| 摘要: |
| 为了了解苯并[a]芘（BaP）对鱼类细胞色素P4501A1（CYP1A1）表达的影响，以褐菖鲉（Sebasticus marmoratus）为实验材料，采用体内实验，研究其在经过不同浓度（0.1、1、10、20、50mg/kg鱼体重量）的BaP诱导后，鱼体肝脏研究CYP1A1基因表达的情况，筛选出后续时间-效应实验中BaP注射的最佳浓度，研究BaP诱导6h、12h、1d、3d、7d后（质量浓度为20mg/kg鱼体重量）鱼体肝脏CYP1A1酶活性、基因表达和蛋白表达的情况。结果表明：剂量-效应实验中，20mg/kg鱼体重量为最佳浓度，此浓度下，基因表达在各组中变化最显著。时间-效应实验中，较空白对照组而言，染毒6h、12h和1d后，EROD酶活性显著增加。3d后开始下降，与对照组相比变化不大，7d后酶活性又发生上调。半定量RT-PCR结果表明，各染毒组与对照组相比，CYP1A1基因表达量都发生了上调，呈现先上升后下降的趋势。其中，6h和12h组相对表达量极显著增加，1d后开始下降且与3d和7d组相比变化不明显。Western blot结果表明，蛋白表达量在染毒12h后表现出显著的诱导效应，随着时间的延长略有回落，但与对照组相比仍有显著性差异。研究表明：BaP对褐菖鲉CYP1A1具有较强的诱导作用。一定质量浓度的BaP注射于褐菖鲉不同的时间后，能诱导褐菖鲉活体EROD酶活性、CYP1A1基因mRNA表达及蛋白表达，并随着时间的延长呈现先诱导后抑制的趋势。这说明BaP作为诱导剂对CYP1A1酶活性和蛋白表达的作用机制可能与调控CYP1A1的转录水平有关。 |
| 关键词: 苯并[a]芘(BaP) 褐菖鲉 CYP1A1 酶活性 基因表达 蛋白表达 |
| DOI：10.11693/hyhz20180300055 |
| 分类号:Q789 |
| 基金项目:浙江省科技计划项目，2017C37073号；浙江海洋大学人才引进项目。 |
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| EFFECTS OF BENZO[A]PYRENE ON EROD ACTIVITY, mRNA EXPRESSION, AND PROTEIN EXPRESSION OF CYP1A1 IN THE LIVER OF SEBASTICUS MARMORATUS |
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SUN Wen-Jing, WANG Xiao-Yan, QI Peng-Zhi, ZHANG Jian-She, WU Chang-Wen
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Marine Science College of Zhejiang Ocean University, National Engineering Research Center of Marine Facilities Aquaculture, Zhoushan 316022, China
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| Abstract: |
| We investigated the effects of benzo[a]pyrene (BaP) on the expression of cytochrome P4501A1 (CYP1A1) in fish. Sebasticus marmoratus was used as the experimental material and it was injected intraperitoneally BaP at different concentrations (0.1, 1, 10, 20, and 50mg/kg) on the mRNA expression of CYP1A1 in the fish liver BaP to determine the best concentration used in the follow-up experiment. In addition, the enzyme activity, mRNA expression of CYP1A1, and protein expression in the liver after the BaP induction was read 6h, 12h, 1d, 3d, and 7d after the injection. The results show that the 20mg/kg concentration was the best in the dose-effect experiment, under which the gene expression varied most significantly in all the groups. The enzyme activity in the 6h, 12h, and 1d groups showed a significant increase compared with the 0h group, the control. The enzyme activity of group began to decrease on Day 3 to the similar level of the control and increased obviously again on Day 7. Results of semi-quantitative RT-PCR show that CYP1A1 expression in all experimental groups was up-regulated from that of the control, showing a trend of increase first and then decrease. The relative expression increased significantly in 6 to 12h, declined in 1d, and stabilized in 3d and 7d. The Western blot result shows that the protein expression was boosted significantly in 12 h of the toxin injection and then slightly decreased over time, which was very different from that of the control. Moreover, the injection of BaP in a certain concentration BaP at different times in S. marmoratus could induce ethoxyresorufin-O-deethylase (EROD) activity, CYP1A1 mRNA expression, and protein expression. Therefore, the function of BaP as an inducer of CYP1A1 EROD activity and protein expression may be related to the regulation of CYP1A1 transcription. |
| Key words: benzo[a]pyrene Sebasticus marmoratus CYP1A1 enzyme activity gene expression protein expression |
