本文關(guān)鍵詞： 磷酸核糖焦磷酸合成酶1 急性白血病 兒童 危險(xiǎn)因素 B細(xì)胞急性白血病 磷酸核糖焦磷酸合成酶1 Bcl-2 增殖 凋亡　出處：《重慶醫(yī)科大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：目的:探討磷酸核糖焦磷酸合成酶1(PRPS1)基因在兒童急性白血病(AL)中的表達(dá),分析其表達(dá)水平與AL臨床指標(biāo)的相關(guān)性,初步探討PRPS1在AL中的臨床意義。方法:采用實(shí)時(shí)熒光定量PCR和免疫組化(IH)的方法檢測(cè)176例兒童AL(初診126例,完全緩解50例)骨髓標(biāo)本中PRPS1的表達(dá)水平,收集患兒病程中診斷結(jié)果,臨床表現(xiàn)以及治療反應(yīng)等臨床特征信息,分析其PRPS1表達(dá)水平與臨床特征的相關(guān)性。以21例非惡性血液病患兒骨髓作為對(duì)照組患兒。結(jié)果:(1)在B-ALL組中,PRPS1初診組表達(dá)水平顯著高于對(duì)照組,同時(shí)高于完全緩解組(均P0.0001);在T-ALL及AML組中,僅初診組與完全緩解組表達(dá)有顯著差異(均P0.0001)。(2)在B-ALL組中,PRPS1表達(dá)水平隨危險(xiǎn)度升高而增加(P0.01);在T-ALL組中各危險(xiǎn)度分組之間則無(wú)明顯差異(P0.05);在AML組中,僅低危組與高危組差異具有統(tǒng)計(jì)學(xué)意義(P0.05)。(3)PRPS1高表達(dá)與B-ALL患兒的高WBC計(jì)數(shù)及MRD陽(yáng)性相關(guān)(P=0.020,P=0.026)。(4)PRPS1與之前證實(shí)的復(fù)發(fā)相關(guān)基因NT5C2表達(dá)水平呈正相關(guān)(P0.05)。結(jié)論:PRPS1是兒童B-ALL預(yù)后不良相關(guān)危險(xiǎn)因素,有望成為判斷預(yù)后,指導(dǎo)個(gè)性化化療的指標(biāo)。目的:利用慢病毒上調(diào)PRPS1基因的表達(dá),研究其對(duì)Sup-b15及Raji細(xì)胞株增殖、凋亡及細(xì)胞周期的調(diào)控作用及機(jī)制。方法:采用PRPS1過表達(dá)慢病毒感染Sup-b15及Raji細(xì)胞株構(gòu)建實(shí)驗(yàn)組PRPS1過表達(dá)細(xì)胞;采用GFP過表達(dá)慢病毒感染Sup-b15及Raji細(xì)胞株作為對(duì)照組細(xì)胞;使用CCK-8法檢測(cè)細(xì)胞增殖;流式細(xì)胞術(shù)檢測(cè)細(xì)胞周期及凋亡;采用q RT-PCR、Western-blot方法檢測(cè)PRPS1及增殖、凋亡相關(guān)基因c-myc,cyclin E1,Bcl-2,CDK2,and caspase-3基因和蛋白表達(dá)水平的變化。結(jié)果:(1)與對(duì)照組細(xì)胞相比,實(shí)驗(yàn)組細(xì)胞PRPS1 m RNA和蛋白表達(dá)水平均顯著上升,P0.05;(2)CCK-8法檢測(cè)細(xì)胞增殖結(jié)果顯示,實(shí)驗(yàn)組Sup-b15及Raji細(xì)胞的增殖率均上升,與對(duì)照組相比,P0.001;(3)流式細(xì)胞學(xué)檢測(cè)細(xì)胞凋亡和周期結(jié)果顯示,與對(duì)照組相比,實(shí)驗(yàn)組Sup-b15及Raji細(xì)胞的凋亡率均下降,P0.01;細(xì)胞周期差異無(wú)統(tǒng)計(jì)學(xué)意義,P0.05。(4)與對(duì)照組相比,實(shí)驗(yàn)組Sup-b15及Raji細(xì)胞的Bcl-2,CDK2基因和蛋白表達(dá)水平均顯著提高,P0.001;Caspase-3蛋白水平表達(dá)下調(diào)P0.01;C-myc及Cyclin E1未見明顯變化。結(jié)論:PRPS1基因過表達(dá)對(duì)Sup-b15及Raji細(xì)胞有促進(jìn)增殖、抑制凋亡的作用,其機(jī)制可能與上調(diào)Bcl-2,增強(qiáng)細(xì)胞的抗凋亡作用相關(guān)。
[Abstract]:Objective: to investigate the expression of ribonucleotide pyrophosphate synthase 1 (PRPS1) gene in children with acute leukemia (ALL) and to analyze the correlation between the expression level and the clinical parameters of AL. To explore the clinical significance of PRPS1 in AL. Methods: real time fluorescence quantitative PCR and immunohistochemistry were used to detect 176 children with ALS (126 cases first diagnosed). The expression level of PRPS1 was collected in 50 cases of complete remission), and the clinical characteristics, such as diagnosis, clinical manifestation and therapeutic response were collected. The correlation between the expression of PRPS1 and clinical features was analyzed. The bone marrow of 21 children with non-malignant hematologic disease was used as control group. Results: 1) in B-ALL group. The expression level of PRPS1 in the newly diagnosed group was significantly higher than that in the control group and in the complete remission group (all P 0.0001). In T-ALL and AML group, the expression of T-ALL was significantly different from that of complete remission group (all P0.0001. 2) in B-ALL group. The expression of PRPS1 increased with the increase of risk. In T-ALL group, there was no significant difference among the risk groups (P 0.05). In the AML group. The high expression of PRPS1 in low risk group and high risk group was significantly higher than that in high risk group. The high expression of PRPS1 was associated with high WBC count and MRD positive correlation with P0. 020 in children with B-ALL. There was a positive correlation between the expression level of PRPS1 and the previously confirmed recurrence related gene NT5C2 (P0.05). Conclusion 1: PRPS1 is a risk factor for poor prognosis of B-ALL in children. Objective: to study the effect of lentivirus on the proliferation of Sup-b15 and Raji cell lines by up-regulating the expression of PRPS1 gene. Methods: PRPS1 overexpression lentivirus was used to infect Sup-b15 and Raji cell lines to construct PRPS1 overexpression cells in experimental group. GFP overexpression lentivirus was used to infect Sup-b15 and Raji cell lines as control cells. Cell proliferation was detected by CCK-8 assay. Cell cycle and apoptosis were detected by flow cytometry. PRPS1, proliferation and apoptosis related gene c-myc cyclin E1Bcl-2 CDK2 were detected by Q RT-PCR Western-blot. Results compared with the control group, the expression of PRPS1 m RNA and protein in the experimental group was significantly higher than that in the control group. P0.05; The proliferation rate of Sup-b15 and Raji cells in the experimental group was higher than that in the control group (P 0.001). Flow cytometric analysis showed that the apoptotic rate of Sup-b15 and Raji cells in the experimental group was lower than that in the control group (P 0.01). There was no significant difference in cell cycle between the experimental group and the control group (P 0.05. 05. 4) the Bcl-2 of Sup-b15 and Raji cells in the experimental group was higher than that in the control group. The expression level of CDK2 gene and protein increased significantly (P 0.001). The expression of Caspase-3 protein down-regulated P0.01; C-myc and Cyclin E1 showed no significant changes. Conclusion the overexpression of the 1 / PRPS1 gene can promote the proliferation and inhibit the apoptosis of Sup-b15 and Raji cells. The mechanism may be related to the up-regulation of BCL-2 and the enhancement of the anti-apoptotic effect of BCL-2.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R733.71

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