本文關(guān)鍵詞： 肺癌 高糖 高脂 侵襲轉(zhuǎn)移　出處：《北京協(xié)和醫(yī)學(xué)院》2017年博士論文　論文類型：學(xué)位論文

【摘要】：背景肺癌是一種患病率和死亡率均較高的惡性疾病,影響肺癌患者預(yù)后的因素包括TNM分期、臨床參數(shù)、分子學(xué)特征等。有臨床研究顯示代謝因素如合并有糖尿病、血脂異常也可能影響肺癌患者的預(yù)后。考慮到代謝性疾病在人群中的高患病率,我們十分有必要明確代謝因素對(duì)肺癌細(xì)胞侵襲轉(zhuǎn)移能力的影響。材料和方法本研究使用非小細(xì)胞肺癌細(xì)胞系A(chǔ)549,使用含不同濃度葡萄糖的培養(yǎng)基以及含不同種類不同濃度脂肪酸的培養(yǎng)基以分別模擬高糖和高脂環(huán)境來培養(yǎng)A549細(xì)胞。通過transwell實(shí)驗(yàn)和檢測(cè)細(xì)胞轉(zhuǎn)移相關(guān)蛋白MMP9和CD147表達(dá)水平的變化來反映高糖或高脂培養(yǎng)后A549細(xì)胞侵襲轉(zhuǎn)移能力的變化。以MTS實(shí)驗(yàn)檢測(cè)高糖或高脂對(duì)A549細(xì)胞活性的影響。之后使用高通量轉(zhuǎn)錄組測(cè)序技術(shù)研究高糖和高脂條件處理后A549細(xì)胞轉(zhuǎn)錄組的變化,分析這兩種條件影響A549細(xì)胞侵襲轉(zhuǎn)移能力和細(xì)胞活性的背后機(jī)制。結(jié)果我們通過transwell實(shí)驗(yàn)發(fā)現(xiàn)高葡萄糖環(huán)境使A549細(xì)胞侵襲能力增強(qiáng),western blot實(shí)驗(yàn)顯示高糖培養(yǎng)使A549細(xì)胞轉(zhuǎn)移相關(guān)蛋白MMP9、CD147的表達(dá)稍有增加,但變化不顯著。棕櫚酸和油酸會(huì)減弱A549細(xì)胞的侵襲能力,亞油酸和A549細(xì)胞侵襲能力無明顯相關(guān)。這3種脂肪酸中棕櫚酸可使A549細(xì)胞轉(zhuǎn)移相關(guān)蛋白MMP9和CD147表達(dá)顯著減少,而不同濃度的油酸和亞油酸未使A549細(xì)胞轉(zhuǎn)移相關(guān)蛋白的表達(dá)發(fā)生顯著變化。另外棕櫚酸培養(yǎng)使A549細(xì)胞活性降低,而高糖培養(yǎng)以及油酸和亞油酸在400μmol/L以內(nèi)培養(yǎng)一代細(xì)胞,細(xì)胞活性無顯著變化。之后我們對(duì)不同濃度葡萄糖和不同濃度棕櫚酸處理過的A549細(xì)胞進(jìn)行轉(zhuǎn)錄組測(cè)序,在前者的3個(gè)樣本中找到梯度表達(dá)的差異表達(dá)基因396個(gè),主要富集在細(xì)胞粘附、應(yīng)激反應(yīng)、有絲分裂等過程和NO-cGMP-PKG、腫瘤相關(guān)蛋白多糖通路等。在后者的4個(gè)樣本中找到梯度表達(dá)的差異表達(dá)基因254個(gè),主要富集在細(xì)胞凋亡、細(xì)胞遷移、炎癥反應(yīng)等過程和凋亡、炎癥反應(yīng)、MAPK、粘附相關(guān)信號(hào)通路。結(jié)論高糖環(huán)境使A549細(xì)胞侵襲轉(zhuǎn)移能力增強(qiáng)。使用棕櫚酸模擬高脂環(huán)境可以使A549細(xì)胞侵襲轉(zhuǎn)移能力和細(xì)胞活性均下降。油酸也可能使A549細(xì)胞侵襲轉(zhuǎn)移能力下降,而亞油酸對(duì)A549細(xì)胞的侵襲轉(zhuǎn)移能力無顯著影響。轉(zhuǎn)錄組測(cè)序分析顯示高糖環(huán)境對(duì)A549細(xì)胞的影響可能是通過細(xì)胞粘附、NO-cGMP-PKG、腫瘤相關(guān)蛋白多糖通路,棕櫚酸對(duì)A549細(xì)胞的影響可能是通過凋亡、炎癥、MAPK、粘附相關(guān)通路。
[Abstract]:Background Lung cancer is a malignant disease with high morbidity and mortality. The prognostic factors of lung cancer include TNM staging and clinical parameters. Molecular characteristics. Clinical studies have shown that metabolic factors such as diabetes mellitus, dyslipidemia may also affect the prognosis of lung cancer patients. Considering the high prevalence of metabolic diseases in the population. It is necessary for us to clarify the effect of metabolic factors on the invasion and metastasis of lung cancer cells. Materials and methods the non-small cell lung cancer cell line A549 was used in this study. A549 cells were cultured in medium containing different concentrations of glucose and different kinds of fatty acids to simulate high glucose and high fat environment respectively. Transwell experiments and detection were carried out. The expression levels of MMP9 and CD147 in A549 cells were detected by MTS assay. The changes of invasion and metastasis ability of A549 cells after high glucose or high fat culture were detected by MTS assay. High throughput transcriptome sequencing was used to study the transcriptional changes of A549 cells treated with high glucose and high fat. The underlying mechanism of these two conditions affecting the invasion and metastasis ability and cell activity of A549 cells was analyzed. Results We found that high glucose environment enhanced the invasion ability of A549 cells by transwell assay. . Western blot assay showed that high glucose culture increased the expression of metastasis associated protein MMP9 / CD147 in A549 cells. Palmitic acid and oleic acid decreased the invasion ability of A549 cells. There was no significant correlation between linoleic acid and invasion ability of A549 cells. Palmitic acid could significantly reduce the expression of MMP9 and CD147 in A549 cells. However, different concentrations of oleic acid and linoleic acid did not significantly change the expression of metastasis related protein in A549 cells. In addition, palmitic acid culture decreased the activity of A549 cells. The cells were cultured in high glucose culture, oleic acid and linoleic acid within 400 渭 mol/L. There was no significant change in cell activity. Then we sequenced the transcriptional sequence of A549 cells treated with different concentrations of glucose and palmitic acid. A total of 396 differentially expressed genes were found in the three samples of the former, mainly concentrated in cell adhesion, stress response, mitosis and NO-cGMP-PKG. In the latter 4 samples, 254 differentially expressed genes were found, mainly concentrated in apoptosis, cell migration, inflammatory reaction and apoptosis, inflammatory reaction. MAPK. Conclusion High glucose environment can enhance the invasion and metastasis ability of A549 cells. Using palmitic acid to simulate high fat environment can decrease the invasion and metastasis ability and cell activity of A549 cells. Oleic acid can also increase the invasion and metastasis ability of A549 cells. The ability of invasion and metastasis of A549 cells was decreased. However, linoleic acid had no significant effect on the invasion and metastasis of A549 cells. Transcriptome sequencing showed that the effect of high glucose environment on A549 cells was probably through cell adhesion to NO-cGMP-PKG. The effect of palmitic acid on A549 cells may be mediated by apoptosis, inflammatory MAPK and adhesion pathway.
【學(xué)位授予單位】：北京協(xié)和醫(yī)學(xué)院
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：R734.2

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