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標題: The Mechanisms Underlying the Leptin-Mediated Inhibition of Neutrophil Infiltration into Ischemic Brain
作者: 葛其梅
關鍵字: 腦缺血;Cerebral ischemia;基礎醫學類;瘦素;嗜中性白血球;內皮細胞;星狀細胞;血腦障壁;黏著蛋白;緊密連結蛋白;趨化性;基礎研究;Leptin;Neutrophil;Endothelial cell;Astrocyte;BBB;Cell adhesion molecule (CAM);Tight junction protein;Chemotaxis
The primary goal of the study is to assess the underlying mechanisms responsible forleptin-mediated inhibition of neutrophil infiltration into ischemic brain that in partcontributes to the leptin-mediated brain protection against cerebral ischemia. This isa three-year project with divided approaches including: 1) would leptin inhibit thechemotaxis (or migration) and activation of neutrophils under ischemic condition; 2)would leptin inhibit the neutrophil adherence to ischemia/reperfusion-inflamedendothelium by altering the cell adhesion molecule (CAM) patterns on both sides;and 3) would leptin reduce the ischemia/reperfusion-induced BBB permeability byincreasing the expression of endothelial tight junction proteins and astrocytic SSeCKS.In the study, the in vivo ischemic animal model and in vitro GOSD (glucose- oxygenandserum-deprivation) model are both applied to examine the impacts of leptin onthe extravasation of blood borne neutrophils, the permeability of endothelial cells andBBB, and the regulatory activity of astrocytes. Techniques applied include: bloodvessel occlusion technique, TTC stain, Evens blue extraction assay, cell migrationassay, Western blotting, immunohistochemistry, immunocytochemistry, andneutrophil activity analyses such as, intracellular Ca+2 measurement, the release ofsuperoxide, nitric oxide and lysozyme, and phagocytic analysis. Results from thestudy can provide valuable information for us to understand the underlyingmechanisms and the clinical applications of leptin-mediated inhibition of neutrophilinfiltration into ischemic brain. In addition, the therapeutic value of leptin in strokecan be further verified and new strategies in the control of this disease may also beevolved from the results of this study.

本計畫的研究目標主要在探討『瘦素如何抑制嗜中性白血入侵缺血之腦組織』。已知嗜中性白血球穿越血腦障壁(blood brain barrier or BBB)進入缺血之腦組織,會對腦組織造成發炎性的二次傷害;而瘦素對缺血之腦組織不但具保護性,且明顯抑制血液中嗜中性白血球滲入腦組織。本計畫因此想進一步探究瘦素對嗜中性白血球之抑制機轉,以便更深入了解瘦素在臨床中風治療上的潛能。計畫主要分三年進行,每年各有一研究議題,彼此相互牽引,互為佐證。三個研究議題分別探討:1) 瘦素對嗜中性白血球的化學吸引力是否因缺血性的壓力而消失,進而減少缺血之腦組織中嗜中性白血球的數目;2) 瘦素是否可藉由抑制嗜中性白血球及血管內皮細胞表面黏著蛋白(cell adhesion molecules or CAMs)的表現,來干擾嗜中性白血球對血管內皮細胞的吸附性,進而阻止嗜中性白血球進入缺血之腦組織;3) 瘦素是否可藉由提升血管內皮細胞緊密連結蛋白(tight junctionproteins and adherens junction proteins)及星狀細胞SSeCKS 蛋白的表現,來降低腦血管障壁的通透性,進而抑制嗜中性白血球滲入缺血之腦組織中。實驗中主要以腦血管結紮的動物缺血模式,及體外缺糖、缺氧、及缺血清(GOSD)的細胞研究模式,做為分析平台,並以不同的實驗方法評估在缺血環境下,瘦素對血腦障壁,嗜中性白血球,血管內皮細胞,及星狀細胞特性之影響。分析技術包括:血管結紮手術, TTC 染色法,Evens blue 淬取法, 細胞移動分析法, 西方墨點法,組織免疫染色法,細胞免疫染色法,及嗜中性白血球活性分析包括:細胞內鈣離子的變化,superoxide、NO , 及lysozyme 的釋放,和細胞之吞噬活性分析等。研究結果將可強化我們對瘦素的認知,能更深入了解瘦素對嗜中性白血球移動力及活性的牽制,對血腦障壁組成細胞之影響,及其對缺血腦組織的保護機轉。據此將可進一步評估,瘦素在中風治療上之潛能,並有助於未來新藥物或治療方法之研發。
其他識別: NSC99-2320-B005-006-MY3
Appears in Collections:生命科學系所

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