含雙色胺酸功能區氧化還原酶(WW domain-containing oxidoreductase, WWOX)是一個新發現的腫瘤抑制基因(tumor suppressor gene)，研究顯示WWOX蛋白質的表現量在乳癌、食道癌、肺癌、卵巢癌、直腸癌、前列腺癌、胃癌以及肝細胞癌(hepatocellular carcinoma, HCC)中有減少的情形。有研究報導指出WWOX參與了Wnt/β-catenin訊息傳遞路徑，在HCC中，β-catenin經常發生突變，進而在細胞質中累積，接著進入細胞核內和T cell factor/lymphoid enhancer factor (TCF/LEF)轉錄因子結合並調控下游基因表現。為了要探討WWOX以及Wnt/β-catenin路徑在HCC中的表現情形，我們利用免疫組織化學染色來分析WWOX、β-catenin以及TCF4蛋白質在HCC組織中的表現並且利用統計方法分析其和臨床特徵之間的關係。當和非腫瘤組織比較時，HCC組織中WWOX的表現有明顯的下降，β-catenin累積在細胞質中，TCF4則是強烈表現在HCC細胞核內。分析臨床特徵的關係時，WWOX的表現和HCC的分級(grade)相關，TCF4則是和分級和分期(stage)有關。我們的結論為HCC中WWOX表現下降可能導致β-catenin在細胞質中的累積，進而活化Wnt/β-catenin路徑。

WW domain-containing oxidoreductase (WWOX) is a novel tumor suppressor gene discovered few years ago. Many researches indicate that expression of WWOX is reduced in a variety of cancers including heptocellular carcinoma (HCC). A recent report suggests that WWOX is implicated in Wnt/β-catenin pathway which is frequently affected in HCC. In this study, we used immunohistochemical (IHC) staining to analyze the expression of WWOX and Wnt/β-catenin pathway components in HCC and adjacent non-tumor tissues. Our result showed that WWOX was significantly downregulated in poor differentiated HCC. In addition, downregulation of WWOX was significantly correlated with cytoplasmic β-catenin expression. We also found that TCF4 was strongly expressed in HCC tissues and the expression was associated with tumor grade and stage. Consequently, our result implied that downregulation of WWOX in HCC might lead to accumulation of β-catenin in the cytoplasm and the subsequent activation of Wnt/β-catenin signaling pathway.

致謝 i
目錄 iv
圖次 viii
表次 ix
摘要 x
Abstract xi
一、序言 1
1. 肝細胞癌(Hepatocellular carcinoma, HCC) 1
1.1 HCC之分期 2
1.2 HCC之分化等級 3
1.3導致HCC的風險因子 5
2. 含雙色胺酸功能區氧化還原酶(WW domain-containing oxidoreductase, WWOX) 7
3. Wnt/β-catenin訊息調控路徑 10
3.1 β-catenin 11
3.2 TCF4 12
3.3 Wnt/β-catenin pathway和肝細胞癌生成 13
二、實驗目的 14
三、材料與方法 16
1. 檢體收集 16
2. 組織前處理 17
2.1 組織固定及包埋 17
2.2 建立組織微陣列 (Tissue microarray, TMA) 18
2.3 組織切片 18
3. 蘇木紫-伊紅染色 (Hematoxylin and Eosin Staining, HE staining) 19
4. 免疫組織化學染色 (Immunohistochemical staining, IHC) 20
4.1 WWOX的染色評估 21
4.2 β-catenin的染色評估 22
4.3 TCF4的染色評估 23
5. 統計方法 24
四、結果 25
1. 研究樣本之臨床病理特徵 25
2. WWOX在HCC中的表現 26
2.1 WWOX之免疫組織化學染色結果 26
2.2 WWOX和臨床病理特徵之間的相關性 26
2.3 WWOX的表現和HCC患者存活率之間的關係 26
3. β-catenin在HCC中的表現情形 27
3.1 β-catenin之免疫組織化學染色結果 27
3.2 β-catenin和臨床病理特徵之間的相關性 27
3.3 β-catenin的表現和HCC病患存活率之間的關係 27
4. TCF4在HCC中的表現情形 28
4.1 TCF4之免疫組織化學染色結果 28
4.2 TCF4和臨床病理特徵之間的關聯性 28
4.3 TCF4的表現和HCC患者存活率之間的關係 28
5. WWOX、β-catenin以及TCF4之間的關聯性 29
5.1 WWOX和β-catenin兩者表現之間的關係 29
5.2 β-catenin和TCF4兩者表現之間的關係 29
5.3 WWOX和TCF4兩者表現之間的關係 29
五、討論 30
六、圖 35
七、表 42
附表一 初級抗體資訊 56
附圖一 WWOX在Wnt/β-catenin路徑中的可能扮演角色 57
八、參考文獻 58

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