【摘要】：背景:長鏈非編碼RNA(long non-coding RNAs,lnc RNAs)是一類轉(zhuǎn)錄本長度超過200nt的RNA分子,它們?nèi)狈Φ鞍拙幋a的潛能。近年來,大量證據(jù)表明lnc RNAs能以RNA的形式在多種層面上(表觀遺傳調(diào)控、轉(zhuǎn)錄調(diào)控以及轉(zhuǎn)錄后調(diào)控等)調(diào)控基因的表達(dá),從而在細(xì)胞增殖、細(xì)胞周期、分化和凋亡等多種生物學(xué)進(jìn)程中發(fā)揮重要的作用。假基因轉(zhuǎn)錄得到的RNA也屬于lnc RNA的一類,以前被認(rèn)為沒有生物學(xué)功能,然而最近一些假基因被證實(shí)在腫瘤的發(fā)生中發(fā)揮著關(guān)鍵的作用。PTENP1被發(fā)現(xiàn)在幾種腫瘤細(xì)胞中發(fā)揮著抑癌作用,然而它在胃癌中的表達(dá)和生物學(xué)功能仍不清楚。目的:研究胃癌組織和細(xì)胞系中PTENP1的表達(dá)水平及其與臨床病理資料的關(guān)系,并探索PTENP1在胃癌細(xì)胞中發(fā)揮的生物學(xué)功能及相關(guān)機(jī)制。方法:通過實(shí)時定量逆轉(zhuǎn)錄聚合酶鏈?zhǔn)椒磻?yīng)(q RT-PCR)檢測68例胃癌組織及4個胃癌細(xì)胞系中PTENP1的表達(dá)水平;通過去甲基化藥物處理,研究PTENP1啟動子異常甲基化與其表達(dá)的關(guān)系;分別轉(zhuǎn)染p LJM-3’UTR或p LJM空質(zhì)粒進(jìn)入胃癌細(xì)胞系MGC-803及SGC-7901,通過CCK-8及平板克隆實(shí)驗(yàn)研究PTENP1對胃癌細(xì)胞增殖能力的影響,通過流式細(xì)胞分析及Hoechst staining實(shí)驗(yàn)研究PTENP1對細(xì)胞凋亡的影響,通過細(xì)胞劃痕實(shí)驗(yàn)和transwell研究PTENP1對細(xì)胞遷移和侵襲能力的影響;通過q RT-PCR及Western blot分析質(zhì)粒轉(zhuǎn)染后PTEN的表達(dá)變化。結(jié)果:q RT-PCR檢測結(jié)果顯示PTENP1表達(dá)在胃癌組織和細(xì)胞系中顯著下調(diào),其表達(dá)下調(diào)可能部分與DNA超甲基化有關(guān)。而且,PTENP1的低表達(dá)與腫瘤的大小、胃癌病人臨床分期、胃癌浸潤深度及淋巴結(jié)轉(zhuǎn)移有關(guān)。另外,結(jié)果顯示PTENP1能夠調(diào)節(jié)胃癌細(xì)胞的增殖、凋亡、遷移和侵襲。并且,PTENP1 3’UTR的表達(dá)升高能夠提高PTEN蛋白的表達(dá)水平。結(jié)論:PTENP1在胃癌中表達(dá)顯著下調(diào),并能通過調(diào)節(jié)PTEN蛋白的表達(dá)發(fā)揮抑癌作用。
[Abstract]:Background: long-chain non-coding RNA (long non-coding RNAs,lnc RNAs) is a class of RNA molecules whose transcripts are longer than 200nt. They lack the potential of protein coding. In recent years, there has been a great deal of evidence that lnc RNAs can regulate the expression of genes at various levels (epigenetic regulation, transcriptional regulation and post-transcriptional regulation) in the form of RNA, thus, in cell proliferation and cell cycle, Differentiation and apoptosis play an important role in many biological processes. Pseudogene transcribed RNA also belongs to the lnc RNA family, which was previously thought to have no biological function. However, some pseudogenes have recently been proved to play a key role in tumorigenesis. PTENP1 has been found to play an anti-cancer role in several tumor cells, but its expression and biological function in gastric cancer are still unclear. Aim: to study the expression level of PTENP1 in gastric cancer tissues and cell lines and its relationship with clinicopathological data, and to explore the biological function and related mechanism of PTENP1 in gastric cancer cells. Methods: real-time quantitative reverse transcription polymerase chain reaction (Q-RT-PCR) was used to detect the expression of PTENP1 in 68 cases of gastric cancer tissues and 4 gastric cancer cell lines. The relationship between aberrant methylation of PTENP1 promoter and its expression was studied by demethylation drug treatment. The effects of PTENP1 on the proliferation of gastric cancer cell lines MGC-803 and SGC-7901, were studied by CCK-8 and plate cloning assay, respectively. The empty plasmids of p-LJM-3'UTR or p-LJM were transfected into gastric cancer cell line MGC-803 and SGC-7901, respectively. The effect of PTENP1 on cell apoptosis was studied by flow cytometry and Hoechst staining assay. The effects of PTENP1 on cell migration and invasion were studied by cell scratch test and transwell. The expression of PTEN was analyzed by Q-RT-PCR and Western blot. Results: the results of Q-RT-PCR showed that the expression of PTENP1 was significantly down-regulated in gastric cancer tissues and cell lines, and the down-regulation of the expression may be partly related to DNA hypermethylation. Furthermore, the low expression of PTENP1 was associated with tumor size, clinical stage, depth of invasion and lymph node metastasis. In addition, the results showed that PTENP1 can regulate the proliferation, apoptosis, migration and invasion of gastric cancer cells. In addition, the increased expression of PTENP1 3'UTR could increase the expression level of PTEN protein. Conclusion: the expression of PTENP1 is significantly down-regulated in gastric cancer, and can play an anti-cancer role by regulating the expression of PTEN protein.
【學(xué)位授予單位】：南京醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：R735.2

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