本文選題：OSM + 腺病毒��； 參考：《蘇州大學(xué)》2012年碩士論文

【摘要】：研究背景及目的：鼻咽癌(nasopharyngeal carcinoma，NPC)是我國南方、越南、菲律賓等其它南亞國家高發(fā)腫瘤之一，廣東、廣西、福建、湖南等省為國內(nèi)高發(fā)區(qū)，，部分地區(qū)高達(dá)30/10萬～50/10萬，調(diào)整死亡率分別為6.47/10萬、4.92/10萬、3.28/10萬、3.22/10萬，亦居世界首位（調(diào)整死亡率2.88/10萬）。男性發(fā)病率約為女性的2～3倍，40～50歲為高發(fā)年齡組。鼻咽癌的發(fā)病率和死亡率均隨年齡增長(zhǎng)而上升。鼻咽癌的發(fā)生發(fā)展是一個(gè)多因素作用、多基因參與、多階段病變發(fā)生的極其復(fù)雜的癌變過程，基因治療作為一種高效性、特異性、靶向性的治療手段已是醫(yī)學(xué)界廣泛關(guān)注的熱點(diǎn)。近年來研究發(fā)現(xiàn)，抑瘤M（Oncostatin M,OSM）基因?qū)δ[瘤細(xì)胞具有生長(zhǎng)抑制作用和靶向誘導(dǎo)凋亡的功能。本項(xiàng)實(shí)驗(yàn)擬采用以腺病毒為載體的OSM基因，觀察其對(duì)該鼻咽癌細(xì)胞及裸鼠移植瘤的抑癌效應(yīng)，并對(duì)其可能機(jī)理進(jìn)行了探討，為將來運(yùn)用Ad OSM應(yīng)用于臨床提供實(shí)驗(yàn)依據(jù)。 方法：體外：將Ad-OSM體外感染CNE-2Z鼻咽癌細(xì)胞，運(yùn)用RT-PCR及Westernblot檢測(cè)OSM基因在CNE-2Z細(xì)胞中的轉(zhuǎn)錄及表達(dá)；MTT法和Annexin-V-PE/7-AAD雙染色的FCM檢測(cè)PBS、Ad-GFP、Ad-OSM體外抑制鼻咽癌細(xì)胞生長(zhǎng)和誘導(dǎo)凋亡的抑癌增效效應(yīng)；PI染色的FCM檢測(cè)Ad-OSM對(duì)鼻咽癌細(xì)胞周期的影響；運(yùn)用RT-PCR檢測(cè)P21、P27、Bcl-2、Survivin等以分析Ad-OSM誘導(dǎo)腫瘤細(xì)胞凋亡可能的分子機(jī)制；Western blot檢測(cè)Ad-OSM對(duì)CNE-2Z細(xì)胞中cleaved Caspase-3的表達(dá)的影響。體內(nèi)：用CNE-2Z細(xì)胞構(gòu)建裸鼠移植瘤，在鼻咽癌的裸鼠移植瘤動(dòng)物模型上，運(yùn)用免疫組化方法檢測(cè)CNE-2Z鼻咽癌移植瘤生長(zhǎng)相關(guān)因子的表達(dá)：（1）P21、P53、Bax、Caspase-3、等促凋亡因子；（2）Bcl-2、Cox-2等凋亡抑制因子。 結(jié)果：RT-PCR和Western blot檢測(cè)證實(shí)腺病毒介導(dǎo)的OSM基因可以在CNE-2Z細(xì)胞中穩(wěn)定轉(zhuǎn)錄及表達(dá)；Ad-OSM體外能明顯抑制CNE-2Z鼻咽癌癌細(xì)胞的生長(zhǎng)，引起G1期阻滯和誘導(dǎo)細(xì)胞凋亡（P0.05）。體內(nèi)也能明顯抑制CNE-2Z裸鼠鼻咽癌移植瘤的生長(zhǎng)。分子機(jī)制檢測(cè)結(jié)果表明：Ad-OSM能明顯上調(diào)CNE-2Z鼻咽癌細(xì)胞P21、P27、P53、Bax、Caspase-3和下調(diào)Cox-2、Bcl-2、Survivin等細(xì)胞周期和凋亡相關(guān)蛋白的表達(dá)。 結(jié)論： 1、Ad-OSM能抑制CNE-2Z人鼻咽癌細(xì)胞及其移植瘤的生長(zhǎng)和誘導(dǎo)其凋亡（P＜0.05）。 2、Ad-OSM能顯著上調(diào)P21、P27、P53、Bax、Caspase-3和下調(diào)Survivin、Cox-2、Bcl-2等細(xì)胞周期和凋亡相關(guān)蛋白表達(dá)的效應(yīng)，最終引起cleaved Caspase-3表達(dá)，可能是OSM表達(dá)對(duì)鼻咽癌細(xì)胞體內(nèi)外生長(zhǎng)具有抑癌效應(yīng)的重要機(jī)制之一。
[Abstract]:Background and objective: nasopharyngeal carcinoma (NPC) is one of the most prevalent tumors in South China, Vietnam, the Philippines and other South Asian countries. Guangdong, Guangxi, Fujian and Hunan provinces are high incidence areas in China, with some areas as high as 30 to 100, 000 to 50, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000, 000. The adjusted mortality rate was 647 / 100, 000 / 100, 000 / 100, 000 / 100, 000 or 32, 200 / 100, 000 respectively, ranking first in the world (adjusted mortality rate is 28. 88 / 100, 000). The incidence of male was about 2 times as high as that of female, and 4050 years old was the age group with high incidence. The morbidity and mortality of nasopharyngeal carcinoma increased with age. The occurrence and development of nasopharyngeal carcinoma (NPC) is an extremely complicated process of carcinogenesis involving many genes and multistage lesions. Gene therapy is a kind of high efficiency and specificity. Targeted therapy has become a hot topic in the medical field. In recent years, it has been found that the tumor suppressor M(Oncostatin Mastoma gene can inhibit the growth of tumor cells and induce apoptosis. The aim of this study was to observe the inhibitory effect of adenovirus OSM gene on nasopharyngeal carcinoma cells and xenografts in nude mice, and to explore its possible mechanism, and to provide experimental evidence for the application of Ad OSM in clinical practice in the future. Methods: CNE-2Z nasopharyngeal carcinoma cells were infected with Ad-OSM in vitro. RT-PCR and Westernblot were used to detect the transcription and expression of OSM gene in CNE-2Z cells. Annexin-V-PE/7-AAD double staining FCM was used to detect the inhibitory effect of Ad-GFP-Ad-OSM on the growth and apoptosis of nasopharyngeal carcinoma cells in vitro. FCM staining and Pi staining were used to detect the effect of Ad-OSM on the cell cycle of nasopharyngeal carcinoma. In order to analyze the possible molecular mechanism of apoptosis induced by Ad-OSM, RT-PCR was used to detect the expression of cleaved Caspase-3 in CNE-2Z cells by using P21 P27, Bcl-2 and survivin. Western blot was used to detect the effect of Ad-OSM on the expression of cleaved Caspase-3 in CNE-2Z cells. In vivo: CNE-2Z cells were used to construct xenografts in nude mice. The expression of growth related factors (GRF) in transplanted tumor of CNE-2Z nasopharyngeal carcinoma (NPC) was detected by immunohistochemical method. The apoptotic inhibitory factors, such as P53-P53-BaxCaspase-3, and the apoptosis-promoting factor, Bcl-2Cox-2, were detected by immunohistochemical method in nude mice model of transplanted tumor of nasopharyngeal carcinoma (NPC). Results RT-PCR and Western blot analysis showed that adenovirus mediated OSM gene could stably transcribe and express in CNE-2Z cells. Ad-OSM could significantly inhibit the growth of CNE-2Z nasopharyngeal carcinoma cells in vitro and induce G1 phase arrest and apoptosis. The growth of nasopharyngeal carcinoma xenografts in nude mice with CNE-2Z was also inhibited in vivo. The results of molecular mechanism analysis showed that the cell cycle and apoptosis-related protein expression of CNE-2Z NPC cell line P21P27P27P53P53P5axCaspase-3 and Cox-2Bcl-2survivin could be down-regulated. Conclusion: 1Ad-OSM could inhibit the growth and induce apoptosis of CNE-2Z human nasopharyngeal carcinoma cells and its transplanted tumor (P < 0.05). (2) Ad-OSM could significantly up-regulate the expression of Caspase-3 and down-regulate the expression of cell cycle and apoptosis-related proteins such as survivvin-Cox-2OSM, which may be one of the important mechanisms of inhibiting the growth of nasopharyngeal carcinoma cells in vivo and in vitro.
【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R739.63

【參考文獻(xiàn)】

相關(guān)期刊論文 前6條

1 李先平;高美華;石學(xué)香;張蓓;程穎;;CD59位點(diǎn)短肽封條對(duì)HeLa細(xì)胞凋亡相關(guān)基因caspase-3和survivin作用[J];青島大學(xué)醫(yī)學(xué)院學(xué)報(bào);2008年03期

2 胡志清;繆競(jìng)誠;;抑瘤素M的研究進(jìn)展[J];上海醫(yī)學(xué);2008年08期

3 孫志清;項(xiàng)鋒鋼;王文華;;人腦星形細(xì)胞瘤組織Survivin與VEGF表達(dá)及意義[J];齊魯醫(yī)學(xué)雜志;2008年06期

4 郝春秋;馮志華;聶青和;;腺病毒載體的特點(diǎn)及其在HCV研究中的應(yīng)用[J];世界華人消化雜志;2003年06期

5 夏曉波,周霞,許惠卓,陳勤;制瘤素通過其受體gp130/OSMRβ誘發(fā)轉(zhuǎn)基因鼠視網(wǎng)膜變性[J];中華眼底病雜志;2005年03期

6 汪毅;乳腺癌裸鼠移植模型的制作概況[J];重慶醫(yī)科大學(xué)學(xué)報(bào);2003年03期

本文編號(hào)：1937409