本文選題：鼻咽癌 + 放射治療　； 參考：《大連大學(xué)》2017年碩士論文

【摘要】：目的:探索IGF-1R抑制劑OSI-906增強(qiáng)鼻咽癌細(xì)胞(NPC)放射敏感性的作用機(jī)制。方法:應(yīng)用蛋白免疫印跡法(Western blot)檢測X線照射(IR)后5種鼻咽癌細(xì)胞pIGF-1R基礎(chǔ)表達(dá)水平;應(yīng)用四甲基偶氮唑藍(lán)(MTT)實(shí)驗(yàn)觀察單獨(dú)應(yīng)用胰島素樣生長因子-1(IGF-1)、IGF-1R抑制劑OSI-906(Linsitinib)以及IGF-1聯(lián)合OSI-906對NPC增殖的影響;應(yīng)用Western blot檢測IR、IGF-1、OSI-906、OSI-906聯(lián)合IGF-1、OSI-906聯(lián)合IR后IGF-1/IGF-1R信號轉(zhuǎn)導(dǎo)通路和相關(guān)蛋白表達(dá)活化情況;應(yīng)用流式細(xì)胞術(shù)(FACS)檢測單獨(dú)應(yīng)用OSI-906、IR、OSI-906聯(lián)合IR后NPC周期和凋亡情況;應(yīng)用細(xì)胞克隆形成實(shí)驗(yàn)觀察不同IR后IGF-1、OSI-906單獨(dú)應(yīng)用以及OSI-906聯(lián)合IGF-1后細(xì)胞克隆形成的差異;最后應(yīng)用gH2AX焦點(diǎn)成型實(shí)驗(yàn),觀察單獨(dú)應(yīng)用OSI-906和OSI-906聯(lián)合X線照射后細(xì)胞DNA損傷情況。以上實(shí)驗(yàn)數(shù)據(jù)采用GraphPad Prism 5進(jìn)行統(tǒng)計(jì)學(xué)分析。結(jié)果:在鼻咽癌細(xì)胞系中,pIGF-1R的表達(dá)水平高低不一,其中CNE-2、SUNE-1細(xì)胞pIGF-1R蛋白表達(dá)屬于中游水平,且對OSI-906敏感性較高,作為本研究對象。MTT結(jié)果提示,IGF-1促進(jìn)NPC增殖,OSI-906明顯抑制細(xì)胞增殖,且兩者的作用呈劑量依賴性。同時,OSI-906能夠逆轉(zhuǎn)IGF-1的促增殖作用;Western blot結(jié)果顯示,IR、IGF-1和OSI-906分別激活和抑制IGF-1/IGF-1R增殖信號轉(zhuǎn)導(dǎo)通路,pIGF-1R、pAKT、pERK表達(dá)明顯增高和降低,總IGF-1R、AKT、ERK蛋白均無明顯變化;同時,OSI-906可以逆轉(zhuǎn)IGF-1、IR介導(dǎo)的IGF-1/IGF-1R信號通路的激活;細(xì)胞周期結(jié)果顯示,單獨(dú)應(yīng)用OSI-906、IR后細(xì)胞G2/M期比例增加不明顯,且細(xì)胞S期比例升高,而OSI-906聯(lián)合X線照射后顯著增加細(xì)胞G2/M期比例,細(xì)胞S期比例降低;單獨(dú)應(yīng)用OSI-906、IR可以促進(jìn)細(xì)胞凋亡,OSI-906聯(lián)合IR后進(jìn)一步增加細(xì)胞凋亡率;克隆形成實(shí)驗(yàn)結(jié)果表明,IR后,IGF-1提高了NPC的生存數(shù),而OSI-906明顯地降低了NPC的生存數(shù)并且可以逆轉(zhuǎn)IGF-1的作用;gH2AX焦點(diǎn)成形實(shí)驗(yàn)提示OSI-906可以顯著增加X線照射細(xì)胞的gH2AX焦點(diǎn)數(shù),細(xì)胞DNA斷裂蛋白標(biāo)記物gH2AX表達(dá)升高。結(jié)論:OSI-906增加NPC放射敏感性,其機(jī)制可能與抑制細(xì)胞增殖信號傳導(dǎo)通路的激活、減少DNA損傷的修復(fù)、改變細(xì)胞周期、促進(jìn)細(xì)胞凋亡、誘導(dǎo)細(xì)胞基因組不穩(wěn)定性、且逆轉(zhuǎn)IGF-1介導(dǎo)的鼻咽癌放射抵抗有關(guān)。
[Abstract]:Objective: to explore the mechanism of IGF-1R inhibitor OSI-906 in enhancing radiosensitivity of nasopharyngeal carcinoma (NPC) cells. Methods: Western blotting was used to detect the basic expression of pIGF-1R in 5 kinds of nasopharyngeal carcinoma cells after X-ray irradiation. The effects of insulin-like growth factor-1 IGF-1R inhibitor OSI-906 Linsitinib and IGF-1 combined with OSI-906 on the proliferation of NPC were observed by MTT. Western blot was used to detect the activation of IGF-1/IGF-1R signal transduction pathway and related protein expression after IRI-906 combined with IGF-1OSI-906 combined with IR, and flow cytometry was used to detect NPC cycle and apoptosis after using OSI-906 combined with IR alone. The difference of cell clone formation between IGF-1OSI-906 and OSI-906 combined with IGF-1 was observed by cell clone formation assay, and the cell DNA damage after OSI-906 and OSI-906 combined with X-ray irradiation was observed by gH2AX focus molding experiment. The above experimental data were statistically analyzed by GraphPad Prism 5. Results: the expression level of pIGF-1R in nasopharyngeal carcinoma cell line was different, and the expression of pIGF-1R protein in CNE-2SUNE-1 cell line was middle level, and the sensitivity to OSI-906 was high. The results showed that IGF-1 promoted NPC proliferation and OSI-906 significantly inhibited the proliferation of NPC cells. The effect of both was dose dependent. At the same time, OSI-906 could reverse the proliferative effect of IGF-1. The results of Western blot showed that IRI IGF-1 and OSI-906 activated and inhibited the expression of pIGF-1RnpAK-pERK respectively, but the total IGF-1RnAK-AKTERK protein did not change. At the same time, OSI-906 could reverse the activation of IGF-1IR mediated IGF-1/IGF-1R signaling pathway, and the cell cycle results showed that the proportion of G _ 2 / M phase was not significantly increased and the S phase ratio of cells was increased after the treatment of OSI-906 IR alone, while the ratio of G _ 2 / M phase was significantly increased after OSI-906 combined with X-ray irradiation. The ratio of S phase was decreased, the apoptosis rate was further increased by OSI-906 combined with IR alone, the results of clone formation showed that IGF-1 increased the survival rate of NPC after IR. However, OSI-906 significantly reduced the survival of NPC and reversed the role of IGF-1. The results showed that OSI-906 could significantly increase the number of gH2AX focal points of the cells irradiated by X-ray, and the gH2AX expression of DNA break protein marker was increased. Conclusion the mechanism of NPC radiosensitivity may be related to inhibiting the activation of cell proliferation signal transduction pathway, reducing the repair of DNA damage, changing cell cycle, promoting cell apoptosis and inducing cell genomic instability. And reversal of IGF-1-mediated radiation resistance in nasopharyngeal carcinoma.
【學(xué)位授予單位】：大連大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R739.63

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相關(guān)期刊論文 前3條

1 Shih-Hung Yang;Ting-Chun Kuo;Hsu Wu;Jhe-Cyuan Guo;Chiun Hsu;Chih-Hung Hsu;Yu-Wen Tien;Kun-Huei Yeh;Ann-Lii Cheng;Sung-Hsin Kuo;;Perspectives on the combination of radiotherapy and targeted therapy with DNA repair inhibitors in the treatment of pancreatic cancer[J];World Journal of Gastroenterology;2016年32期

2 Wei Chen;Guo-Hua Hu;;Biomarkers for enhancing the radiosensitivity of nasopharyngeal carcinoma[J];Cancer Biology & Medicine;2015年01期

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本文編號：1811402