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題名 | The Influence of Extract of Reishi Polysaccharides on Macrophage Measured by optical Tweezers=藉由光鉗探討靈芝多醣體對於巨噬細胞的影響 |
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作者 | 魏名佐; 花國鋒; 許喬威; 柯孟揚; 許先業; 邱爾德; | 書刊名 | 光學工程 |
卷期 | 97 2007.03[民96.03] |
頁次 | 頁65-81 |
分類號 | 414.33 |
關鍵詞 | 光鉗; 四象限位置感測器; 光學彈力常數; 光學力場; 布朗運動; 震動光鉗; 巨噬細胞; 膜蛋白接受器; 大腸桿菌; 靈芝多醣體; Optical tweezers; Quadrant photodiode; QPD; Optical spring constant; Optical force field; Brownian motion; Oscillatory optical tweezers; Optical forced oscillation; Macrophage; Lipopolysaccharide; LPS; Escherichia coli; CD14; Toll-like receptor 4; TLR4; Extract of reishi polysaccharides; EORP; |
語文 | 英文(English) |
中文摘要 | 我們成功建立了光鉗微操控系統,並利用鉗住微粒時因為熱擾動所造成的布朗運動或是利用光學震盪力所造成的微力震動強度及相差定義光鉗光學彈力常數。如果我們利用水鏡(數值孔鏡為1.0)聚焦光強2mW且波長為1,064nm的電射光束,所量測出的光鉗光學彈力常數為7.6pN/μm。基於光鉗光學力場的測量,我們利用光鉗(使用值值孔鏡1.0的水鏡聚焦一波長為1064nm的雷射高斯光束)鉗住鍍有lipopolysaccharide (LPS)的聚苯乙烯微粒(直徑為1.5μm)量測,當巨噬細胞(J774A.1 cell)受到靈芝多醣體(EORP)24小時刺激後,細胞與鍍有LPS微粒間的作用力。並藉由追蹤鉗住鍍有LPS微粒時所造成的熱擾動,分析當巨噬細受靈芝多醣體刺激時,其微粒貼附到的細胞上的時間為1.18秒;相較於前者,未經刺激的巨噬細胞,其粘附時間為4.8秒。此外,我們也利用震盪光鉗鉗住鍍有LPS的微粒,並量測當被paraformaldehyde固定的巨噬細胞,比較當細胞在固定前受到靈芝多醣體刺激時所測量出LPS黏附作用彈力為364.63pN/μm;相較於沒有受到靈芝多醣體刺激的黏附作用彈力則為51.9oN/μm。我們提出了靈芝多醣體不只促使巨噬細胞結合以及內吞LPS的能力;並量測出巨噬細胞與鍍有LPS的微粒結合時間的縮短以及黏附作用彈力的增加。不論我們用光鉗、共軛焦顯微鏡或是流式細胞義所量測出LPS與受靈芝多醣體刺激過後的巨噬細胞,其LPS與細胞間結合以及內吞能力均明顯上升。 |
英文摘要 | We have established optical tweezers and manipulation systems, and analyzed the optical spring constant by detecting Brownian motion or optical forced oscillation of the trapped particle. For example, the optical spring constant was determined to be 7.6pN/μm with 2mW trapping laser power and 1,064nm laser wavelength focused by a water-immersed objective lens with N.A. = 1.0. Based on the measurement of optical force field in optical tweezers and the frequency response of optical forced oscillation, we have trapped the lipopolysaccharide (LPS) conjugated polystyrene particle (diameter = 1.5μm) via optical tweezers (λ=1064nm Gaussian beam focused by water-immersed objective lens with N.A.=1.0) and measured its interaction with a macrophage (J774A..1 cell) pre-treated with the extract of Reishi polysaccharides (EORP). By tracking and analyzing the Brownian motion of trapped LPS conjugated particle, the time constant characterizing the adhesion of LPS particle with a macrophage was measured to be around 4.8s when the macrophage was not pretreated with EOPR compared with 1.18s in cases when the macrophage was pre-treated with EOPR. In addition, we also measured, via optical forced oscillation of a LPS conjugated particle, the elasticity of such ligand-recdeptor interactions on the plasma membrane of macrophages pre-treated with paraformaldehyde; the elastic constant was measured to be around 51.9pN/μm when the macrophage was not pre-treated with EOPR compared with 364.63pN/μm in pre-treated cases. We demonstrated, by tracking the dynamics of trapped LPS conjugated particle, that EOPR not only enhanced macrophages surface expression of TLR4 and CD14 as well as LPS binding and internalization, but also reduced the adhesion time constant and increased the elasticity of the binding interaction. Our experimental results based on optical tweezers are consistent with the observation, via confocal microscopy and flow cytometry, of the binding and internalization (phagocytosis) reaction of macrophages. |
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