葡萄糖濃度對(duì)子宮內(nèi)膜癌細(xì)胞生長(zhǎng)的影響及其機(jī)制研究
發(fā)布時(shí)間：2018-08-29 18:38
【摘要】：目的:子宮內(nèi)膜癌(EC)是最常見的女性惡性腫瘤之一,其發(fā)病率和病死率逐年升高。肥胖是促使Ⅰ型EC進(jìn)展的高危因素,與EC的高病死率密切相關(guān)。多項(xiàng)研究結(jié)果顯示,細(xì)胞代謝異常和胰腺癌、結(jié)直腸癌和乳腺癌等多種腫瘤的發(fā)病密切相關(guān),與EC的關(guān)系也日益獲得關(guān)注。糖代謝異常是超重、肥胖和糖尿病的病理特征之一,也是EC發(fā)病的重要危險(xiǎn)因素。但糖代謝異常促進(jìn)EC發(fā)生的作用機(jī)制尚未明了。因此,在本研究中,我們應(yīng)用已建立的EC細(xì)胞系(Ishikawa、HEC-1-B),觀察在體外不同葡萄糖濃度培養(yǎng)條件下EC細(xì)胞增殖情況及其機(jī)制探討。方法:通過體外培養(yǎng)EC細(xì)胞株Ishikawa和HEC-1-B,用含不同糖濃度(無(wú)糖組:0mmol/L、低糖組:2.5mmol/L、正常糖濃度組:5.5mmol/L、高糖組:25.5mmol/L)的DMEM完全培養(yǎng)液處理細(xì)胞,使用CCK-8法來評(píng)價(jià)不同糖濃度對(duì)EC細(xì)胞增殖的影響;使用PI染色法和Annexin V/PI法檢測(cè)不同糖濃度對(duì)EC細(xì)胞周期和凋亡的影響;使用DCFH-DA熒光探針來測(cè)定不同糖濃度引起的EC細(xì)胞活性氧(ROS)的改變;使用ATP發(fā)光檢測(cè)試劑盒來檢測(cè)不同糖濃度對(duì)EC細(xì)胞產(chǎn)生ATP的影響;通過Western蛋白印跡實(shí)驗(yàn)檢測(cè)上述EC細(xì)胞在不同糖濃度干預(yù)下,AMPK和AKT/mTOR/S6信號(hào)通路相關(guān)蛋白的表達(dá)變化,以明確在不同的代謝環(huán)境下細(xì)胞代謝的信號(hào)轉(zhuǎn)導(dǎo)通路。結(jié)果:在不同糖濃度條件下培養(yǎng)EC細(xì)胞系Ishikawa和HEC-1-B,發(fā)現(xiàn)2株EC細(xì)胞的增殖均依賴于培養(yǎng)基中的葡萄糖濃度,且呈劑量依賴模式;低糖濃度通過G1期阻滯抑制Ishikawa和HEC-1-B細(xì)胞增殖,高糖濃度抑制細(xì)胞凋亡;低糖濃度改變糖酵解活性,引起Ishikawa和HEC-1-B細(xì)胞ATP生成減少,ROS產(chǎn)生增加;Western蛋白印跡實(shí)驗(yàn)結(jié)果顯示,在Ishikawa和HEC-1-B細(xì)胞中,低糖增加p-AMPK的表達(dá),降低p-AKT及p-S6的表達(dá),證實(shí)糖代謝異常通過調(diào)節(jié)AMPK/mTOR和Akt/mTOR/S6信號(hào)通路促進(jìn)EC細(xì)胞增殖。結(jié)論:發(fā)現(xiàn)EC細(xì)胞的增殖以劑量依賴性方式依賴于培養(yǎng)基中的葡萄糖濃度,糖代謝異常通過調(diào)節(jié)AMPK/mTOR和Akt/mTOR/S6信號(hào)通路促進(jìn)EC細(xì)胞增殖,靶向糖代謝異常及其相關(guān)信號(hào)通路可能為肥胖型EC患者個(gè)體化治療提供新思路。
[Abstract]:Objective: (EC) is one of the most common malignant tumors in women. Obesity is a high risk factor for the progression of type I EC and is closely related to the high mortality of EC. Many studies have shown that abnormal cell metabolism is closely related to pancreatic cancer, colorectal cancer, breast cancer and other tumors, and the relationship with EC has been paid more and more attention. Abnormal glucose metabolism is one of the pathological features of overweight, obesity and diabetes, and is also an important risk factor of EC. However, the mechanism of abnormal glucose metabolism promoting the occurrence of EC is not clear. Therefore, in this study, we used the established EC cell line (Ishikawa,HEC-1-B) to observe the proliferation of EC cells under different glucose concentrations in vitro and its mechanism. Methods: EC cell lines Ishikawa and HEC-1-B, were treated with DMEM complete culture medium containing different sugar concentrations (no sugar group: 0 mmol / L, low sugar group: 2.5 mmol / L, normal glucose concentration group: 5. 5 mmol / L, high glucose group: 25. 5 mmol / L). The effect of different glucose concentrations on the proliferation of EC cells was evaluated by CCK-8 method. PI staining and Annexin V/PI assay were used to detect the effect of different glucose concentrations on the cell cycle and apoptosis of EC cells, and DCFH-DA fluorescence probe was used to detect the changes of reactive oxygen (ROS) (Ros) in EC cells induced by different glucose concentrations. The effects of different glucose concentrations on the production of ATP in EC cells were detected by ATP luminescence assay kit, and the expression of AMPK and AKT/mTOR/S6 signaling pathway related proteins in EC cells were detected by Western Western blot. In order to clarify the signal transduction pathway of cell metabolism in different metabolic environment. Results: the proliferation of EC cell lines Ishikawa and HEC-1-B, in different glucose concentration was found to be dependent on glucose concentration in culture medium, and low glucose concentration inhibited the proliferation of Ishikawa and HEC-1-B cells through G1 phase arrest. High glucose concentration inhibited cell apoptosis, low glucose concentration changed glycolysis activity, reduced ATP production in Ishikawa and HEC-1-B cells, and increased Ros production. Western blot analysis showed that low glucose increased p-AMPK expression in Ishikawa and HEC-1-B cells. Decreasing the expression of p-AKT and p-S6 showed that abnormal glucose metabolism promoted the proliferation of EC cells by regulating the signal pathway of AMPK/mTOR and Akt/mTOR/S6. Conclusion: the proliferation of EC cells depends on glucose concentration in a dose-dependent manner. Abnormal glucose metabolism promotes the proliferation of EC cells by regulating AMPK/mTOR and Akt/mTOR/S6 signaling pathways. The abnormal glucose metabolism and its related signaling pathway may provide a new idea for individual treatment of obese EC patients.
【學(xué)位授予單位】：濟(jì)南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R737.33
本文編號(hào)：2212065
[Abstract]:Objective: (EC) is one of the most common malignant tumors in women. Obesity is a high risk factor for the progression of type I EC and is closely related to the high mortality of EC. Many studies have shown that abnormal cell metabolism is closely related to pancreatic cancer, colorectal cancer, breast cancer and other tumors, and the relationship with EC has been paid more and more attention. Abnormal glucose metabolism is one of the pathological features of overweight, obesity and diabetes, and is also an important risk factor of EC. However, the mechanism of abnormal glucose metabolism promoting the occurrence of EC is not clear. Therefore, in this study, we used the established EC cell line (Ishikawa,HEC-1-B) to observe the proliferation of EC cells under different glucose concentrations in vitro and its mechanism. Methods: EC cell lines Ishikawa and HEC-1-B, were treated with DMEM complete culture medium containing different sugar concentrations (no sugar group: 0 mmol / L, low sugar group: 2.5 mmol / L, normal glucose concentration group: 5. 5 mmol / L, high glucose group: 25. 5 mmol / L). The effect of different glucose concentrations on the proliferation of EC cells was evaluated by CCK-8 method. PI staining and Annexin V/PI assay were used to detect the effect of different glucose concentrations on the cell cycle and apoptosis of EC cells, and DCFH-DA fluorescence probe was used to detect the changes of reactive oxygen (ROS) (Ros) in EC cells induced by different glucose concentrations. The effects of different glucose concentrations on the production of ATP in EC cells were detected by ATP luminescence assay kit, and the expression of AMPK and AKT/mTOR/S6 signaling pathway related proteins in EC cells were detected by Western Western blot. In order to clarify the signal transduction pathway of cell metabolism in different metabolic environment. Results: the proliferation of EC cell lines Ishikawa and HEC-1-B, in different glucose concentration was found to be dependent on glucose concentration in culture medium, and low glucose concentration inhibited the proliferation of Ishikawa and HEC-1-B cells through G1 phase arrest. High glucose concentration inhibited cell apoptosis, low glucose concentration changed glycolysis activity, reduced ATP production in Ishikawa and HEC-1-B cells, and increased Ros production. Western blot analysis showed that low glucose increased p-AMPK expression in Ishikawa and HEC-1-B cells. Decreasing the expression of p-AKT and p-S6 showed that abnormal glucose metabolism promoted the proliferation of EC cells by regulating the signal pathway of AMPK/mTOR and Akt/mTOR/S6. Conclusion: the proliferation of EC cells depends on glucose concentration in a dose-dependent manner. Abnormal glucose metabolism promotes the proliferation of EC cells by regulating AMPK/mTOR and Akt/mTOR/S6 signaling pathways. The abnormal glucose metabolism and its related signaling pathway may provide a new idea for individual treatment of obese EC patients.
【學(xué)位授予單位】：濟(jì)南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R737.33
【參考文獻(xiàn)】
相關(guān)期刊論文 前2條
1 JI Cheng Ye;CHEN Tian Jiao;;Empirical Changes in the Prevalence of Overweight and Obesity among Chinese Students from 1985 to 2010 and Corresponding Preventive Strategies[J];Biomedical and Environmental Sciences;2013年01期
2 楊華;譚先杰;郎景和;;代謝綜合征與子宮內(nèi)膜癌相關(guān)性研究進(jìn)展[J];中國(guó)實(shí)用婦科與產(chǎn)科雜志;2012年11期
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