的NADPH浓度进行微粒体孵育，导致HRN样品中7-苄氧基喹啉周转的实际消除。这些数据为细胞色素B（5）/细胞色素B（5）还原酶在体外和体内可作为P450系统的唯一电子供体提供了强有力的证据。
We previously described the development of genetic models to study the in vivo functions of the hepatic cytochrome P450 (P450) system, through the hepatic deletion of either cytochrome P450 oxidoreductase [POR; HRN (hepatic reductase null) line] or cytochrome b5 [HBN (hepatic cytochrome b(5) null) line]. However, HRN mice still exhibit low levels of mono-oxygenase activity in spite of the absence of detectable reductase protein. To investigate whether this is because cytochrome b(5) and cytochrome b(5) reductase can act as the sole electron donor to the P450 system, we crossed HRN with HBN mice to generate a line lacking hepatic expression of both electron donors (HBRN). HBRN mice exhibited exacerbation of the phenotypic characteristics of the HRN line: liver enlargement, hepatosteatosis, and increased expression of certain P450s. Also, drug metabolizing activities in vitro were further reduced relative to the HRN model, in some cases to undetectable levels. Pharmacokinetic studies in vivo demonstrated that midazolam half-life, C-max, and area under the concentration-time curve were increased, and clearance was decreased, to a greater extent in the HBRN line than in either the HBN or HRN model. Microsomal incubations using NADPH concentrations below the apparent K-m of cytochrome b(5) reductase, but well above that for POR, led to the virtual elimination of 7-benzyloxyquinoline turnover in HRN samples. These data provide strong evidence that cytochrome b(5)/cytochrome b(5) reductase can act as a sole electron donor to the P450 system in vitro and in vivo.