扇贝脑啡肽能神经系统介导的免疫调节作用

Enkephalin has become one of the most noticeable molecules as a member of neuropeptides in the peptidergic neuron system, which comprised a large portion in the neuro-endocrine-immuno-modulation meshwork. Our project is performed based on techniques from immunology, biochemistry and histology, aiming to (1) clone and characterize enkephalinergic neuron system related genes and enkephalin receptor genes, recombine the proteins, and verify counterpart biological functions, thus to understand the basal molecule characterizations; (2) explore the distributions of enkephalinergic neurons in all kinds of ganglia and tissues, to grasp the structure constitutes of this system in scallops; (3) investigate the alterations of enkephalin concentration and temporal expression of related genes, and distributions of TNF receptor (TNFR) over the peptidergic neurons of scallops in response to pathogen invasions, thus to clarify the immunity activation mechanisms of enkephalinergic neuron system; (4) detect the immune parameter changes and transcriptome level in stimulation of two enkephalins; (5) detect the immune parameters and expressions of related genes, concentrations of Ca2+ and cAMP in haemocytes and NF-κB in nucleus in scallops infected by pathogens, when critic components in enkephalinergic neuron system are inhibited using interference techniques such as RNAi; (6) explore the pathways mediated and regulations involved in the scallop neuro-endocrine-immunity network by enkephalin receptors and enkephalinergic neuron system, to further enrich and develop the neuro-endocrine-immunology of marine invertebrates.

肽能神经系统是神经内分泌免疫调节网络的重要组成部分，脑啡肽作为重要的神经肽成员而备受关注。本课题拟采用分子免疫学、生物化学、组学等技术手段，从扇贝中克隆脑啡肽能神经系统相关基因及脑啡肽受体基因，获得体外重组蛋白，验证其相应生物学功能，检测扇贝脑啡肽能神经元在各神经节及组织中的分布，了解扇贝脑啡肽能神经系统的结构组成及基本分子特征；分析病原感染后扇贝脑啡肽含量变化及相应基因时序表达，及肽能神经元上TNFR的分布，阐明扇贝脑啡肽能神经系统的免疫激活机制；分析两种脑啡肽诱导下扇贝免疫学指标和转录组水平变化，应用RNAi等技术阻断脑啡肽能系统关键成分，检测扇贝在病原感染中免疫学指标及免疫基因表达量，分析血淋巴细胞中Ca2+和cAMP等相应分子的浓度及胞核中活性NF-kB的含量变化，探讨脑啡肽受体及脑啡肽能系统在扇贝神经内分泌免疫中的介导通路及调控作用，丰富和发展海洋无脊椎动物神经内分泌免疫学内容。

肽能神经系统是神经内分泌免疫调节网络的重要组成部分，脑啡肽作为重要的神经肽成员而成为近年来的研究焦点。本课题采用分子免疫学、生物化学、多组学等技术手段，查明了脑啡肽能神经系统的组织结构和分子特征，探索了扇贝/牡蛎脑啡肽能神经系统的免疫调节作用及脑啡肽受体介导的信号转导途径和免疫转录调控机制，明确了软体动物Met-ENK能通过其受体，进一步调节胞内二级信使Ca2+和cAMP的浓度介导下游的细胞免疫和体液免疫应答，证明软体动物已经拥有结构和功能完善的（甲硫氨酸）脑啡肽能神经内分泌免疫系统。利用组学技术系统研究了脑啡肽与ENK与ACh/NE 的协同调控作用，发现牡蛎AP/NF-kB/TNF等细胞因子和转录因子参与了ENK/ACh/NE 的调控过程，提出了软体动物血淋巴细胞可作为免疫细胞，与神经系统组成原始的“神经—血淋巴细胞”类神经内分泌免疫调节轴的创新性理论，丰富和发展了海洋无脊椎动物神经内分泌免疫学内容。按计划完成了研究任务，发表SCI收录论文6篇。