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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 鄭永銘(Yung-Ming Jeng) | |
dc.contributor.author | Fang-I Huang | en |
dc.contributor.author | 黃芳儀 | zh_TW |
dc.date.accessioned | 2021-05-20T21:04:38Z | - |
dc.date.available | 2016-10-07 | |
dc.date.available | 2021-05-20T21:04:38Z | - |
dc.date.copyright | 2011-10-07 | |
dc.date.issued | 2011 | |
dc.date.submitted | 2011-07-07 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/10136 | - |
dc.description.abstract | 肝細胞生長因子(Hepatocyte growth factor, HGF)與其接受體c-Met在癌症的生長與遷移扮演重要的角色。HGF由纖維組織母細胞(fibroblast)分泌,是一個癌症生長與發育的重要中間因子。然而,這其中的機轉尚未清楚。Wnt/β-catenin/Tcf/LEF1 是一條經常在癌細胞中被活化的路徑。利用β-catenin相關路徑的報導基因表現分析,我們發現在肝癌細胞株,肝細胞生長因子會活化β-catenin的路徑。利用微陣列基因表現分析,我們發現LEF1mRNA的表現量被肝細胞生長因子增加了。給予MDA-MB-231細胞株Wortmannin 會抑制HGF誘導的LEF1表現,這代表HGF 誘導LEF1是藉由PI3K/Akt的路徑。我們也給予MDA-MB-231細胞株BAY11-7082及SN50二種專一抑制NF-κB 的抑制劑,也發現可以減少HGF誘導的LEF1表現。利用shRNA敲毀LEF1,會抑制HGF造成的腫瘤侵襲。根據組織免疫染色,我們發現LEF1在各種不同的癌症表現。我們研究顯示,藉由LEF1的轉譯調控,HGF/Met與Wnt/β-catenin/Tcf/Lef之間的訊息傳遞是HGF誘導侵襲的必要條件。 | zh_TW |
dc.description.abstract | Hepatocyte growth factor (HGF) and its receptor c-Met play important roles in cancer growth and metastasis. HGF is an important fibroblast-secreted protein which is a mediator of cancer development and progression. However, the mechanism is still poorly characterized. Wnt/β-catenin/Tcf/LEF1 pathway is frequently activated in cancers. Using the reporter of β-catenin pathway, we found HGF activates β-catenin pathway in liver cancer cell line HepG2. Using microarray analysis, we found LEF1 expression is enhanced by HGF. Treatment of the MDA-MB-231 cells with wortmannin suppressed the induction of LEF1, indicating that the effect was mediated by signaling through phosphatidylinositol-3-kinase (PI3K)/Akt pathway. We also treated MDA-MB-231 cells with BAY11-7082 and SN50, two specific NF-κB translocation inhibitors, and also found that it could decrease LEF1 expression after HGF treatment. Knockdown of LEF1 by shRNA in MDA-MB-231 cells inhibited tumor invasion induced by HGF treatment. According to IHC staining data, we found that LEF1 is expressed in various cancers. Our study suggests that cross-talk of HGF/Met and Wnt/β-catenin/Tcf/Lef by transcriptional regulation of LEF1 is essential for the invasion-inducing effect of HGF. | en |
dc.description.provenance | Made available in DSpace on 2021-05-20T21:04:38Z (GMT). No. of bitstreams: 1 ntu-100-R98444002-1.pdf: 11982429 bytes, checksum: a692c5e1525be4f3d83b0ad67785c61f (MD5) Previous issue date: 2011 | en |
dc.description.tableofcontents | 口試委員審定書 II
中文摘要 III Abstract IV Contents V 1. Introduction 1 1.1 Structure of MET and HGF-------------------------------------------------------------------1 1.2 Major MET-regulated signaling pathways--------------------------------------------------2 1.3 HGF/MET induce EMT-----------------------------------------------------------------------4 1.4 Wnt/β-catenin pathway------------------------------------------------------------------------6 1.5 Lymphoid enhancer factor (LEF1) ----------------------------------------------------------7 1.6 Breast cancer------------------------------------------------------------------------------------9 1.7 The study aim---------------------------------------------------------------------------------10 2. Materials and Methods 11 2.1 Cell culture------------------------------------------------------------------------------------11 2.2 RNA interference-----------------------------------------------------------------------------11 2.3 RNA isolation---------------------------------------------------------------------------------12 2.4 RT-PCR ----------------------------------------------------------------------------------------12 2.5 Western Blot-----------------------------------------------------------------------------------12 2.6 In vitro Boyden chamber Invasion Assay -------------------------------------------------14 2.7 Wound-healing Assay------------------------------------------------------------------------14 2.8 Scattering assay-------------------------------------------------------------------------------15 2.9 Chromatin immunoprecipitation assay----------------------------------------------------15 2.10 Immunohistochemistry---------------------------------------------------------------------16 2.11 Reporter assay-------------------------------------------------------------------------------17 3. Results 20 3.1 HGF induces cell scattering-----------------------------------------------------------------20 3.2 Identification of differential gene expression between HGF treated or untreated HepG2 cell line------------------------------------------------------------------------------20 3.3 HGF induces LEF1 expression in HepG2 and MDA-MB-231-------------------------21 3.4 LEF1 is required for HGF-induced cell invasion and migration. ----------------------21 3.5 HGF induces LEF1 expression independently of β-catenin activity. ----------------- 22 3.6 HGF-induced LEF1 expression depends on PI3K/AKT pathway--------------------- 22 3.7 HGF induces LEF1 expression via NF-κB------------------------------------------------23 3.8 LEF1 is a target gene of NF-κB in MDA-MB-231-------------------------------------- 23 3.9 Knockdown LEF1 in MDA-MB-231 reduce epithelial-mesenchymal transition and tumor migration----------------------------------------------------------------------------- 24 3.10 The expression degree of LEF1 in various cancers------------------------------------ 24 4. Discussion 25 4.1 The role of HGF in cancer invasion------------------------------------------------------- 25 4.2 HGF induces LEF1 through PI3K/AKT/NF-κB pathway ----------------------------- 26 4.3 HGF induces LEF1 expression in HepG2 and MDA-MB-231------------------------ 30 4.4LEF1 is required for HGF-induced invasion and migration---------------------------- 32 4.5 Knockdown of LEF1 in MDA-MB-231 reduced epithelial-mesenchymal transition and tumor migration. ---------------------------------------------------------------------- 33 5. Figures and Tables 34 Figure 1. HGF induces cell scattering -------------------------------------------------------- 34 Figure 2.Veridation of the genes upregulated by HGF in microarray data by RT-PCR.-37 Figure 3. HGF induces LEF1 expression in HepG2 and MB231. ------------------------ 39 Figure 4. LEF1 is required for HGF-induced cell invasion. ------------------------------- 40 Figure 5. LEF1 is required for HGF-induced wound-healing migration ----------------- 42 Figure 6. HGF induces TCF/ LEF1 reporter activation in various cell lines------------- 44 Figure 7. LEF1 is regulated by HGF through PI3K/AKT and NF-kB pathway--------- 47 Figure 8. NF-κB binds to its predicted upstream binding site on LEF1 locus----------- 48 Figure 9. Knockdown of LEF1 reduce the expression of Snail and ZEB2. The down expression was not due to activation of GSK3β. -------------------------------- 51 Figure 10. Immunohistochemical detection of LEF1 in breast cancer. ------------------- 54 Figure 11. Immunohistochemical detection of LEF1 in renal cell carcinoma. ---------- 55 Figure 12. Immunohistochemical detection of LEF1 in lung cancer. --------------------- 56 Figure 13. Immunohistochemical detection of LEF1 in ovary cancer -------------------- 57 Figure 14. Immunostaining of LEF1 in oral mucosa and OSCC-------------------------- 58 Figure 15. Molecular mechanisms underlying HGF-induced cell scattering------------- 59 Table 1. The primers used for PCR reaction. -------------------------------------------------18 Table 2. The primers used for ChIP analysis. -------------------------------------------------18 Table 3. Detect the genes HGF induced by microarray--------------------------------------36 Table 4. Expression of LEF1 andβ-catenin in human breast cancer ---------------------45 Table 5. The expression of LEF1 in various cancers ----------------------------------------52 7. Reference 61 | |
dc.language.iso | en | |
dc.title | HGF藉由活化LEF1轉錄而活化Wnt路徑以促進腫瘤侵襲 | zh_TW |
dc.title | HGF activates Wnt pathway by transcriptional activation of LEF1 to facilitate tumor invasion | en |
dc.type | Thesis | |
dc.date.schoolyear | 99-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 張正琪,林佼穎,黃祥博 | |
dc.subject.keyword | 肝細胞生長因子, | zh_TW |
dc.subject.keyword | HGF,Wnt,LEF1, | en |
dc.relation.page | 73 | |
dc.rights.note | 同意授權(全球公開) | |
dc.date.accepted | 2011-07-07 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 病理學研究所 | zh_TW |
顯示於系所單位: | 病理學科所 |
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