水生植物化感物质抑藻作用下蓝藻细胞内一氧化氮（NO）信号分子的产生及其作用机理研究

The explosive growth of planktonic algae is one of the problems caused by eutrophication. Allelochemicals released by aquatic plants can inhibit the growth of algae, which has been widely accepted as a safe and efficient ecological technology to control algae. Studies have shown that nitric oxide (NO) molecule can be produced in algal cells under allelopathic stress. However, it is not clear that whether NO as an important bioactive molecule, has been involved in the process of information transmission, cell growth inhibition, and even cell death of algae cells. In allusion to the above-mentioned problems, the project is based on the phenomenon that the excessive nitric oxide could be produced under the allelopathic stress. Firstly, the concentration of NO produced by algal cells under allelopathic stress was quantified indirectly, and the effects of NO on the growth of algal cells were explored by combining the physiological and biochemical processes of algal cells. Meanwhile, the main pathways and effects of NO production in algae cells were explored. By analyzing the relationship between NO production and reactive oxygen species (ROS) metabolism in cells, the mechanism of NO production in algae cells on allelopathic algae inhibition was determined. The scientific significance of this project is to elucidate the allelopathic mechanism of inhibiting the growth of algae by aquatic plants, and to reveal the mechanism of chemical ecological action between organisms in natural water bodies. In addition, the project also plays an important role in the ecological management of eutrophic lakes and the reconstruction of aquatic vegetation.

浮游藻类的爆发性生长是水体富营养化带来的问题之一，利用水生植物化感作用抑制浮游藻类生长被认为是一种高效安全的生态控藻技术，对水华的生态防治具有重要意义。已有研究表明化感抑藻会诱导藻细胞产生一氧化氮(NO)，而NO作为重要的生物活性小分子，是否参与了化感作用下藻细胞间的信息传递、抑制细胞生长，甚至诱导细胞死亡等过程，目前还未见报道。针对上述问题，本项目以藻细胞所产生的NO为研究对象，首先间接定量化感胁迫下藻细胞产生的NO浓度，结合藻细胞生理生化过程，探索NO对藻细胞生长的影响。同时，通过探讨藻细胞内NO产生的主要途径和作用，并分析细胞内NO与活性氧(ROS)代谢的关系，明确藻细胞内NO对水生植物化感抑藻的影响机制。本项目的科学意义在于阐明水生植物分泌的化感物质对藻类生长的抑制机理，揭示自然水体中生物间的化学生态作用机制。此外，本项目也为富营养化湖泊的生态治理和水生植被重建提供重要的理论依据。

水体富营养化和有害藻华会对水生态系统以及公共卫生安全造成严重的危害。水生植物的化感效应作为一种安全高效的控藻技术，备受学者广泛关注。本项目以铜绿微囊藻作为典型水华藻种，研究在化感物质暴露下，藻细胞内一氧化氮(NO)、活性氧(ROS)等生化指标的变化，阐述化感胁迫下藻细胞内源NO的产生规律、产生途径以及其变化对藻细胞生长调控作用，和NO诱导藻细胞细胞程序性死亡机制。研究结果对于控制蓝藻水华有重要意义。主要结论如下：.（1）明确了典型水华蓝藻铜绿微囊藻在典型化感物质（焦性没食子酸PA、β-紫罗兰酮和N-苯基-1-萘胺）的化感抑制下，胞内NO水平变化随化感物质暴露浓度的升高而出现两个阶段的变化。同时，藻细胞内NO水平变化和化感物质种类不存在相关性。.（2）通过添加抑制剂的方式，发现铜绿微囊藻细胞同时存在硝酸还原酶（NR）途径和一氧化氮合成酶（NOS）途径产生NO。且两种途径产生的NO所起功能存在差异。其中，NOS途径产生的NO起到促进细胞生长作用，NR途径所产生NO在藻细胞处于环境胁迫下起到重要调控作用。.（3）表明NO对铜绿微囊藻调控作用和藻细胞所受胁迫压力相关。低浓度PA（0.4-0.8 mg/L）胁迫下，胞内NO主要调控细胞内氧化还原稳态；在高浓度PA（1.6-16.0 mg/L）胁迫下，藻细胞处于抑制状态，NO起到加速细胞抑制甚至死亡的作用生理特性发生了明显的变化。.（4）研究结果显示NO的作用功能取决于其在细胞内存在的水平，200 μM SNP暴露下，低藻密度铜绿微囊藻胞内NO大量增加，SOD和MDA上调，氧化应激显著，诱导藻细胞PCD发生。中间藻密度组NO在前48 h上升，ROS下调，SOD和MDA均有增加，干扰氧化还原平衡，导致细胞氧化损伤；48 h后NO和ROS平衡，细胞逐渐恢复稳态。而高藻密度铜绿微囊藻细胞内NO水平的变化仅起到调节ROS水平的左右，藻细胞基本处于正常生长状态。.（5）验证同浓度SNP可以在水生态系统中对大/小生物体产生正面/负面作用。同浓度SNP能诱导铜绿微囊藻发生细胞程序性死亡，而对水生植物浮萍起调节生长状态的正面作用。