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Protective role of red blood cells PAF-acetylhydrolase against oxidative siess

红细胞PAF-乙酰水解酶对氧化酶的保护作用

基本信息

批准号：
15590069
负责人：
KARASAWA Ken
金额：
$ 2.18万
依托单位：
Teikyo University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

项目摘要

Three kinds of subtypes have been identified as PAF-acetylhydrolase (PAF-AH) which degrades PAF ; mediator of inflammation and allergy. Those consist of plasma-type, distributed in body fluids such as plasma, and intracellular type I and type II. Type I PAF-AH forms oligomer consisting of catalytic subunits termed α_1 or α_2. In the human blood, 70 % of enzyme activity is detected in plasma, while 30 % of enzyme activity is detected in red blood cells. Total enzyme activity of white blood cells such as polymorphonuclear leukocytes and platelets was less than 1 %. Despite high occupancy of enzyme activity, subtype of this enzyme and physiological role remain unknown. In this study, enzyme purification was undertaken from easily available pig blood. As a result, it was clarified that pig red blood cells contained α_1/ α_2 heterodimer and α2/α2 homodimer. Although ai subunit is highly expressed in fetal brain, expression of this subunit is very low in the adult except spermatogonium. By contrast, white blood cells such as polymorphonuclear leukocytes and platelets contained α_2/α_2 homodimer alone. This fact suggests that this subunit plays an important role in regulation of red blood cells function. In addition, Western blotting analysis using specific antibodies to these subunits showed that human red blood cells contained α_1/α_2 heterodimer and α_2/α_2 homodimer, as well as human red blood cells.Since red blood cells do not produce PAF and lack PAF receptor, physiological substrate towards this enzyme is not thought to be PAR Thus, we added recombinant PAF-AH to the lipids extracted from red blood cells membranes and analyzed the lipid constituents by mass-spectrometry. As a result, it was shown that recombinant PAF-AH degraded oxidatively modified lipids of the red blood cells membranes exposed by Cu^<2+> as the substrates.

PAF-乙酰水解酶（PAF-AH）是一种降解炎症和变态反应介质PAF的酶，目前已鉴定出三种亚型。这些包括血浆型，分布在体液中，如血浆，和细胞内的I型和II型。Ⅰ型PAF-AH形成由称为α_1或α_2的催化亚基组成的寡聚体。在人血液中，70%的酶活性在血浆中检测到，而30%的酶活性在红细胞中检测到。多形核白细胞和血小板等白色血细胞的总酶活性小于1%。尽管酶活性的高占有率，这种酶的亚型和生理作用仍然未知。在这项研究中，酶纯化进行了从容易获得的猪血液。结果表明，猪红细胞含有α_1/α_2异二聚体和α_2/α_2同二聚体。虽然ai亚基在胎儿脑中高表达，但在成人中除精原细胞外表达很低。而白色血细胞如多形核白细胞和血小板仅含有α_2/α_2同源二聚体。这一事实表明，该亚基在调节红细胞功能中起着重要作用。另外，用特异性抗体进行的Western印迹分析表明，人红细胞中含有α_1/α_2异二聚体和α_2/α_2同二聚体，而人红细胞中也含有α_1/α_2异二聚体和α_2/α_2同二聚体。我们将重组PAF-AH加入到从红细胞膜提取的脂质中，并通过质谱分析脂质成分。结果表明，重组PAF-AH可降解以Cu ^<2 +>为底物的红细胞膜的氧化修饰脂质。