本文選題：宮頸腺癌 + TGF-β1　； 參考：《瀘州醫(yī)學(xué)院》2014年碩士論文

【摘要】：目的：宮頸癌是女性婦科三大惡性腫瘤之一，宮頸腺癌是宮頸癌第二多見(jiàn)的組織學(xué)類型，約占宮頸癌的10%，宮頸腺癌較宮頸鱗癌更易于發(fā)生轉(zhuǎn)移，預(yù)后差。近年來(lái)研究發(fā)現(xiàn)上皮間質(zhì)轉(zhuǎn)化（Epithelial-MesenchymalTransitions，EMT）與腫瘤的發(fā)生發(fā)展有重要聯(lián)系。轉(zhuǎn)化生長(zhǎng)因子β1（Transforming growth factor beta1，TGF-β1）已被證實(shí)是上皮間質(zhì)轉(zhuǎn)化的重要誘導(dǎo)劑，本實(shí)驗(yàn)擬通過(guò)體外培養(yǎng)人宮頸腺癌Hela細(xì)胞，在TGF-β1誘導(dǎo)下體外構(gòu)建Hela細(xì)胞上皮間質(zhì)轉(zhuǎn)化模型，研究其對(duì)Hela細(xì)胞增殖、凋亡及遷移能力的影響。方法：體外培養(yǎng)人宮頸腺癌Hela細(xì)胞，分為實(shí)驗(yàn)組：用不同濃度TGF-β1（0.01、0.1、1、10ng/ml）刺激Hela細(xì)胞，對(duì)照組：用不含TGF-β1的無(wú)血清培養(yǎng)基培養(yǎng)Hela細(xì)胞。①用不同濃度TGF-β1（0.01、0.1、1、10ng/ml）刺激Hela細(xì)胞后于不同時(shí)間點(diǎn)觀察其細(xì)胞形態(tài)變化；②不同濃度TGF-β1刺激Hela細(xì)胞72h后，用半定量RT-PCR法檢測(cè)上皮間質(zhì)轉(zhuǎn)化相關(guān)標(biāo)記物E-cadherin及Vimentin mRNA表達(dá)情況；③不同濃度TGF-β1刺激Hela細(xì)胞后，細(xì)胞免疫組織化學(xué)方法檢測(cè)E-cadherin，Vimentin蛋白的表達(dá)變化；④用CCK8法檢測(cè)不同濃度TGF-β1刺激Hela細(xì)胞后對(duì)其增殖影響；⑤用流式細(xì)胞儀檢測(cè)不同濃度TGF-β1刺激Hela細(xì)胞72h時(shí)對(duì)其凋亡的影響；⑥用10ng/ml TGF-β1刺激Hela細(xì)胞后用劃痕實(shí)驗(yàn)檢測(cè)其細(xì)胞遷移能力的變化。結(jié)果：①倒置顯微鏡下觀察到不同濃度TGF-β1刺激Hela細(xì)胞24h時(shí)與對(duì)照組相比形態(tài)未發(fā)生明顯變化；刺激48h時(shí)就開始出現(xiàn)變化，細(xì)胞伸出許多小突出，部分失去多邊形或立方形形態(tài)變?yōu)樗笮危淮碳?2h后變化更明顯，與對(duì)照組比較，實(shí)驗(yàn)組大部分細(xì)胞呈現(xiàn)明顯的間質(zhì)細(xì)胞形態(tài)，表現(xiàn)為細(xì)胞拉長(zhǎng)呈梭形或紡錘形，細(xì)胞的間隙變寬，細(xì)胞與細(xì)胞之間的連接變得松散，細(xì)胞由鋪路石樣分布變?yōu)椴灰?guī)則的分布；②RT-PCR檢測(cè)結(jié)果顯示：TGF-β1刺激后，Hela細(xì)胞的上皮標(biāo)記物E-cadherin mRNA表達(dá)下調(diào)，間質(zhì)標(biāo)記物Vimentin mRNA表達(dá)上調(diào)且均呈濃度依賴性，，與對(duì)照組相比差異有統(tǒng)計(jì)學(xué)差異（P0.05）；③細(xì)胞免疫組織化學(xué)結(jié)果顯示：TGF-β1刺激后，隨著TGF-β1濃度的增加E-cadherin蛋白的表達(dá)逐漸降低，Vimentin蛋白的表達(dá)則逐漸升高，與對(duì)照組相比差異有統(tǒng)計(jì)學(xué)意義（P0.05）；④CCK-8檢測(cè)結(jié)果顯示：TGF-β1刺激后，24h時(shí)不同濃度TGF-β1對(duì)Hela細(xì)胞增殖無(wú)明顯作用，與對(duì)照組相比差異無(wú)統(tǒng)計(jì)學(xué)意義（P0.05），48h和72h后對(duì)Hela細(xì)胞生長(zhǎng)增殖起抑制作用，與對(duì)照組相比差異有統(tǒng)計(jì)學(xué)意義（P0.05），且呈時(shí)間和濃度依賴性；⑤流式細(xì)胞儀檢測(cè)結(jié)果顯示：TGF-β1刺激人宮頸腺癌Hela細(xì)胞72h時(shí)，隨著TGF-β1濃度的增加Hela細(xì)胞的凋亡率也逐漸增加，與對(duì)照組相比差異有統(tǒng)計(jì)學(xué)意義（P0.05）；⑥10ng/ml TGF-β1刺激Hela細(xì)胞后其遷移能力增強(qiáng)。結(jié)論：①TGF-β1可以誘導(dǎo)人宮頸腺癌Hela細(xì)胞發(fā)生上皮間質(zhì)轉(zhuǎn)化；②TGF-β1對(duì)人宮頸腺癌Hela細(xì)胞的生長(zhǎng)增殖起抑制作用并能誘導(dǎo)其發(fā)生凋亡；③TGF-β1可增強(qiáng)子宮頸腺癌Hela細(xì)胞的遷移能力。
[Abstract]:Objective : To investigate the effect of transforming growth factor - 尾1 ( 0.01 , 0.1 , 1 , 10 ng / ml ) on Hela cells .
( 2 ) The expression of E - cadherin and vimentin was detected by semi - quantitative RT - PCR after 72 h of Hela cells stimulated by TGF - 尾1 at different concentrations .
( 3 ) After stimulated Hela cells with different concentrations of TGF - 尾1 , the expression of E - cadherin and Vimentin was detected by immunohistochemical method .
( 4 ) The effect of TGF - 尾1 stimulated Hela cells on the proliferation of Hela cells was detected by CC8 method .
( 5 ) The effect of TGF - 尾1 stimulated by different concentrations of TGF - 尾1 on the apoptosis of Hela cells was detected by flow cytometry .
( 6 ) After stimulated Hela cells with 10 ng / ml TGF - 尾1 , the changes of cell migration ability were detected by scratch test . Results : 鈶

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