Please use this identifier to cite or link to this item:
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/32994
Title: | 變異型脂蛋白元AV功能之研究 Functional Study of Apolipoprotein AV Variant |
Authors: | Ching-Tzu Yen 顏靜慈 |
Advisor: | 高照村 |
Keyword: | 脂蛋白元AV, apolipoprotein AV, |
Publication Year : | 2006 |
Degree: | 碩士 |
Abstract: | 高三酸甘油脂血症是一種相當常見、致因多樣化的代謝性異常疾病,導因於製造過多、或無法有效清除血液中富含三酸甘油脂的脂蛋白,且高三酸甘油酯血症亦為心血管疾病的一項獨立危險因子。血液中的脂質是由脂蛋白所攜帶以進行代謝;同時,在脂蛋白表面有各式各樣且功能各異的脂蛋白元調節著脂質和脂蛋白的代謝。脂蛋白元AV為近年新發現的一個脂蛋白元,根據先前的研究中指出,在基因改造小鼠體內,血中脂蛋白元AV的含量和三酸甘油脂的濃度成負相關;同時在人類脂蛋白元A5基因的多型性研究中,顯示了許多SNPs與高三酸甘油酯血症具有統計學上的相關性。而近期所發表的研究結果指出脂蛋白元AV能以直接活化或是間接影響的方式來提升脂蛋白解脂酶的水解能力,藉此降低血液中的三酸甘油酯。
本實驗室所發現的c.553G>T多型性,會造成脂蛋白元AV的第185個胺基酸由甘胺酸(glycine)變為半胱胺酸(cysteine),為瞭解此變異型脂蛋白元AV影響三酸甘油酯正常代謝之機制,在本篇論文中,建構了此變異型之質體,利用細菌表現、並純化出重組脂蛋白元AV。在體外脂蛋白解脂酶活性分析中,野生型和變異型脂蛋白元AV皆能提升脂蛋白解脂酶的活性,但後者的能力較差(較野生型降低26%, p<0.001)。只是這樣的實驗結果在以其他批次純化的重組脂蛋白元AV進行分析時,無法重複呈現;同時,也發現到實驗結果會受到尿素濃度、溫度以及脂蛋白元AV活性的穩定度等其他因子的影響。此外,為模擬生理環境,更進一步將脂蛋白解脂酶連結在HSPG上進行活性分析,雖然兩者皆能促進HSPG連結脂蛋白解脂酶的水解能力,但彼此之間並無統計學上的差異。 目前實驗的結果,仍無法證明變異型脂蛋白元AV影響三酸甘油酯代謝的機轉,未來若能以apoa5基因剔除小鼠為動物模式做更進一步的探討,將能給予更多的解答。 Hypertriglyceridemia(HTG)is a common metabolic disorder resulted from multiple determinants. It may arise from either over-production or defective clearance of triglyceride-rich lipoproteins in circulation. Furthermore, HTG is also an independent risk factor for coronary heart disease. Plasma lipid is known to be carried by lipoproteins and to undergo further hydrolysis. Meanwhile, diverse-functioned apolipoproteins on lipoproteins modulate metabolism of lipid and lipoprotein. ApoAV was discovered as a new member of the apolipoprotein. Early studies indicated that the concentration of apoAV exerted an inverse correlation with the level of plasma triglyceride(TG)in genetically engineered mice. The studies also revealed that the APOA5 gene polymorphism in human was statistically correlated with the incidence of HTG. Recently studies have shown that apoAV can modulate the triacylglycerol hydrolase activity of lipoprotein lipase(LPL)through direct activation or indirect effects to reduce the plasma TG. We have identified a c.553G>T polymorphism, resulting in the substitution of cysteine for glycine at residue 185. To gain insight into the mechanism of the effect of APOA5 185C variant, we constructed the vector carrying APOA5 185C variant cDNA, and then expressed and purified the recombinant apoAV. Using in vitro LPL activity assay, we initially showed that both apoAV wild type and 185C variant significantly increased LPL activity, but the later was less effective(26% lower compared to apoAV wild type. p<0.001). But this result could not be reproduced using recombinant apoAV purified at different batch. Meanwhile, we also found other factors, such as urea, temperature and the stability of apoAV activity, would cause variation of results. Furthermore, in order to reflect the physiological situation in which LPL is bound to cell surface through HSPG, we examined the effect of apoAV on the LPL activity in the presence of HSPG. Although both wild type and variant significantly increased lipolysis ability of HSPG-bound LPL, there was no statistical difference between them. According to results of this study, the mechanism of the effect of APOA5 185C variant on triglyceride metabolism remains uncertain. Further animal studies using apoa5 knockout mice will be necessary in order to explore the mechanism in the future. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/32994 |
Fulltext Rights: | 有償授權 |
Appears in Collections: | 醫學檢驗暨生物技術學系 |
Files in This Item:
File | Size | Format | |
---|---|---|---|
ntu-95-1.pdf Restricted Access | 827.54 kB | Adobe PDF |
Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.