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研究生:蘇惠宇
研究生(外文):HUEI-YU SU
論文名稱:促發炎物質IL6促進SMAD4缺乏的胰臟癌細胞增生跟轉移的能力
論文名稱(外文):Inflammatory cytokine IL6 promotes in vitro cell proliferation and cell migration in SMAD4 null pancreatic cancer cells.
指導教授:鄭光宏鄭光宏引用關係
指導教授(外文):Kuang-hung Cheng
學位類別:碩士
校院名稱:國立中山大學
系所名稱:生物醫學研究所
學門:生命科學學門
學類:生物化學學類
論文種類:學術論文
論文出版年:2014
畢業學年度:102
語文別:英文
論文頁數:58
中文關鍵詞:白細胞介素-6EMT現象胰臟癌NestinTGF-β1/Smad4
外文關鍵詞:EMTTGF-β1/Smad4NestinInterleukin-6PDAC
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早期胰腺癌的症狀難以被檢測到,以致於病患發現罹患胰臟癌時,通常已經是癌症的晚期了。另外,研究指出患者的胰腺導管腺癌(PDAC)的預後極差,五年內的存活率低於百分之五。研究發現晚期或腫瘤已轉移的癌症患者的血液中,具有較高的白細胞介素-6(Interleukin-6)蛋白表現量。Interleukin-6(IL-6)是一種多功能的細胞因子,在宿主的免疫系統中扮演重要的角色。這些效果是由多種訊息路徑(signaling pathway)所介導的,通常經由活化transcription 3 (STAT3)去影響基因的轉錄。因此,我們想知道IL-6是否在胰腺癌缺失SMAD4時,對EMT的誘導是否有調節的作用。我們的研究發現IL-6在Smad4缺失的情況下會增加TGF-β蛋白的表現量。而實驗室先前的研究發現,在胰臟癌中,Nestin是被TGF-β1/Smad4訊息路徑所調控,並且對癌症細胞的EMT現象中扮演重要的角色。我們使用IL-6的siRNA轉染胰臟癌細胞,發現Nestin和各種EMT相關蛋白質SMA、E-cadherin和vimentin的蛋白表現量同時被抑制。因此,IL-6對Nestin所調控的EMT現象具有顯著的影響力。在Western blot和Wound healing assay中,缺失SMAD4的胰臟癌細胞在處理IL-6生長激素後使得EMT相關的蛋白質表現量增加並且促進細胞的遷移能力(cell migration)。並且Western blot的結果表明,過度表達或敲除Smad4基因會顯著的影響IL-6在胰腺癌細胞株中的表現量。總而言之,我們的研究指出,IL-6在缺失SMAD4的胰臟癌細胞中可經由磷酸化STAT3並且通過TGF-β的訊息傳導途徑,達到促進EMT的效果。另外,在MTT assay中顯示,缺失SMAD4的胰臟癌細胞處理IL-6後增加了細胞的生長增殖能力。然而,轉染了IL-6 siRNA的胰臟癌細胞,在Smad4缺失的情況下會使得癌症細胞的增殖能力受到抑制。
關鍵字 胰臟癌 白細胞介素-6 Nestin TGF-β1/Smad4 EMT現象
Early cancers of the pancreas often cause no symptoms leading to pancreatic cancer often can''t be detected until the cancer already reach to advanced stage. The prognosis of patients with pancreatic ductal adenocarcinoma (PDAC) is extremely poor and 5 year survival rate only 4%. Many studies indicated that advanced or metastatic pancreatic cancer patientscan be detected higher levels of Interleukin-6 (IL-6) in their blood. IL-6 is a multifunctional cytokine which plays a central role in host defense due to its wide range of immune and biologic activities in different types of cells including tumor cells. These effects of IL-6 can be transduced by multiple signaling pathways, in particular signal transducer and activator of transcription 3 (STAT3). Hence we want to figure out whether IL-6 plays an important role in mediating EMT in Smad4-null pancreatic cancers. We assessed the overall effect of IL-6 on TGF-β1 signaling in human PDAC cells and found that Smad4 null PDAC cells could increase the TGF-β protein expression level after IL-6 treatment in vitro. In our laboratory’s previous study, Nestin was involved in TGF-β1/Smad4 signaling pathway to induce EMT in PDAC. Transfection of IL-6 siRNA markedly decreased Nestin mRNA and protein levels compared to control cells, which lead to reduce various EMT-related marker SMA, E-cadherin and vimentin expression. Thereby, the treatment of IL-6 may regulate Nestin protein expression in Smad4 null PDAC cells. In Western blot and Wound healing analysis, we alos observed that IL-6 could induce EMT and promoted cell migration in Smad4-deficient PDAC cells. Moreover, Western blots revealed that overexpression or knockdown of SMAD4 had an impact on the regulation of IL- 6 expression in pancreatic cancer cell lines. Our research suggest that IL-6 could induce P-Stat3 to promote EMT may through TGF-β signaling pathway with ablation of SMAD4 gene in pancreatic cancer. Furthermore, MTT assays showed IL-6 increased the ability of cell proliferation in Smad4-deficient PDAC cells. Silimar results were obtained when we, transfected with IL-6 siRNA to silence IL-6 protein expression to led to reduce the ability of cell growth in Smad4-deficient PDAC cells.
Keywords: PDAC, Interleukin-6, Nestin ,TGF-β1/Smad4 ,EMT
Verification letter from the Oral Examination Committee ------------------- i
Abstract in Chinese ------------------------------------------------- ii
Abstract in English -------------------------------------------------- iii
Introduction ------------------------------------------------------------ 1
Materials and Methods --------------------------------------------- 5
Results ---------------------------------------------------------------- 12
Discussion ------------------------------------------------------------ 18
Figures and Tables -------------------------------------------------- 21
References ------------------------------------------------------------ 45
Appendix ------------------------------------------------------------ 50
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