臺灣博碩士論文加值系統

Atorvastatin (Lipitor®) 是三氫氧基三甲基穀胱胺基輔酶還原酶的抑制劑(HMG-CoA reductase inhibitor)，除了能夠降低血漿中低密度脂蛋白與膽固醇的濃度外，亦具有多項功能，在臨床上常被用來治療心血管疾病。血栓調節蛋白(thrombomodulin, TM)是位於血管內皮細胞上的醣蛋白，可與血液中的血栓蛋白結合，進而抑制血液凝血機制。在本實驗主要探討以atorvastatin處理，是否會影響TNF-α刺激人類主動脈內皮細胞表現TM與ICAM-1的表現之影響以及其相關機制。
實驗結果顯示，以atorvastatin處理人類主動脈內皮細胞之後，可回復受到TNF-α所抑制的TM表現，同時與控制組相比較，TM的表現有增加的趨勢；使用共軛焦顯微鏡進行觀察，以atorvastatin處理組，其細胞質內的TM表現有增加的趨勢。另外atorvastatin也可減少由TNF-α所誘發的細胞間黏附分子-1(Intercellular adhesion molecule-1, ICAM-1)表現，與控制組相比，ICAM-1的表現亦顯著受到抑制。以TNF-α刺激內皮細胞，可以誘導JNK (c-Jun N-terminal kinases)、ERK (extracellular signal-regulated protein kinases)及p38 MAPK (p38 mitogen activated protein kinase)的磷酸化，比較控制組與atorvastatin處理組發現atorvastatin能同時抑制JNK、ERK及p38MAPKs的活化；而加入ERK、JNK及p38MAPK抑制劑不僅能增加TM的表現，同時也抑制ICAM-1的表現。若加入抑制劑後再以atorvastatin處理，加入ERK inhibitor後，TM的表現增加但ICAM-1的表現受到抑制；而加入JNK及p38MAPK inhibitor後，其TM表現與控制組相比並無明顯差別，然而ICAM-1表現卻會受到抑制。此外，進一步以adhesion assay來觀察人類主動脈內皮細胞與單核球的黏附情形，發現給予atorvastatin處理，單核球黏附的情形有減少的趨勢，若以anti-TM抗體處理細胞，則單核球黏附內皮細胞的情形大量增加。以 NF-κB的p65抗體進行免疫螢光染色後，發現atorvastatin能減少受到TNF-α所活化而進入細胞核的NF-κB p65蛋白。
由以上實驗結果，可推論TM與ICAM-1之表現呈現負相關的情形，而atorvastatin可同時經由ERK、JNK和p38調控ICAM-1的表現，並經由ERK調控TM的表現；因此atorvastatin可以藉由增加TM表現，減少ICAM-1表現，並抑制NF-κB的活化，保護血管內皮細胞，減少受到發炎因子刺激。

Expression of functionally active thrombomodulin (TM) on endothelial cells is critical for vascular thromboresistance. The 3-hydroxyl-3-methyl coenzyme A reductase inhibitor, atorvastatin (Lipitor®), can protect the vasculature that is independent on its lipid-lowering activity. In the present study, the effects of atorvastatin on the expression of TM and ICAM-1 and their underlying mechanisms in human aortic endothelial cells (HAECs) with or without tumor necrosis-alpha (TNF-α) treatment were investigated. Atorvastatin upregulated TM levels in both control and (TNF-α) treatment were investigated. Atorvastatin upregulated TM levels in both control and TNFα-treated HAECs by Western blotting and by immunohistochemical staining. In contrast, atorvastatin decreased ICAM-1 expression in both control and TNFα-treated HAECs by Western blotting as well as the adherence of monocytes to TNFα-treated HAECs. Atorvastatin attenuated the phosphorylation of ERK, JNK and p38 MAPKs with or without TNF-α treatment. Atorvastatin upregulated the expression of TM with pretreatment of ERK inhibitor but not with JNK and p38MAPK inhibitors.
Atorvastatin downregulated the expression of ICAM-1 with ERK, JNK and p38MAPK inhibitors. Atorvastatin also inhibited the translocation of p65 subunit of NF-κB from the cytosol to the nucleus. These results suggested that atorvastatin mediated the increase of TM expression and the decrease of ICAM-1 expression which may contribute the beneficial effects to endothelial functions. The increase in endothelial TM activity and the decrease of ICAM-1 in response to atorvastatin constitute the novel pleiotropic effects of the commonly used compound and may be of clinical significance in diseases in which deficient endothelial TM plays a pathophysiological role.

中文摘要 2
英文摘要 4
緒論 5
一、前言 5
二、動脈硬化(atherosclerosis)與內皮細胞(endothelial cells)的關係 5
三、動脈硬化(atherosclerosis)與腫瘤壞死因子(TNF-α)的關係 7
四、血栓調節蛋白(Thrombomodulin, TM) 8
五、動脈硬化與細胞黏附分子的關係 9
六、動脈硬化和Mitogen Activated Protein Kinase (MAPK)的關係 11
七、動脈硬化與轉錄因子NF-κB的關係 12
八、三氫氧基三甲基穀胱胺基輔酶A還原酶抑制劑 (Statins) 13
九、研究目的 15
貳、材料與方法 16
叁、實驗結果 28
肆、討論與結論 36
伍、附圖 40
陸、參考文獻 69

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