以微卡計量測細胞膜不同含量膽固醇和神經節?脂之 PC12細胞與不同型態之類澱粉胜?作用後的反應熱與其 細胞活性之研究 ; Microcalorimetric and cytotoxic measurements of different cholesterol and ganglioside content plasma membrane PC12 cells exposed to various form of amyloid-beta

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題名:	以微卡計量測細胞膜不同含量膽固醇和神經節?脂之 PC12細胞與不同型態之類澱粉胜?作用後的反應熱與其細胞活性之研究;Microcalorimetric and cytotoxic measurements of different cholesterol and ganglioside content plasma membrane PC12 cells exposed to various form of amyloid-beta
作者:	洪士耘;Shih-yun Hung
貢獻者:	化學工程與材料工程研究所
關鍵詞:	細胞膜;細胞毒性;神經節苷脂;膽固醇;類澱粉胜肽;阿茲海默症;plasma membrane;microcalorimetry;cytotoxic;ganglioside;cholesterol;amyloid-beta;Alzheimer's disease
日期:	2009-11-01
上傳時間:	2010-06-10 16:49:43 (UTC+8)
出版者:	國立中央大學圖書館
摘要:	類澱粉蛋白(amyloid-?，A?)被認為是引起阿茲海默症的主因，當A?由單體在細胞膜上聚集成纖維狀結構時就會毒殺細胞，但是A?引起細胞毒性及導致疾病的機制目前還不清楚。近年研究指出，細胞膜上的神經節苷脂?GM1)和膽固醇的含量與A?的貼附和聚集有關。本研究以恆溫滴定微卡計作為即時熱流量之量測工具，測量細胞膜不同含量膽固醇和神經節苷脂之PC12細胞與不同聚集度之類澱粉蛋白作用後的反應熱，熱流數據計錄從細胞放入恆溫滴定微卡計到注射A?後並反應兩到三天。同時我們也在96孔盤內進行與微卡計內相同條件的實驗，透過這兩個實驗的結合期望能夠了解細胞在類澱粉蛋白的作用下其熱流量與細胞活性的關係。在96孔盤內的實驗結果顯示在48小時內，降低細胞膜GM1可抑制A?對細胞的毒性；另外降低細胞膜膽固醇也可抑制A?對細胞的毒性，而增加細胞膜膽固醇則會加強A?對細胞的毒性。將細胞培養在組織培養皿內並改變細胞膜上的膽固醇和GM1的含量後，取下培養皿上細胞放入微卡計內待平衡和細胞貼附後注射入不同聚集度的A??monomer rich, oligomer rich, fiber rich)，結果顯示熱流量與細胞活性及死亡率有高度相關，證明微卡計熱流量可做為細胞狀態的參考，而透過積分注射後之總放熱量並除以死亡細胞數，發現當系統內細胞毒性較強時，單位細胞死亡所放出的熱量會增加。 ?-amyloid(A?) was believed to cause the prime factor of Alzheimer’s disease. The deposition and aggregation of A? on cell membrane led to cytotoxicity. However, the detail mechanism?was still unclear. Recent studies demonstrated that certan components of plasma membrane, ganglioside(GM1) and cholesterol play an influential role in the accumulation of A? on the plasma membranes. In this study, we applied isothermal titration calorimetry (ITC) as an “on-line” heat probe to identify the living cell systems. We changed the cholesterol and GM1 amount on PC12 cells plasma membrane in vitro and traced the thermodynamic activity of the interaction between A? and different contents plasma membrane PC12 cells by ITC. The heat flux data contained not only metabolic heat of cells but olso other reaction heat between A? and cells. Base on different experiment designs, different forms of A??(monomer rich, oligomer rich and fiber rich) were chosen. All heat flow is recorded by ITC from cell seeding to the end of experiment. We also redo the control experiment in 96 wells as a parallel experiment. Our results demonstrated that the reduction of cholesterol or GM1 contents on plasma membrane could diminish the A??cytotoxicity?in12, 24 and 48 hours but higher cholesterol content could cause higher cytotoxicity. The heat flow data from ITC was highly correlated to the MTT reduction and LDH release of parallel experiment. We demonstrated the heat flow from our ITC system could be a good indication of cell viability. By integrating the total heat dissipation after the injection of A?, the exothermic heat per dead cell enhanced when the system condition was more toxic to cell.
顯示於類別:	[化學工程與材料工程研究所] 博碩士論文

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