亞砷酸鈉對於SV-40人類泌尿道上皮細胞及泌尿道上皮細胞癌Caveolin-1、eNOS、IKKβ及COX-2蛋白質表現之影響

Effect of sodium arsenite on Caveolin-1, eNOS, IKKβ and COX-2 expression in SV-40 immortalized human uroepithelial cells and urothelial carcinomas

劉萱蘋(Xuan-Ping Liu)

指導教授 ： 蔡志仁

高雄醫學大學/醫學院/醫學研究所/碩士(2009年)

摘要

目的：砷是大自然中常見的一種元素，可分為有機砷及無機砷二種形
式，其中以無機砷最具毒性。在台灣主要的砷中毒地區分別是西南沿
海與東北沿海等區域，其中最引起注意的是因砷暴露而引起之烏腳
病；此外，該地區之泌尿道上皮細胞癌的發生率也偏高。許多癌症研
究已證實砷與及其化合物屬於致癌物質，並且其致癌性與暴露的時間
長短和劑量呈現相關性。砷之致癌機轉被認為和致癌基因活化、腫瘤
抑制基因的失調、干擾DNA 的複製及訊息傳遞蛋白異常表現等多種
因素有關。本研究之目的是觀察亞砷酸鈉對於人類泌尿道上皮細胞及
泌尿道上皮培養細胞癌之腫瘤相關蛋白質之影響。
材料與方法：泌尿道上皮培養細胞經由不同濃度的砷劑處理後，利用
細胞染色法及西方墨點法觀察細胞內Caveolin-1、eNOS、IKKβ及
COX-2之表現量，同時本研究也利用免疫組織化學染色觀察烏腳病
及非烏腳病地區泌尿道上皮細胞癌之Caveolin-1、eNOS、IKKβ 及
COX-2的表現情形，以便進一步探討砷化物如何影響蛋白質的作用
機轉，使得細胞生長調節失去控制進而導致腫瘤的形成。
結果：本研究結果顯示，低濃度亞砷酸鈉可以誘導泌尿道上皮培養細
胞Caveolin-1、IKKβ 及COX-2之高度表現及抑制eNOS 之表現，而
泌尿道上皮癌之免疫組織化學染色結果亦顯示Caveolin-1、IKKβ 及
COX-2之過度表現以及eNOS之低下表現，並藉由統計分析顯示
Caveolin-1-eNOS-IKKβ-COX-2蛋白質彼此之間的相互作用具有顯著
相關，而這些訊號蛋白異常表現，將有助於我們對砷於泌尿道上皮癌
致病機轉有更進一步的了解。

關鍵字

亞砷酸鈉 ； SV-40人類泌尿道上皮細胞 ； 泌尿道上皮癌 ； Caveolin-1 ； eNOS ； IKKβ ； COX-2

並列摘要

Purpose: Arsenic is a known human carcinogen, and extend to every part of the earth's crust. Cancer risk assessments have been based on epidemiological studies of a large population exposed to arsenic in south western Taiwan. It is well known that there was significantly correlation between urothelial cancer and blackfoot disease with arsenic. Arsenic exposure can accumulate in the liver, spleen, kidneys, and gastrointestinal tract, further result in the development of various cancers. Caveolin-1 scaffolding domain bind to eNOS and thereby inhibit NO production. NO represses IKK through S-nitrosylation in the immune system. IKK-mediated NF-κB activation can upregulate COX-2 expression in oncogenesis. The aim of this study is to determine if sodium arsenite can affect Caveolin-1 and downstream signaling molecules expression(eNOS, IKKβ and COX-2) in low dose condition. In addition, we also performed immunohistochemistry staining of Caveolin-1, eNOS, IKKβ and COX-2 in human urothelial carcinoma specimens in both the blackfoot disease and non-blackfoot disease endemic areas in Taiwan.
Material and method: SVHUC-1 cells were treated in culture with low concentration sodium arsenite. The expression of Caveolin-1, eNOS, IKKβ and COX-2 were examined by Immunocytochemistry and western blotting. Furthermore, we collected 34 paraffin-embedded specimens of urothelial carcinoma with blackfoot disease and non blackfoot disease areas. The sections were examined immunohistochemistry for Caveolin-1, eNOS, IKKβ and COX-2 expression. The correlations with clinicopathologic features were evaluated statistically.
Results: Immunocytochemical staining and Western blotting results revealed increased expression of Caveolin-1, IKKβ and COX-2 and decreased eNOS in low concentration of arsenite treated SV-HUC-1 cells. Our findings indicate that U0126 inhibitor significantly attenuated arsenite-induced expression of Caveolin-1, IKKβ and COX-2, and restored the reduced eNOS expression. Immunohistochemistry staining revealed cytoplasmic higher expression of Caveolin-1, IKKβ and COX-2 in patients with urothelial carcinoma blackfoot disease(P=0.037, P=0.01, P=0.0001), but decrease of eNOS expression in blackfoot disease endemic area(P=0.0001).
Conclusion: Our present result provide new aspects for the correlation among Caveolin-1 and downstream signaling molecules expression, which may play important roles in the mechanisms of arsenic-induced urothelial carcinogenesis. By U0126 inhibitor treatment, we found the inhibitor could be a potentially therapeutic agent for urothelial cancer.

並列關鍵字

sodium arsenite ； urothelial carcinoma ； SV-40 immortalized human uroepithelial cells ； Caveolin-1 ； eNOS ； IKKβ ； COX-2

參考文獻

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