本文選題：前列腺癌 + 雄激素受體　； 參考：《上海交通大學(xué)》2015年博士論文

【摘要】：目的:前列腺癌是男性泌尿生殖系統(tǒng)中最常見的惡性腫瘤之一。在前列腺癌的發(fā)生、發(fā)展過程中雄激素受體(androgen receptor,AR)信號(hào)通路發(fā)揮重要的作用。AR通過介導(dǎo)下游多種編碼及非編碼基因(例如microRNA)的表達(dá),調(diào)控前列腺癌的增殖。已有研究表明,microRNA-124(miR-124)在前列腺癌中發(fā)揮抑癌基因的功能,但是其上游調(diào)控基因及相關(guān)分子機(jī)制尚不明確。本論文以AR及miR-124為研究對(duì)象,基于已有的研究成果通過解析兩者之間的調(diào)控模式,探索AR及mi R-124參與調(diào)控前列腺癌增殖的可能機(jī)制。方法:在AR陰性的前列腺癌細(xì)胞系PC3中構(gòu)建AR過表達(dá)亞克隆細(xì)胞系PC3/AR和對(duì)照PC3/neo;在AR陽(yáng)性的前列腺癌細(xì)胞系LNCaP中使用包含AR靶向shRNA(或?qū)φ?的慢病毒感染獲得AR持續(xù)敲降細(xì)胞系LNCaP-sh-AR和對(duì)照LNCaP-sh-control;在該細(xì)胞系中使用包含成熟miR-124(或?qū)φ?的慢病毒感染獲得穩(wěn)定過表達(dá)miR-124細(xì)胞系LNCaP-miR-124和對(duì)照LNCaP-control。采用實(shí)時(shí)定量反轉(zhuǎn)錄PCR方法檢測(cè)AR表達(dá)量改變對(duì)miR-124表達(dá)產(chǎn)生的影響及miR-124表達(dá)變化對(duì)AR表達(dá)產(chǎn)生的影響。通過雄激素應(yīng)答實(shí)驗(yàn),檢測(cè)雄激素能否引起miR-124的表達(dá)應(yīng)答。通過細(xì)胞增殖實(shí)驗(yàn)、細(xì)胞侵潤(rùn)實(shí)驗(yàn)、裸鼠皮下植瘤實(shí)驗(yàn),在體外及體內(nèi)驗(yàn)證過表達(dá)miR-124對(duì)前列腺癌細(xì)胞增殖的抑制作用。通過重亞硫酸鹽測(cè)序法檢測(cè)pri-miR-124-3啟動(dòng)子區(qū)域的甲基化程度。使用Prism GraphPad5軟件進(jìn)行統(tǒng)計(jì)分析和作圖。采用學(xué)生式t檢驗(yàn)比較不同組間差異,當(dāng)p0.05時(shí)認(rèn)為兩者具有統(tǒng)計(jì)學(xué)意義上的顯著差異。結(jié)果:本研究中,我們發(fā)現(xiàn)AR與miR-124之間存在環(huán)式互調(diào)控,即AR激活后可以促進(jìn)miR-124的轉(zhuǎn)錄表達(dá),同時(shí)miR-124則在轉(zhuǎn)錄后水平上抑制AR的表達(dá),在正常生理狀態(tài)下實(shí)現(xiàn)穩(wěn)態(tài)。在前列腺癌細(xì)胞中,由于pri-miR-124-3啟動(dòng)子區(qū)域高度甲基化阻礙了AR與啟動(dòng)子的互作,導(dǎo)致miR-124的表達(dá)降低及互調(diào)控穩(wěn)態(tài)的破壞。通過外源恢復(fù)miR-124的表達(dá),在體外及體內(nèi)均能抑制前列腺癌細(xì)胞的增殖及成瘤能力。結(jié)論:AR與miR-124通過環(huán)式互調(diào)控方式實(shí)現(xiàn)的穩(wěn)態(tài),在前列腺癌細(xì)胞中由于啟動(dòng)子區(qū)域甲基化遭到破壞。通過外源恢復(fù)miR-124表達(dá)可以有效抑制前列腺癌的增殖。
[Abstract]:Objective: prostate cancer is one of the most common malignant tumors in male genitourinary system. Androgen receptor AR-signaling pathway plays an important role in the pathogenesis and development of prostate cancer. AR regulates the proliferation of prostate cancer by mediating the expression of multiple coding and non-coding genes (such as microRNAs) downstream. It has been shown that microRNA-124miR-124) plays a role as a tumor suppressor gene in prostate cancer, but its upstream regulatory gene and its related molecular mechanisms are not clear. In this thesis, AR and miR-124 were taken as the research objects. Based on the existing research results, the possible mechanism of AR and miR-124 involved in the regulation of prostate cancer proliferation was explored by analyzing the regulatory models between them. Methods: construction of AR overexpression subclone cell line PC3/AR and control PC3 / neoc in AR negative prostate cancer cell line PC3 and AR positive prostate cancer cell line LNCaP using lentivirus containing AR targeted shRNAs (or controls) to obtain AR persistence LNCaP-sh-AR and control LNCaP-sh-control cell lines were used to obtain stable expression of miR-124 LNCaP-miR-124 and control LNCaP-control cells by using lentivirus infection containing mature miR-124 (or control). Real time quantitative reverse transcription PCR was used to detect the effect of AR expression on miR-124 expression and miR-124 expression on AR expression. Androgen response test was used to determine whether androgen can induce miR-124 expression response. The inhibitory effect of overexpression of miR-124 on the proliferation of prostate cancer cells in vitro and in vivo was verified by cell proliferation assay, cell infiltration test and subcutaneous tumor implantation in nude mice. The methylation of pri-miR-124-3 promoter region was detected by bisulfite sequencing. Use Prism GraphPad5 software for statistical analysis and mapping. Student t test was used to compare the difference between different groups. When p0.05, the difference between the two groups was statistically significant. Results: in this study, we found that there is cyclic regulation between AR and miR-124, that is, AR activation can promote the transcription expression of miR-124, while miR-124 inhibits AR expression at posttranscriptional level and achieves homeostasis in normal physiological state. In prostate cancer cells, the hypermethylation of pri-miR-124-3 promoter region hinders the interaction between AR and promoter, which leads to the decrease of miR-124 expression and the destruction of the stable state of mutual regulation. By restoring the expression of miR-124, the proliferation and tumorigenesis of prostate cancer cells were inhibited in vitro and in vivo. Conclusion the homeostasis of miR-124 and AR is destroyed by promoter region methylation in prostate cancer cells. Restoring miR-124 expression by exogenous can effectively inhibit the proliferation of prostate cancer.
【學(xué)位授予單位】：上海交通大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2015
【分類號(hào)】：R737.25

【相似文獻(xiàn)】

相關(guān)會(huì)議論文 前1條

1 梁燕妮;羅學(xué)群;柯志勇;黃禮彬;;miR-124與糖皮質(zhì)激素受體之間在兒童急性淋巴細(xì)胞白血病的相關(guān)性研究[A];中華醫(yī)學(xué)會(huì)第十七次全國(guó)兒科學(xué)術(shù)大會(huì)論文匯編（上冊(cè)）[C];2012年

相關(guān)博士學(xué)位論文 前2條

1 彭小紅;miR-124調(diào)控靶基因Foxq1抑制鼻咽癌增殖及轉(zhuǎn)移的分子機(jī)制研究[D];南方醫(yī)科大學(xué);2015年

2 常云麗;雄激素受體-miR-124環(huán)式互調(diào)控抑制前列腺癌的增殖[D];上海交通大學(xué);2015年

相關(guān)碩士學(xué)位論文 前2條

1 周金懿;miR-124甲基化異常與去甲基化橋接治療對(duì)骨髓增生異常綜合征的預(yù)后影響[D];蘇州大學(xué);2015年

2 岳慶;miR-124對(duì)間充質(zhì)干細(xì)胞定向遷移的調(diào)控研究[D];蘇州大學(xué);2015年

，

本文編號(hào)：1967630