Qu X W, Wang H, Rozenfeld R A, Huang W, Hsueh W
Department of Pathology, Children's Memorial Hospital, Northwestern University Medical School, Chicago, IL 60614, USA.
Biochim Biophys Acta. 1999 Aug 12;1451(1):211-7. doi: 10.1016/s0167-4889(99)00076-2.
Constitutive nitric oxide synthase (cNOS) may play an important protective role in the intestine, since our previous study has shown that the degree of bowel injury induced by platelet-activating factor (PAF), a potent inflammatory mediator, is inversely related to the cNOS content of the intestine. This study aims to examine the composition of the cNOS system in rat small intestine, and its regulation by PAF. We found that an approximately 120 kDa NOS I (neuronal NOS) is the predominant NOS in rat intestine, as evidenced by the following: (a) immunoblotting with specific antibodies detected a NOS I of approximately 120 kDa, but little NOS III; (b) the Ca(2+)-dependent, constitutive NOS (cNOS) activity of the rat intestine was removed by immunoprecipitation with the anti-NOS I, but not anti-NOS II or anti-NOS III antibodies; (c) RT-PCR revealed constitutive expression of NOS I in the intestinal tissue, but only a minute amount of NOS III. Immunofluorescent staining with anti-NOS I located NOS in the Auerbach plexus and nerve fibers in the muscle layer. We also found that this 120 kDa NOS I is rapidly (within 1 h) down-regulated in response to PAF administration. The protein level, enzyme activity as well as mRNA of nNOS were all decreased in the intestine.
组成型一氧化氮合酶（cNOS）可能在肠道中发挥重要的保护作用，因为我们之前的研究表明，血小板激活因子（PAF，一种强效炎症介质）诱导的肠道损伤程度与肠道的cNOS含量呈负相关。本研究旨在检测大鼠小肠中cNOS系统的组成及其受PAF的调节情况。我们发现，一种约120 kDa的一氧化氮合酶I（神经元型一氧化氮合酶）是大鼠肠道中的主要一氧化氮合酶，依据如下：（a）用特异性抗体进行免疫印迹检测到一条约120 kDa的一氧化氮合酶I条带，但几乎没有一氧化氮合酶III条带；（b）用抗一氧化氮合酶I抗体进行免疫沉淀可去除大鼠肠道中依赖钙离子的组成型一氧化氮合酶（cNOS）活性，但抗一氧化氮合酶II或抗一氧化氮合酶III抗体则不能；（c）逆转录聚合酶链反应（RT-PCR）显示肠道组织中一氧化氮合酶I组成性表达，但仅有微量的一氧化氮合酶III。用抗一氧化氮合酶I抗体进行免疫荧光染色显示一氧化氮合酶位于奥尔巴赫神经丛和肌层的神经纤维中。我们还发现，给予PAF后，这种120 kDa的一氧化氮合酶I会迅速（1小时内）下调。肠道中神经元型一氧化氮合酶（nNOS）的蛋白质水平、酶活性以及信使核糖核酸（mRNA）均降低。
