NDRG2基因?qū)ξ赴┘毎麗盒员硇偷挠绊?/H1>
發(fā)布時間：2018-05-18 11:36
本文選題：胃癌 + NDRG2�。� 參考：《鄭州大學(xué)》2015年碩士論文
【摘要】：胃癌是世界范圍內(nèi)最常見的惡性腫瘤之一,其發(fā)病率和死亡率在我國均居第二位,遠遠高于世界水平。大多數(shù)患者在有癥狀就診時已進入中晚期,失去了外科手術(shù)機會,這主要是由于胃癌的發(fā)生發(fā)展機制尚不清楚,臨床上缺乏有效的早期診斷指標及早期有效的治療。惡性腫瘤的局部浸潤、遠處轉(zhuǎn)移、再次復(fù)發(fā)和多藥耐藥是胃癌患者晚期死亡的主要原因,因此研究其惡性表型的機制十分有必要,同時對于胃癌的早期診斷及治療也有一定的指導(dǎo)意義。人類NDRG家族由NDRG1、NDRG2、NDRG3、NDRG4共同組成,各成員之間約57%~65%的氨基酸具有同源性,其中NDRG2是由我國第四軍醫(yī)大學(xué)生物化學(xué)與分子生物學(xué)教研室首先克隆并報道的。眾多研究表明,NDRG2在大多數(shù)腫瘤中表達下調(diào),在胃癌中也有類似報道,但其對胃癌的細胞學(xué)水平影響及其相關(guān)分子機制尚不完全清楚,有待進一步深入的研究。目的:NDRG2是一個潛在的腫瘤抑制基因,在胃癌組織中低表達。本課題旨在探討NDRG2基因?qū)ξ赴┘毎麗盒员硇偷挠绊憽７椒?1.通過Western Blot、Real-Time PCR驗證NDRG2在永生化的正常胃上皮細胞GES及胃癌細胞系(MKN-28、MKN-45、AGS、SGC-7901、BGC-823)中的表達水平。2.通過瞬時轉(zhuǎn)染使pc DNA3.1-NDRG2、pc DNA3.1、si RNA-NDRG2、si RNA-negetive進入AGS細胞內(nèi),使用RT-PCR及Western Blot驗證轉(zhuǎn)染效率;將實驗設(shè)置為pc DNA3.1-NDRG2(上調(diào)表達載體組)、pc DNA3.1(對照組)、si RNA-NDRG2(下調(diào)表達載體組)、si RNA-negetive(對照組)四組,并采用MTT及平板克隆實驗檢測細胞的體外增殖能力、采用流式細胞儀分析細胞的凋亡。3.轉(zhuǎn)染pc DNA3.1-NDRG2、pc DNA3.1、si RNA-NDRG2、si RNA-negetive后通過Western Blot檢測NDRG2與Bcl-2之間的關(guān)系;通過購買組織芯片運用免疫組織化學(xué)技術(shù)初步檢測NDRG2和Bcl-2在胃癌組織中的表達模式,再次通過收集臨床標本及相關(guān)病理資料,擴大樣本量運用免疫組織化學(xué)技術(shù)檢測NDRG2和Bcl-2在胃癌組織中的表達模式,并利用統(tǒng)計學(xué)軟件分析NDRG2和Bcl-2與臨床病理資料間的關(guān)系。結(jié)果:1.通過Western Blot、Real-Time PCR發(fā)現(xiàn)NDRG2在胃癌細胞中低表達,在AGS細胞中表達量居中,可用于后續(xù)構(gòu)建過表達載體及低表達載體的研究。2.通過Western Blot、RT-PCR發(fā)現(xiàn)AGS細胞轉(zhuǎn)染pc DNA3.1-NDRG2、si RNA-NDRG2后與對照組相比NDRG2 m RNA及蛋白水平會分別增高與降低;MTT發(fā)現(xiàn)pc DNA3.1-NDRG2組與pc DNA3.1組相比細胞增殖能力明顯減弱,si RNA-NDRG2組與si RNA-negetive組相比細胞增殖能力明顯增強;平板克隆實驗發(fā)現(xiàn)pc DNA3.1-NDRG2組與pc DNA3.1組相比細胞克隆形成能力明顯減弱,si RNA-NDRG2組與si RNA-negetive組相比細胞克隆形成能力明顯增強;流式細胞儀檢測細胞凋亡發(fā)現(xiàn)pc DNA3.1-NDRG2組與pc DNA3.1組相比細胞凋亡明顯增加,si RNA-NDRG2組與si RNA-negetive組相比細胞凋亡明顯減少。3.通過Western Blot發(fā)現(xiàn)轉(zhuǎn)染pc DNA3.1-NDRG2后Bcl-2明顯減少,而轉(zhuǎn)染si RNA-NDRG2后Bcl-2明顯增加;利用免疫組織化學(xué)技術(shù)發(fā)現(xiàn)NDRG2在胃癌組織中與胃正常組織相比低表達,而Bcl-2在胃癌組織中與胃正常組織相比高表達,二者并呈負相關(guān),通過統(tǒng)計相關(guān)臨床病理資料發(fā)現(xiàn)NDRG2和Bcl-2在胃癌組織中的表達陽性率與年齡、性別無關(guān),而與腫瘤分化程度、臨床分期、淋巴結(jié)有無轉(zhuǎn)移相關(guān)。結(jié)論:NDRG2能夠抑制胃癌細胞的增殖、促進胃癌細胞的凋亡,初步發(fā)現(xiàn)NDRG2在胃癌中發(fā)揮抑癌作用可能與Bcl-2有關(guān),但具體兩者之間的機制還有待進一步深入的研究。
[Abstract]:Gastric cancer is one of the most common malignant tumors in the world. Its incidence and mortality rate are second in China, which is far higher than the world level. Most patients have entered the middle and late stages of symptomatic treatment and lost the chance of surgical operation. This is mainly due to the unclear mechanism of the occurrence and development of gastric cancer and the lack of effective early clinical practice. The main cause of late death of gastric cancer patients is local invasion, distant metastasis, recurrence and multidrug resistance, so it is necessary to study the mechanism of malignant phenotype, and it also has certain guiding significance for the early diagnosis and treatment of gastric cancer. The human NDRG family is NDRG1, N DRG2, NDRG3, and NDRG4 are common, and the amino acids of approximately 57%~65% are homologous among members. NDRG2 is first cloned and reported by the Department of Biochemistry and molecular biology at the The Fourth Military Medical University. Many studies have shown that NDRG2 has been downregulated in most tumors and has similar reports in gastric cancer, but it is fine for gastric cancer. NDRG2 is a potential tumor suppressor gene and low expression in gastric cancer tissue. The purpose of this study is to explore the effect of NDRG2 gene on the malignant phenotype of gastric cancer cells. Methods: 1. through Western Blot, Real-Time PCR, NDRG2 is proved to be immortalized. The expression level of the normal gastric epithelial cells GES and gastric cancer cell lines (MKN-28, MKN-45, AGS, SGC-7901, BGC-823).2. through transient transfection makes PC DNA3.1-NDRG2, PC DNA3.1, Si, into the cells. PC DNA3.1 (control group), Si RNA-NDRG2 (down regulated expression vector group) and Si RNA-negetive (control group) were used to detect the proliferation of cells in vitro by MTT and flat clones. The apoptotic.3. was transfected to PC DNA3.1-NDRG2, PC DNA3.1. The relationship between NDRG2 and Bcl-2; using immunohistochemical technique to detect the expression pattern of Bcl-2 in gastric cancer tissue by using immunohistochemical technique, and by collecting the clinical specimens and related pathological data, expanding the sample size and using immunohistochemical technique to detect the expression pattern of NDRG2 and Bcl-2 in gastric cancer tissue, and use the system. Analysis of the relationship between NDRG2 and Bcl-2 and clinicopathological data. Results: 1. through Western Blot and Real-Time PCR, the expression of NDRG2 in gastric cancer cells is low, and the expression of NDRG2 in AGS cells is middle, and the study of.2. through Western Blot can be used for subsequent construction of overexpression vector and low expression vector. Compared with the control group, the levels of NDRG2 m RNA and protein were increased and decreased respectively after DRG2 and Si RNA-NDRG2, and MTT found that the proliferation ability of PC DNA3.1-NDRG2 group was significantly lower than that of PC DNA3.1 group. Compared with the cell clone formation ability, the cell clone formation ability of Si RNA-NDRG2 group was obviously enhanced compared with that of Si RNA-negetive group. The apoptosis of PC DNA3.1-NDRG2 group was obviously increased compared with that of PC DNA3.1 group, and the apoptosis of Si RNA-NDRG2 group was significantly reduced than that of Si RNA-negetive group. Western Blot found that Bcl-2 decreased significantly after transfection of PC DNA3.1-NDRG2, and Bcl-2 increased obviously after transfection of Si RNA-NDRG2, and the expression of NDRG2 in gastric cancer tissues was lower than that of normal gastric tissue, while Bcl-2 was highly expressed in gastric carcinoma tissue compared with normal gastric tissue, and the two were negatively correlated, by statistical correlation. It is found that the positive rate of NDRG2 and Bcl-2 in gastric cancer tissues is not related to age and sex, but is related to the degree of differentiation, clinical stage, and lymph node metastasis. Conclusion: NDRG2 can inhibit the proliferation of gastric cancer cells and promote the apoptosis of gastric cancer cells. It is found that NDRG2 may play a role in inhibiting cancer in gastric cancer and Bcl-2 in gastric cancer. But the mechanism between them should be further studied.
【學(xué)位授予單位】：鄭州大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：R735.2
【共引文獻】
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本文選題：胃癌 + NDRG2�。� 參考：《鄭州大學(xué)》2015年碩士論文
【摘要】：胃癌是世界范圍內(nèi)最常見的惡性腫瘤之一,其發(fā)病率和死亡率在我國均居第二位,遠遠高于世界水平。大多數(shù)患者在有癥狀就診時已進入中晚期,失去了外科手術(shù)機會,這主要是由于胃癌的發(fā)生發(fā)展機制尚不清楚,臨床上缺乏有效的早期診斷指標及早期有效的治療。惡性腫瘤的局部浸潤、遠處轉(zhuǎn)移、再次復(fù)發(fā)和多藥耐藥是胃癌患者晚期死亡的主要原因,因此研究其惡性表型的機制十分有必要,同時對于胃癌的早期診斷及治療也有一定的指導(dǎo)意義。人類NDRG家族由NDRG1、NDRG2、NDRG3、NDRG4共同組成,各成員之間約57%~65%的氨基酸具有同源性,其中NDRG2是由我國第四軍醫(yī)大學(xué)生物化學(xué)與分子生物學(xué)教研室首先克隆并報道的。眾多研究表明,NDRG2在大多數(shù)腫瘤中表達下調(diào),在胃癌中也有類似報道,但其對胃癌的細胞學(xué)水平影響及其相關(guān)分子機制尚不完全清楚,有待進一步深入的研究。目的:NDRG2是一個潛在的腫瘤抑制基因,在胃癌組織中低表達。本課題旨在探討NDRG2基因?qū)ξ赴┘毎麗盒员硇偷挠绊憽７椒?1.通過Western Blot、Real-Time PCR驗證NDRG2在永生化的正常胃上皮細胞GES及胃癌細胞系(MKN-28、MKN-45、AGS、SGC-7901、BGC-823)中的表達水平。2.通過瞬時轉(zhuǎn)染使pc DNA3.1-NDRG2、pc DNA3.1、si RNA-NDRG2、si RNA-negetive進入AGS細胞內(nèi),使用RT-PCR及Western Blot驗證轉(zhuǎn)染效率;將實驗設(shè)置為pc DNA3.1-NDRG2(上調(diào)表達載體組)、pc DNA3.1(對照組)、si RNA-NDRG2(下調(diào)表達載體組)、si RNA-negetive(對照組)四組,并采用MTT及平板克隆實驗檢測細胞的體外增殖能力、采用流式細胞儀分析細胞的凋亡。3.轉(zhuǎn)染pc DNA3.1-NDRG2、pc DNA3.1、si RNA-NDRG2、si RNA-negetive后通過Western Blot檢測NDRG2與Bcl-2之間的關(guān)系;通過購買組織芯片運用免疫組織化學(xué)技術(shù)初步檢測NDRG2和Bcl-2在胃癌組織中的表達模式,再次通過收集臨床標本及相關(guān)病理資料,擴大樣本量運用免疫組織化學(xué)技術(shù)檢測NDRG2和Bcl-2在胃癌組織中的表達模式,并利用統(tǒng)計學(xué)軟件分析NDRG2和Bcl-2與臨床病理資料間的關(guān)系。結(jié)果:1.通過Western Blot、Real-Time PCR發(fā)現(xiàn)NDRG2在胃癌細胞中低表達,在AGS細胞中表達量居中,可用于后續(xù)構(gòu)建過表達載體及低表達載體的研究。2.通過Western Blot、RT-PCR發(fā)現(xiàn)AGS細胞轉(zhuǎn)染pc DNA3.1-NDRG2、si RNA-NDRG2后與對照組相比NDRG2 m RNA及蛋白水平會分別增高與降低;MTT發(fā)現(xiàn)pc DNA3.1-NDRG2組與pc DNA3.1組相比細胞增殖能力明顯減弱,si RNA-NDRG2組與si RNA-negetive組相比細胞增殖能力明顯增強;平板克隆實驗發(fā)現(xiàn)pc DNA3.1-NDRG2組與pc DNA3.1組相比細胞克隆形成能力明顯減弱,si RNA-NDRG2組與si RNA-negetive組相比細胞克隆形成能力明顯增強;流式細胞儀檢測細胞凋亡發(fā)現(xiàn)pc DNA3.1-NDRG2組與pc DNA3.1組相比細胞凋亡明顯增加,si RNA-NDRG2組與si RNA-negetive組相比細胞凋亡明顯減少。3.通過Western Blot發(fā)現(xiàn)轉(zhuǎn)染pc DNA3.1-NDRG2后Bcl-2明顯減少,而轉(zhuǎn)染si RNA-NDRG2后Bcl-2明顯增加;利用免疫組織化學(xué)技術(shù)發(fā)現(xiàn)NDRG2在胃癌組織中與胃正常組織相比低表達,而Bcl-2在胃癌組織中與胃正常組織相比高表達,二者并呈負相關(guān),通過統(tǒng)計相關(guān)臨床病理資料發(fā)現(xiàn)NDRG2和Bcl-2在胃癌組織中的表達陽性率與年齡、性別無關(guān),而與腫瘤分化程度、臨床分期、淋巴結(jié)有無轉(zhuǎn)移相關(guān)。結(jié)論:NDRG2能夠抑制胃癌細胞的增殖、促進胃癌細胞的凋亡,初步發(fā)現(xiàn)NDRG2在胃癌中發(fā)揮抑癌作用可能與Bcl-2有關(guān),但具體兩者之間的機制還有待進一步深入的研究。
[Abstract]:Gastric cancer is one of the most common malignant tumors in the world. Its incidence and mortality rate are second in China, which is far higher than the world level. Most patients have entered the middle and late stages of symptomatic treatment and lost the chance of surgical operation. This is mainly due to the unclear mechanism of the occurrence and development of gastric cancer and the lack of effective early clinical practice. The main cause of late death of gastric cancer patients is local invasion, distant metastasis, recurrence and multidrug resistance, so it is necessary to study the mechanism of malignant phenotype, and it also has certain guiding significance for the early diagnosis and treatment of gastric cancer. The human NDRG family is NDRG1, N DRG2, NDRG3, and NDRG4 are common, and the amino acids of approximately 57%~65% are homologous among members. NDRG2 is first cloned and reported by the Department of Biochemistry and molecular biology at the The Fourth Military Medical University. Many studies have shown that NDRG2 has been downregulated in most tumors and has similar reports in gastric cancer, but it is fine for gastric cancer. NDRG2 is a potential tumor suppressor gene and low expression in gastric cancer tissue. The purpose of this study is to explore the effect of NDRG2 gene on the malignant phenotype of gastric cancer cells. Methods: 1. through Western Blot, Real-Time PCR, NDRG2 is proved to be immortalized. The expression level of the normal gastric epithelial cells GES and gastric cancer cell lines (MKN-28, MKN-45, AGS, SGC-7901, BGC-823).2. through transient transfection makes PC DNA3.1-NDRG2, PC DNA3.1, Si, into the cells. PC DNA3.1 (control group), Si RNA-NDRG2 (down regulated expression vector group) and Si RNA-negetive (control group) were used to detect the proliferation of cells in vitro by MTT and flat clones. The apoptotic.3. was transfected to PC DNA3.1-NDRG2, PC DNA3.1. The relationship between NDRG2 and Bcl-2; using immunohistochemical technique to detect the expression pattern of Bcl-2 in gastric cancer tissue by using immunohistochemical technique, and by collecting the clinical specimens and related pathological data, expanding the sample size and using immunohistochemical technique to detect the expression pattern of NDRG2 and Bcl-2 in gastric cancer tissue, and use the system. Analysis of the relationship between NDRG2 and Bcl-2 and clinicopathological data. Results: 1. through Western Blot and Real-Time PCR, the expression of NDRG2 in gastric cancer cells is low, and the expression of NDRG2 in AGS cells is middle, and the study of.2. through Western Blot can be used for subsequent construction of overexpression vector and low expression vector. Compared with the control group, the levels of NDRG2 m RNA and protein were increased and decreased respectively after DRG2 and Si RNA-NDRG2, and MTT found that the proliferation ability of PC DNA3.1-NDRG2 group was significantly lower than that of PC DNA3.1 group. Compared with the cell clone formation ability, the cell clone formation ability of Si RNA-NDRG2 group was obviously enhanced compared with that of Si RNA-negetive group. The apoptosis of PC DNA3.1-NDRG2 group was obviously increased compared with that of PC DNA3.1 group, and the apoptosis of Si RNA-NDRG2 group was significantly reduced than that of Si RNA-negetive group. Western Blot found that Bcl-2 decreased significantly after transfection of PC DNA3.1-NDRG2, and Bcl-2 increased obviously after transfection of Si RNA-NDRG2, and the expression of NDRG2 in gastric cancer tissues was lower than that of normal gastric tissue, while Bcl-2 was highly expressed in gastric carcinoma tissue compared with normal gastric tissue, and the two were negatively correlated, by statistical correlation. It is found that the positive rate of NDRG2 and Bcl-2 in gastric cancer tissues is not related to age and sex, but is related to the degree of differentiation, clinical stage, and lymph node metastasis. Conclusion: NDRG2 can inhibit the proliferation of gastric cancer cells and promote the apoptosis of gastric cancer cells. It is found that NDRG2 may play a role in inhibiting cancer in gastric cancer and Bcl-2 in gastric cancer. But the mechanism between them should be further studied.
【學(xué)位授予單位】：鄭州大學(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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