摘要
本研究以Cu^(2+)(0、100μg·L^(-1))对罗氏沼虾Macrobrachium rosenbergii进行处理,分别在胁迫后的0 h、3 h、6 h、12 h、24 h和48 h收集血淋巴,检测血细胞中的细胞凋亡率、活性氧(ROS)含量、酯酶活性、一氧化氮(NO)含量以及细胞凋亡执行分子caspase-3和免疫因子α-2M基因的表达。结果显示,血细胞凋亡率和ROS含量的变化趋势一致,在胁迫后的6~48 h显著升高(P<0.05)。酯酶活性在胁迫后的3~48 h均显著下降(P<0.05),最低值出现在48 h。在胁迫后的6~48 h,NO含量持续增加,并与对照组的差异有统计学意义(P<0.05)。caspase-3的相对基因表达量在胁迫后的6~48 h均显著升高(P<0.05),并在24 h达到最高值。在胁迫后的12 h,α-2M基因的表达量显著升高(P<0.05)并达到峰值,在24 h有短暂降低,但与对照组相比仍显著增加(P<0.05),在48 hα-2M基因的表达量显著降低(P<0.05)。相关性分析显示,血细胞凋亡率和NO以及ROS都呈显著的正相关(P<0.001)。实验表明,Cu^(2+)胁迫显著抑制了机体的酯酶活性,诱导NO和ROS的产生以及免疫相关基因α-2M的表达来对抗Cu^(2+)胁迫带来的损伤。随着胁迫时间的延长,过高浓度的NO和ROS对机体自身造成了伤害,并诱导凋亡相关基因caspase-3的表达,最终导致细胞凋亡,说明Cu^(2+)对虾类具有免疫抑制性和毒性。NO和ROS的升高可能是细胞凋亡的主要诱因。
The present study analyzed the toxic effects of Cu^2＋ on apoptotic cell ratio, ROS production, esterase activi-ties, NO production, and the expression levels of caspase-3 and α-2M in hemocytes of Macrobrachium rosenbergii. The re-sults showed that the changes of apoptotic cell ratio were consistent with ROS production which was increased significantly （P 〈 0. 05 ） after exposure for 6 -48 h. Esterase activities were significantly decreased （ P 〈 0. 05 ） at 3 -48 h and the min-imum was observed at48 h. The NO production continued increasing and the level became significant difference （P 〈0. 05） after exposure for 6 - 48 h. The relative expression levels of caspase-3 were significantly induced （ P 〈 0. 05 ） after exposure for 6 -48 h,and reached the peak at 24 h. The relative expression of α-2M was significantly increased （P 〈 0. 05 ） and reached the peak at 12 h. Moreover, the expression level of α-2M reduced after exposure for 24 h and became significantly different （P 〈0. 05） at 48 h. Correlation analysis showed that significant positive correlation （P 〈0. 001） between apoptot-ic cell ratio and ROS production was observed, as well as apoptotic cell ratio and NO production. All these results suggested that esterase activities were significantly inhibited after Cu2＋ exposure, while NO and ROS production as well as α-2M ex-pression were induced against Cu^2＋ exposure. Along with stress time increasing, overfull production of ROS and NO caused oxidative stress to prawn and then induced the expression level of caspase-3 which finally resulted in apoptosis, indicating tha t Cu^2＋ s tress may have immunosuppressive and toxic effects to M. rosenbergii, an d the in crea sed product ion of NO and ROS might be the main cause of apoptosis.
出处
《四川动物》
北大核心
2017年第3期293-299,共7页
Sichuan Journal of Zoology
基金
国家自然科学基金项目(31600321)
广东省自然科学基金项目(2015A030310438)
广东海洋大学创新强校工程科研项目(GDOU2016050253)
广东海洋大学优秀青年骨干教师特别资助计划项目(HDYQ2015003)
