出版年:

2001

研究生:

莊國賓

研究生(英文姓名):

kuo-pin chuang

論文名稱:

氧活性中間產物在嗜咿紅性白血球黏著所扮演的角色

英文論文名稱:

The role of reactive oxygen intermediated on integrin mediated eosinophil adhesion

指導教授:

謝奇璋

學位類別:

碩士

校院名稱:

國立成功大學　

系所名稱:

微生物暨免疫學研究所

學號:

S46881107

學年度:

89

語文別:

中文

論文頁數:

77

關鍵詞:

嗜咿紅性白血球 ; 一氧化氮 ; 過氧化物

被引用次數:

0

[ 摘要 ]

氣喘被認為是一種呼吸道的慢性發炎反應，而氣喘最主要的特徵是會有多量的嗜咿紅性白血球（Eosinophil；EOS）堆積於呼吸道的上皮組織中。這些嗜咿紅性白血球會釋放非常大量的有害物質進而對呼吸道上皮細胞造成非常大的傷害。所以如何引起嗜咿紅性白血球從血液中穿透血管上皮細胞，移動到氣管上皮細胞上是引起氣喘的最重要步驟。其中位於血管上皮細胞表面的黏著分子（adhesion molecules），Vascular cell adhesion molecular-1----VCAM-1。這個黏著分子（adhesion molecular）已經被認為與嗜咿紅性白血球從血管內移動到呼吸道中有很大的關係，因為嗜咿紅性白血球細胞表面具有VCAM-1的受器 ---very lateactivation antigen 4（VLA-4）可與其結合進而使嗜咿紅性白血球從血管移動到呼吸道。
近年來有人指出在氣喘病患呼出的氣體中含有大量的一氧化氮（NO）。另外也發現在氣喘病人的肺泡沖洗液（bronchoalveolar lavage fluid；BALF）含有高量的superoxide（O2-）。所以我們想要研究這些自由基對於嗜咿紅性白血球和VCAM-1 這個黏著分子之間黏著能力的影響。
首先我們建構了VCAM-1 第1到第3 domain的重組蛋白，因為VCAM-1 的第1到第3 domain即具有這個黏著分子的全部功能，接著利用嗜咿紅性白血球細胞株（HL-60 clone 15），及利用一些NO的提供者例如：sodium nitroprusside (SNP)，S-nitroso-L-glutathione (GSNO)，產生NO，並且利用phorbol 12-myristate 13-acetate (PMA)刺激後的多形性顆粒球提供superoxide，用以研究NO、 ROS對於嗜咿紅性白血球和黏著分子之間結合力的影響。
結果顯示NO、和H2O2會降低eosinophil與VCAM-1之間的黏著能力，但是相反的superoxide卻是有可使黏著力上升的能力。Superoxide具有可以將因NO所造成黏著力下降的情形救援回來的能力。
我們的這些結果顯示，在組織的微環境中，NO和ROS之間的平衡可以決定adhesion moleculars 和白血球之間的黏著力，而這種影響可能也是引起在氣喘病人的白血球堆積在氣管內皮的最主要因素。

[ 英文摘要 ]

Asthma has been recognized as a chronic inflammatory disease in the airway. A characteristic feature of allergic inflammation is the accumulation of eosinophils in the respiratory epithelium. Eosinophils release various mediators that may damage bronchial epithelia cells. The recruitment of eosinophils from blood to the bronchial epithelium thus is regarded a key event in allergic inflammation. Vascular cell adhesion molecular-1 (VCAM-1) has been recognized as an important molecule for selective eosinophils migration into the airway, because vary late antigen-4 (VLA-4), a integrin ligand for VCAM-1, is expressed in eosinophils but not in neutrophils. Integrin-mediated adhesion of eosinophil to VCAM-1 may contribute to the selective recruitment of eosinophils to tissue with allergic inflammation.
In recent years, many reports have indicated that reactive oxygen species (ROS) and nitric oxide (NO) are important modulators of leukocyte functions. Patients with bronchial asthma have increased levels of NO in exhaled air, and the bronchoalveolar lavage fluid (BALF) from asthma patients also has high level of superoxide. In this investigation, we studied the effect of inflammation mediators, including ROS and NO, on integrin-mediated eosinophil adhesion.
We constructed a recombinant soluble VCAM-1 (rsVCAM-1) and ICAM-1 (rsICAM-1) and used these proteins and an eosinophil cell line (HL60 Clone15) to investigate the role of NO and ROS on VCAM-1 and ICAM-1 mediated eosinophil adhesion. We used NO donors (sodium nitroprusside (SNP) and S-nitroso-L-glutathione (GSNO)) to generate NO, and PMA stimulated PMN to generate superoxide, and used a static cell-to-protein adhesion assay to analyze their effect on adhesion. Our results showed that both H2O2 and NO decreased eosinophil binding to VCAM-1, yet superoxide (O2-) increased the specific adhesion. In the presence of superoxide generating leukocytes, the suppressive effect of NO was reversed.
Our results suggested that the balance between ROS and NO in the tissue microenvironment may determine the integrin-mediated leukocyte adhesion and is likely to be a key factor in leukocyte recruitment in allergic reactions.