海参自溶初期线粒体相关凋亡信号蛋白分子的激活机理研究

项目名称： 海参自溶初期线粒体相关凋亡信号蛋白分子的激活机理研究

项目编号： No.31301430

项目类型： 青年科学基金项目

立项/批准年度： 2014

项目学科： 农业科学

项目作者： 杨静峰

作者单位： 大连工业大学

项目金额： 24万元

中文摘要： 海参在捕捞、运输及加工过程中常发生强烈的自溶现象，导致"化皮、溶解"， 这一现象困扰着海参的加工业。迄今对海参自溶的研究主要集中于海参自溶酶的酶学研究。海参自溶是由细胞凋亡中的信号蛋白分子激活自溶酶所导致。而在海参自溶酶激活前到底由哪些信号蛋白分子参与凋亡的发生，以及它们如何参与自溶酶的激活等问题均未见有研究。本项目拟从凋亡信号传导的角度研究海参自溶现象。首先采用免疫组化法研究海参细胞内Ca2+的分布变化和线粒体跨膜电位(△Ψm)变化，了解线粒体凋亡的起始；其次采用Western Blotting法对线粒体相关信号分子Bcl-2、细胞色素c、AIF、Apaf-1、Caspase-9相互激活进行研究，探明线粒体参与海参凋亡的分子机制；最后通过对凋亡关键效应分子Caspase-3酶的激活，明确海参凋亡分子信号对自溶酶激活的影响。以期从细胞凋亡的角度，以线粒体凋亡的研究为切入点揭示海参自溶机理。

中文关键词： 刺参；自溶；细胞凋亡；细胞色素c；Bcl-2

英文摘要： Sea cucumber would initiate the autolysis which lead to skin melting and hydralysis during fishing,transport and processing. The sea cucumber processors was puzzled by this phenomenon. Up to now, the studies of autolysis in sea cucumber mainly focus on enzymology field. By means of cell apoptosis, the signal protein molecules triggered the precursor autolysis enzyme, finally the autolytic enzyme result in autolysis. But which signal moleculars involved in the autolysis activitation, and how they worked were still unknown. This project aimed to find out the signal pathway that ignited the sea cucumber apoptosis and finally result in autolysis. Firstly, the immunohistochemical staining method was adopt to discover the Ca2+ distribution and the changing of Mitochondrial transmembrane potential (△Ψm) in initial stage. Secondly, the Western Blotting was adopt to explicit the apoptosis activition relationship of Bcl-2、cytochromeC、AIF、Apaf-1、Caspase-9. Finally, we would detected the activation of Caspase-3 that the key effector molecule of the apoptosis. We hope the detection can help to reveal the mechanism of the sea cucumber autolysis in the angle of mitochondria apoptosis.

英文关键词： Stichopus japonicas；autolysis；apoptosis；Cyt c；Bcl-2