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Analyses of the induction of gene expressions by sonication and their signal transductions

超声诱导基因表达及其信号转导分析

基本信息

批准号：
16500314
负责人：
OGAWA Ryohei
金额：
$ 1.73万
依托单位：
University of Toyama (2005)Toyama Medical and Pharmaceutical University (2004)
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

项目摘要

Induction of the heme oxygenase 1(HO1) gene expression of DU145 cells derived from human prostate cancer was shown to be dependent upon intensity and duration of sonication applied, in addition, associated were decrease in mitochondria membrane potential and increase in intracellular superoxide concentration. When free radical scavengers that could enter a cell were added before or immediately after sonication, HO1 expression was suppressed, suggesting that reactive oxygen species secondarily generated in the cell was involved in the induction. At the same time, out of 3 subcloned DNA fragments of 10 to 5.8 kb,5.8 to 4.5 kb and 4.5 to 0 kb relative to the mRNA start site of the HO1 gene, the 10 to 5.8 kb fragment and the 4.5 to 0 kb fragment were found responsive to sonication. The either fragment contains multiple numbers of a nucleotide sequence motif, Stress Response Element (StRE), that may be involved in oxidative stress. When mutations or deletions were applied to the motifs, the promoter activity responsive to sonication was significantly affected. Thus, it was suggested that activation of Nrf2,a transcription factor associating with StRE, was facilitated by the intracellular oxidative stress generated from mitochondria, resulting in induction of HO1 gene expression.We cloned DNA fragments containing promoter regions of cycloxigenase-2,flt-1,bmp-7 and c-fos genes that are involved in osteogenesis and linked to the luciferase gene, constructing gene cassettes. The gene cassettes were introduced into human osteoblast cells and sonicated with 1 MHz ultrasound at intensities of 0.1 to 0.5 W/cm^2, duty cycles of 5 to 10%, for 5 to 30 min. After incubation for 6 to 48 hrs, activations of the promoters were accessed. However, any induction of luciferase activity was confirmed including the cycloxigenase-2 promoter that previously showed induction after sonication in a similar condition.

血红素加氧酶1（HO 1）基因表达的诱导来自人前列腺癌的DU 145细胞被证明是依赖于强度和持续时间的超声波处理，此外，相关的是线粒体膜电位的降低和细胞内超氧化物浓度的增加。当自由基清除剂，可以进入细胞前或后立即加入超声，HO 1的表达被抑制，表明活性氧在细胞中产生的二次参与了诱导。同时，在相对于HO 1基因mRNA起始位点的10 ~ 5.8kb、5.8 ~ 4.5kb和4.5 ~ 0 kb的3个亚克隆DNA片段中，发现10 ~ 5.8kb和4.5 ~ 0 kb的片段对超声有反应。任一片段含有多个核苷酸序列基序，应激反应元件（StRE），其可能参与氧化应激。当突变或缺失应用于基序时，响应于超声处理的启动子活性受到显著影响。因此，这表明，激活Nrf 2，与StRE相关的转录因子，是由线粒体产生的细胞内的氧化应激，导致诱导HO 1基因的expression.We克隆的DNA片段包含环氧化酶-2，flt-1，bmp-7和c-fos基因的启动子区，参与骨生成和连接到荧光素酶基因，构建基因盒。将基因盒导入人成骨细胞中，用1 MHz的超声波以0.1 ~ 0.5 W/cm ^2的强度、5 ~ 10%的占空比超声处理5 ~ 30 min。孵育6 ~ 48 h后，检测启动子的激活情况。然而，确认了荧光素酶活性的任何诱导，包括先前在类似条件下超声处理后显示诱导的环氧化酶-2启动子。

项目成果

期刊论文数量（46）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Expression of heme oxygenase-1 due to intracellular reactive oxygen species induced by ultrasound

DOI：
10.1016/j.ultsonch.2005.09.004
发表时间：
2006-07-01
期刊：
ULTRASONICS SONOCHEMISTRY
影响因子：
8.4
作者：
Kagiya, Go;Ogawa, Ryohei;Kondo, Takashi
通讯作者：
Kondo, Takashi

Basic studies on gene therapy with ultrasound.

超声基因治疗的基础研究。

DOI：
发表时间：
2005
期刊：
Medical and Biological Engineering 43
影响因子：
0
作者：
Ogawa;Ryohei;et al.
通讯作者：
et al.

Identification of genes responsive to low intensity pulsed ultrasound in a human leukemia cell line Molt-4.

DOI：
10.1016/j.canlet.2006.02.011
发表时间：
2007-02
期刊：
Cancer letters
影响因子：
9.7
作者：
Y. Tabuchi;H. Ando;I. Takasaki;L. B. Feril;Qing‐Li Zhao;R. Ogawa;N. Kudo;K. Tachibana;T. Kondo
通讯作者：
Y. Tabuchi;H. Ando;I. Takasaki;L. B. Feril;Qing‐Li Zhao;R. Ogawa;N. Kudo;K. Tachibana;T. Kondo

Bio-effects of ultrasound at a molecular level and their application in medicine

超声在分子水平上的生物效应及其在医学中的应用