本文關鍵詞：促炎癥消退介質D類消退素對肺癌細胞遷移及侵襲力的影響　出處：《揚州大學》2016年碩士論文　論文類型：學位論文

  更多相關文章： 上皮細胞間質轉化 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D類消退素AT-RvD1對肺癌細胞A549轉移能力的影響,同時探索核轉錄因子(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移過程和炎癥相關腫瘤的分子機制提供實驗理論依據(jù)。方法：1、腫瘤的轉移與上皮細胞間質轉化(EMT)的關系極其密切,利用不同濃度轉化生長因子TGF-β1構建EMT肺癌轉移模型,分別利用熒光探針技術檢測細胞內(nèi)活性氧(ROS)水平,MTT法檢測細胞增殖,劃痕實驗測遷移,Transwell法測侵襲,同時利用實時熒光定量PCR技術和免疫印跡技術對上皮細胞表型E-鈣黏蛋白、間質細胞表型波形蛋白、Nrf2及Pkm2的表達進行檢測。2、利用不同濃度AT-RvD1和哺乳動物雷帕霉素靶蛋白(mTOR)激活劑預刺激細胞30分鐘,觀察其對TGF-β1誘導的肺癌EMT過程的影響,分別比較細胞增殖性、遷移性,侵襲性的變化,檢測細胞內(nèi)ROS水平及相關基因蛋白的表達情況。結果：1、轉化生長因子TGF-β1能明顯下調E-鈣黏蛋白和Nrf2的表達,上調波形蛋白和Pkm2的表達,增加細胞內(nèi)ROS水平并呈濃度依賴,增強A549細胞的遷移及侵襲能力,但對細胞增殖無明顯影響。2、D類消退素AT-RvD1能抑制TGF-β1誘導的EMT過程,上調E-鈣黏蛋白和下調波形蛋白的表達,并且對A549細胞的遷移及侵襲能力有明顯的抑制作用,但不影響細胞的增殖,研究結果提示,AT-RvD1的抑制作用可能與下調Nrf2和Pkm2的表達相關。3、哺乳動物雷帕霉素靶蛋白(mTOR)激活劑能逆轉AT-RvD1抑制Pkm2的作用,并且下調E-鈣黏蛋白的表達影響侵襲性,但尚未對細胞的增殖能力有明確影響。結論：1、D類消退素AT-RvD1能抑制TGF-β1誘導的肺癌EMT過程,抑制細胞的遷移及侵襲能力,但對細胞增殖無顯著影響。2、AT-RvD1可能部分通過抑制mTOR信號進而下調Pkm2的表達來抑制TGF-β1誘導EMT過程,Nrf2表達降低可能與之相關。
[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.

【學位授予單位】：揚州大學
【學位級別】：碩士
【學位授予年份】：2016
【分類號】：R734.2

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