本文選題：前列腺癌　切入點(diǎn)：低氧誘導(dǎo)因子　出處：《武漢大學(xué)》2015年博士論文　論文類型：學(xué)位論文

【摘要】：目的：前列腺癌是男性泌尿生殖系統(tǒng)中的惡性腫瘤。在世界范圍內(nèi),前列腺癌已經(jīng)成為在男性中最為高發(fā)的惡性腫瘤之一,在美國每七個(gè)男性就有一個(gè)會被診斷為前列腺癌。而低氧是腫瘤的重要特征之一,在前列腺癌中低氧誘導(dǎo)因子(HIF1a)的高表達(dá)與前列腺癌的惡性程度和去勢抵抗有著十分密切的聯(lián)系。此外,近年的研究發(fā)現(xiàn)在腫瘤中脂代謝高表達(dá)和自噬的異常會促進(jìn)腫瘤的發(fā)生發(fā)展。本研究著重探討低氧誘導(dǎo)因子(HIFla)對前列腺癌細(xì)胞脂代謝和自噬的作用及其調(diào)控機(jī)制。方法：通過實(shí)時(shí)定量-聚合酶鏈反應(yīng)(Real Time- Polymerase Chain Reaction, RT-PCR)與免疫印跡法(Western blot)比較不同前列腺癌細(xì)胞中HIF1a與脂肪合成基因的表達(dá)。在此基礎(chǔ)上通過低氧環(huán)境(1%O2)培養(yǎng)與轉(zhuǎn)染HIF1a突變質(zhì)粒,分析脂肪合成基因的表達(dá)。并且通過熒光素酶報(bào)告基因與染色質(zhì)免疫沉淀方法探究該變化的分子機(jī)制,’通過裸鼠實(shí)驗(yàn)證實(shí)HIF1a可以促進(jìn)脂合成基因的表達(dá)。另一方面,成功利用低氧環(huán)境誘導(dǎo)前列腺癌細(xì)胞發(fā)生自噬后,通過實(shí)時(shí)定量-聚合酶鏈反應(yīng)(Real Time-Polymerase Chain Reaction, RT-PCR)與免疫印跡法(Western blot)檢測自噬相關(guān)基因的表達(dá),并通過熒光素酶報(bào)告基因與染色質(zhì)免疫沉淀方法詳細(xì)解答內(nèi)深層次的分子機(jī)制。結(jié)果：前列腺癌小鼠模型TRAMP的前列腺組織中,HIF1a高表達(dá)。在前列腺癌細(xì)胞中,缺氧誘導(dǎo)因子(HIF1a)與脂肪合成基因的表達(dá)存在一定相關(guān)性。前列腺癌細(xì)胞在缺氧條件下以及轉(zhuǎn)染HIF1a突變質(zhì)粒的條件下均可促進(jìn)脂肪合成基因的表達(dá)。缺氧誘導(dǎo)因子(HIF1a)可以結(jié)合到SREBP-1c的啟動子區(qū)域促進(jìn)其轉(zhuǎn)錄。與對照組相比(1.000± 0.095,1.000±0.042), HIFla可以顯著升高SREBP-1c的mRNA(2.992±0.159)與蛋白質(zhì)(2.337±0.113)的表達(dá)水平。在裸鼠成瘤的結(jié)果中,由于HIF1a的過表達(dá)也呈現(xiàn)出脂合成基因SREBP-1c,ACC1和FASN都上調(diào)的結(jié)果,并伴隨腫瘤體積的增大。此外,在前列腺癌細(xì)胞中,低氧的環(huán)境可以誘導(dǎo)前列腺癌發(fā)生自噬,自噬相關(guān)基因在RNA水平和蛋白質(zhì)水平均顯著升高。并且HIFla可以通過促進(jìn)自噬關(guān)鍵元件ATG5的轉(zhuǎn)錄達(dá)到促進(jìn)細(xì)胞自噬的效果。結(jié)論：缺氧的環(huán)境中,HIFla會通過激活SREBP-1c轉(zhuǎn)錄,從而促進(jìn)前列腺癌細(xì)胞的從頭脂肪合成；激活A(yù)TG5轉(zhuǎn)錄,促進(jìn)前列腺癌的自噬。
[Abstract]:Objective: prostate cancer is a malignant tumor in the male genitourinary system. In the world, prostate cancer has become one of the most common malignant tumors in men. One in seven men in the United States is diagnosed with prostate cancer, and hypoxia is an important feature of the tumor. The high expression of hypoxia inducible factor (HIF1a) in prostate cancer is closely related to the malignancy and castration resistance of prostate cancer. Recent studies have found that hyperlipidemia and abnormal autophagy may promote the development of tumor. This study focuses on the effect of hypoxia inducible factor HIFla-induced lipid metabolism and autophagy in prostate cancer cells and its regulatory mechanism. Methods: real time Polymerase Chain reaction (RT-PCRR) was used to compare the expression of HIF1a and fat synthesis genes in different prostate cancer cells by real time Polymerase Chain reaction (RT-PCRR). To analyze the expression of fat synthesis gene, and to explore the molecular mechanism of this change by luciferase reporter gene and chromatin immunoprecipitation. After successfully induced autophagy in prostate cancer cells by hypoxia, real Time-Polymerase Chain reaction (RT-PCR) and Western blotting were used to detect the expression of autophagy related genes. The deep molecular mechanism was explained in detail by luciferase reporter gene and chromatin immunoprecipitation. Results: HIF1a was overexpressed in prostate tissue of prostate cancer mouse model TRAMP. Hypoxia inducible factor (HIF1a) is related to the expression of adipose synthase gene. Prostate cancer cells can promote the expression of adipose synthesis gene under hypoxia and transfection of HIF1a mutant plasmid. Hypoxia inducible factor (HIF1a) can promote the expression of adipose synthase gene. HIF1a) can bind to the promoter region of SREBP-1c to promote its transcription. Compared with the control group, HIFla can significantly increase the expression levels of mRNA(2.992 鹵0.159) and 2.337 鹵0.113) of SREBP-1c. In addition, in prostate cancer cells, hypoxia can induce autophagy in prostate cancer cells due to the up-regulation of lipid synthesis gene SREBP-1cACC1 and FASN, and the increase of tumor volume. Autophagy related genes were significantly increased at both RNA and protein levels, and HIFla could promote autophagy by promoting transcription of ATG5, a key component of autophagy. Conclusion: HIFla can activate SREBP-1c transcription in hypoxic environment. This promotes the ab initio fat synthesis of prostate cancer cells, activates ATG5 transcription, and promotes autophagy of prostate cancer.
【學(xué)位授予單位】：武漢大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2015
【分類號】：R737.25

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本文編號：1604954