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The investigation on the expression regulation and the role on chemosensitivity of a xenobiotic enzyme GSTP1 in lung cancer.

肺癌中异生酶GSTP1的表达调控及其对化疗敏感性的研究

基本信息

批准号：
14570559
负责人：
MATSUSE Takeshi
金额：
$ 2.18万
依托单位：
Yokohama city university
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2003
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14570559/
关键词：
lung cencer chemosensitivity xenobiotic enzyme GSTP1 DNA methylation

项目摘要

1.Induction of CpG island methylation of GSTP1 promoter in lung adenocarcinoma cells.The CpG island of GSTP1 promoter was not methylated and GSTP1 was detected by immunoblotting in A549 lung adenocarcinoma cells. When we transduced the methylated oligonucleotides to these cells, methylation of the GSTP1 promoter and reduction of GSTP1 expression was induced.2.Induction of CpG island methylation with DNA methyltransferases (DNMT)It was observed that DNMT1, 3a, and 3b were all expressed in A549 cells. We cloned the cDNA of DNMT3b, and established a stable transfectant of DNMT3b from A549 cells. Now we check the methylation status of GSTP1 CpG island in these transfectants with bisulfite sequenting.3.Association between GSTP1 expression and viability of lung-derived cells (including lung cancer cells)We regulate the expression level of GSTP1 in human lung-derived fibroblast HFL-1 and A549 lung cancer cells, and investigated the role of GSTP1 on spontaneous apoptosis. Depletion of GSTP1 increased spontaneous apoptosis and necrosis in both kinds of cells. Although we investigated the mechanism of this phenomena, we found no relations between this phenomena and JNK pathway or intracellular concentration of glutathione.4.Association between GSTP1 expression and chemosensitivityAs a pilot study, we carried out transfection of GSTP1 sense and antisense vectors to examine effects of GSTP1 on apoptosis induced by camptothecin in HeLa cells. Camptothecin-induced apoptosis was attenuated by transfection of GSTP1 sense vector in HeLa cells.We also observed that GSTP1 depletion with siRNA increased camptothecin-induced cell death in A549 lung cancer cells.

1.肺腺癌细胞GSTP1启动子CpG岛甲基化的诱导：免疫印迹法检测到A549肺腺癌细胞GSTP1启动子的CpG岛未发生甲基化。2.DNA甲基转移酶(DNMT)诱导CpG岛甲基化A549细胞均表达DNMT1、3a和3b。我们克隆了Dnmt3b的基因，并从A549细胞中建立了稳定的Dnmt3b转染体。3.GSTP1表达与肺源性细胞(包括肺癌细胞)活力的关系我们调节人肺源性成纤维细胞HFL-1和A549肺癌细胞中GSTP1的表达水平，并探讨GSTP1在自发性凋亡中的作用。GSTP1缺失增加了两种细胞的自发性凋亡和坏死。尽管我们研究了这种现象的机制，但我们没有发现这种现象与JNK途径或细胞内谷胱甘肽浓度之间的关系。4.GSTP1表达与化疗敏感性的关系作为一项初步研究，我们进行了GSTP1正义和反义载体的转染，以检测GSTP1对喜树碱诱导的HeLa细胞凋亡的影响。GSTP1正义载体可减弱喜树碱诱导的HeLa细胞的凋亡。同时，我们还观察到GSTP1的缺失和siRNA增加了喜树碱诱导的A549肺癌细胞的死亡。