在翼狀贅片進展中與上皮鈣黏著素訊息相關之蛋白質表現之研究

E-cadherin Signaling Associated Protein Expression in Pterygium Progression

楊啟賢(Chi-Hsien Young)

指導教授 ： 鄭雅文

中山醫學大學/醫學院/醫學研究所/博士(2010年)

https://doi.org/10.6834/CSMU.2010.00106

摘要

背景：翼狀贅片(Pterygium)長久以來認為是一種慢性退化性疾病，不過隨著有些研究發現翼狀贅片之上皮細胞有p53 蛋白表現異常，而認為翼狀贅片可能是一種腫瘤，不過仍需更多的證據來證實。許多研究已顯示在許多腫瘤的研究中發現上皮鈣黏素(E-cadherin)相關的蛋白質表現減少可能是腫瘤移轉及預後重要的指標之一，在翼狀贅片的研究中亦發現此蛋白的減少，但並未有相關研究去探討翼狀贅片組織中E-cadherin蛋白質不活化的原因。本研究目的主要分析在翼狀贅片組織內上皮鈣黏素轉錄起始區高度甲基化與蛋白質不活化的相關性，並進一步了解E-cadherin及其相關蛋白α-連環素(α-catenin)、β-連環素 (β-catenin) 、及p120-連環素(p120-catenin)表現的相關性。
方法：本研究分析120個翼狀贅片樣本及30正常結膜樣本，以組織免疫化學染色法(immunohistochemistry)進行分析E-cadherin、α-連環素、β-連環素、及p120-連環素之蛋白質表現。另外並使用methylation-specific PCR(MS-PCR)技術來分析E-cadherin甲基化狀態。
結果：組織免疫化學染色顯示在120個翼狀贅片樣本中，有32個(26.7%)檢測到E-cadherin基因轉錄起始區的高度甲基化。而在這些樣本中有79個(65.8%)樣本具有E-cadherin蛋白質表現，有41個(34.2%)則未表現。而E-cadherin染色則局限於表皮層的膜上。在這些樣本中，進一步分析E-cadherin基因轉錄起始區的高度甲基化與E-cadherin蛋白質表現的關係則呈負相關(p<0.0001)。且在未具有E-cadherin蛋白表現之樣本中，發現β-連環素表現位置改變，其表現位置由細胞膜移至細胞質及細胞核。
結論：本研究證實E-cadherin基因轉錄起始區的高度甲基化抑制E-cadherin蛋白質表現。E-cadherin蛋白質不表現，可能導致β-連環素表現位置改變進而促進細胞增生。本研究結果除證明E-cadherin在翼狀贅片組織中的不表現是因轉錄起始區過度甲基化所造成外，易發現上皮鈣黏素訊息路徑的活化可能參與翼狀贅片的形成。

關鍵字

翼狀贅片 ； 鈣黏素 ； 連環素 ； 高度甲基化 ； 組織免疫化學染色 ； 甲基化專一性聚合酶鏈反應

並列摘要

Background: Our recent reports indicated that the molecular changes of pterygia are similar to tumor cells. We believe that pterygia may have a similar mechanism in oncogenesis. Many studies have revealed that E-cadherin associated protein expression decreases in many tumors and pterygia E-cadherin may be a marker for both tumor metastasis and prognosis. However, no studies have examined the reason for E-cadherin protein inactivation in pterygia. Therefore, this study aimed to analyze the association of E-cadherin promoter hypermethylation with protein inactivation in pterygial tissues.
Methods: E-cadherin methylation-status and the expression of E-cadherin and β-catenin protein were studied using methylation-specific PCR and immunohistochemistry, respectively, on 120 pterygial specimens and 30 normal conjunctivas.
Results: Hypermethylation of E-cadherin gene promoter was detected in 32 (26.7%) of the 120 pterygial specimens. A total of 79 (65.8%) pterygial specimens tested positive for E-cadherin protein expression and 41 (34.2%) specimens tested negative. The E-cadherin staining was limited to the membrane of the epithelial layer. There was a reverse correlation between E-cadherin gene promoter hypermethylation and E-cadherin protein expression (p<0.0001). Aberrant localization of β-catenin was higher in the E-cadherin negative group than in E-cadherin positive group.
Conclusions: Our study demonstrates E-cadherin gene promoter hypermethylation were associated with low or absent expression of E-cadherin. Moreover, loss of E-cadherin protein may contribute to aberrant localization of β-catenin. These data provide evidence that methylation exists in pterygia and may play a role in their development.

並列關鍵字

Pterygium ； Cadherin ； Catenin ； Hypermethylation ； Immunohistolochemistry ； Methylation-specific PCR

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