本文選題：胰腺癌 + DAB2IP��； 參考：《華中科技大學(xué)》2016年博士論文

【摘要】：目的：胰腺癌作為轉(zhuǎn)移侵襲能力極強(qiáng)、致死率極高的腫瘤,患者的預(yù)后往往不佳。DAB2IP作為一種RasGAP蛋白家族成員之一,在多種腫瘤中證明其發(fā)揮抑癌基因的作用。本研究旨在探討DAB2IP對(duì)胰腺癌生長(zhǎng)、轉(zhuǎn)移及化療抗性的作用及機(jī)制,從而為治療胰腺癌探索新的靶點(diǎn)。方法：1.探討DAB2IP影響胰腺癌生長(zhǎng)能力的體內(nèi)外研究：首先,臨床胰腺癌標(biāo)本中分析DAB2IP表達(dá)的差異及與臨床病理學(xué)資料的相關(guān)性,重點(diǎn)分析其與胰腺癌預(yù)后的關(guān)系；然后,利用胰腺癌內(nèi)源性表達(dá)DAB2IP不同的細(xì)胞系分別轉(zhuǎn)染過(guò)表達(dá)及低表達(dá)質(zhì)粒,構(gòu)建穩(wěn)定高、低表達(dá)DAB2IP的細(xì)胞；其次,分別應(yīng)用細(xì)胞增殖、凋亡實(shí)驗(yàn)、平板克隆及軟瓊脂克隆實(shí)驗(yàn)證明其對(duì)胰腺癌細(xì)胞體外生長(zhǎng)的作用；最后,利用胰腺癌皮下移植瘤模型,分析DAB2IP對(duì)腫瘤生長(zhǎng)的影響,并分析其可能的機(jī)制。2.探討DAB2IP影響胰腺癌轉(zhuǎn)移能力的體內(nèi)外研究：首先,我們應(yīng)用構(gòu)建好的DAB2IP穩(wěn)定高/低表達(dá)細(xì)胞系,利用劃痕實(shí)驗(yàn)、Transwell侵襲遷移實(shí)驗(yàn)檢測(cè)細(xì)胞體外轉(zhuǎn)移能力的改變；然后,分別利用Western-blot和PCR技術(shù)檢測(cè)EMT相關(guān)蛋白及相關(guān)轉(zhuǎn)錄因子的變化；其次,分析DAB2IP對(duì)調(diào)控轉(zhuǎn)移的關(guān)鍵信號(hào)通路的作用：最后,利用尾靜脈注射胰腺癌細(xì)胞建立的肺轉(zhuǎn)移模型,分析DAB2IP對(duì)腫瘤體內(nèi)轉(zhuǎn)移情況的影響。3.探討DAB2IP對(duì)胰腺癌化療敏感性的研究：首先,檢測(cè)常規(guī)一線化療藥物吉西他濱在胰腺癌中的IC50,以此濃度作用于上述兩組穩(wěn)定株細(xì)胞,檢測(cè)細(xì)胞活力的改變分析DAB2IP是否增強(qiáng)胰腺癌對(duì)吉西他濱藥物的敏感性；其次,我們檢測(cè)新型化療藥物鹽霉素的IC50,同樣作用于上述兩種穩(wěn)定株細(xì)胞分析DAB2IP與鹽霉素對(duì)胰腺癌細(xì)胞的活力的影響；然后,我們利用Realtime-PCR檢測(cè)DAB2IP與化療多重耐藥相關(guān)基因的作用；最后,我們利用Western-blot分析DAB2IP對(duì)胰腺癌干細(xì)胞相關(guān)標(biāo)記分子的影響。結(jié)果：1.DAB2IP抑制胰腺癌體內(nèi)外生長(zhǎng)情況：①利用免疫組化分析96例胰腺癌患者標(biāo)本中DAB2IP的表達(dá)情況顯示,胰腺癌組織中DAB2IP的表達(dá)明顯低于癌旁組織；124例胰腺癌患者臨床標(biāo)本中DAB2IP的表達(dá)與腫瘤的病理分級(jí)、淋巴結(jié)轉(zhuǎn)移的有無(wú)存在明顯的相關(guān)性：我們進(jìn)一步首次分析99例胰腺癌患者預(yù)后與DAB2IP表達(dá)的關(guān)系后發(fā)現(xiàn),DAB2IP的表達(dá)缺失是胰腺癌預(yù)后較差的重要預(yù)測(cè)因子。②檢測(cè)胰腺癌細(xì)胞檢測(cè)內(nèi)源性DAB2IP表達(dá)情況,轉(zhuǎn)染已有DAB2IP干擾質(zhì)粒及過(guò)表達(dá)質(zhì)粒構(gòu)建穩(wěn)定改變DAB2IP表達(dá)的細(xì)胞,發(fā)現(xiàn)DAB2IP可以抑制細(xì)胞增殖、促進(jìn)細(xì)胞凋亡,同時(shí)DAB2IP可以抑制細(xì)胞貼壁性及非貼壁性克隆的形成能力。③建立胰腺癌皮下移植瘤模型,發(fā)現(xiàn)DAB2IP可以抑制胰腺癌細(xì)胞體內(nèi)生長(zhǎng),其可能是通過(guò)誘導(dǎo)腫瘤組織細(xì)胞凋亡、抑制增殖來(lái)實(shí)現(xiàn)的。2.DAB2IP對(duì)胰腺癌體內(nèi)外轉(zhuǎn)移情況的影響：①細(xì)胞劃痕、Trans well侵襲遷移實(shí)驗(yàn)顯示,DAB2IP可以抑制胰腺癌細(xì)胞體外遷移和侵襲能力。②WB實(shí)驗(yàn)提示DAB2IP在胰腺癌可能通過(guò)抑制EMT的過(guò)程發(fā)揮抑制轉(zhuǎn)移的功能的,并與轉(zhuǎn)錄因子Slug表達(dá)改變密切相關(guān)：Luciferase實(shí)驗(yàn)進(jìn)一步驗(yàn)證了DAB2IP可以抑制Slug的轉(zhuǎn)錄活性。③檢測(cè)調(diào)控EMT發(fā)生的重要信號(hào)通路關(guān)鍵蛋白表達(dá)發(fā)現(xiàn),DAB2IP可以抑制STAT3、Akt的激活,并首次發(fā)現(xiàn)其可以抑制Smad2/3的磷酸化水平繼而參與TGF-β信號(hào)通路的調(diào)控。3.DAB2IP對(duì)胰腺癌化療敏感性的作用：①發(fā)現(xiàn)DAB2IP可以抑制胰腺癌細(xì)胞對(duì)一線化療藥物的吉西他濱的抗性,也可以協(xié)同促進(jìn)對(duì)新型化療藥物鹽霉素對(duì)胰腺癌細(xì)胞的敏感性。②DAB2IP可以抑制多重耐藥相關(guān)基因特別是ABCC1基因的表達(dá)。③DAB2IP可以抑制胰腺癌干性標(biāo)記分子,特別是CD133蛋白的表達(dá)。結(jié)論：DAB2IP作為一種腫瘤抑制基因,分別通過(guò)抑制胰腺癌的生長(zhǎng)、轉(zhuǎn)移、化療抗性等方面發(fā)揮作用,其可能與抑制Slug的轉(zhuǎn)錄或激活、ABCC1等耐藥基因、CD133等干性基因的表達(dá)有關(guān),可以為未來(lái)臨床治療胰腺癌提出新的思路。
[Abstract]:Objective : To investigate the role and mechanism in the treatment of pancreatic cancer with pancreatic cancer as one of the members of a family of RasGAP protein . The aim of this study was to investigate the effect and mechanism of the gene on growth , metastasis and chemotherapy resistance of pancreatic cancer .
then , the cell lines with different expression and low expression plasmids are respectively transfected with different cell lines of the endogenous expression of the pancreatic cancer , so that the cells with high stability and low expression are constructed ;
Secondly , cell proliferation , apoptosis experiment , plate cloning and soft agar cloning were used to prove its effect on the growth of pancreatic cancer cells in vitro .
Finally , by using the model of subcutaneous transplantation of pancreatic cancer , the influence on tumor growth was analyzed and the possible mechanism was analyzed .
Then , the changes of EMT - related protein and related transcription factors were detected by Western - blot and PCR .
Secondly , the key signal pathway for the control and metastasis of pancreatic cancer was analyzed by analyzing the influence on the metastasis of pancreatic cancer by using the model of lung metastasis established by the tail vein injection of pancreatic cancer .
Secondly , we tested the IC50 of the new chemotherapeutic drug salt , which was also applied to the analysis of the effects of the above two stable cell lines on the viability of pancreatic cancer cells .
Then , we used Realtime - PCR to detect the effect of multi - drug resistance related genes in vitro .
Finally , we used Western - blot to analyze the effect of the on - vitro growth of pancreatic cancer stem cells . Results : 1 . The growth of pancreatic cancer cells was inhibited by 1 . 2 IP .
In this paper , we can inhibit the proliferation of pancreatic cancer cells and inhibit the proliferation of pancreatic cancer cells .
【學(xué)位授予單位】：華中科技大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類號(hào)】：R735.9

【相似文獻(xiàn)】

相關(guān)博士學(xué)位論文 前1條

1 許琮;DAB2IP對(duì)胰腺癌生長(zhǎng)、轉(zhuǎn)移及化療耐藥的作用及機(jī)制研究[D];華中科技大學(xué);2016年

，

本文編號(hào)：2049410