本文關(guān)鍵詞：促炎癥消退介質(zhì)D類消退素對(duì)肺癌細(xì)胞遷移及侵襲力的影響　出處：《揚(yáng)州大學(xué)》2016年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 上皮細(xì)胞間質(zhì)轉(zhuǎn)化 D類消退素 M2型丙酮酸激酶 活性氧 肺癌

【摘要】：目的：觀察促炎癥消退介質(zhì)D類消退素AT-RvD1對(duì)肺癌細(xì)胞A549轉(zhuǎn)移能力的影響,同時(shí)探索核轉(zhuǎn)錄因子(Nrf2)及M2型丙酮酸激酶(Pkm2)在此過程中的作用,為更深入研究肺癌轉(zhuǎn)移過程和炎癥相關(guān)腫瘤的分子機(jī)制提供實(shí)驗(yàn)理論依據(jù)。方法：1、腫瘤的轉(zhuǎn)移與上皮細(xì)胞間質(zhì)轉(zhuǎn)化(EMT)的關(guān)系極其密切,利用不同濃度轉(zhuǎn)化生長(zhǎng)因子TGF-β1構(gòu)建EMT肺癌轉(zhuǎn)移模型,分別利用熒光探針技術(shù)檢測(cè)細(xì)胞內(nèi)活性氧(ROS)水平,MTT法檢測(cè)細(xì)胞增殖,劃痕實(shí)驗(yàn)測(cè)遷移,Transwell法測(cè)侵襲,同時(shí)利用實(shí)時(shí)熒光定量PCR技術(shù)和免疫印跡技術(shù)對(duì)上皮細(xì)胞表型E-鈣黏蛋白、間質(zhì)細(xì)胞表型波形蛋白、Nrf2及Pkm2的表達(dá)進(jìn)行檢測(cè)。2、利用不同濃度AT-RvD1和哺乳動(dòng)物雷帕霉素靶蛋白(mTOR)激活劑預(yù)刺激細(xì)胞30分鐘,觀察其對(duì)TGF-β1誘導(dǎo)的肺癌EMT過程的影響,分別比較細(xì)胞增殖性、遷移性,侵襲性的變化,檢測(cè)細(xì)胞內(nèi)ROS水平及相關(guān)基因蛋白的表達(dá)情況。結(jié)果：1、轉(zhuǎn)化生長(zhǎng)因子TGF-β1能明顯下調(diào)E-鈣黏蛋白和Nrf2的表達(dá),上調(diào)波形蛋白和Pkm2的表達(dá),增加細(xì)胞內(nèi)ROS水平并呈濃度依賴,增強(qiáng)A549細(xì)胞的遷移及侵襲能力,但對(duì)細(xì)胞增殖無(wú)明顯影響。2、D類消退素AT-RvD1能抑制TGF-β1誘導(dǎo)的EMT過程,上調(diào)E-鈣黏蛋白和下調(diào)波形蛋白的表達(dá),并且對(duì)A549細(xì)胞的遷移及侵襲能力有明顯的抑制作用,但不影響細(xì)胞的增殖,研究結(jié)果提示,AT-RvD1的抑制作用可能與下調(diào)Nrf2和Pkm2的表達(dá)相關(guān)。3、哺乳動(dòng)物雷帕霉素靶蛋白(mTOR)激活劑能逆轉(zhuǎn)AT-RvD1抑制Pkm2的作用,并且下調(diào)E-鈣黏蛋白的表達(dá)影響侵襲性,但尚未對(duì)細(xì)胞的增殖能力有明確影響。結(jié)論：1、D類消退素AT-RvD1能抑制TGF-β1誘導(dǎo)的肺癌EMT過程,抑制細(xì)胞的遷移及侵襲能力,但對(duì)細(xì)胞增殖無(wú)顯著影響。2、AT-RvD1可能部分通過抑制mTOR信號(hào)進(jìn)而下調(diào)Pkm2的表達(dá)來抑制TGF-β1誘導(dǎo)EMT過程,Nrf2表達(dá)降低可能與之相關(guān)。
[Abstract]:Objective: To observe the inflammatory resolution medium class D AT-RvD1 regression effect on metastasis of lung cancer cells A549, and explore the nuclear transcription factor (Nrf2) and type M2 pyruvate kinase (Pkm2) in this process, provide theoretical basis for further study on molecular mechanism of lung cancer metastasis related tumor and inflammation. Methods: 1, the metastasis of tumor and epithelial mesenchymal transition (EMT) extremely close relationship, transforming growth factor beta 1 TGF- to construct the EMT model of lung cancer metastasis using different concentrations of reactive oxygen, respectively using fluorescence probe technique in the detection of cell (ROS) level, MTT cell proliferation assay, scratch test migration, invasion test method Transwell at the same time, using real-time fluorescence quantitative PCR and Western blot on the epithelial cell phenotype of E- cadherin, interstitial cell phenotype of vimentin, expression of Nrf2 and Pkm2 were detected using different concentrations of.2, AT -RvD1 and mammalian target of rapamycin (mTOR) cell activator pre stimulation for 30 minutes, to observe the effects of TGF- beta 1 induced lung cancer EMT process were compared, cell proliferation, migration, changes of invasion, detection of intracellular ROS levels and relative gene expression. Results: 1, transformation the expression of growth factor beta 1 TGF- could downregulate E- cadherin and Nrf2, upregulation of the expression of vimentin and Pkm2, increase the level of ROS cells and in a concentration dependent enhancement, the migration and invasion of A549 cells, but had no obvious effect on cell proliferation of.2, D type of resolvin AT-RvD1 can inhibit TGF- beta 1 EMT by the upregulation of the expression of E- cadherin and downregulation of vimentin, and the migration and invasion ability of A549 cells was significantly inhibited, but does not affect cell proliferation, results suggest that the inhibition of AT-RvD1 might be associated with the down-regulation of Nrf Expression of.3 2 and Pkm2, the mammalian target of rapamycin (mTOR) activator AT-RvD1 can reverse the inhibitory effect of Pkm2, and the effects of down regulated expression of E- cadherin and aggressive, but there is a clear impact yet on cell proliferation. Conclusion: 1. D resolvin AT-RvD1 can inhibit TGF- beta 1 lung cancer EMT the process of induction, inhibition of migration and invasion of cells, but no significant effect of.2 on cell proliferation, AT-RvD1 inhibited TGF- induced EMT expression of beta 1 may part through inhibition of mTOR signaling and down-regulation of Pkm2, decreased Nrf2 expression may be associated with it.

【學(xué)位授予單位】：揚(yáng)州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類號(hào)】：R734.2

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