本文選題：胃癌 + SGC-7901細胞��； 參考：《安徽醫(yī)科大學》2015年碩士論文

【摘要】：目的：研究miR-34a在人胃腺癌及配對癌旁正常組織、人正常胃粘膜上皮細胞及多種胃癌細胞株中的表達水平；通過構建過表達miR-34a慢病毒載體并感染人胃癌SGC-7901細胞,觀察其對細胞增殖、細胞周期和細胞凋亡的影響；預測miR-34a可能的腫瘤相關靶基因,探討miR-34a在胃癌發(fā)生、發(fā)展中的作用及其調(diào)控機制。方法：1.通過Real-time PCR檢測34例人胃腺癌及配對的癌旁正常組織,人正常胃黏膜上皮細胞GES及人胃癌細胞株AGS、SGC-7901、MKN-45、BGC-823中miR-34a的表達水平。2.構建過表達hsa-miR-34a慢病毒載體并體外感染人胃癌SGC-7901細胞,同時設對照組。熒光顯微鏡觀察評估慢病毒細胞感染情況,MTT實驗、流式細胞術檢測感染后胃癌SGC-7901細胞的細胞增殖、周期及細胞凋亡情況。3.采用microRNA在線靶基因預測軟件miRanda、TargetScan、PICTAR共同預測并篩選miR-34a可能腫瘤相關靶基因。4.應用SPSS 19.0軟件對所有實驗數(shù)據(jù)進行統(tǒng)計學分析。結(jié)果：1.胃癌組織的miR-34a相對表達量為0.65+0.69,明顯低于配對癌旁組織的相對表達量1.02±0.04(t=3.085,P0.01),但miR-34a的表達水平與胃癌的分化程度、浸潤深度、局部淋巴結(jié)轉(zhuǎn)移無明顯相關性(P0.05)。2.胃癌細胞AGS、SGC-7901、MKN-45、BGC-823中miR-34a的相對表達量分別為0.24±0.01,0.03±0.00,0.31±0.04和0.25±0.05,均分別明顯低于miR-34a在正常人胃黏膜上皮細胞GES的相對表達量(0.98±0.07),差異均具有統(tǒng)計學意義(t=17.522、22.879、13.836、14.393,P0.05),其中miR-34a在胃癌SGC-7901細胞中的相對表達量降低最為明顯(P0.01)。3. miR-34a慢病毒感染組在檢測第3到5天的相對增殖倍數(shù)分別為1.39±0.19、1.82±0.16和2.37±0.04,均明顯低于陰性對照慢病毒感染組第3到5天的相對增殖倍數(shù)(分別為1.844±0.10、2.46±0.11和3.144±0.38),差異有統(tǒng)計學意義(t=4.567、7.265、4.500,P0.01)。4. miR-34a慢病毒感染組G1/G0期細胞比例為58.72%,顯著高于陰性對照組的56.36%(t=-4.804,P0.05)。5. miR-34a慢病毒感染組細胞凋亡率為6.44%,顯著高于陰性對照組的3.06%(t=-13.226,P0.01)。6.通過對miRanda、TargetScain、PICTAR三個在線數(shù)據(jù)庫檢索交集的分析,篩選出與腫瘤增殖/凋亡/細胞周期/侵襲轉(zhuǎn)移相關的7個靶基因：E2F3、MET、NOTCH1、 SIRT1、Bcl-2、YY1、CCNE2。結(jié)論：1.miR-34a在人胃癌組織及多種人胃癌細胞中呈低表達,提示miR-34a與胃癌的發(fā)病關系密切。2.miR-34a可以抑制胃癌SGC7901細胞的增殖,誘導細胞G1/G0期停滯及細胞凋亡,在胃癌SGC-7901細胞中可能發(fā)揮抑癌基因的作用；miR-34a可能通過抑制E2F3、MET、NOTCH1、SIRT1、Bcl-2、YY1、CCNE2等靶基因發(fā)揮促進胃癌細胞增值抑制、細胞周期停滯及細胞凋亡的抗腫瘤作用。
[Abstract]:Objective: to study the expression of miR-34a in human gastric adenocarcinoma and adjacent normal tissues, normal gastric mucosal epithelial cells and gastric cancer cell lines, and to construct and infect human gastric cancer SGC-7901 cells by overexpression of miR-34a lentivirus vector. The effects of miR-34a on cell proliferation, cell cycle and apoptosis were observed, the possible tumor-related target genes of miR-34a were predicted, and the role and regulatory mechanism of miR-34a in carcinogenesis and development of gastric cancer were discussed. Method 1: 1. Real-time PCR was used to detect the expression of miR-34a in 34 cases of human gastric adenocarcinoma and matched adjacent normal tissues, the expression level of miR-34a in human gastric epithelial cells (GES) and human gastric cancer cell line AGSN SGC-7901 (MKN-45) BGC-823. Expression of hsa-miR-34a lentivirus vector was constructed and transfected into human gastric cancer SGC-7901 cells in vitro. MTT assay and flow cytometry were used to detect the proliferation, cycle and apoptosis of SGC-7901 cells after infection. MicroRNA online target gene prediction software miRandaScant PICTAR was used to predict and screen miR-34a possible tumor-related target gene. 4. All experimental data were statistically analyzed by SPSS 19.0 software. The result is 1: 1. The relative expression of miR-34a in gastric cancer was 0.65 0.69, which was significantly lower than that in matched paracancerous tissues (1.02 鹵0.04). However, the expression level of miR-34a was not correlated with the differentiation, depth of invasion and local lymph node metastasis of gastric carcinoma. The relative expression of miR-34a was 0.24 鹵0.01 鹵0.03 鹵0.001 鹵0.31 鹵0.04 and 0.25 鹵0.05 in gastric cancer cell line AGSC-7901MKN-45-BGC-823, respectively, which was significantly lower than that of miR-34a in normal gastric epithelial cells (0.98 鹵0.07). The difference was statistically significant (P 0.05), and the miR-34a expression in SGC-7901 cells was significantly lower than that in normal gastric cancer SGC-7901 cells. There were significant differences in the expression of miR-34a in gastric cancer SGC-7901 cells. There were significant differences in the expression of miR-34a in gastric cancer SGC-7901 cells. There were significant differences in the relative expression of miR-34a in gastric cancer SGC-7901 cells. There were significant differences in the expression of miR-34a in SGC-7901 cells. The relative proliferative times of lentivirus infection group were 1.39 鹵0.19 鹵1.82 鹵0.16 and 2.37 鹵0.04, respectively, which were significantly lower than those of the negative control group on the 3rd to 5th day (1.844 鹵0.102.46 鹵0.11, respectively). The difference was statistically significant between 3.144 鹵0.38 and 4.5677.265A4.500P0.01n.4miR-34a lentivirus infection group, the cell ratio of G1 / G0 phase was 58.722.It was significantly higher than that of negative control group (56.36t-4.804P0.05.5.MiR-34a lentivirus infection group), which was significantly higher than that of negative control group (3.06tti-13.226P0.01p0.01. 6.) By analyzing the intersections of three online databases of MiRanda TargetScainTar, seven target genes: E2F3METTCH1, SIRT1 Bcl-2OY1 (YY1) and CCNE2 were screened out, which were related to tumor proliferation / apoptosis / cell cycle / invasion and metastasis. Conclusion the low expression of miR-34a in human gastric cancer tissues and various human gastric cancer cells suggests that miR-34a is closely related to gastric cancer. 2.miR-34a can inhibit the proliferation of gastric cancer SGC7901 cells and induce G1 / G0 phase arrest and apoptosis of SGC7901 cells. The inhibitory effect of miR-34a on the proliferation, cell cycle arrest and apoptosis of gastric cancer cells may be promoted by inhibiting the target genes, such as E2F3METNTCH1, SIRT1, Bcl-2Y1 and CCNE2, in SGC-7901 cells.
【學位授予單位】：安徽醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：R735.2

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相關期刊論文 前2條

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本文編號：2041965