草鱼细胞色素P4503A和孕烷X受体基因表达分析及其功能初步研究
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摘要
细胞色素P450(CYP)是动物体内代谢药物的重要酶系,其亚型细胞色素P4503A(CYP3A)是动物肝脏、小肠和肾脏中含量最多的P450酶系,参与大多数药物的生物转化;其亚型CYP1A也参与部分药物的生物转化, CYP1A和CYP3A均能被药物诱导或者抑制。孕烷X受体(PXR)属于核受体超家族(NR)的NR1Ⅰ家族,其作为药物代谢的关键转录调控因子,参与CYP3A的诱导表达。关于鱼类CYP3A表达稳定阶段研究,CYP3A经由其转录因子PXR调控共同参与药物代谢研究报道甚少。
本文首先通过生物化学和分子生物学相结合的方法,先对CYP3A基因进行克隆并分析其结构功能域,检测其组织中表达丰度;再克隆了CYP3A转录因子PXR基因部分序列,检测其组织中表达丰度;其次,研究在草鱼体外最佳诱导模型中,CYP3A在mRNA和酶活性水平与时间动态变化的相关性,确定了CYP3A在鱼类中稳定表达阶段。在药物处理条件下,对PXR基因与CYP3A表达量差异进行研究,分析了CYP3A酶活性-CYP3A mRNA-PXR mRNA三者间相关性。用初步建立的诱导平台,氧氟沙星(OFLX)作为CYP3A的底物和诱导剂,验证CYP3A参与OFLX代谢过程中,其稳定表达阶段和CYP3A酶活性-CYP3A mRNA-PXR mRNA三者间相关性。全文主要研究结果如下:
一草鱼CYP3A和鲫鱼CYP1A基因克隆、序列分析及mRNA表达差异
根据GenBank中斑马鱼和虹鳟的CYP3A基因序列,设计保守区域的简并引物,通过PCR扩增获取CYP3A基因部分保守序列。根据CYP3A已获序列再设计引物,获取开放阅读框,然后,采用3' RACE法获取CYP3A序列末端;设计qRT-PCR引物,用SYBR荧光染料检测CYP3A mRNA组织表达差异。在此基础上,以鲫鱼CYP1A为参照,分析不同鱼类不同亚型基因结果特征。实验结果表明:⑴本研究得到草鱼CYP3A部分cDNA核苷酸序列为1849bp,包含保守编码区、3'UTR序列。开放阅读框序列长1542bp,编码513个氨基酸和一个终止密码(TAA),编码区包括信号肽(29aa)。⑵分析草鱼CYP3A主要结构域,其包含3个跨膜螺旋区,9个磷酸化位点,6个底物结合位点、1个血红素蛋白结合区和含1个半胱氨酸(Cys,C—448aa)的亚铁-血红素配体结合位点;而鲫鱼CYP1A基因结构特征,仅包含1个跨膜螺旋区,其含6个磷酸化位点,1个血红素蛋白结合区和含1个半胱氨酸(Cys,C—454aa)的亚铁-血红素配体结合位点。⑶应用实时荧光定量PCR(qRT-PCR)检测草鱼9种组织CYP3A mRNA相对含量,其表达丰度依次如下:前肠﹥肝脏﹥肾脏﹥鳃﹥肌肉﹥后肠﹥性腺﹥心脏﹥脾脏。其中,前肠、肝脏和肾脏是其主要表达部位,而其他各组织表达量较弱。同样方法检测鲫鱼9种组织CYP1A分布情况,其表达丰度依次如下:肝脏﹥前肠﹥鳃﹥肾脏﹥后肠﹥性腺﹥心脏﹥肌肉﹥脾脏,其中,肝脏、前肠和鳃是其主要表达部位,而其他各组织表达量较弱,CYP1A基因主要表达组织与参与药物代谢部位相同。药物代谢酶各亚型在组织中的分布与其参与药物代谢的主要部位相关。
二草鱼PXR基因克隆、序列分析及mRNA表达差异
根据GenBank中斑马鱼和虹鳟鱼类的PXR基因序列,设计保守区域的简并引物,通过PCR扩增获取PXR基因部分保守序列。设计qRT-PCR引物,用SYBR荧光染料检测PXR mRNA组织表达差异。实验结果表明:⑴本研究得到草鱼PXR部分cDNA核苷酸序列为509bp和推导169aa序列。⑵应用实时荧光定量PCR(qRT-PCR)检测9种组织PXR mRNA相对含量,其表达丰度依次如下:前肠﹥肝脏﹥肾脏﹥鳃﹥肌肉﹥后肠﹥性腺﹥心脏﹥脾脏。其中,前肠、肝脏和肾脏是其主要表达部位,而其他各组织表达量较弱。根据结果显示PXR与CYP3A基因在鱼体组织中表达部位基本一致。
三利福平和氧氟沙星对草鱼肾细胞的毒性实验
细胞的传代培养、冻存与复苏采用实验室常规方法。细胞中蛋白含量检测采用BCA法。采用MTT法分析了利福平在浓度为10-、20-、40和80μM,孵育时间为1-、2-、4 -、6-、8-、10-、12-、24-、48-和72 h;氧氟沙星在浓度分别为25-、50-、100-、200-、400和800μM,对草鱼肾细胞作用1-、2-、4-、6 -、8 -、10 -、12-、24和48 h之后,其结果表明肾细胞中选择RIF浓度为40μM和细胞孵育时间为1～24 h,作为最佳诱导模型进行后续实验,OFLX选择50μM和细胞孵育时间为1～24 h作为最佳条件进行后续实验,能对后续研究利福平和氧氟沙星在肾细胞中最佳代谢时间和浓度提供依据。
四最佳诱导模型CYP3A基因转录和酶活性检测
为了确定CYP3A在草鱼肾细胞中主要诱导及稳定表达阶段,本研究从CYP3A基因转录水平和酶活表达水平进行测定并评价。实验中使用CYP3A指示酶红霉素-N-脱甲基酶(ERND),采用分光光度计法检测CYP3A酶活性;设计qRT-PCR引物,采用SYBR荧光染料检测CYP3A mRNA表达差异。实验结果表明:CYP3A转录水平在2h诱导量提高,而酶活性则在4 h开始升高,两种水平间具有时间差异性,而CYP3A转录水平先于酶活2 h表达;RIF对CYP3A转录水平诱导效应显著强于酶活性水平诱导,这很可能反映了RIF诱导CYP3A表达是作用于转录水平的基因调控来影响酶活性。
五CYP3A与PXR mRNA表达及CYP3A酶活性三者相关性
RIF诱导后,CYP3A转录水平在2 h被诱导基因上调,而PXR则至10 h开始上调,此结果显示出CYP3A和PXR的基因转录水平上调呈现时间先后顺序。推断出PXR与药物结合后,受到药物刺激经时间发生反应,再参与后续的一系列作用;也显示出RIF与PXR结合后能激活PXR,并经由PXR这一转录调控因子参与激活并诱导CYP3A mRNA表达,不直接作用于翻译后酶活性,CYP3A酶活性主要是通过转录水平上调而达到活性增强的效果。
六氧氟沙星、CYP3A和PXR相关性
氧氟沙星(OFLX)孵育CIK,研究其时间动态过程中CYP3A与PXR量变相关性。用分光光度法测定CYP3A酶活性,用实时定量PCR检测CYP3A和PXR基因表达量。实验结果表明:⑴CYP3A酶活性在氧氟沙星处理后4 h开始提高,达致平台期后处于稳定阶段,而CYP3A mRNA在孵育10 h后开始开始上调。⑵PXR mRNA基底水平表达量较低,在OFLX孵育6～8 h后,逐渐上调,其mRNA开始上调时间先于CYP3A mRNA发生。由此推断整个药物代谢过程首先是OFLX经与PXR结合,然后其结合复合物作为转录调控调控因子参与激活CYP3A诱导表达。⑶结果显示氧氟沙星既是CYP3A代谢底物又是CYP3A一种诱导剂。
Cytochrome P450 (CYP) is an important animal drug metabolism enzyme. Cytochrome P450 3A (CYP3A), responsible for the metabolism of the majority of drugs, comprises the largest portion of the liver, small intestinal and kidney P450s in animals. CYP1A are responsible for the biotransformation of drug, CYP1A and CYP3A are induced or inhibited by drug. Pregnane X receptor (PXR) belongs to the nuclear receptor superfamily (NR) of the NR1Ⅰfamily, and PXR are responsible in the regulatory regions of CYP3A genes. However, little information is available about the stable stage on CYP3A expression and available on the correlation between PXR and CYP3A involved in the metabolic processes of medicines in fish.
In the paper, We used biochemistry and molecular biology methods. Firstly, We cloned CYP3A gene, analysed the gene structure and function domains, and detected the gene expression level in grass carp tissues. We cloned the PXR gene and detected the expression level in tissues. Secondly, We analysed the correlation of CYP3A mRNA and enzyme activity depending on temporal dynamics for determination of the stable expression in optional induction of grass carp in vitro model. In drug treatment conditions, we studied the different expression and analyzed the correlation between the levels of CYP3A enzyme, CYP3A and PXR mRNA. In initial induced platform, ofloxacin (OFLX) is both a substrate and inducer for CYP3A, we verified the CYP3A stable phase and the correlation of CYP3A activity -CYP3A mRNA-PXR mRNA in the OFLX metabolism process. The major results are as follows:
1. Cloning, sequencing and different mRNA expression of CYP3A gene in Grass carp
According to the sequences of CYP3A gene in zebra fish and rainbow trout from the GenBank, we designed the degenerate primers using premier primer 5.0, then we used primer for polymerase chain reaction (PCR) amplification and gained the partial conserved sequences of CYP3A. According to the conserved sequences, we re-designed the primer to amplify the ORF, then we designed primers for 3' Rapid Amplification of cDNA Ends followed by the conserved sequences. We designed qRT-PCR primers to detect CYP3A mRNA expression in different tissues by fluorescent dye SYBR. On the basis of crucian carp CYP1A, we analyzed the results of different genetic characteristics of different subtypes in fish. The results showed that: (1) The partial CYP3A gene cDNA nucleotide sequences of grass carp is 1847 bp, including conserved coding region and 3'–untranslated region(3' UTR). The CYP3A gene has an open reading frame (ORF) of 1542bp. The ORF encodes a 513 amino acid (aa) protein, which has a signal peptide (29aa) and a stop codon. (2) The main domain of grass carp CYP3A is composed of 3 transmembrane helix, 9 phosphorylation sites, 6 substrate binding site, a heme protein binding and containing a cysteine (Cys, C-448aa) of iron-heme ligand binding site. (3) The quantitative real time polymerase chain reaction (qRT-PCR) assay was developed to estimate the mRNA expression levels in 9 tissues, its expression abundance in turn are as follows: anterior intestine> liver> kidney> gills> muscle> hindgut> gonad> heart> spleen. The Foregut, liver and kidney are the main expression sites,however weak expression was detected in the other tissues. CYP1A expression in curcian carp was assayed with the same method, the results were as follows: liver> foregut> gills> kidney> hindgut > gonads> heart> muscle> spleen, where liver, intestine and gills are the main expression sites. However weak expression were detected in other organs. The distribution of Cytochome P450 isoforms in tissues consist with the main site of drug metabolism.
2. Cloning, sequencing and different mRNA expression of PXR gene in Grass carp
According to the sequences of PXR gene in zebra fish and rainbow trout from the GenBank, we designed the degenerate primers using premier primer 5.0, then we used primer for polymerase chain reaction (PCR) amplification and gained the partial conserved sequences of PXR. We designed qRT-PCR primers to detect PXR mRNA expression in different tissues using fluorescent dye SYBR. The results showed that: (1) The partial PXR gene cDNA nucleotide sequences was 509 bp with the deduced 169 aa for grass carp. (2) the results of qRT-PCR assay showed high similarity with CYP3A gene,that is, anterior intestine> liver> kidney> gills> muscle> hindgut> gonad> heart> spleen.
3. Toxicity test by RIF and OFLX in CIK
Conventional methods were employed for cell continuous culture, resuscitation and cryopreservation. Protein contents was determined by BCA method. MTT reduction was used to optimize the sublethal concentrations at 10-,20-,40-,80 uM, and time for 1-, 2-, 4-, 6-, 12-, 24-, 48-,72 h for RIF test. MTT reduction was used to optimize concentrations at 25 -, 50 -, 100 -, 200 -, 400 and 800μM, and time for 1 -, 2 -, 4 -, 6 -, 8 -, 10 -, 12 -, 24 and 48 h for ofloxacin test. The relative viability of CIK Sharply decreased when the concentrations were set at 200 uM and with incubation time for 48h above incubation with OFLX. The results showed that RIF incubation at 40 uM and 24 h was the best induction model for experiment. The OFLX incubation at 50 uM and 24 h for the further study. It can offer the information to further study RIF and OFLX in CIK.
4. Assay of CYP3A gene transcription and CYP3A enzyme activity with optimal induction model in CIK
In order to evaluate the CYP3A expression in kidney of grass carp, the study determined the CYP3A gene transcription and enzyme activity. CYP3A gene expression after induction by RIF in grass carp kidney cells (CIK) was assayed by quantitative real-time PCR. CYP3A-dependent erythromycin N-demethylase (ERND) activity was determined by spectrophotometry, and CYP3A activity was assessed by measuring the formation of formaldehyde. In the induced group, CYP3A mRNA expression reached a plateau at 8h, while the highest level of CYP3A activity occurred at 10h. The results indicated that CYP3A mRNA expression and enzyme activity were significantly higher than those in the control group (p<0.05). The highest level of CYP3A mRNA expression appeared 2 hours before the maximum of enzyme activity. In addition, the transcription level of CYP3A was apparently higher than the enzyme activity, implying that induction by RIF affect enzyme activity by transcriptional regulation of the CYP3A gene.
5. Establishment of the model for CYP3A activity, CYP3A and PXR mRNA correlation in fish
CYP3A mRNA transcription was up-regulated after 2 h, while PXR was up-regulated after 10 h. The results indicated that the expression of CYP3A was prior to PXR. In addition, the transcription level of CYP3A was apparently higher than the enzyme activity. The PXR transcription level was involved in the activation and induction of CYP3A mRNA expression, but not directly on the enzyme activity. Therefore, we hypothesized that the induction of CYP3A activity in grass carp was dependent on PXR. That induction by RIF affected enzyme activity by transcriptional regulation of the CYP3A gene.
6. The correlation between OFLX ,CYP3A and PXR
We study the correlation between CYP3A and PXR mRNA after incubation by OFLX in CIK against time. CYP3A gene expression was assayed by quantitative real-time PCR, and CYP3A activity was assessed by measuring the formation of formaldehyde. The results showed that: (1) The CYP3A transcription level was up-regulated after 4 h but acted as a weak induction of CYP3A enzyme activities. (2) CYP3A mRNA transcript was up-regulated after 12 h. PXR mRNA have been down-regulated from 1-10 h, and up-regulated after 12 h treated by OFLX. (3) Thus, our work confirmed that OFLX is both the substrate and inducer of CYP3A.
本文首先通过生物化学和分子生物学相结合的方法,先对CYP3A基因进行克隆并分析其结构功能域,检测其组织中表达丰度;再克隆了CYP3A转录因子PXR基因部分序列,检测其组织中表达丰度;其次,研究在草鱼体外最佳诱导模型中,CYP3A在mRNA和酶活性水平与时间动态变化的相关性,确定了CYP3A在鱼类中稳定表达阶段。在药物处理条件下,对PXR基因与CYP3A表达量差异进行研究,分析了CYP3A酶活性-CYP3A mRNA-PXR mRNA三者间相关性。用初步建立的诱导平台,氧氟沙星(OFLX)作为CYP3A的底物和诱导剂,验证CYP3A参与OFLX代谢过程中,其稳定表达阶段和CYP3A酶活性-CYP3A mRNA-PXR mRNA三者间相关性。全文主要研究结果如下:
一草鱼CYP3A和鲫鱼CYP1A基因克隆、序列分析及mRNA表达差异
根据GenBank中斑马鱼和虹鳟的CYP3A基因序列,设计保守区域的简并引物,通过PCR扩增获取CYP3A基因部分保守序列。根据CYP3A已获序列再设计引物,获取开放阅读框,然后,采用3' RACE法获取CYP3A序列末端;设计qRT-PCR引物,用SYBR荧光染料检测CYP3A mRNA组织表达差异。在此基础上,以鲫鱼CYP1A为参照,分析不同鱼类不同亚型基因结果特征。实验结果表明:⑴本研究得到草鱼CYP3A部分cDNA核苷酸序列为1849bp,包含保守编码区、3'UTR序列。开放阅读框序列长1542bp,编码513个氨基酸和一个终止密码(TAA),编码区包括信号肽(29aa)。⑵分析草鱼CYP3A主要结构域,其包含3个跨膜螺旋区,9个磷酸化位点,6个底物结合位点、1个血红素蛋白结合区和含1个半胱氨酸(Cys,C—448aa)的亚铁-血红素配体结合位点;而鲫鱼CYP1A基因结构特征,仅包含1个跨膜螺旋区,其含6个磷酸化位点,1个血红素蛋白结合区和含1个半胱氨酸(Cys,C—454aa)的亚铁-血红素配体结合位点。⑶应用实时荧光定量PCR(qRT-PCR)检测草鱼9种组织CYP3A mRNA相对含量,其表达丰度依次如下:前肠﹥肝脏﹥肾脏﹥鳃﹥肌肉﹥后肠﹥性腺﹥心脏﹥脾脏。其中,前肠、肝脏和肾脏是其主要表达部位,而其他各组织表达量较弱。同样方法检测鲫鱼9种组织CYP1A分布情况,其表达丰度依次如下:肝脏﹥前肠﹥鳃﹥肾脏﹥后肠﹥性腺﹥心脏﹥肌肉﹥脾脏,其中,肝脏、前肠和鳃是其主要表达部位,而其他各组织表达量较弱,CYP1A基因主要表达组织与参与药物代谢部位相同。药物代谢酶各亚型在组织中的分布与其参与药物代谢的主要部位相关。
二草鱼PXR基因克隆、序列分析及mRNA表达差异
根据GenBank中斑马鱼和虹鳟鱼类的PXR基因序列,设计保守区域的简并引物,通过PCR扩增获取PXR基因部分保守序列。设计qRT-PCR引物,用SYBR荧光染料检测PXR mRNA组织表达差异。实验结果表明:⑴本研究得到草鱼PXR部分cDNA核苷酸序列为509bp和推导169aa序列。⑵应用实时荧光定量PCR(qRT-PCR)检测9种组织PXR mRNA相对含量,其表达丰度依次如下:前肠﹥肝脏﹥肾脏﹥鳃﹥肌肉﹥后肠﹥性腺﹥心脏﹥脾脏。其中,前肠、肝脏和肾脏是其主要表达部位,而其他各组织表达量较弱。根据结果显示PXR与CYP3A基因在鱼体组织中表达部位基本一致。
三利福平和氧氟沙星对草鱼肾细胞的毒性实验
细胞的传代培养、冻存与复苏采用实验室常规方法。细胞中蛋白含量检测采用BCA法。采用MTT法分析了利福平在浓度为10-、20-、40和80μM,孵育时间为1-、2-、4 -、6-、8-、10-、12-、24-、48-和72 h;氧氟沙星在浓度分别为25-、50-、100-、200-、400和800μM,对草鱼肾细胞作用1-、2-、4-、6 -、8 -、10 -、12-、24和48 h之后,其结果表明肾细胞中选择RIF浓度为40μM和细胞孵育时间为1～24 h,作为最佳诱导模型进行后续实验,OFLX选择50μM和细胞孵育时间为1～24 h作为最佳条件进行后续实验,能对后续研究利福平和氧氟沙星在肾细胞中最佳代谢时间和浓度提供依据。
四最佳诱导模型CYP3A基因转录和酶活性检测
为了确定CYP3A在草鱼肾细胞中主要诱导及稳定表达阶段,本研究从CYP3A基因转录水平和酶活表达水平进行测定并评价。实验中使用CYP3A指示酶红霉素-N-脱甲基酶(ERND),采用分光光度计法检测CYP3A酶活性;设计qRT-PCR引物,采用SYBR荧光染料检测CYP3A mRNA表达差异。实验结果表明:CYP3A转录水平在2h诱导量提高,而酶活性则在4 h开始升高,两种水平间具有时间差异性,而CYP3A转录水平先于酶活2 h表达;RIF对CYP3A转录水平诱导效应显著强于酶活性水平诱导,这很可能反映了RIF诱导CYP3A表达是作用于转录水平的基因调控来影响酶活性。
五CYP3A与PXR mRNA表达及CYP3A酶活性三者相关性
RIF诱导后,CYP3A转录水平在2 h被诱导基因上调,而PXR则至10 h开始上调,此结果显示出CYP3A和PXR的基因转录水平上调呈现时间先后顺序。推断出PXR与药物结合后,受到药物刺激经时间发生反应,再参与后续的一系列作用;也显示出RIF与PXR结合后能激活PXR,并经由PXR这一转录调控因子参与激活并诱导CYP3A mRNA表达,不直接作用于翻译后酶活性,CYP3A酶活性主要是通过转录水平上调而达到活性增强的效果。
六氧氟沙星、CYP3A和PXR相关性
氧氟沙星(OFLX)孵育CIK,研究其时间动态过程中CYP3A与PXR量变相关性。用分光光度法测定CYP3A酶活性,用实时定量PCR检测CYP3A和PXR基因表达量。实验结果表明:⑴CYP3A酶活性在氧氟沙星处理后4 h开始提高,达致平台期后处于稳定阶段,而CYP3A mRNA在孵育10 h后开始开始上调。⑵PXR mRNA基底水平表达量较低,在OFLX孵育6～8 h后,逐渐上调,其mRNA开始上调时间先于CYP3A mRNA发生。由此推断整个药物代谢过程首先是OFLX经与PXR结合,然后其结合复合物作为转录调控调控因子参与激活CYP3A诱导表达。⑶结果显示氧氟沙星既是CYP3A代谢底物又是CYP3A一种诱导剂。
Cytochrome P450 (CYP) is an important animal drug metabolism enzyme. Cytochrome P450 3A (CYP3A), responsible for the metabolism of the majority of drugs, comprises the largest portion of the liver, small intestinal and kidney P450s in animals. CYP1A are responsible for the biotransformation of drug, CYP1A and CYP3A are induced or inhibited by drug. Pregnane X receptor (PXR) belongs to the nuclear receptor superfamily (NR) of the NR1Ⅰfamily, and PXR are responsible in the regulatory regions of CYP3A genes. However, little information is available about the stable stage on CYP3A expression and available on the correlation between PXR and CYP3A involved in the metabolic processes of medicines in fish.
In the paper, We used biochemistry and molecular biology methods. Firstly, We cloned CYP3A gene, analysed the gene structure and function domains, and detected the gene expression level in grass carp tissues. We cloned the PXR gene and detected the expression level in tissues. Secondly, We analysed the correlation of CYP3A mRNA and enzyme activity depending on temporal dynamics for determination of the stable expression in optional induction of grass carp in vitro model. In drug treatment conditions, we studied the different expression and analyzed the correlation between the levels of CYP3A enzyme, CYP3A and PXR mRNA. In initial induced platform, ofloxacin (OFLX) is both a substrate and inducer for CYP3A, we verified the CYP3A stable phase and the correlation of CYP3A activity -CYP3A mRNA-PXR mRNA in the OFLX metabolism process. The major results are as follows:
1. Cloning, sequencing and different mRNA expression of CYP3A gene in Grass carp
According to the sequences of CYP3A gene in zebra fish and rainbow trout from the GenBank, we designed the degenerate primers using premier primer 5.0, then we used primer for polymerase chain reaction (PCR) amplification and gained the partial conserved sequences of CYP3A. According to the conserved sequences, we re-designed the primer to amplify the ORF, then we designed primers for 3' Rapid Amplification of cDNA Ends followed by the conserved sequences. We designed qRT-PCR primers to detect CYP3A mRNA expression in different tissues by fluorescent dye SYBR. On the basis of crucian carp CYP1A, we analyzed the results of different genetic characteristics of different subtypes in fish. The results showed that: (1) The partial CYP3A gene cDNA nucleotide sequences of grass carp is 1847 bp, including conserved coding region and 3'–untranslated region(3' UTR). The CYP3A gene has an open reading frame (ORF) of 1542bp. The ORF encodes a 513 amino acid (aa) protein, which has a signal peptide (29aa) and a stop codon. (2) The main domain of grass carp CYP3A is composed of 3 transmembrane helix, 9 phosphorylation sites, 6 substrate binding site, a heme protein binding and containing a cysteine (Cys, C-448aa) of iron-heme ligand binding site. (3) The quantitative real time polymerase chain reaction (qRT-PCR) assay was developed to estimate the mRNA expression levels in 9 tissues, its expression abundance in turn are as follows: anterior intestine> liver> kidney> gills> muscle> hindgut> gonad> heart> spleen. The Foregut, liver and kidney are the main expression sites,however weak expression was detected in the other tissues. CYP1A expression in curcian carp was assayed with the same method, the results were as follows: liver> foregut> gills> kidney> hindgut > gonads> heart> muscle> spleen, where liver, intestine and gills are the main expression sites. However weak expression were detected in other organs. The distribution of Cytochome P450 isoforms in tissues consist with the main site of drug metabolism.
2. Cloning, sequencing and different mRNA expression of PXR gene in Grass carp
According to the sequences of PXR gene in zebra fish and rainbow trout from the GenBank, we designed the degenerate primers using premier primer 5.0, then we used primer for polymerase chain reaction (PCR) amplification and gained the partial conserved sequences of PXR. We designed qRT-PCR primers to detect PXR mRNA expression in different tissues using fluorescent dye SYBR. The results showed that: (1) The partial PXR gene cDNA nucleotide sequences was 509 bp with the deduced 169 aa for grass carp. (2) the results of qRT-PCR assay showed high similarity with CYP3A gene,that is, anterior intestine> liver> kidney> gills> muscle> hindgut> gonad> heart> spleen.
3. Toxicity test by RIF and OFLX in CIK
Conventional methods were employed for cell continuous culture, resuscitation and cryopreservation. Protein contents was determined by BCA method. MTT reduction was used to optimize the sublethal concentrations at 10-,20-,40-,80 uM, and time for 1-, 2-, 4-, 6-, 12-, 24-, 48-,72 h for RIF test. MTT reduction was used to optimize concentrations at 25 -, 50 -, 100 -, 200 -, 400 and 800μM, and time for 1 -, 2 -, 4 -, 6 -, 8 -, 10 -, 12 -, 24 and 48 h for ofloxacin test. The relative viability of CIK Sharply decreased when the concentrations were set at 200 uM and with incubation time for 48h above incubation with OFLX. The results showed that RIF incubation at 40 uM and 24 h was the best induction model for experiment. The OFLX incubation at 50 uM and 24 h for the further study. It can offer the information to further study RIF and OFLX in CIK.
4. Assay of CYP3A gene transcription and CYP3A enzyme activity with optimal induction model in CIK
In order to evaluate the CYP3A expression in kidney of grass carp, the study determined the CYP3A gene transcription and enzyme activity. CYP3A gene expression after induction by RIF in grass carp kidney cells (CIK) was assayed by quantitative real-time PCR. CYP3A-dependent erythromycin N-demethylase (ERND) activity was determined by spectrophotometry, and CYP3A activity was assessed by measuring the formation of formaldehyde. In the induced group, CYP3A mRNA expression reached a plateau at 8h, while the highest level of CYP3A activity occurred at 10h. The results indicated that CYP3A mRNA expression and enzyme activity were significantly higher than those in the control group (p<0.05). The highest level of CYP3A mRNA expression appeared 2 hours before the maximum of enzyme activity. In addition, the transcription level of CYP3A was apparently higher than the enzyme activity, implying that induction by RIF affect enzyme activity by transcriptional regulation of the CYP3A gene.
5. Establishment of the model for CYP3A activity, CYP3A and PXR mRNA correlation in fish
CYP3A mRNA transcription was up-regulated after 2 h, while PXR was up-regulated after 10 h. The results indicated that the expression of CYP3A was prior to PXR. In addition, the transcription level of CYP3A was apparently higher than the enzyme activity. The PXR transcription level was involved in the activation and induction of CYP3A mRNA expression, but not directly on the enzyme activity. Therefore, we hypothesized that the induction of CYP3A activity in grass carp was dependent on PXR. That induction by RIF affected enzyme activity by transcriptional regulation of the CYP3A gene.
6. The correlation between OFLX ,CYP3A and PXR
We study the correlation between CYP3A and PXR mRNA after incubation by OFLX in CIK against time. CYP3A gene expression was assayed by quantitative real-time PCR, and CYP3A activity was assessed by measuring the formation of formaldehyde. The results showed that: (1) The CYP3A transcription level was up-regulated after 4 h but acted as a weak induction of CYP3A enzyme activities. (2) CYP3A mRNA transcript was up-regulated after 12 h. PXR mRNA have been down-regulated from 1-10 h, and up-regulated after 12 h treated by OFLX. (3) Thus, our work confirmed that OFLX is both the substrate and inducer of CYP3A.
引文
Amold, K, Bordoli.L, Kopp.J, Schwede.T. 2006. The SWISS-MODEL workspace: a web-based environment for protein structure homology modelling [J]. Bioinformatics, 22(2): 195-201.
Andersen V, Pedersen N, Larsen NE, Sonne J, Larsen S.2002. Intestinal first pass metabolism of midazolam in liver cirrhosis-effect of grapefruit juice [J]. Br J Clin Pharmacol. 54:120–4.
Anzenbacher P, Anzenbacherova E. 2001. Cytochromes P450 and metabolism of xenobiotics [J]. Cellular and Molecular Life Sciences, 58: 737-747
Araujo CSA, Marques SAF, Carrondo MJT. 2000. In vitro response of the brown bullhead catfish (BB) and rainbow trout (RTG-2) cell lines to benzopyrene [J]. The Science of the Total Environment, 247: 127-135.
Arnold G E, Ornstein R L. 1997. Molecular dynamics study of time-correlated protein domain motions and molecular flexibility: cytochrome P450BM-3 [J]. Biophys J, 73 (3): 1147-1159.
Arnold, K, Bordoli, L, Kopp, J, Schwede, T. 2006. The SWISS-MODEL workspace: a web-based environment for protein structure homology modelling [J]. Bioinformatics, 22(2): 195-201.
Babín M, Casado S, Chana A, et al.2005. Cytochrome P4501A induction caused by the imidazole derivative Prochloraz in a rainbow trout cell line [J]. Toxicol In Vitro, 19(7): 899-902.
BarralloA., Gonzalez-SarmentoI R., Garcia-Isidoro M., Cidad P., Porteros A., and Rodriguez R.E., 1999, Differential brain expression of a new beta-actin gene from zebrafish (Danio rerio) [J]. Eur. J. Neurosci., 11(1):369-372.
BauerB, HartzAM S, Fricker G, MillerD S .2004. Pregnane X receptor upregulation of P-glycop rotein expression and transport function at the blood-brain barrier[J]. Mol Pharmacol, 66: 413- 9
BauerB, HartzAM S, Fricker G, MillerD S. 2004. Pregnane X receptor upregulation of P-glycoprotein expression and transport function at the blood-brain barrier[J]. Mol.Pharmacol, 66: 413- 9.
Billiard S M, Bols N C, Hodson P V. 20040. In vitro and in vivo comparisons of fish-specific CYP1A induction relative potency factors for selected polycyclic aromatic hydrocarbons[J]. Ecotoxicology and Environmental Safety, 59: 292-299.
Bork R W, Muto T, Beaune P H, et al. 1989. Characterization of mRNA species related to human liver cytochrome P450 nifedipine oxidase and the regulation of catalytic activity[J]. The Journal of Biological Chemistry, 264: 910-919.
Brandon EFA, Raap CD, Meijerman I, et al. 2003. An update on in vitro test methods in human hepatic drug biotransformation research: pros and cons [J]. Toxicol Appl Pharmacol, 189: 233-246.
Bresolin T, Rebelo M D F, Bainy A C D. 2005. Expression of PXR, CYP3A and MDR1 genes inliver of zebrafish[J]. Comparative Biochemistry and Physiology C, 140: 403-407.
Britt Wassmur, Johanna Gr?ns, Peter Kling, Malin C. Celander. 2010. Interactions of pharmaceuticals and other xenobiotics on hepatic pregnane X receptor and cytochrome P450 3A signaling pathway in rainbow trout(Oncorhynchus mykiss) [J]. Aquatic Toxicology. 100:91–100.
Buhler D R, Miranda C L, Henderson M C, et al. 2000. Effects of 17β-estradiol and testosterone on hepatic mRNA/protein levels and catalytic activities of CYP2M1, CYP2K1, and CYP3A27 in rainbow trout (Oncorhynchus mykiss) [J]. Toxicology and Applied Pharmacology, 168: 91-101.
Bustin S.A., 2000. Absolute quantification of rnRNAusing rea1-time reverse transcription polymerase chain reaction assays[J]. J.Mol.Endocrinol. 25(2):169-193.
Caminada D, Escher C, Fent K. 2006. Cytotoxicity of pharmaceuticals found in aquatic systems: Comparison of PLHC-1 and RTG-2 fish cell lines[J]. Aquatic Toxicology. (79):114–123.
Cawley G F, Batie C J, Backs W L. 1995. Structure and Function, and Enzymol Relot Areas[J]. Mol, Biol, 60:35-87.
Chen Q, Lu H, Xu X Y et al.1992. Pharmacokinetic and relative bioavailability of ofloxacin after a single oral administration in Chinese healthy volunteers[J]. Chin J Clin Pharmacol, 8:193-197.
Chen Y H, Wang J P, Wang H, et al. 2005. Lipopolysaccharide treatment downregulates the expression of the pregnane X receptor, cyp3a11 and mdr1a genes in mouse placenta [ J ]. Toxicology, 211: 242– 52.
Chirulli V, Longo V, Marini S, et al. 2005. CAR and PXR expressionand inducibility of CYP2B and CYP3A activities in rat and rabbitlungs[J]. Life Sci, 76:2535-46.
ChuW.Y., ShiC.Y.,HuangR, LZhangH, Fand Yin Y.L.2009.Molecular cloning,tissue distribution and ontogenetie expression of the amino acid transporter ba+cDNA in the small intestine of Tibetan suckling piglets,154(1): 157-164.
Coon M J, Vaz A D , Bestervelt L L.1996. Cytochrome P450 2: Peroxidative reactions of diversozymes[J]. FASEB J,10:428-434.
Coon MJ, Ding X, Pernecky SJ et al. 1992.Cytochrome P450: Progress and predictions, FASEB J.6:669-673.
Courtenay, S., Grunwald, C., Kreamer, G, et al.1993. Induction and clearance of cytochrome P4501A mRNA in Atlantic tomcod caged in bleached kraft mill effluent in the Miramichi River [J]. Aquatic Toxicology, 27: 225–244.
Cravedi J P, Perdu-Durand E, Paris A. 1998. Cytochrome P450-dependent metabolic pathways and glucuronidation in trout liver slices[J]. Comparative Biochemistry and Physiology C, 121: 267-275.
Eichelbaum M, Burk O.2001. CYP3A genetics in drug metabolism[J].NatureMed, 2001,7 (3):285- 287.
Ekins,S., Reschly, E.J.,Hagey, L.R., Krasowski,M.D., 2008. Evolution of pharmacologic specificity in the pregnane X receptor[J]. BMC Evol. Biol. 8:103.
Elcombe CR, Lech JJ. 1979. Induction and characterization of hemoprotein(s) P450 and monoxyge-nation in rainbow trout (Salmo Gairdneri). Toxicol Appl Pharmacol, 49(3): 437-450.
Estabrook R W. 1996. The remarkable P450s: A historical overview of the versatile heme protein catalysts[J]. FASEB J,10:202-204.
Fent K. 2001. Fish cell lines as versatile tools in ecotoxicology: Asssessment of cytotoxicity, cytochrome P450 1A induction potential and estrogenic activity of chemicals and environmental samples [J]. Toxicol In Vitro, 15: 477-488.
F(o|¨)rlin L, Celander M. 1995. Studies of the inducibility of P450 1A in perch from the PCB-contaminated lake J?rnsj?n in Sweden. Marine Environmental Research, 39: 85-88.
Gasteiger E., Hoogland C., Gattiker A., Duvaud S., Wilkins M.R., Appel R.D., A.;, B, 2005. Protein Identification and Analysis Tools on the ExPASy Server. In: Walker, J M (Ed.), The Proteomics Protocols Handbook. Humana Press.
Gibson GG, Skett P.1994. Introduction to Drug Metabolism, (2nd edition), Chapman & Hall, Londin.
Giorgi, M., Marini, S., Longo,V., et al. 2000. Cytochrome P450-dependent monooxygenase activities and their inducibility by classic P450 inducers in the liver, kidney, and nasal mucosa of male adult ring-necked pheasants [J]. Toxicology and Applied Pharmacology, 167: 237–245.
Gotoh, O., 1992. Substrate recognition sites in cytochrome P450 family 2 (CYP2) proteins inferred from comparative analyses of amino acid and coding nucleotide sequences[J]. J. Biol. Chem. 267:83– 90.
Graham S E, Peterson J A.1999. How similar are P450s and what can their differences teach us. Arch. Biochem. Biophysics. 369:24-29.
Graham-Lorence S E, Amarneh B, White R E et al. 1995. A three-dimensional model of aromatase cytochrome P 450. Protein Sci. 4 (6):1065-1080.
Graham-Lorence S, Peterson J A.1996. Cytochrome P450 1: structural similarities and functional differences. FASEB J,10:206-214.
Gravato C, Teles M, Oliveira M, et al. 2006. Oxidative stress, liver biotransformation and genotoxic effects induced by copper in Anguilla anguilla L. the influence of pre-exposure to β-naphthoflavone. Chemosphere, 65: 1821-1830.
Guengerich F P, Turvy C G.1991. Comparison of levels of several human microsomal cytochrome P450 enzymes and epoxide hydroxylase in normal and disease states using immunochemialanalysis of surgical liver samples[J]. J. Pharmacol.Exp. Ther., 256:1189-1194.
Guex, N, Peitsch, MC. 1997b. SWISS-MODEL and the Swiss-PdbViewer: An environment for comparative protein modeling [J]. Electrophoresis 18: 2714-2723.
Gui hong Fu, Xian le Yang, Hai Xin Zhang, Wen Juan Yu, Kun Hu. 2011. Effects of cytochrome P450 1A substrate (Difloxacin) on enzyme gene expression and pharmacokinetics in crucian carp (hybridized Prussian carp). Environmental Toxicology and Pharmacology. 31(1):307-313.
Handschin C, Podvinec M, Meyer U A.2000. CXR, a chicken xenobiotic-sensing orphan nuclear receptor, is related to both mammalian pregnane X receptor (PXR) and constitutive androstane receptor (CAR) [J]. ProcNatlAcad SciUSA, 97:10769 - 74.
Hasselberg L, Meier S, Svardal A, et al.2004. Effects of alkylphenols on CYP1A and CYP3A expression in first spawning Atlantic cod (Gadus morhua)[J]. Aquatic Toxicology, 67: 303-313
Hegelund T, Celander M C. 2003. Hepatic versus extrahepatic expression of CYP3A30 and CYP3A56 in adult killifish (Fundulus heteroclitus). Aquatic Toxicology, 64: 277-291.
Henrik, N, Jacob, E, S?ren, B, Gunnar, vH. 1997. Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites.
Hines RN, Piechocki MP, Boncher PD. 1994. Molecular mechanisms controlling CYP1A gene expression[J]. Frontiers in Biotransformation.9:85-110.
Hodgson E.1958. Microsomal monooxygenase. In:Kurkut G A, GilbertL I(eds), Comprehesive insect physiology, biochemistry and pharmacology 11[J], Oxford: Pergamon, 225-321.
Hoover K L(ed). 1984. Use of small fish species in carcinogenicity testing. National Cancer Institute Monograph.65.
Hua-Pin Tsenga, Tzong-Hsiung Hseua, Donald R. Buhler, Wen-Der Wangc, Chin-Hwa Huc. 2005. Constitutive and xenobiotics-induced expression of a novel CYP3A gene from zebrafish larva. Toxicology and Applied Pharmacology. 205:247- 258.
Hui Wang, Zhang-Xiu Liao, Man Chen, Xiao-Ling Hu. 2006. Effects of hepatic fibrosis on ofloxacin pharmacokinetics in rats. Pharmacological Research. 53: 28–34.
James M O, Lou Z, Rowland-Faux L, et al.2005. Properties and regional expression of a CYP3A-like protein in channel catfish intestine. Aquatic Toxicology.72: 361-371.
Jannick, DB, Henrik, N, Gunnar, vH, S?ren, B. 2004. Improved prediction of signal peptides SignalP 3.0 [J]. J. Mol. Biol., 304: 783-795.
Jansson I. 1993. Post-transcriptional modification of cytochrome P450. In Cytochrome P450 Schenkam, JB and Griem H, (ends). Berlin: Springer-Verlag, 361-380.
Jian Yang, Chunshu Hao, Dongfang Yang, et al. 2010. Pregnane X receptor is required for interleukin-6-mediated down-regulation of cytochrome P450 3A4 in human hepatocytes [J].Toxicology Letters. 197: 219–226.
Jones S A, Moore L B, Shenk J L, et al . 2000. The p regnane X receptor : a p romiscuous xenobi otic recep t or that has diverged during evolution[ J ]. Mol Endocrinol, 14: 27 - 39.
Kashiwada S, Hinton D E, Kullman S W. 2005. Functional characterization of medaka CYP3A38 and CYP3A40: kinetics and catalysis by expression in a recombinant baculovirus system. Comparative Biochemistry and Physiology C, 141: 338-348.
Kashiwada S, Hinton D E, Kullman S W. 2005. Functional characterization of medaka CYP3A38 and CYP3A40: kinetics and catalysis by expression in a recombinant baculovirus system. Comparative Biochemistry and Physiology C, 141: 338-348.
Kashiwada, S., Kameshiro, M., Tatsuta, et al.2007. Estrogenic modulation of CYP3A38, CYP3A40, and CYP19 in mature male medaka (Oryzias latipes) [J]. Comp. Biochem. Physiol.C,145:370–378.
Kiefer, F, Arnold, K, Künzli, M, Bordoli, L, Schwede, T. 2009. The SWISS-MODEL Repository and associated resources[J]. Nucleic Acids Research, 37(suppl 1): D387-D392.
Kliewer S A, Moore J T, Wade L, et al . 1998. An or phan nuclear re2cep t or activated by p regnanes defines a novel steroid signalingpathway[ J ]. Cell, 92: 73 - 82.
Krasowski, M.D., Yasuda, K., Hagey, L.R., Schuetz, E.G., 2005. Evolutionary selection across the nuclear hormone receptor superfamily with a focus on the NR1I subfamily (vitamin D, pregnane X, and constitutive androstane receptors)[J]. Nucl.Recept. 3, 2.
Kullman S W, Hinton D E. 2001. Identification, characterization, and ontogeny of a second cytochrome P450 3A gene from the fresh water teleost medaka (Oryzias latipes) [J]. Molecular Reproduction and Development, 58: 149-158.
Kullman, S.W., Hamm, J.T., Hinton, D.E, et al. 2000. Identification and characterization of a cDNA encoding cytochrome P450 3A from the fresh water teleost medaka (Oryzias latipes) [J]. Arch. Biochem. Biophys, 380: 29– 38.
Lee S J, Buhler D R.2003. Cloning, tissue distribution, and functional studies of a new cytochrome P450 3A subfamily member, CYP3A45, from rainbow trout (Oncorhynchus mykiss) intestinalceca. Archives of Biochemistry and Biophysics, 412: 77-89.
Lee S J, Wang-Buhler J L, Cok I, et al. 1998. Cloning, sequencing, and tissue expression of CYP3A27, a new member of the CYP3A subfamily from embryonic and adult rainbow trout livers[J]. Archives of Biochemistry and Biophysics, 1998, 360(1): 53-61.
Lee Y M, Seo J S, Kim I C, et al.2006. Endocrine disrupting chemicals (bisphenol A, 4-nonylphenol, 4-tert-octylphenol) modulate expression of two distinct cytochrome P450 aromatase genes differently in gender types of the hermaphroditic fish Rivulus marmoratus[J].Biochemical and Biophysical Research Communications, 345: 894-903.
Lehmann J M, McKee D D, Watson M A, et al. 1998. The human orphan nuclear receptor PXR is activated by compounds that regulateCYP3A4 gene expression and cause drug interactions[J]. J ClinInvest, 102: 1016 - 23.
Lewis DFV. 1996 Cytochromes P 450 Structure, Function and Mechanism. Taylor& Francis: London,79-88.
Li A P, Kaminski D L, 1995. Rasmussen A. Substrates of human hepatic cytochrome P450 3A4. Toxicology, 104: l-8.
Li D, Yang XL, Zhang SJ, et al.2008. Effects of mammalian CYP3A inducers on CYP3A-related enzyme activities in the freshwater teleost grass carp (Ctenopharyngodon idellus): possible implications for the establishment of a fish CYP3A induction model[J]. Comparative Biochemistry and Physiology, part C. 147,17-2.
Li H, Poulos T L.1996. Conformational dynamics in cytochrome P450-substrate interaction. Biochimie. 78(8-9):695-699.
Livak, KJ, Schmittgen, TD. 2001. Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2-△△CT Method [J]. Methods, 25(4): 402-408.
Lown K, Kolars J, Turgeon K et al. 1992. The erythromycin breath test selectively measures P450ⅢA in patients with severe liver disease. Clin. Pharmacol. Ther.,51:229-238.
Luo G, CUNNINGHAMM, KIM S, et al. 2002. CYP3A4 induction by drugs: correlation between apregnaneX receptor reportergene assay and CYP3A4 expressing in human hepatocytes[ J ]. Drug MetabDispos, 30(7):795-804.
LUO G, GUENTHNER T, GAN LS, et al. 2004. CYP3A4 induction by xenobiotics: biochemistry, experimental methods and impact on drug discovery and development[J]. CurrDrug Metab, 5 (6):483 - 505.
Ma X, Shah Y, CheungC, et al. 2007. The pregnane X receptor gene humanized mouse: a model for investigating drug-drug interactionsmediated by cytochromes P450 3A[J]. Drug Metab Dispos, 35(2):194 - 200.
Maenesh M D, Anders M W. 1973. Characterization of the heme of cytochrome P450 using gas chromatography-masspectrometry. Arch Biochem Biophys. 159(1):201-205.
Maglich J M, Caravella J A, Lambert M H, et al.2003. The first completed genome sequence from a teleost fish (Fugu rubripes) adds significant diversity to the nuclear receptor superfamily. Nucleic Acids Research, 31: 4051-4058.
Marangos M N, Zhu Z, Nicolau D P. 1997.Disposition of ofloxacin in femal New Zealand White rabbits [J]. J. Vet. Pharmacol. Therap.20:17-20.
Martel, P.H., Kovacs, T.G., O’Connor, B.I., et al. 1994. A survey of pulp and paper efffluents for their potential to induce mixed function oxidase enzyme activity in fish [J]. Water Research, 28: 1835–1844.
Masuyama H, Hiramatsu Y, Mizutani Y, et al.2001. The expression of pregnane X receptor and its target gene, cytochrome P450 3A1, in perinatalmouse[J]. Mol Cell Endocrinol, 172:47-56.
McArthur, A.G., Hegelund, T., Cox, R.L.2003.Phylogenetic analysis of the cytochrome P4503 (CYP3) gene family[J]. J. Mol. Evol, 57: 200-211.
Meldgaard M., Fenger C., LambertsenK. L., Pedersen M.D., Ladeby R., Finsen B., 2006, Validation of two reference genes for mRNA level studies of routine disease models in neurobiology journal of neuroscience methods[J], Journal of Neuroscience Methods,156(1-2):10l-110.
Meucci V, Arukwe A. 2006. The xenoestrogen 4-nonylphenol modulates hepatic gene expression of pregnane X receptor, aryl hydrocarbon receptor, CYP3A and CYP1A1 in juvenile Atlantic salmon (Salmo salar)[J]. Comparative Biochemistry and Physiology C,142:142-150.
Miki Y, Suzuki T, Tazawa C, et al. 2005. Ster oid and xenobi otic receptor ( SXR) , cytochrome P450 3A4 andmultidrug resistance gene 1in human adult and fetal tissues[J]. Mol Cell Endocrinol, 231:75-85.
Miki Y, Suzuki T, Tazawa C, et al. 2005. Steroid and xenobiotic receptor ( SXR) , cytochrome P450 3A4 andmultidrug resistance gene in human adult and fetal tissues[ J ]. Mol Cell Endocrinol,231: 75 - 85.
Miranda C L, Henderson M C, Buhler D R.1998. Evaluation of chemicals as inhibitors of trout cytochrome P450s. Toxicology and Applied Pharmacology, 148: 237-244.
Modi S, Sutcliffe M J, Primrose W U, Lian L Y, Roberts G C.1996. The catalytic mechanism of cytochrome P450 BM3 involves a 6 A movement of the bound substrate on reduction[J]. Nat Struct Biol., 3(5):414-417.
MOORE JT, KLIEWER SA. 2000. Use of the nuclear receptor PXR to predict drug interaction [ J ]. Toxicology, 153(1/3):1-10.
Moore, L.B., Maglich, J.M., McKee, D.D.,Wisely, B.,Willson, T.M., Kliewer, S.A., Lambert,M.H.,Moore, J.T., 2002. Pregnane X receptor (PXR), constitutive androstane receptor (CAR), and benzoate X receptor (BXR) define three pharmacologically distinct classes of nuclear receptors[J]. Mol. Endocrinol. 16: 977-986.
Mosmann T. Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays[J]. J Immunol Methods,1983,65:55-63.
Moutou K A, Burke M D, Houlihan D F. 1998. Hepatic P450 monooxygenase response in rainbowtrout administered aquaculture antibiotics. Fish Physiology and Biochemistry,18(1): 97-106.
Murray BP, Edwards RJ, Davies DS et al.1993a. Conservation of a functionally important surface region between two families of the cytochrome superfamily, Biochemical J. 292:309-312.
Murray BP. 1992. P450 enzymes: Inhibition mechanisms, genetic regulation and effects of liver disease[J], Clinical Pharmacokinetic Concepts.23:132-146.
Natalya Hanlona, Nick Coldhamb, Maurice J. Sauerb and Costas Ioannides.2008. Up-regulation of the CYP1 family in rat and human liver by the aliphatic isothiocyanates erucin and sulforaphane[J]. Toxicology. 252: 92-98.
Nebert D W, Gonzalez F J. 1987. P450 gene:structure, evolution and regulation[J]. Annu Rev. Biochem.369(1):1-10.
Nebert D W, Nelson D R, Coon M J et al. 1991. The P450 superfamily: update on new sequences, gene mapping and recommended nomenclature[J].DNA Cell. Biol. 10:1-14.
Negishi M et al. 1996. Cytochrome P450 4: Structural flexibility and functional versatility of mammalian P450 enzymes. FASEB J,10:683-689.
Nelson D R, Kamataki T, Waxman D J et al. 1993. The P450 superfamily: update on new sequence ,gene mapping, accession numbers, early trivial names of enzymes and nomenclature[J]. DNA Cell Biology., 12(1):1-15.
Nelson, D.R.. 2003. Comparison of P450s from human and fugu: 420 million years of vertebrate P450 evolution[J]. Arch.Biochem.Biophys, 409: 18– 24.
Nobuo Nemoto, and Junko Sakurai,. 1992. Altered regulation of Cyp1a-1 gene expression during cultivation of mouse hepatocytes in primary culture[J]. Biochemical Pharmacology. 44, 51-58.
Nuclear Receptors Nomenclature Committee .1999. A unified nomenclature system for the nuclear receptor superfamily [J]. Cell 97:161–163.
OlsvikP. A., Lie k. k.,Jordal A.E.O., Nilsen T.O., and Hordvik L, 2005. Evaluation of potential reference genes in real-time RT-PCR studies of Atlantic salmon[J]. BMC Mol. Biol., 6:21.
Omata Y, Aibara K, Ueno Y. 1987.Conformation between the substrate-binding site and heme of cytochrome P450 studied by exciation energy transfer, Biochemical et Physic Acta, 912:115-123.
Omura T, Ishimura Y, Fujii-Kuriyama Y (eds.).1993.Cytochrome P450 (2nd edition). Kodansha. VCH Tokyo.
Omura T. 1999. Forty years of cytochrome P450. Biochemical and Biophysical Research Communications. 266(3): 690-698.
Oost R Vander, Satumalay A Lero, K and Vermeulen NPE.2000. Validation of inland water pollution assessment using biomarker responses in caged carp[J].Marine Environmental Research. 50: 431-432.
OransJ, Teotico DG, Redinbo MR.2005. The nuclear xenobiotic receptor pregane X receptor: recent insights and new challenges [J]. Mol Endocrinol, 19(12):2891-2900.
Outman WR, Nightingale CH. 1989. Metabolism and the fluoroquinolones.Am J Med 87(Suppl. 6C):37–42.
PascussiJ M, Drocourt L, Fabre J M, et al.2000. Dexamethasone induces pregnane X receptor and retinoid X receptor-αexpression in human hepatocytes: synergistic increase of CYP3A4 induction by pregnane X receptor activators[J]. Mol Pharmacol, 58:361-72.
Paulsen M D, Ornstein R L. 1995. Ligand partitioning into membranes: It significance in determing Km and Ks values for cytochrome P450 and other membrane bound receptors and enzyme. FEBS letters, 67(2):23-29.
Peng H, Cheng ZN. 2001. Drug interaction related to CYP3A4 [J]. Chin J Clin Pharmarcol, 17(5):379-385.
Piddock L J V. 1989. Mechanisms of resistance to quinolonges anclinical perspectives[J]. J. Antimicrob. Chemother, 23: 475.
Plant N. 2007. The human cytochromeP450 sub-family: transcriptional regulation, inter-individual variation and interaction net works [J].Biochim BiophysActa,1770(3): 478-488.
Porter TD, Coon MJ. 1991. Cytochrome P450: multiplicity of isoforms, substrate and catalytic and regulatory mechanisms[J]. J. Biol. Chem.266:13469-13472.
Pyerin W, Taniguchi H. 1991. Post-transcriptional modification by phosphorylation: Control of cytochrome P450 and associated enzymes. In Molecular Aspects of Monooxygenases and Bioactivation of Toxic Compounds. Arinc, E., Schenkman JB and Hodgson E (ends.). New York: Plenum,199-232.
Quattrochi LC, Guzelian PS.2001. CYP3A regulation: from pharmacology to nuclear receptors [J]. Drug Metab Dispos. 29(5) :615-622.
R.I.弗雷谢尼著.章静波译.动物细胞培养.背景:科学出版社.2004.
Raku Shinkyoa, Toshiyuki Sakakia, Miho Ohtab and Kuniyo Inouye., 2003. Metabolic pathways of dioxin by CYP1A1: species difference between rat and human CYP1A subfamily in the metabolism of dioxins. Archives of Biochemistry and Biophysics. 409, 180-187.
Ravichandran K G, Boddupalli S S, Hasemann C A et al. 1993. Crystal structure of hemoprotein domain of P450 BM-3, a prototype for microsomal P450s. Science., 261:731-736.
Rost, B, Yachdav, G, Liu, J. 2004. The PredictProtein server [J]. Nucleic Acids Research, 32(suppl 2): W321-W326.
Roussel, F., Khan, K.K., Halpert, J.R, et al.2000. The importance of SRS-1 residues in catalytic specificity of human cytochrome P450 3A4 [J]. Arch. Biochem. Biophys, 374: 269-278.
Roussel, F., Khan, K.K., Halpert, J.R., 2000. The importance of SRS-1 residues in catalytic specificity of human cytochrome P450 3A4[J]. Arch. Biochem. Biophys. 374:269– 278.
Sarmiento J., Leal S., Quezada C., 2000. Envrironmental acclimatization of the carp modulates the transription of beta-actin,J.CellBiochem., 80(2):223-228.
Son D S. Ikenoue N. Tagawa Y. 2000. Non-linear pharmacokinetics of ofloxacin after a single intravenous bolus dose in pigs [J]. J. Vet. Pharmacol. Therap, 23:311-315.
Sonoda J, Chong LW, DownesM, et al.2005. Pregnane X receptor prevents hepatorenal toxicity from cholesterol metabolites[J].Proc. Natl. Acad Sci USA. 102(6):2198 - 2203.
SONODA J, ROSENFELD JM, XU L, et al. 2003. A nuclear receptor-mediated xenobiotic response and its implication in drugmetaboism and host protection [J]. Curr Drug Metab. 4 (1):59-72.
SouidiM, Gueguen Y, Linard C, et al. 2005. In vivo effects of chroniccontaminati on with dep leted uraniumon CYP3A and ass ociated nu2clear recep tors PXR and CAR in the rat [J]. Toxicology, 214: 113-22.
SouidiM, Gueguen Y, Linard C, et al. 2005. In vivo effects of chroniccontaminati on with dep leted uranium on CYP3A and ass ociated nuclear receptors PXR and CAR in the rat [J]. Toxicology. 214: 113-22.
Squires EJ, Sueyoshi T, NegishiM.2004. Cytoplasmic localization of pregnane X receptor and ligand2dependent nuclear translocation in mouse liver[J]. J Biol Chem, 279 (47):49307-49314.
SuzukiT., HigginsP.J., and Crawford D.R., 2000. Control selection for RNA quantitation[J]. Bio.Techniques, 29(2):332-337.
Tan F, Wang M, Wang W, et al.2008. Comparative evaluation of the cytotoxicity sensitivity of six fish cell lines to four heavy metals in vitro. Toxicology in Vitro. 22, 164–170.
Tang J, Yang X L, Zheng Z L, et al. 2006. Pharmacokinetics and its active metabolite of enrofloxacin in Chinese mitten-handed crab (Eriocheir sinensis). Aquaculture, 260: 69-76.
Xue, L., Zgoda, V.G., Arison, B. et al. 2003. Structure–function relationships of rat liver CYP3A9 to its human liver orthologs: site-directed active site mutagenesis to a progesterone dihydroxylase [J]. Arch. Biochem. Biophys, 409:113-126.
Teng S, MillerM P.2005. The involvement of the pregnane X receptor in hepatic gene regulation during inflammation in mice [J]. J Pharmacol Exp Ther, 312:841-848.
ThellinO., ZorzjW., LakayeB., de Bonnanc B., Coumansa B., Hennenb G., Grisarc T., Igoutb A., and Heinen E., 1999. Housekeeping genesas internal standards: useand limits[J]. Journal of Biotechnology,75:291-295.
Tompkins LM, Wallace AD.2007. Mechanismsof cytochrome P450 induction[J]. J Biochem MolToxicol, 21(4):176-181.
Tseng H P, Hseu T H, Buhler D R, et al. 2005. Constitutive and xenobiotics-induced expression of a novel CYP3A gene from zebrafish larva. Toxicology and Applied Pharmacology, 205:247-258.
Tukey RH, Johnson EF. 1990. Molecular aspecs of regulation and structure of drug-metabolizing enzymes. In Principles of Drug Action Pratt, WB and Taylor P, (ends.), New York: Churchill-Livingstone, 423-467.
Vaccaro E, Giorgi M, Longo V, et al.2003. Inhibition of cytochrome P450 enzymes by enrofloxacin in the sea bass (Dicentrarchus labrax)[J]. Aquatic Toxicology, 62:27-33.
Vaccaro E, Meucci V, Intorre L, et al. 2005. Effects of 17β-estradiol, 4-nonylphenol and PCB 126 on the estrogenic activity and phase 1 and 2 biotransformation enzymes in male sea bass (Dicentrarchus labrax) [J]. Aquatic Toxicology, 75:293-305.
Vaccaro, E., Giorgi, M., Longo, V, et al. 2003. Inhibition of cytochrome P450 enzymes by enrofloxacin in the sea bass (Dicentrarchus labrax) [J]. Aquat. Toxicol, 62: 27–33.
Tang R.Y., Dodd A., Lai D., Mcnabb W.C., and Love D.R.,2007. Validation of Zebrafish (Danio rerio) reference genes for quantitative real-time RT-PCR normalization[J]. Acta Biochimicaet Biophysica Sinica,39(5):384-390.
Wang JS,Xu ZH,Zhou HH. 1996. Cytochrome P450 3A4 and drug metabolism[J]. Chin J Clin Pharmarcol. 12(4):231-235.
Wang W.C., Gu W.T., TangXl.F., GengM.M., Fan M., LiT.J.,Watkins RE, Wisely GB, Moore LB, et al. 2001. The human nuclear xenobiotic receptor PXR: structural determinants of directed promiscuity [J]. Science, 292: 2329 -2333.
XIE W, EVANS RM.2002. Pharmaceutical use of mouse models humanized for the xenobiotic receptor[ J ]. Drug Discov Today, 7(9):509-515.
XIE W, EVANSRM. 2001. Orphan nudear receptor: the exotics of xenobiotics[J]. J Biol Chem. 276(41):37739-37742.
XieW, Barwick J L, DownesM. 2000. Humanized xenobiotic response in mice expressing nuclear receptor SXR[J]. Nature, 406:435 - 8.
Xu D X, Chen Y H, Wang J P, et al.2005. Perinatal lipopolysaccha ride exposure downregulates pregnane X receptor and Cyp3a11 expression in fetalmouse liver[J]. Toxicol Sci,87:38-45.
Xu DX, Wei W, Sun MF, et al.2004. Kupffer cells and ROS partially mediate LPS induced down-regulation of nuclear receptor Pregnane X receptor and its target gene CYP3A in mouse liver[J ]. Free Radic Bio Med, 37:10 - 22.
XU DX,CHEN YH, WANG JP, et al.2005. Perinatal lipopolysaccharide exposure downregulates pregnane X receptor and Cyp3a11 expression in fetal mouse liver[J]. Toxicol Sci, 87 (1):38 - 45.
Xue, L., Zgoda, V.G., Arison, B. et al. 2003. Structure–function relationships of rat liver CYP3A9 to its human liver orthologs: site-directed active site mutagenesis to a progesterone dihydroxylase [J]. Arch. Biochem. Biophys, 2003, 409: 113– 126.
YanH.Z., and Liou R. F. 2006. Selection of intemal control genes for real-time quantitative RT-PCR assays in the oomycete plant pathogen Phytophthora parasitica, Fungal Genetics and Biology, 43(6):430-438.
Zhang J, Kuehl P, Green E D, et al.2001. The human pregnane X receptor:genomic structure and identification and functional characterization of natural allelic variants [J]. Pharmacogenetics, 11: 555-72.
Zhang LL, Zhang JR, Guo K, et al.2010. Effects of fluoroquinolones on CYP4501A and 3A in male broilers [J]. Research in veterinary Science.
Zhou H T, Lu J Z, Xia X R et al.1994. Pharmacokinetics of ofloxacin through three administration routes[J]. Acts Pharmacologics Sinics,15(5): 411-413.
Zhou Ruixue, Meng Tao, Zhao Falan., Zhang jian she. 2009. Comparative research on the stability of internal control genes based on mRNA expression analysis of MYH gene in the grass carp, Ctenopharyngodon idellus[J].Genomics and Applied Biology. 28:896-900.
安飞云,廖春华,边寰峰. 2006.改良MTT法检测HePG-2细胞增殖的研究[J]..毒理学杂志, 20(3):186-188.
操继跃,葛建,方之平,潘黔生,彭玉芬. 2005.氧氟沙星在鲤体内的药物动力学和残留研究[J].水生生物学报. 29(2):227-229.
陈大健,王加才,张萍等.2007.氯唑沙宗评价氟苯尼考对鲫鱼CYP2E1活性影响[J].扬州大
学学报(农业与生命科学版),28(2): 9-16.
陈大健,杨海峰,覃少华等. 2007.乙醇对鲫鱼CYP2E1活性的影响.西北农林科技大学学报(自然科学版), 35(8): 82-86.
陈国胜,张甬元,徐盈等.1997.鲫鱼肝脏中CYP1A1的诱导作为沉积物中二恶英的毒理学指标的研究[J].中国环境科学, 17(3): 264-167.
程宝鸾.动物细胞培养技术[M].广州:华南理工大学出版社,:74–112.
董武,魏强,寺冈宏树等.2002.二恶英诱发斑马鱼初期胚的循环系统障碍.中国实验动物学报, 10(2): 69-72.
冯仁丰.2002.分析灵敏度(检测限).上海医学检验杂志, 17(3):133-136.
顾海峰.2002.海水养殖污染区鱼肝EROD酶活力分析[J]..台湾海峡, 21(3): 292-295.
韩晓非,刘淑敏,李滨兰.1999.氟喹诺酮类药物研究概况[J].黑龙江医药,12(1):28-29.
洪锡钧. 1995.南方鲶胚胎细胞培养及其细胞周期特性检测[J].水产学报, 21(3):240-245.
胡云珍,姚彤炜.2003.细胞色素P4501A的研究进展[J].中国药学杂志. 38(4)246-248.
黄俊勇,冷欣夫. 1991.细胞色素P450酶系的研究进展.
霍传林,王菊英,韩庚辰等. 2002.鱼体内EROD活性对多氯联苯类的指示作用[J].海洋环境科学, 21(1): 5-8.
姜泽东,周伯文,段晶晶. 2008.鱼类细胞培养的研究与应用.北京水产, 3(112):42-47.
冷欣夫,邱星辉. 2001.细胞色素P450酶系的结构、功能与应用前景.北京:科学出版社[M], 1-28.
冷欣夫,邱星辉.2001.细胞色素P450酶系的结构、功能与应用前景.北京:科学出版社[M], 11-13.
黎雯,徐盈,吴文忠.2000.鱼肝EROD酶活力诱导作为二恶英的水生态毒理学指标[J].水生生物学报, 24(3): 201-206.
李聃.2008.鱼类CYP3A活性体外诱导细胞模型的研究[学位论文].上海海洋大学.
林茂,杨先乐,王翔凌等.2007.草鱼肝细胞中恩诺沙星脱乙基代谢的酶动力学[J].高技术通讯, 17(3): 314-318.
林茂.鱼类药物代谢酶体外诱导细胞模型的建立[学位论文] .上海水产大学,2006.
农业部《新编渔药手会册》编撰委员.2005.新编渔药手册.北京:中国农业出版社:169-218.
钱华金,张四明.1991.鱼类细胞培养在渔业科学上的应用[J].淡水渔业,(5):40-42.
沈慧,王万银,秦海宏等.2004. MTT比色法绘制不同锌浓度下细胞生长曲线[J].广东微量元素科学,1(11): 16-18.
舒焱,周宏灏.细胞色素P450药物氧化代谢酶的遗传药理学进展.北京:科学出版社, 2000: 19-36
司徒镇强,吴军正. 2000.细胞培养.西安:世界图书出版公司:58-187.
孙春华,李可欣,谭玲.氧氟沙星注射液的人体药物动力学[J].中国抗生素杂志,1995,20(1):59-62.
唐振华.1990a.昆虫中的细胞色素P450及其特征.昆虫知识,27(1):52-55.
唐震华,胡刚.1993.细胞色素P450基因的命名及其基因表达调控[J].昆虫知识, 30(5):311-314.
童裳亮. 1999.海洋动物的细胞培养与应用[J].生物工程进展, 6:46-48.
王海黎,陶澎. 1999.生物标志物在水环境研究中的应用.中国环境科学[J]. 19(5): 421- 426.
王翔凌.体内外诱导、抑制草鱼(Ctenopharyngodon idellus)CYP2E1的研究[学位论文].上海海洋大学.2008.
王宇飞,邬红娟,文琛等. 2009.改良MTT法在鱼类细胞毒性实验中的使用条件[J].环境科学与技术, 32(2):36-38.
王宇光,王升启,高月. 2006. PXR受体调控的CYP3A诱导及其在药物代谢中的重要意义[J].药学学报, 41(1):1-6.
徐镜波,王咏,张蕾等. 2003. 9种硝基苯对鱼肝微粒体EROD活性的影晌[J].环境科学研究,16(1): 43-45.
杨先乐,林茂,喻文娟等. 2007. MTT比色法在药物对鱼类细胞的毒性检测中的应用.上海海洋大学学报, 16(2):157-161.
杨先乐. 1999.鱼类组织培养的回顾与展望.水产学报, 23:74–81.
杨智源,郑忠辉,黄耀坚等. 1999.海洋放线菌细胞毒抗肿瘤活性物质的初筛.中国海洋药物, 2:52~55.
于淼,管华诗,郭华荣等. 2002.鱼类细胞培养及其应用.海洋科学[J], 27(3): 4-8.
余帮宁,周宏灏.2005.孕烷受体的研究进展[J].中国临床药理学杂志, 21: 61-4.
曾令兵,李晓莉,张林等. 2009.斑点叉尾鮰肾脏组织细胞系的建立及其生物学特性[J].中国水产科学, 16(1):75-81.
张慧锋,王月鹏,李妍.2005.细胞色素P450的研究进展[J].吉林医药学院学报. 3(26):174-176.
张建清,刘晓波. 2002.噻唑蓝比色法检测细菌生长和存活状况的研究[J].卫生研究,31(5): 361-363.
张铭,陈荪红等. 2001.淡水白鲳细胞系及其生长温度特性的研究.生物工程学报, 17(1):105-108.
张宁. 2007.草鱼(Ctenopharyngodon idellus)CYP2E1诱导体外模型研究.[学位论文].上海.上海水产大学.
张义兵. 2002.白鳍豚组织培养的初步研究[J].水生生物学报, 24(2):193-195.
张柞新,张推斌,刘艳辉,吴永魁,李乾学.2004.氧氟沙星(ofloxacin )在鲤鱼体内的药代动力学. 24(4):383-385.
郑春静,张铭. 2003.鱼类细胞体外培养及其应用[J].广西水产科技.01:18-25.
朱琳,钱芸,刘广良. 2001.细胞色素P450酶系及其在毒理学上的应用[J].上海环境科学, 20(2):88-91.
朱玉贤&李毅.2002.现代分子生物学[M].高等教育出版社,北京.
Andersen V, Pedersen N, Larsen NE, Sonne J, Larsen S.2002. Intestinal first pass metabolism of midazolam in liver cirrhosis-effect of grapefruit juice [J]. Br J Clin Pharmacol. 54:120–4.
Anzenbacher P, Anzenbacherova E. 2001. Cytochromes P450 and metabolism of xenobiotics [J]. Cellular and Molecular Life Sciences, 58: 737-747
Araujo CSA, Marques SAF, Carrondo MJT. 2000. In vitro response of the brown bullhead catfish (BB) and rainbow trout (RTG-2) cell lines to benzopyrene [J]. The Science of the Total Environment, 247: 127-135.
Arnold G E, Ornstein R L. 1997. Molecular dynamics study of time-correlated protein domain motions and molecular flexibility: cytochrome P450BM-3 [J]. Biophys J, 73 (3): 1147-1159.
Arnold, K, Bordoli, L, Kopp, J, Schwede, T. 2006. The SWISS-MODEL workspace: a web-based environment for protein structure homology modelling [J]. Bioinformatics, 22(2): 195-201.
Babín M, Casado S, Chana A, et al.2005. Cytochrome P4501A induction caused by the imidazole derivative Prochloraz in a rainbow trout cell line [J]. Toxicol In Vitro, 19(7): 899-902.
BarralloA., Gonzalez-SarmentoI R., Garcia-Isidoro M., Cidad P., Porteros A., and Rodriguez R.E., 1999, Differential brain expression of a new beta-actin gene from zebrafish (Danio rerio) [J]. Eur. J. Neurosci., 11(1):369-372.
BauerB, HartzAM S, Fricker G, MillerD S .2004. Pregnane X receptor upregulation of P-glycop rotein expression and transport function at the blood-brain barrier[J]. Mol Pharmacol, 66: 413- 9
BauerB, HartzAM S, Fricker G, MillerD S. 2004. Pregnane X receptor upregulation of P-glycoprotein expression and transport function at the blood-brain barrier[J]. Mol.Pharmacol, 66: 413- 9.
Billiard S M, Bols N C, Hodson P V. 20040. In vitro and in vivo comparisons of fish-specific CYP1A induction relative potency factors for selected polycyclic aromatic hydrocarbons[J]. Ecotoxicology and Environmental Safety, 59: 292-299.
Bork R W, Muto T, Beaune P H, et al. 1989. Characterization of mRNA species related to human liver cytochrome P450 nifedipine oxidase and the regulation of catalytic activity[J]. The Journal of Biological Chemistry, 264: 910-919.
Brandon EFA, Raap CD, Meijerman I, et al. 2003. An update on in vitro test methods in human hepatic drug biotransformation research: pros and cons [J]. Toxicol Appl Pharmacol, 189: 233-246.
Bresolin T, Rebelo M D F, Bainy A C D. 2005. Expression of PXR, CYP3A and MDR1 genes inliver of zebrafish[J]. Comparative Biochemistry and Physiology C, 140: 403-407.
Britt Wassmur, Johanna Gr?ns, Peter Kling, Malin C. Celander. 2010. Interactions of pharmaceuticals and other xenobiotics on hepatic pregnane X receptor and cytochrome P450 3A signaling pathway in rainbow trout(Oncorhynchus mykiss) [J]. Aquatic Toxicology. 100:91–100.
Buhler D R, Miranda C L, Henderson M C, et al. 2000. Effects of 17β-estradiol and testosterone on hepatic mRNA/protein levels and catalytic activities of CYP2M1, CYP2K1, and CYP3A27 in rainbow trout (Oncorhynchus mykiss) [J]. Toxicology and Applied Pharmacology, 168: 91-101.
Bustin S.A., 2000. Absolute quantification of rnRNAusing rea1-time reverse transcription polymerase chain reaction assays[J]. J.Mol.Endocrinol. 25(2):169-193.
Caminada D, Escher C, Fent K. 2006. Cytotoxicity of pharmaceuticals found in aquatic systems: Comparison of PLHC-1 and RTG-2 fish cell lines[J]. Aquatic Toxicology. (79):114–123.
Cawley G F, Batie C J, Backs W L. 1995. Structure and Function, and Enzymol Relot Areas[J]. Mol, Biol, 60:35-87.
Chen Q, Lu H, Xu X Y et al.1992. Pharmacokinetic and relative bioavailability of ofloxacin after a single oral administration in Chinese healthy volunteers[J]. Chin J Clin Pharmacol, 8:193-197.
Chen Y H, Wang J P, Wang H, et al. 2005. Lipopolysaccharide treatment downregulates the expression of the pregnane X receptor, cyp3a11 and mdr1a genes in mouse placenta [ J ]. Toxicology, 211: 242– 52.
Chirulli V, Longo V, Marini S, et al. 2005. CAR and PXR expressionand inducibility of CYP2B and CYP3A activities in rat and rabbitlungs[J]. Life Sci, 76:2535-46.
ChuW.Y., ShiC.Y.,HuangR, LZhangH, Fand Yin Y.L.2009.Molecular cloning,tissue distribution and ontogenetie expression of the amino acid transporter ba+cDNA in the small intestine of Tibetan suckling piglets,154(1): 157-164.
Coon M J, Vaz A D , Bestervelt L L.1996. Cytochrome P450 2: Peroxidative reactions of diversozymes[J]. FASEB J,10:428-434.
Coon MJ, Ding X, Pernecky SJ et al. 1992.Cytochrome P450: Progress and predictions, FASEB J.6:669-673.
Courtenay, S., Grunwald, C., Kreamer, G, et al.1993. Induction and clearance of cytochrome P4501A mRNA in Atlantic tomcod caged in bleached kraft mill effluent in the Miramichi River [J]. Aquatic Toxicology, 27: 225–244.
Cravedi J P, Perdu-Durand E, Paris A. 1998. Cytochrome P450-dependent metabolic pathways and glucuronidation in trout liver slices[J]. Comparative Biochemistry and Physiology C, 121: 267-275.
Eichelbaum M, Burk O.2001. CYP3A genetics in drug metabolism[J].NatureMed, 2001,7 (3):285- 287.
Ekins,S., Reschly, E.J.,Hagey, L.R., Krasowski,M.D., 2008. Evolution of pharmacologic specificity in the pregnane X receptor[J]. BMC Evol. Biol. 8:103.
Elcombe CR, Lech JJ. 1979. Induction and characterization of hemoprotein(s) P450 and monoxyge-nation in rainbow trout (Salmo Gairdneri). Toxicol Appl Pharmacol, 49(3): 437-450.
Estabrook R W. 1996. The remarkable P450s: A historical overview of the versatile heme protein catalysts[J]. FASEB J,10:202-204.
Fent K. 2001. Fish cell lines as versatile tools in ecotoxicology: Asssessment of cytotoxicity, cytochrome P450 1A induction potential and estrogenic activity of chemicals and environmental samples [J]. Toxicol In Vitro, 15: 477-488.
F(o|¨)rlin L, Celander M. 1995. Studies of the inducibility of P450 1A in perch from the PCB-contaminated lake J?rnsj?n in Sweden. Marine Environmental Research, 39: 85-88.
Gasteiger E., Hoogland C., Gattiker A., Duvaud S., Wilkins M.R., Appel R.D., A.;, B, 2005. Protein Identification and Analysis Tools on the ExPASy Server. In: Walker, J M (Ed.), The Proteomics Protocols Handbook. Humana Press.
Gibson GG, Skett P.1994. Introduction to Drug Metabolism, (2nd edition), Chapman & Hall, Londin.
Giorgi, M., Marini, S., Longo,V., et al. 2000. Cytochrome P450-dependent monooxygenase activities and their inducibility by classic P450 inducers in the liver, kidney, and nasal mucosa of male adult ring-necked pheasants [J]. Toxicology and Applied Pharmacology, 167: 237–245.
Gotoh, O., 1992. Substrate recognition sites in cytochrome P450 family 2 (CYP2) proteins inferred from comparative analyses of amino acid and coding nucleotide sequences[J]. J. Biol. Chem. 267:83– 90.
Graham S E, Peterson J A.1999. How similar are P450s and what can their differences teach us. Arch. Biochem. Biophysics. 369:24-29.
Graham-Lorence S E, Amarneh B, White R E et al. 1995. A three-dimensional model of aromatase cytochrome P 450. Protein Sci. 4 (6):1065-1080.
Graham-Lorence S, Peterson J A.1996. Cytochrome P450 1: structural similarities and functional differences. FASEB J,10:206-214.
Gravato C, Teles M, Oliveira M, et al. 2006. Oxidative stress, liver biotransformation and genotoxic effects induced by copper in Anguilla anguilla L. the influence of pre-exposure to β-naphthoflavone. Chemosphere, 65: 1821-1830.
Guengerich F P, Turvy C G.1991. Comparison of levels of several human microsomal cytochrome P450 enzymes and epoxide hydroxylase in normal and disease states using immunochemialanalysis of surgical liver samples[J]. J. Pharmacol.Exp. Ther., 256:1189-1194.
Guex, N, Peitsch, MC. 1997b. SWISS-MODEL and the Swiss-PdbViewer: An environment for comparative protein modeling [J]. Electrophoresis 18: 2714-2723.
Gui hong Fu, Xian le Yang, Hai Xin Zhang, Wen Juan Yu, Kun Hu. 2011. Effects of cytochrome P450 1A substrate (Difloxacin) on enzyme gene expression and pharmacokinetics in crucian carp (hybridized Prussian carp). Environmental Toxicology and Pharmacology. 31(1):307-313.
Handschin C, Podvinec M, Meyer U A.2000. CXR, a chicken xenobiotic-sensing orphan nuclear receptor, is related to both mammalian pregnane X receptor (PXR) and constitutive androstane receptor (CAR) [J]. ProcNatlAcad SciUSA, 97:10769 - 74.
Hasselberg L, Meier S, Svardal A, et al.2004. Effects of alkylphenols on CYP1A and CYP3A expression in first spawning Atlantic cod (Gadus morhua)[J]. Aquatic Toxicology, 67: 303-313
Hegelund T, Celander M C. 2003. Hepatic versus extrahepatic expression of CYP3A30 and CYP3A56 in adult killifish (Fundulus heteroclitus). Aquatic Toxicology, 64: 277-291.
Henrik, N, Jacob, E, S?ren, B, Gunnar, vH. 1997. Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites.
Hines RN, Piechocki MP, Boncher PD. 1994. Molecular mechanisms controlling CYP1A gene expression[J]. Frontiers in Biotransformation.9:85-110.
Hodgson E.1958. Microsomal monooxygenase. In:Kurkut G A, GilbertL I(eds), Comprehesive insect physiology, biochemistry and pharmacology 11[J], Oxford: Pergamon, 225-321.
Hoover K L(ed). 1984. Use of small fish species in carcinogenicity testing. National Cancer Institute Monograph.65.
Hua-Pin Tsenga, Tzong-Hsiung Hseua, Donald R. Buhler, Wen-Der Wangc, Chin-Hwa Huc. 2005. Constitutive and xenobiotics-induced expression of a novel CYP3A gene from zebrafish larva. Toxicology and Applied Pharmacology. 205:247- 258.
Hui Wang, Zhang-Xiu Liao, Man Chen, Xiao-Ling Hu. 2006. Effects of hepatic fibrosis on ofloxacin pharmacokinetics in rats. Pharmacological Research. 53: 28–34.
James M O, Lou Z, Rowland-Faux L, et al.2005. Properties and regional expression of a CYP3A-like protein in channel catfish intestine. Aquatic Toxicology.72: 361-371.
Jannick, DB, Henrik, N, Gunnar, vH, S?ren, B. 2004. Improved prediction of signal peptides SignalP 3.0 [J]. J. Mol. Biol., 304: 783-795.
Jansson I. 1993. Post-transcriptional modification of cytochrome P450. In Cytochrome P450 Schenkam, JB and Griem H, (ends). Berlin: Springer-Verlag, 361-380.
Jian Yang, Chunshu Hao, Dongfang Yang, et al. 2010. Pregnane X receptor is required for interleukin-6-mediated down-regulation of cytochrome P450 3A4 in human hepatocytes [J].Toxicology Letters. 197: 219–226.
Jones S A, Moore L B, Shenk J L, et al . 2000. The p regnane X receptor : a p romiscuous xenobi otic recep t or that has diverged during evolution[ J ]. Mol Endocrinol, 14: 27 - 39.
Kashiwada S, Hinton D E, Kullman S W. 2005. Functional characterization of medaka CYP3A38 and CYP3A40: kinetics and catalysis by expression in a recombinant baculovirus system. Comparative Biochemistry and Physiology C, 141: 338-348.
Kashiwada S, Hinton D E, Kullman S W. 2005. Functional characterization of medaka CYP3A38 and CYP3A40: kinetics and catalysis by expression in a recombinant baculovirus system. Comparative Biochemistry and Physiology C, 141: 338-348.
Kashiwada, S., Kameshiro, M., Tatsuta, et al.2007. Estrogenic modulation of CYP3A38, CYP3A40, and CYP19 in mature male medaka (Oryzias latipes) [J]. Comp. Biochem. Physiol.C,145:370–378.
Kiefer, F, Arnold, K, Künzli, M, Bordoli, L, Schwede, T. 2009. The SWISS-MODEL Repository and associated resources[J]. Nucleic Acids Research, 37(suppl 1): D387-D392.
Kliewer S A, Moore J T, Wade L, et al . 1998. An or phan nuclear re2cep t or activated by p regnanes defines a novel steroid signalingpathway[ J ]. Cell, 92: 73 - 82.
Krasowski, M.D., Yasuda, K., Hagey, L.R., Schuetz, E.G., 2005. Evolutionary selection across the nuclear hormone receptor superfamily with a focus on the NR1I subfamily (vitamin D, pregnane X, and constitutive androstane receptors)[J]. Nucl.Recept. 3, 2.
Kullman S W, Hinton D E. 2001. Identification, characterization, and ontogeny of a second cytochrome P450 3A gene from the fresh water teleost medaka (Oryzias latipes) [J]. Molecular Reproduction and Development, 58: 149-158.
Kullman, S.W., Hamm, J.T., Hinton, D.E, et al. 2000. Identification and characterization of a cDNA encoding cytochrome P450 3A from the fresh water teleost medaka (Oryzias latipes) [J]. Arch. Biochem. Biophys, 380: 29– 38.
Lee S J, Buhler D R.2003. Cloning, tissue distribution, and functional studies of a new cytochrome P450 3A subfamily member, CYP3A45, from rainbow trout (Oncorhynchus mykiss) intestinalceca. Archives of Biochemistry and Biophysics, 412: 77-89.
Lee S J, Wang-Buhler J L, Cok I, et al. 1998. Cloning, sequencing, and tissue expression of CYP3A27, a new member of the CYP3A subfamily from embryonic and adult rainbow trout livers[J]. Archives of Biochemistry and Biophysics, 1998, 360(1): 53-61.
Lee Y M, Seo J S, Kim I C, et al.2006. Endocrine disrupting chemicals (bisphenol A, 4-nonylphenol, 4-tert-octylphenol) modulate expression of two distinct cytochrome P450 aromatase genes differently in gender types of the hermaphroditic fish Rivulus marmoratus[J].Biochemical and Biophysical Research Communications, 345: 894-903.
Lehmann J M, McKee D D, Watson M A, et al. 1998. The human orphan nuclear receptor PXR is activated by compounds that regulateCYP3A4 gene expression and cause drug interactions[J]. J ClinInvest, 102: 1016 - 23.
Lewis DFV. 1996 Cytochromes P 450 Structure, Function and Mechanism. Taylor& Francis: London,79-88.
Li A P, Kaminski D L, 1995. Rasmussen A. Substrates of human hepatic cytochrome P450 3A4. Toxicology, 104: l-8.
Li D, Yang XL, Zhang SJ, et al.2008. Effects of mammalian CYP3A inducers on CYP3A-related enzyme activities in the freshwater teleost grass carp (Ctenopharyngodon idellus): possible implications for the establishment of a fish CYP3A induction model[J]. Comparative Biochemistry and Physiology, part C. 147,17-2.
Li H, Poulos T L.1996. Conformational dynamics in cytochrome P450-substrate interaction. Biochimie. 78(8-9):695-699.
Livak, KJ, Schmittgen, TD. 2001. Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2-△△CT Method [J]. Methods, 25(4): 402-408.
Lown K, Kolars J, Turgeon K et al. 1992. The erythromycin breath test selectively measures P450ⅢA in patients with severe liver disease. Clin. Pharmacol. Ther.,51:229-238.
Luo G, CUNNINGHAMM, KIM S, et al. 2002. CYP3A4 induction by drugs: correlation between apregnaneX receptor reportergene assay and CYP3A4 expressing in human hepatocytes[ J ]. Drug MetabDispos, 30(7):795-804.
LUO G, GUENTHNER T, GAN LS, et al. 2004. CYP3A4 induction by xenobiotics: biochemistry, experimental methods and impact on drug discovery and development[J]. CurrDrug Metab, 5 (6):483 - 505.
Ma X, Shah Y, CheungC, et al. 2007. The pregnane X receptor gene humanized mouse: a model for investigating drug-drug interactionsmediated by cytochromes P450 3A[J]. Drug Metab Dispos, 35(2):194 - 200.
Maenesh M D, Anders M W. 1973. Characterization of the heme of cytochrome P450 using gas chromatography-masspectrometry. Arch Biochem Biophys. 159(1):201-205.
Maglich J M, Caravella J A, Lambert M H, et al.2003. The first completed genome sequence from a teleost fish (Fugu rubripes) adds significant diversity to the nuclear receptor superfamily. Nucleic Acids Research, 31: 4051-4058.
Marangos M N, Zhu Z, Nicolau D P. 1997.Disposition of ofloxacin in femal New Zealand White rabbits [J]. J. Vet. Pharmacol. Therap.20:17-20.
Martel, P.H., Kovacs, T.G., O’Connor, B.I., et al. 1994. A survey of pulp and paper efffluents for their potential to induce mixed function oxidase enzyme activity in fish [J]. Water Research, 28: 1835–1844.
Masuyama H, Hiramatsu Y, Mizutani Y, et al.2001. The expression of pregnane X receptor and its target gene, cytochrome P450 3A1, in perinatalmouse[J]. Mol Cell Endocrinol, 172:47-56.
McArthur, A.G., Hegelund, T., Cox, R.L.2003.Phylogenetic analysis of the cytochrome P4503 (CYP3) gene family[J]. J. Mol. Evol, 57: 200-211.
Meldgaard M., Fenger C., LambertsenK. L., Pedersen M.D., Ladeby R., Finsen B., 2006, Validation of two reference genes for mRNA level studies of routine disease models in neurobiology journal of neuroscience methods[J], Journal of Neuroscience Methods,156(1-2):10l-110.
Meucci V, Arukwe A. 2006. The xenoestrogen 4-nonylphenol modulates hepatic gene expression of pregnane X receptor, aryl hydrocarbon receptor, CYP3A and CYP1A1 in juvenile Atlantic salmon (Salmo salar)[J]. Comparative Biochemistry and Physiology C,142:142-150.
Miki Y, Suzuki T, Tazawa C, et al. 2005. Ster oid and xenobi otic receptor ( SXR) , cytochrome P450 3A4 andmultidrug resistance gene 1in human adult and fetal tissues[J]. Mol Cell Endocrinol, 231:75-85.
Miki Y, Suzuki T, Tazawa C, et al. 2005. Steroid and xenobiotic receptor ( SXR) , cytochrome P450 3A4 andmultidrug resistance gene in human adult and fetal tissues[ J ]. Mol Cell Endocrinol,231: 75 - 85.
Miranda C L, Henderson M C, Buhler D R.1998. Evaluation of chemicals as inhibitors of trout cytochrome P450s. Toxicology and Applied Pharmacology, 148: 237-244.
Modi S, Sutcliffe M J, Primrose W U, Lian L Y, Roberts G C.1996. The catalytic mechanism of cytochrome P450 BM3 involves a 6 A movement of the bound substrate on reduction[J]. Nat Struct Biol., 3(5):414-417.
MOORE JT, KLIEWER SA. 2000. Use of the nuclear receptor PXR to predict drug interaction [ J ]. Toxicology, 153(1/3):1-10.
Moore, L.B., Maglich, J.M., McKee, D.D.,Wisely, B.,Willson, T.M., Kliewer, S.A., Lambert,M.H.,Moore, J.T., 2002. Pregnane X receptor (PXR), constitutive androstane receptor (CAR), and benzoate X receptor (BXR) define three pharmacologically distinct classes of nuclear receptors[J]. Mol. Endocrinol. 16: 977-986.
Mosmann T. Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays[J]. J Immunol Methods,1983,65:55-63.
Moutou K A, Burke M D, Houlihan D F. 1998. Hepatic P450 monooxygenase response in rainbowtrout administered aquaculture antibiotics. Fish Physiology and Biochemistry,18(1): 97-106.
Murray BP, Edwards RJ, Davies DS et al.1993a. Conservation of a functionally important surface region between two families of the cytochrome superfamily, Biochemical J. 292:309-312.
Murray BP. 1992. P450 enzymes: Inhibition mechanisms, genetic regulation and effects of liver disease[J], Clinical Pharmacokinetic Concepts.23:132-146.
Natalya Hanlona, Nick Coldhamb, Maurice J. Sauerb and Costas Ioannides.2008. Up-regulation of the CYP1 family in rat and human liver by the aliphatic isothiocyanates erucin and sulforaphane[J]. Toxicology. 252: 92-98.
Nebert D W, Gonzalez F J. 1987. P450 gene:structure, evolution and regulation[J]. Annu Rev. Biochem.369(1):1-10.
Nebert D W, Nelson D R, Coon M J et al. 1991. The P450 superfamily: update on new sequences, gene mapping and recommended nomenclature[J].DNA Cell. Biol. 10:1-14.
Negishi M et al. 1996. Cytochrome P450 4: Structural flexibility and functional versatility of mammalian P450 enzymes. FASEB J,10:683-689.
Nelson D R, Kamataki T, Waxman D J et al. 1993. The P450 superfamily: update on new sequence ,gene mapping, accession numbers, early trivial names of enzymes and nomenclature[J]. DNA Cell Biology., 12(1):1-15.
Nelson, D.R.. 2003. Comparison of P450s from human and fugu: 420 million years of vertebrate P450 evolution[J]. Arch.Biochem.Biophys, 409: 18– 24.
Nobuo Nemoto, and Junko Sakurai,. 1992. Altered regulation of Cyp1a-1 gene expression during cultivation of mouse hepatocytes in primary culture[J]. Biochemical Pharmacology. 44, 51-58.
Nuclear Receptors Nomenclature Committee .1999. A unified nomenclature system for the nuclear receptor superfamily [J]. Cell 97:161–163.
OlsvikP. A., Lie k. k.,Jordal A.E.O., Nilsen T.O., and Hordvik L, 2005. Evaluation of potential reference genes in real-time RT-PCR studies of Atlantic salmon[J]. BMC Mol. Biol., 6:21.
Omata Y, Aibara K, Ueno Y. 1987.Conformation between the substrate-binding site and heme of cytochrome P450 studied by exciation energy transfer, Biochemical et Physic Acta, 912:115-123.
Omura T, Ishimura Y, Fujii-Kuriyama Y (eds.).1993.Cytochrome P450 (2nd edition). Kodansha. VCH Tokyo.
Omura T. 1999. Forty years of cytochrome P450. Biochemical and Biophysical Research Communications. 266(3): 690-698.
Oost R Vander, Satumalay A Lero, K and Vermeulen NPE.2000. Validation of inland water pollution assessment using biomarker responses in caged carp[J].Marine Environmental Research. 50: 431-432.
OransJ, Teotico DG, Redinbo MR.2005. The nuclear xenobiotic receptor pregane X receptor: recent insights and new challenges [J]. Mol Endocrinol, 19(12):2891-2900.
Outman WR, Nightingale CH. 1989. Metabolism and the fluoroquinolones.Am J Med 87(Suppl. 6C):37–42.
PascussiJ M, Drocourt L, Fabre J M, et al.2000. Dexamethasone induces pregnane X receptor and retinoid X receptor-αexpression in human hepatocytes: synergistic increase of CYP3A4 induction by pregnane X receptor activators[J]. Mol Pharmacol, 58:361-72.
Paulsen M D, Ornstein R L. 1995. Ligand partitioning into membranes: It significance in determing Km and Ks values for cytochrome P450 and other membrane bound receptors and enzyme. FEBS letters, 67(2):23-29.
Peng H, Cheng ZN. 2001. Drug interaction related to CYP3A4 [J]. Chin J Clin Pharmarcol, 17(5):379-385.
Piddock L J V. 1989. Mechanisms of resistance to quinolonges anclinical perspectives[J]. J. Antimicrob. Chemother, 23: 475.
Plant N. 2007. The human cytochromeP450 sub-family: transcriptional regulation, inter-individual variation and interaction net works [J].Biochim BiophysActa,1770(3): 478-488.
Porter TD, Coon MJ. 1991. Cytochrome P450: multiplicity of isoforms, substrate and catalytic and regulatory mechanisms[J]. J. Biol. Chem.266:13469-13472.
Pyerin W, Taniguchi H. 1991. Post-transcriptional modification by phosphorylation: Control of cytochrome P450 and associated enzymes. In Molecular Aspects of Monooxygenases and Bioactivation of Toxic Compounds. Arinc, E., Schenkman JB and Hodgson E (ends.). New York: Plenum,199-232.
Quattrochi LC, Guzelian PS.2001. CYP3A regulation: from pharmacology to nuclear receptors [J]. Drug Metab Dispos. 29(5) :615-622.
R.I.弗雷谢尼著.章静波译.动物细胞培养.背景:科学出版社.2004.
Raku Shinkyoa, Toshiyuki Sakakia, Miho Ohtab and Kuniyo Inouye., 2003. Metabolic pathways of dioxin by CYP1A1: species difference between rat and human CYP1A subfamily in the metabolism of dioxins. Archives of Biochemistry and Biophysics. 409, 180-187.
Ravichandran K G, Boddupalli S S, Hasemann C A et al. 1993. Crystal structure of hemoprotein domain of P450 BM-3, a prototype for microsomal P450s. Science., 261:731-736.
Rost, B, Yachdav, G, Liu, J. 2004. The PredictProtein server [J]. Nucleic Acids Research, 32(suppl 2): W321-W326.
Roussel, F., Khan, K.K., Halpert, J.R, et al.2000. The importance of SRS-1 residues in catalytic specificity of human cytochrome P450 3A4 [J]. Arch. Biochem. Biophys, 374: 269-278.
Roussel, F., Khan, K.K., Halpert, J.R., 2000. The importance of SRS-1 residues in catalytic specificity of human cytochrome P450 3A4[J]. Arch. Biochem. Biophys. 374:269– 278.
Sarmiento J., Leal S., Quezada C., 2000. Envrironmental acclimatization of the carp modulates the transription of beta-actin,J.CellBiochem., 80(2):223-228.
Son D S. Ikenoue N. Tagawa Y. 2000. Non-linear pharmacokinetics of ofloxacin after a single intravenous bolus dose in pigs [J]. J. Vet. Pharmacol. Therap, 23:311-315.
Sonoda J, Chong LW, DownesM, et al.2005. Pregnane X receptor prevents hepatorenal toxicity from cholesterol metabolites[J].Proc. Natl. Acad Sci USA. 102(6):2198 - 2203.
SONODA J, ROSENFELD JM, XU L, et al. 2003. A nuclear receptor-mediated xenobiotic response and its implication in drugmetaboism and host protection [J]. Curr Drug Metab. 4 (1):59-72.
SouidiM, Gueguen Y, Linard C, et al. 2005. In vivo effects of chroniccontaminati on with dep leted uraniumon CYP3A and ass ociated nu2clear recep tors PXR and CAR in the rat [J]. Toxicology, 214: 113-22.
SouidiM, Gueguen Y, Linard C, et al. 2005. In vivo effects of chroniccontaminati on with dep leted uranium on CYP3A and ass ociated nuclear receptors PXR and CAR in the rat [J]. Toxicology. 214: 113-22.
Squires EJ, Sueyoshi T, NegishiM.2004. Cytoplasmic localization of pregnane X receptor and ligand2dependent nuclear translocation in mouse liver[J]. J Biol Chem, 279 (47):49307-49314.
SuzukiT., HigginsP.J., and Crawford D.R., 2000. Control selection for RNA quantitation[J]. Bio.Techniques, 29(2):332-337.
Tan F, Wang M, Wang W, et al.2008. Comparative evaluation of the cytotoxicity sensitivity of six fish cell lines to four heavy metals in vitro. Toxicology in Vitro. 22, 164–170.
Tang J, Yang X L, Zheng Z L, et al. 2006. Pharmacokinetics and its active metabolite of enrofloxacin in Chinese mitten-handed crab (Eriocheir sinensis). Aquaculture, 260: 69-76.
Xue, L., Zgoda, V.G., Arison, B. et al. 2003. Structure–function relationships of rat liver CYP3A9 to its human liver orthologs: site-directed active site mutagenesis to a progesterone dihydroxylase [J]. Arch. Biochem. Biophys, 409:113-126.
Teng S, MillerM P.2005. The involvement of the pregnane X receptor in hepatic gene regulation during inflammation in mice [J]. J Pharmacol Exp Ther, 312:841-848.
ThellinO., ZorzjW., LakayeB., de Bonnanc B., Coumansa B., Hennenb G., Grisarc T., Igoutb A., and Heinen E., 1999. Housekeeping genesas internal standards: useand limits[J]. Journal of Biotechnology,75:291-295.
Tompkins LM, Wallace AD.2007. Mechanismsof cytochrome P450 induction[J]. J Biochem MolToxicol, 21(4):176-181.
Tseng H P, Hseu T H, Buhler D R, et al. 2005. Constitutive and xenobiotics-induced expression of a novel CYP3A gene from zebrafish larva. Toxicology and Applied Pharmacology, 205:247-258.
Tukey RH, Johnson EF. 1990. Molecular aspecs of regulation and structure of drug-metabolizing enzymes. In Principles of Drug Action Pratt, WB and Taylor P, (ends.), New York: Churchill-Livingstone, 423-467.
Vaccaro E, Giorgi M, Longo V, et al.2003. Inhibition of cytochrome P450 enzymes by enrofloxacin in the sea bass (Dicentrarchus labrax)[J]. Aquatic Toxicology, 62:27-33.
Vaccaro E, Meucci V, Intorre L, et al. 2005. Effects of 17β-estradiol, 4-nonylphenol and PCB 126 on the estrogenic activity and phase 1 and 2 biotransformation enzymes in male sea bass (Dicentrarchus labrax) [J]. Aquatic Toxicology, 75:293-305.
Vaccaro, E., Giorgi, M., Longo, V, et al. 2003. Inhibition of cytochrome P450 enzymes by enrofloxacin in the sea bass (Dicentrarchus labrax) [J]. Aquat. Toxicol, 62: 27–33.
Tang R.Y., Dodd A., Lai D., Mcnabb W.C., and Love D.R.,2007. Validation of Zebrafish (Danio rerio) reference genes for quantitative real-time RT-PCR normalization[J]. Acta Biochimicaet Biophysica Sinica,39(5):384-390.
Wang JS,Xu ZH,Zhou HH. 1996. Cytochrome P450 3A4 and drug metabolism[J]. Chin J Clin Pharmarcol. 12(4):231-235.
Wang W.C., Gu W.T., TangXl.F., GengM.M., Fan M., LiT.J.,Watkins RE, Wisely GB, Moore LB, et al. 2001. The human nuclear xenobiotic receptor PXR: structural determinants of directed promiscuity [J]. Science, 292: 2329 -2333.
XIE W, EVANS RM.2002. Pharmaceutical use of mouse models humanized for the xenobiotic receptor[ J ]. Drug Discov Today, 7(9):509-515.
XIE W, EVANSRM. 2001. Orphan nudear receptor: the exotics of xenobiotics[J]. J Biol Chem. 276(41):37739-37742.
XieW, Barwick J L, DownesM. 2000. Humanized xenobiotic response in mice expressing nuclear receptor SXR[J]. Nature, 406:435 - 8.
Xu D X, Chen Y H, Wang J P, et al.2005. Perinatal lipopolysaccha ride exposure downregulates pregnane X receptor and Cyp3a11 expression in fetalmouse liver[J]. Toxicol Sci,87:38-45.
Xu DX, Wei W, Sun MF, et al.2004. Kupffer cells and ROS partially mediate LPS induced down-regulation of nuclear receptor Pregnane X receptor and its target gene CYP3A in mouse liver[J ]. Free Radic Bio Med, 37:10 - 22.
XU DX,CHEN YH, WANG JP, et al.2005. Perinatal lipopolysaccharide exposure downregulates pregnane X receptor and Cyp3a11 expression in fetal mouse liver[J]. Toxicol Sci, 87 (1):38 - 45.
Xue, L., Zgoda, V.G., Arison, B. et al. 2003. Structure–function relationships of rat liver CYP3A9 to its human liver orthologs: site-directed active site mutagenesis to a progesterone dihydroxylase [J]. Arch. Biochem. Biophys, 2003, 409: 113– 126.
YanH.Z., and Liou R. F. 2006. Selection of intemal control genes for real-time quantitative RT-PCR assays in the oomycete plant pathogen Phytophthora parasitica, Fungal Genetics and Biology, 43(6):430-438.
Zhang J, Kuehl P, Green E D, et al.2001. The human pregnane X receptor:genomic structure and identification and functional characterization of natural allelic variants [J]. Pharmacogenetics, 11: 555-72.
Zhang LL, Zhang JR, Guo K, et al.2010. Effects of fluoroquinolones on CYP4501A and 3A in male broilers [J]. Research in veterinary Science.
Zhou H T, Lu J Z, Xia X R et al.1994. Pharmacokinetics of ofloxacin through three administration routes[J]. Acts Pharmacologics Sinics,15(5): 411-413.
Zhou Ruixue, Meng Tao, Zhao Falan., Zhang jian she. 2009. Comparative research on the stability of internal control genes based on mRNA expression analysis of MYH gene in the grass carp, Ctenopharyngodon idellus[J].Genomics and Applied Biology. 28:896-900.
安飞云,廖春华,边寰峰. 2006.改良MTT法检测HePG-2细胞增殖的研究[J]..毒理学杂志, 20(3):186-188.
操继跃,葛建,方之平,潘黔生,彭玉芬. 2005.氧氟沙星在鲤体内的药物动力学和残留研究[J].水生生物学报. 29(2):227-229.
陈大健,王加才,张萍等.2007.氯唑沙宗评价氟苯尼考对鲫鱼CYP2E1活性影响[J].扬州大
学学报(农业与生命科学版),28(2): 9-16.
陈大健,杨海峰,覃少华等. 2007.乙醇对鲫鱼CYP2E1活性的影响.西北农林科技大学学报(自然科学版), 35(8): 82-86.
陈国胜,张甬元,徐盈等.1997.鲫鱼肝脏中CYP1A1的诱导作为沉积物中二恶英的毒理学指标的研究[J].中国环境科学, 17(3): 264-167.
程宝鸾.动物细胞培养技术[M].广州:华南理工大学出版社,:74–112.
董武,魏强,寺冈宏树等.2002.二恶英诱发斑马鱼初期胚的循环系统障碍.中国实验动物学报, 10(2): 69-72.
冯仁丰.2002.分析灵敏度(检测限).上海医学检验杂志, 17(3):133-136.
顾海峰.2002.海水养殖污染区鱼肝EROD酶活力分析[J]..台湾海峡, 21(3): 292-295.
韩晓非,刘淑敏,李滨兰.1999.氟喹诺酮类药物研究概况[J].黑龙江医药,12(1):28-29.
洪锡钧. 1995.南方鲶胚胎细胞培养及其细胞周期特性检测[J].水产学报, 21(3):240-245.
胡云珍,姚彤炜.2003.细胞色素P4501A的研究进展[J].中国药学杂志. 38(4)246-248.
黄俊勇,冷欣夫. 1991.细胞色素P450酶系的研究进展.
霍传林,王菊英,韩庚辰等. 2002.鱼体内EROD活性对多氯联苯类的指示作用[J].海洋环境科学, 21(1): 5-8.
姜泽东,周伯文,段晶晶. 2008.鱼类细胞培养的研究与应用.北京水产, 3(112):42-47.
冷欣夫,邱星辉. 2001.细胞色素P450酶系的结构、功能与应用前景.北京:科学出版社[M], 1-28.
冷欣夫,邱星辉.2001.细胞色素P450酶系的结构、功能与应用前景.北京:科学出版社[M], 11-13.
黎雯,徐盈,吴文忠.2000.鱼肝EROD酶活力诱导作为二恶英的水生态毒理学指标[J].水生生物学报, 24(3): 201-206.
李聃.2008.鱼类CYP3A活性体外诱导细胞模型的研究[学位论文].上海海洋大学.
林茂,杨先乐,王翔凌等.2007.草鱼肝细胞中恩诺沙星脱乙基代谢的酶动力学[J].高技术通讯, 17(3): 314-318.
林茂.鱼类药物代谢酶体外诱导细胞模型的建立[学位论文] .上海水产大学,2006.
农业部《新编渔药手会册》编撰委员.2005.新编渔药手册.北京:中国农业出版社:169-218.
钱华金,张四明.1991.鱼类细胞培养在渔业科学上的应用[J].淡水渔业,(5):40-42.
沈慧,王万银,秦海宏等.2004. MTT比色法绘制不同锌浓度下细胞生长曲线[J].广东微量元素科学,1(11): 16-18.
舒焱,周宏灏.细胞色素P450药物氧化代谢酶的遗传药理学进展.北京:科学出版社, 2000: 19-36
司徒镇强,吴军正. 2000.细胞培养.西安:世界图书出版公司:58-187.
孙春华,李可欣,谭玲.氧氟沙星注射液的人体药物动力学[J].中国抗生素杂志,1995,20(1):59-62.
唐振华.1990a.昆虫中的细胞色素P450及其特征.昆虫知识,27(1):52-55.
唐震华,胡刚.1993.细胞色素P450基因的命名及其基因表达调控[J].昆虫知识, 30(5):311-314.
童裳亮. 1999.海洋动物的细胞培养与应用[J].生物工程进展, 6:46-48.
王海黎,陶澎. 1999.生物标志物在水环境研究中的应用.中国环境科学[J]. 19(5): 421- 426.
王翔凌.体内外诱导、抑制草鱼(Ctenopharyngodon idellus)CYP2E1的研究[学位论文].上海海洋大学.2008.
王宇飞,邬红娟,文琛等. 2009.改良MTT法在鱼类细胞毒性实验中的使用条件[J].环境科学与技术, 32(2):36-38.
王宇光,王升启,高月. 2006. PXR受体调控的CYP3A诱导及其在药物代谢中的重要意义[J].药学学报, 41(1):1-6.
徐镜波,王咏,张蕾等. 2003. 9种硝基苯对鱼肝微粒体EROD活性的影晌[J].环境科学研究,16(1): 43-45.
杨先乐,林茂,喻文娟等. 2007. MTT比色法在药物对鱼类细胞的毒性检测中的应用.上海海洋大学学报, 16(2):157-161.
杨先乐. 1999.鱼类组织培养的回顾与展望.水产学报, 23:74–81.
杨智源,郑忠辉,黄耀坚等. 1999.海洋放线菌细胞毒抗肿瘤活性物质的初筛.中国海洋药物, 2:52~55.
于淼,管华诗,郭华荣等. 2002.鱼类细胞培养及其应用.海洋科学[J], 27(3): 4-8.
余帮宁,周宏灏.2005.孕烷受体的研究进展[J].中国临床药理学杂志, 21: 61-4.
曾令兵,李晓莉,张林等. 2009.斑点叉尾鮰肾脏组织细胞系的建立及其生物学特性[J].中国水产科学, 16(1):75-81.
张慧锋,王月鹏,李妍.2005.细胞色素P450的研究进展[J].吉林医药学院学报. 3(26):174-176.
张建清,刘晓波. 2002.噻唑蓝比色法检测细菌生长和存活状况的研究[J].卫生研究,31(5): 361-363.
张铭,陈荪红等. 2001.淡水白鲳细胞系及其生长温度特性的研究.生物工程学报, 17(1):105-108.
张宁. 2007.草鱼(Ctenopharyngodon idellus)CYP2E1诱导体外模型研究.[学位论文].上海.上海水产大学.
张义兵. 2002.白鳍豚组织培养的初步研究[J].水生生物学报, 24(2):193-195.
张柞新,张推斌,刘艳辉,吴永魁,李乾学.2004.氧氟沙星(ofloxacin )在鲤鱼体内的药代动力学. 24(4):383-385.
郑春静,张铭. 2003.鱼类细胞体外培养及其应用[J].广西水产科技.01:18-25.
朱琳,钱芸,刘广良. 2001.细胞色素P450酶系及其在毒理学上的应用[J].上海环境科学, 20(2):88-91.
朱玉贤&李毅.2002.现代分子生物学[M].高等教育出版社,北京.