Objective To analyze the upstream region of radiation-induced junB gene. Methods Four plasmids containing 250 bp, 590 bp, 900 bp and 1650 bp, and CAT reporter gene were constructed separately and introduced to L8704 c...Objective To analyze the upstream region of radiation-induced junB gene. Methods Four plasmids containing 250 bp, 590 bp, 900 bp and 1650 bp, and CAT reporter gene were constructed separately and introduced to L8704 cells. The cells were irradiated with 2 Gy X-rays and incubated at different intervals. Total RNA was extracted from the cells and fluctuation of the CAT mRNA level was assessed by the RNA ratio of CAT/β-actin measured by quantitative Northern blot hybridization. Results CAT mRNA expression containing 900 bp and 1560 bpjunB promoter remarkably and rapidly increased, and reached its peak 30min after 2 Gy X-my irradiation. Conclusions 590-900 bp fragments located in the upstream region of junB gene play an important role in the early process of cells against radiation.展开更多
即刻早期反应基因IEX-1(immediate early response gene X-1)作为即刻早期基因家族之一,参与细胞的多种生物学效应,其转录和表达受多种因素的调控如NF-κB、P53、SP1、AP2、1a-25(OH)2D3、TNF-α及电离辐射等。研究发现,IEX-1在多种肿...即刻早期反应基因IEX-1(immediate early response gene X-1)作为即刻早期基因家族之一,参与细胞的多种生物学效应,其转录和表达受多种因素的调控如NF-κB、P53、SP1、AP2、1a-25(OH)2D3、TNF-α及电离辐射等。研究发现,IEX-1在多种肿瘤细胞中转录及表达降低或缺失,而电离辐射等则可以激活诱导其在多种类型肿瘤细胞和正常细胞中快速的表达,其激活表达与辐射后细胞的生长、周期、凋亡等的改变密切相关。辐射可诱导特性使其具有潜在的临床和基因工程生产应用前景。展开更多
AIM To observe the growth suppression effect of exogenous introduction of early growth response gene-1 (Egr-1 gene) on esophageal carcinoma tissue as well as on esophageal carcinoma cell line Eca109 and to explore the...AIM To observe the growth suppression effect of exogenous introduction of early growth response gene-1 (Egr-1 gene) on esophageal carcinoma tissue as well as on esophageal carcinoma cell line Eca109 and to explore the potential application of Egr-1 gene in gene therapy of tumor.METHODS Eukaryotic expression vector of PCMV-Egr-1 plasmid was introduced into Eca109 cell line which expressed no Egr-1 protein originally with lipofectamine transfection method. The introduction and expression of PCMV-Egr-1 plasmid into Eca109 cell line was confirmed by G418 selection culture, PCR amplification of neogene contained in the vector, Western blot analysis and immunocytochemical analysis. The cell growth curve,soft agar colony formation rate and tumorigenicity in SClD mice were examined to demonstrate the growth suppression effect of exogenous Egr-1 gene on Eca109 cell line. The Egr-1 mRNA and Egr-1 protein were also detected in 50 surgical specimens of esophageal carcinoma by in situ hybridization and immunohistochemistry.RESULTS Exogenous Egr-1 gene was introduced successfully into Eca109 cell line and expressed Egr-1 protein stably. The transfected Eca109 cell line grew more slowly than control Eca109 as shown by cell growth curves, the soft agar colony formation rate (4.0% vs 6.9%, P＜0.01) and the average growth rate of tumor in SCID mice (35.5 ± 7.6 vs 65.8 ± 7.6, P＜0.05). The expression level of Egr-1 mRNA and protein significantly increased in dysplastic epithelia adjacent to cancer rather than in cancer tissues (65.8% vs 20.0% by ISH and 57.9% vs 14.0% by IHC, P＜0.01).CONCLUSION Exogenous Egr-1 gene shows the strong effect of growth inhibition in Eca109 cell line. Egr-1 in the cancer tissue shows down-regulated expression that supports the inhibited function of Egr-1 in cancer growth and suggests Egr-1 may have an important role in gene therapy of esophageal carcinoma.展开更多
目的:探讨即刻早期反应基因-1(immediate early response gene X-1,IEX-1)转染人卵巢癌细胞SKOV3后,对顺铂药物敏感性的影响。方法:提取并鉴定重组质粒pEGFP-N1-IEX-1,采用脂质体介导将重组质粒转染到人卵巢癌细胞SKOV3,RT-PCR检测转染...目的:探讨即刻早期反应基因-1(immediate early response gene X-1,IEX-1)转染人卵巢癌细胞SKOV3后,对顺铂药物敏感性的影响。方法:提取并鉴定重组质粒pEGFP-N1-IEX-1,采用脂质体介导将重组质粒转染到人卵巢癌细胞SKOV3,RT-PCR检测转染前后IEX-1mRNA和蛋白的表达情况,采用四甲基偶氮唑蓝(MTT)法检测转染后卵巢癌细胞SKOV3增殖情况以及对顺铂敏感性的变化。结果:成功转染IEX-1基因后可检测到IEX-1mRNA的高表达;转染组细胞的增殖速度增快(P<0.05),但对顺铂的敏感性明显增强(P<0.05)。结论:IEX-1可增强卵巢癌细胞的体外增殖能力,并增强卵巢癌细胞体外对顺铂化疗的敏感性,为肿瘤的有效干预和治疗提供新的靶点。展开更多
基金This project was supported by Japanese Research Development Corporation (JRDC).
文摘Objective To analyze the upstream region of radiation-induced junB gene. Methods Four plasmids containing 250 bp, 590 bp, 900 bp and 1650 bp, and CAT reporter gene were constructed separately and introduced to L8704 cells. The cells were irradiated with 2 Gy X-rays and incubated at different intervals. Total RNA was extracted from the cells and fluctuation of the CAT mRNA level was assessed by the RNA ratio of CAT/β-actin measured by quantitative Northern blot hybridization. Results CAT mRNA expression containing 900 bp and 1560 bpjunB promoter remarkably and rapidly increased, and reached its peak 30min after 2 Gy X-my irradiation. Conclusions 590-900 bp fragments located in the upstream region of junB gene play an important role in the early process of cells against radiation.
文摘即刻早期反应基因IEX-1(immediate early response gene X-1)作为即刻早期基因家族之一,参与细胞的多种生物学效应,其转录和表达受多种因素的调控如NF-κB、P53、SP1、AP2、1a-25(OH)2D3、TNF-α及电离辐射等。研究发现,IEX-1在多种肿瘤细胞中转录及表达降低或缺失,而电离辐射等则可以激活诱导其在多种类型肿瘤细胞和正常细胞中快速的表达,其激活表达与辐射后细胞的生长、周期、凋亡等的改变密切相关。辐射可诱导特性使其具有潜在的临床和基因工程生产应用前景。
基金Supported by the National Natural Science Foundation of China,No.39670298.
文摘AIM To observe the growth suppression effect of exogenous introduction of early growth response gene-1 (Egr-1 gene) on esophageal carcinoma tissue as well as on esophageal carcinoma cell line Eca109 and to explore the potential application of Egr-1 gene in gene therapy of tumor.METHODS Eukaryotic expression vector of PCMV-Egr-1 plasmid was introduced into Eca109 cell line which expressed no Egr-1 protein originally with lipofectamine transfection method. The introduction and expression of PCMV-Egr-1 plasmid into Eca109 cell line was confirmed by G418 selection culture, PCR amplification of neogene contained in the vector, Western blot analysis and immunocytochemical analysis. The cell growth curve,soft agar colony formation rate and tumorigenicity in SClD mice were examined to demonstrate the growth suppression effect of exogenous Egr-1 gene on Eca109 cell line. The Egr-1 mRNA and Egr-1 protein were also detected in 50 surgical specimens of esophageal carcinoma by in situ hybridization and immunohistochemistry.RESULTS Exogenous Egr-1 gene was introduced successfully into Eca109 cell line and expressed Egr-1 protein stably. The transfected Eca109 cell line grew more slowly than control Eca109 as shown by cell growth curves, the soft agar colony formation rate (4.0% vs 6.9%, P＜0.01) and the average growth rate of tumor in SCID mice (35.5 ± 7.6 vs 65.8 ± 7.6, P＜0.05). The expression level of Egr-1 mRNA and protein significantly increased in dysplastic epithelia adjacent to cancer rather than in cancer tissues (65.8% vs 20.0% by ISH and 57.9% vs 14.0% by IHC, P＜0.01).CONCLUSION Exogenous Egr-1 gene shows the strong effect of growth inhibition in Eca109 cell line. Egr-1 in the cancer tissue shows down-regulated expression that supports the inhibited function of Egr-1 in cancer growth and suggests Egr-1 may have an important role in gene therapy of esophageal carcinoma.
文摘目的:探讨即刻早期反应基因-1(immediate early response gene X-1,IEX-1)转染人卵巢癌细胞SKOV3后,对顺铂药物敏感性的影响。方法:提取并鉴定重组质粒pEGFP-N1-IEX-1,采用脂质体介导将重组质粒转染到人卵巢癌细胞SKOV3,RT-PCR检测转染前后IEX-1mRNA和蛋白的表达情况,采用四甲基偶氮唑蓝(MTT)法检测转染后卵巢癌细胞SKOV3增殖情况以及对顺铂敏感性的变化。结果:成功转染IEX-1基因后可检测到IEX-1mRNA的高表达;转染组细胞的增殖速度增快(P<0.05),但对顺铂的敏感性明显增强(P<0.05)。结论:IEX-1可增强卵巢癌细胞的体外增殖能力,并增强卵巢癌细胞体外对顺铂化疗的敏感性,为肿瘤的有效干预和治疗提供新的靶点。
