探討p53與PKC對於TRAIL誘導非小細胞肺癌細胞凋亡之角色

王川銘(Chuan-Ming Wang)

指導教授 ： 吳文俊

中山醫學大學/醫學院/醫學分子毒理學研究所/碩士(2005年)

摘要

在肺癌細胞株中，PI3K/Akt訊息傳遞路徑在細胞對TRAIL誘導細胞凋亡中扮演重要的角色。已知p53可藉由調節PTEN進而抑制PI3K/Akt路徑。首先，本研究欲探討是否在肺癌細胞中p53會調節PI3K/Akt的活性進而影響對TRAIL的感受性。我們發現以shRNA干擾方式將p53抑制時會降低細胞對TRAIL誘導的細胞凋亡；而轉染wild type p53至某些細胞則會增加對TRAIL的敏感性。在老鼠實驗中也得到相似的結果。另外，當p53存在時能夠增加PTEN的表現，在p53存在的條件下，我們以LY294002抑制PI3K的活性發現細胞對TRAIL的感受性有加乘的作用。因此我們推測，在肺癌細胞株中會藉由p53/PTEN調節PI3K/Akt路徑進而影響對TRAIL誘導細胞凋亡的情形。許多研究已經證實PKC活化對於保護細胞對抗TRAIL引起的細胞凋亡扮演著重要的角色。本研究我們除了利用PKC抑制劑GF109203X及PKC-delta抑制劑Rottlerin抑制PKC及PKC-delta的活性外，也利用antisense RNA及siRNA抑制PKC-delta及PKC-epsilon的表現，並觀察其對TRAIL誘導細胞凋亡的情形。研究結果顯示，在肺癌細胞中，當以高劑量GF109203X處理細胞時，除了H1299外，均會增加細胞對於TRAIL的感受性；另外，除了H1355外，p53能透過藉由GF109203X抑制PKC的活性，進而增加細胞對TRAIL誘導細胞凋亡的情形。當以低劑量GF109203X(針對抑制PKC-alpha的活性)處理細胞時並不會增加細胞對於TRAIL的感受性。當以Rottlerin處理細胞時，均會增加細胞對於TRAIL的感受性。除了H1299細胞，p53能透過藉由Rottlerin抑制PKC-delta的活性，進而增加細胞對TRAIL誘導細胞凋亡的情形。然而，當轉染PKC-delta antisense RNA時，細胞對於TRAIL誘導的細胞凋亡並沒有明顯的變化。當細胞轉染PKC-epsilon siRNA時，發現對於TRAIL具有抗性的H1355及H1299細胞，會增加對於TRAIL誘導的細胞凋亡。綜合以上結果我們推測，PKC-epsilon的活性與肺癌細胞對TRAIL的感受性可能有直接的關係，這也許是肺癌細胞株對TRAIL具有抗藥性的原因之一。

關鍵字

非小細胞肺癌細胞

並列摘要

PI3K/Akt signal transduction pathway plays an important role in TRAIL- induced apoptosis. It has been suggested that p53 could suppress PI3K/Akt activity through upregulation of PTEN expression. In this study, we plan to investigate if this phenomenon will occur in lung cancer cells. First of all, TRAIL-induced apoptosis was decreased when cells treated with shRNA p53. Also, TRAIL-induced apoptosis was increased when cells overexpress wild type p53. The similar results were observed in mouse tumor model. We found p53 increased PTEN expression in lung cancer cells. There is an additive effect on TRAIL-induced apoptosis when cells treated with LY294002 in the presence of p53. The results suggest that p53 may regulate PI3K/Akt activity by increasing PTEN expression resulted in enhanced apoptotic rate induced by TRAIL. Some reports indicated that PKC activation was an important factor for cells to protect apoptosis from TRAIL. Finally, we used GF109203X, Rottlerin, PKC-delta antisense RNA and PKC-epsilon siRNA to inhibit PKC activation or expression, and then observed their effects on TRAIL-induced apoptosis. Our results demonstrated that the activities of PKC-alpha and -delta may be not associated with TRAIL sensitivity but PKC-epsilon activity is associated with the sensitivity of lung cancers to TRAIL. This mechanism may be one of the resistant factors for lung cancer cells against TRAIL attack.

並列關鍵字

non-small cell lung cancer clee

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