本文選題：HOTAIR + 急性髓系白血病��； 參考：《安徽醫(yī)科大學(xué)》2017年碩士論文

【摘要】：研究背景與目的:急性髓系白血病(Acute myeloid leukemia,AML)是一類髓系造血干祖細(xì)胞的惡性克隆性疾病,其在各個(gè)年齡段均可發(fā)病,且病程進(jìn)展迅速、臨床預(yù)后差。近年來,隨著靶向藥物全反式維甲酸(All-transretinoic acid,ATRA)的應(yīng)用,AML-M3即急性早幼粒白血病(Acute promyelocytic leukemia,APL)的治療及預(yù)后取得了極大進(jìn)展。然而其他類型的AML治療效果仍不佳,因此,需要更加深入的發(fā)病機(jī)制研究來為臨床治療方案提供新的思路和理論依據(jù)。長鏈非編碼RNA(long non-coding RNA,lnc RNA)是一類長度大于200bp、不具有或者很少具有編碼蛋白質(zhì)功能的RNA,其不僅參與了機(jī)體細(xì)胞正常增殖、分化、凋亡等生理過程,還參與了腫瘤發(fā)生發(fā)展等病理過程。HOTAIR為一長度為2158bp,位于12號(hào)染色體HOXC區(qū)域的lnc RNA,其作為促癌基因通過調(diào)控細(xì)胞分化、組蛋白修飾等多種方式參與了腫瘤的發(fā)生發(fā)展,但在AML中的作用及機(jī)制尚不明確。鑒于此,本研究旨在探討HOTAIR在全反式維甲酸誘導(dǎo)AML細(xì)胞分化過程中的作用及機(jī)制,為AML的診治提供新的思路及實(shí)驗(yàn)基礎(chǔ)。方法:收集我院臨床確診AML患者45例,缺鐵性貧血患者15例的骨髓標(biāo)本;實(shí)時(shí)定量PCR(q PCR)檢測HOTAIR在AML患者,缺鐵性貧血患者骨髓組織以及AML細(xì)胞株(HL-60,NB4,U937)中的表達(dá)水平;全反式維甲酸誘導(dǎo)分化AML細(xì)胞株,q PCR檢測和分析HOTAIR經(jīng)ATRA分化干預(yù)前后HOTAIR表達(dá)變化,流式檢測AML細(xì)胞分化及細(xì)胞周期;si RNA沉默HOTAIR基因,研究HOTAIR在AML細(xì)胞分化過程中的作用及機(jī)制;進(jìn)一步Western Blot檢測HOTAIR對細(xì)胞周期相關(guān)蛋白的水平變化;此外,采用Kaplan-meier法分析HOTAIR與患者臨床預(yù)后的相關(guān)性。結(jié)果:我們研究發(fā)現(xiàn):全反式維甲酸處理AML細(xì)胞后,細(xì)胞分化增多,CD11b及HOTAIR表達(dá)水平上調(diào),HL-60細(xì)胞周期G1/G0期上升,S期下降;而沉默HOTAIR基因,細(xì)胞分化減少,HL-60細(xì)胞周期G1/G0期下降,S期上升,提示HOTAIR可部分逆轉(zhuǎn)全反式維甲酸誘導(dǎo)分化作用。而且,全反式維甲酸可下調(diào)HL-60細(xì)胞CDK4,cylin D1蛋白水平,上調(diào)p21蛋白水平;沉默HOTAIR基因,HL-60細(xì)胞CDK4,cylin D1蛋白水平表達(dá)上升,p21蛋白表達(dá)下降。此外,臨床骨髓標(biāo)本的q PCR結(jié)果顯示,HOTAIR表達(dá)水平在AML-M2型患者組中較缺鐵性貧血患者組低,相應(yīng)結(jié)果在AML細(xì)胞株也得到驗(yàn)證;進(jìn)一步統(tǒng)計(jì)分析還發(fā)現(xiàn)HOTAIR與AML患者WBC高低相關(guān),而與預(yù)后無明顯相關(guān)。結(jié)論:1.HOTAIR在ATRA誘導(dǎo)AML細(xì)胞分化過程中,表達(dá)上調(diào);2.HOTAIR通過調(diào)控p21,cylin D1及CDK4的表達(dá),調(diào)控細(xì)胞周期,參與AML細(xì)胞分化;3.HOTAIR表達(dá)與AML患者高WBC數(shù)存在相關(guān),HOTAIR可能為AML診治提供新靶點(diǎn)。
[Abstract]:Background & objective: acute myeloid leukemia (myeloid) is a kind of malignant clonal disease of hematopoietic stem progenitor cells of myeloid system. In recent years, great progress has been made in the treatment and prognosis of the target drug All-Transretinoic Acid ATRAA (AML-M3). However, other types of AML still do not work well. Therefore, more in-depth research on pathogenesis is needed to provide new ideas and theoretical basis for clinical treatment. Long-chain noncoding RNAs (LNRNAs) are a class of RNAs with length larger than 200bpand with little or no function of encoding proteins, which not only participate in the physiological processes of normal cell proliferation, differentiation and apoptosis, but also participate in the physiological processes of cell proliferation, differentiation and apoptosis. HOTAIR is also involved in tumorigenesis and development. HOTAIR is a lnc RNAs located in the HOXC region of chromosome 12, which is involved in tumor development by regulating cell differentiation and histone modification. However, the role and mechanism in AML is unclear. In view of this, the purpose of this study was to explore the role and mechanism of HOTAIR in the differentiation of AML cells induced by all-trans retinoic acid, and to provide a new idea and experimental basis for the diagnosis and treatment of AML. Methods: bone marrow samples of 45 patients with AML and 15 patients with iron deficiency anemia were collected, and the expression of HOTAIR in bone marrow tissue of AML patients, patients with iron deficiency anemia and AML cell line HL-60NB4U937was detected by real-time quantitative PCRQ PCR. All trans retinoic acid (ATRA) induced differentiation AML cell line was detected and analyzed by qPCR. The expression of HOTAIR was detected before and after ATRA. Flow cytometry was used to detect the differentiation of AML cells and the silencing of HOTAIR gene by cell cycle siRNA. The role and mechanism of HOTAIR in the differentiation of AML cells were studied. In addition, Kaplan-meier method was used to analyze the correlation between HOTAIR and clinical prognosis. Results: in AML cells treated with all-trans retinoic acid, the expression of CD11b and HOTAIR increased and the expression of CD11b and HOTAIR upregulated and decreased in the G _ 1 / G _ 0 phase of HL-60 cell cycle, while the HOTAIR gene was silenced. The cell differentiation decreased and the G1 / G0 phase decreased and the S phase increased, suggesting that HOTAIR could partially reverse the differentiation induced by all-trans retinoic acid (ATRA). In addition, all trans retinoic acid could down-regulate the level of CDK4cylin D1 and up-regulate the level of p21 protein in HL-60 cells, while silencing the expression of CDK4 cylin D1 in HL-60 cells of HOTAIR gene could increase the expression of CDK4cylin D1 protein and decrease the expression of p21 protein. In addition, the Q PCR results of clinical bone marrow samples showed that the expression level of HOTAIR was lower in AML-M2 group than in iron deficiency anemia group, and the corresponding results were also verified in AML cell line. Further statistical analysis also found that HOTAIR was correlated with WBC level in AML patients. There was no significant correlation between prognosis and prognosis. Conclusion: 1. The expression of HOTAIR is up-regulated during ATRA-induced AML cell differentiation. Secondly, HOTAIR regulates the cell cycle by regulating the expression of p21 cylin D1 and CDK4. The expression of HOTAIR may provide a new target for the diagnosis and treatment of AML. 3. The expression of HOTAIR is related to the high WBC number of AML patients.
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R733.71

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本文編號(hào)：2036861