【摘要】：背景和目的： 腦膠質(zhì)瘤是中樞神經(jīng)系統(tǒng)最常見(jiàn)的原發(fā)性腫瘤,由于惡性膠質(zhì)瘤有侵襲性生長(zhǎng)、增殖速度快等特征,應(yīng)用目前的治療措施如外科手術(shù)切除、術(shù)后放療、化療和免疫治療等不能治愈,患者復(fù)發(fā)率很高,臨床預(yù)后甚差。造成這種現(xiàn)狀的原因,很大部位是對(duì)腦膠質(zhì)瘤的病因,發(fā)病機(jī)制的認(rèn)識(shí)不充分。為此,必須尋找與膠質(zhì)瘤發(fā)生、發(fā)展相關(guān)的基因和信號(hào)通路,進(jìn)一步深入了解膠質(zhì)瘤發(fā)生發(fā)展的分子病理機(jī)制,尋找膠質(zhì)瘤治療的新靶點(diǎn),在此基礎(chǔ)上開(kāi)拓膠質(zhì)瘤治療的新策略和新手段。 有研究證實(shí),Nemo-like kinase(NLK)是一種抑癌基因,在體外的惡性腫瘤細(xì)胞株中的研究發(fā)現(xiàn)它可以通過(guò)促進(jìn)腫瘤細(xì)胞的凋亡而起到抑癌作用,研究表明NLK能夠促進(jìn)人的結(jié)腸癌細(xì)胞DLD-1細(xì)胞株,乳腺癌細(xì)胞株和膠質(zhì)瘤細(xì)胞株的凋亡[1-3]。但NLK在腦膠質(zhì)瘤中的研究甚少,故可以初步認(rèn)定NLK是一個(gè)膠質(zhì)瘤抑癌基因。此外,為了尋找可調(diào)控NLK表達(dá)的microRNA,我們搜索了預(yù)測(cè)microRNA靶基因網(wǎng)站,包括TARGETSCAN, MIRANDA等,發(fā)現(xiàn)有很多的microRNA,包括niR-92b在NLK mRNA3'UTR上有潛在的結(jié)合位點(diǎn),故本課題重點(diǎn)研究了miR-92b I抑制膠質(zhì)瘤增殖、細(xì)胞周期進(jìn)展、凋亡和侵襲的影響及其可能的作用機(jī)制。 方法和結(jié)果： 本課題研究分為以下兩個(gè)部分： 第一部分中應(yīng)用Real time PCR的方法,檢測(cè)了5個(gè)膠質(zhì)瘤細(xì)胞系和38例不同級(jí)別人腦膠質(zhì)瘤組織中miR-92b的表達(dá)水平。結(jié)果表明miR-92b在膠質(zhì)瘤組織和細(xì)胞系中的表達(dá)較正常組織上調(diào),并與腫瘤級(jí)別呈正相關(guān)。 在課題第二部分,miR-92b Ⅰ抑制膠質(zhì)瘤惡性表型的體外研究。將niR-92b Ⅰ轉(zhuǎn)染至人腦膠質(zhì)瘤細(xì)胞系SNB19和LN229中。采用Real time PCR檢測(cè)轉(zhuǎn)染后膠質(zhì)瘤細(xì)胞的miR-92b表達(dá)水平以鑒定抑制效果；四甲基偶氮唑藍(lán)(MTT)實(shí)驗(yàn)評(píng)價(jià)細(xì)胞的增殖率；流式細(xì)胞術(shù)檢測(cè)細(xì)胞周期變化；Annexin Ⅴ法檢測(cè)細(xì)胞早期凋亡;Transwell實(shí)驗(yàn)檢測(cè)細(xì)胞侵襲能力變化。Real time PCR檢測(cè)結(jié)果顯示轉(zhuǎn)染miR-92b Ⅰ后,腫瘤細(xì)胞miR-92b表達(dá)下降,miR-92b Ⅰ組細(xì)胞增殖活性降低,細(xì)胞侵襲能力明顯受到抑制,細(xì)胞周期S期縮短,阻滯在G0/G1期以及早期凋亡增加。該實(shí)驗(yàn)說(shuō)明miR-92b是癌微RNA(OncomiRs),可能成為膠質(zhì)瘤治療的潛在靶點(diǎn)。 結(jié)論： miR-92b在膠質(zhì)瘤中表達(dá)上調(diào),低級(jí)別腫瘤與高級(jí)別腫瘤比較,其差異具有顯著性,但是正常組織與低級(jí)別腫瘤比較,差異沒(méi)有顯著性。轉(zhuǎn)染miR-92b Ⅰ至人膠質(zhì)瘤細(xì)胞系,可抑制膠質(zhì)瘤細(xì)胞的增殖活性和侵襲能力,細(xì)胞周期S期縮短,阻滯在G0/G1期,并促進(jìn)細(xì)胞凋亡,由此,初步認(rèn)定miR-92b為癌微RNA,可成為膠質(zhì)瘤基因治療的候選靶標(biāo)。這是首次有關(guān)miR-92b在人腦膠質(zhì)瘤中的表達(dá)以及對(duì)膠質(zhì)瘤惡性表型的影響的研究報(bào)道,為進(jìn)一步認(rèn)識(shí)腦膠質(zhì)瘤的分子病理機(jī)制及新一類的靶向治療提供實(shí)驗(yàn)依據(jù)。
[Abstract]:Background and objective: glioma is the most common primary tumor in the central nervous system. Chemotherapy and immunotherapy can not be cured, the recurrence rate is very high, the clinical prognosis is very poor. Most of the causes of this situation are the etiology of glioma, and the pathogenesis of glioma is not well understood. Therefore, it is necessary to search for genes and signal pathways related to glioma development, to further understand the molecular pathological mechanism of glioma development, and to find new targets for the treatment of glioma. On this basis, a new strategy and a new method for the treatment of glioma were developed. Some studies have confirmed that Nemo-like kinase (NLK) is a tumor suppressor gene. In vitro, it has been found that Nemo-like kinase (NLK) can inhibit cancer by promoting apoptosis of tumor cells. It has been shown that NLK can promote human colon cancer cell DLD-1 cell line. Apoptosis of breast cancer cell line and glioma cell line [1-3]. However, there are few studies of NLK in gliomas, so it can be preliminarily concluded that NLK is a tumor suppressor gene. In addition, in order to find microRNA, that can regulate the expression of NLK, we searched microRNA target gene websites, including TARGETSCAN, MIRANDA, and found that many microRNA, including niR-92b, had potential binding sites on NLK mRNA3'UTR. Therefore, this study focused on the inhibition of glioma proliferation by miR-92b I. Cell cycle progression, the effects of apoptosis and invasion and their possible mechanisms. Methods and results: the study was divided into the following two parts: in the first part, the expression of miR-92b in 5 glioma cell lines and 38 cases of human gliomas of different grades were detected by the method of Real time PCR. The results showed that the expression of miR-92b in glioma tissues and cell lines was higher than that in normal tissues, and there was a positive correlation between the expression of miR-92b and tumor grade. In the second part of the study, miR-92b 鈪，

本文編號(hào)：2275836