| 亚硝酸盐对凡纳滨对虾血细胞毒性及p53基因表达的影响 |
| Effects of Nitrite Stress on Haemocyte Toxicity and p53 Gene Expression of Litopenaeus vannamei |
| 投稿时间：2014-10-21 修订日期：2014-12-10 |
| DOI：10.15928/j.1674-3075.2015.02.010 |
| 中文关键词:凡纳滨对虾 亚硝酸盐 血细胞 基因表达 氧化胁迫 |
| 英文关键词:Litopenaeus vannamei nitrite haemocyte gene expression oxidative stress |
| 基金项目:广东省科技计划项目(2012A020602050)；广东省自然科学基金项目(S2012040008093)。 |
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| 中文摘要: |
| 随着水产动物集约化养殖的发展，水质污染加剧，环境中亚硝酸盐的含量不断上升，成为水产养殖中诱发爆发性疾病的重要环境因子。为探讨亚硝酸盐对凡纳滨对虾（Litopenaeus vannamei）血细胞的毒性以及p53 mRNA的影响，通过预试验以及前期资料获取亚硝酸盐对凡纳滨对虾的毒性效应，选取0 mg/L对照组和 20 mg/L浓度组，用流式细胞术检测血细胞活性氧（ROS）、一氧化氮（NO）含量和非特异性酯酶活性，荧光定量PCR技术检测p53基因的表达量变化。结果显示，在亚硝酸盐应激12 h后，对虾血细胞的NO含量与对照组相比有显著升高（P<0.05），在应激24、48、72 h后有极显著的升高（P<0.01）；ROS含量在应激12、48、72 h时有极显著升高（P<0.01），在应激24 h时与对照组相比有显著的升高（P<0.05）。非特异性酯酶活性在应激24、48、72 h显著下降（P<0.05），p53的表达水平在应激48 h和72 h后显著升高（P<0.05）。研究结果表明，亚硝酸盐应激诱导对虾血细胞产生了过量的NO和ROS，造成氧化胁迫作用，诱导凋亡相关基因的表达，最终导致细胞凋亡。 |
| 英文摘要: |
| Intensive aquaculture leads to serious water pollution, including high concentrations of nitrite. The latter has become an important environmental variable inducing explosive disease outbreak in aquaculture. This study aimed to explore the effects of nitrite stress on haemocyte toxicity and p53 gene expression in Litopenaeus vannamei. Litopenaeus vannamei with initial weight（4.41 ± 1.80）g were cultured at a salinity of 5 g/Kg, pH 7.9-8.0 and temperature 26 ± 2°C for two weeks with aeration. Moulting Litopenaeus vannamei of similar size and body color were then transferred into culture buckets for stress testing. Based on the results of a preliminary investigation and previous studies, two levels of nitrite concentrations, 0 and 20mg NO2-N\L, were tested. 20 Litopenaeus vannamei were placed in each container with 180 L of water at salinity 5 g/Kg and each treatment was conducted in three replicates. Test solutions in the buckets were replaced every 24 h with 50% replacement. At 0, 4, 8, 12, 24, 48 and 72 h of exposure, three Litopenaeus vannamei in each container were sampled. Reactive oxygen species (ROS) production, nitric oxide (NO) production and esterase activity of Litopenaeus vannamei haemocytes were evaluated using flow cytometry and p53 gene expression level was evaluated by real-time fluorescent quantitative PCR. The results show that NO production in Litopenaeus vannamei haemocytes increased significantly after 12 h of NO2-N exposure (P<0.05) and the differences compared to the control group was highly significant after 24, 48 and 72 h NO2-N exposure (P<0.01). ROS production showed a significant increase at 24 h (P<0.05) compared with the control group, and a more significant increase at 12, 48 and 72 h (P<0.01). Esterase activity declined significantly after 24, 48 and 72 h NO2-N exposure (P<0.01). The expression level of p53 Gene increased significantly after 48 h and 72 h NO2-N exposure (P<0.05). The results of our study reveal that nitrite stress firstly induces ROS and NO production and an increase in p53 Gene expression level of the haemocytes to reduce the damage by nitrite exposure. With extended exposure time, excessive ROS and NO production causes oxidative stress, subsequently inducing the expression of apoptosis-inducing genes leading to apoptosis. In conclusion, nitrite causes immune suppression and toxicity to Litopenaeus vannamei, which increases with exposure time, and increasing ROS and NO production is probably the primary reason for apoptosis. |
| 郭慧,王安利,冼健安.2015.亚硝酸盐对凡纳滨对虾血细胞毒性及p53基因表达的影响[J].水生态学杂志,36(2):61-67. |
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