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Angiogenesis and lymphangiogenesis as molecular targets for oral cancer therapy

血管生成和淋巴管生成作为口腔癌治疗的分子靶点

基本信息

批准号：
15592107
负责人：
SUMI Tetsuro
金额：
$ 2.24万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15592107/
关键词：
Teratocarcinoma Cisplatin p34^<cdc2>リンパ管新生リンパ節転移

项目摘要

(1)F9 cells arrested at G21M with CDDPSince the treatment with CDDP causes complete inhibition of growth in F9 cells, the effect of CDDP on cell cycle progression was analyzed using fiow cytometry. At various times after treatment with 5 x 10〜6 M CDDP, aliquots of cells were removed for flow cytometric analysis to determine the cell cycle stage. From these findings, the cell cycle was arrested at the late S + G2/M phase.(2)Decrease in p34cdc2 by treatment with CDDPThe level of p34^<cdc2> in CDDP treated F9 cells was evaluated by Western blot analysis using anti- p34^<cdc2> antibody. Total cell lysates were prepared from cells collected at various times after treatment with CDDP. These lysates were assayed at equal cell numbers or at equal volumes of total protein. After the treatment with CDDP for 36 h, the level of p34^<cdc2> was decreased in the examination assay using equal cell numbers. In the assay using equal total proteins, a similar finding was observed after 36 h, but the level of p34^<cdc2> was increased for 24 h treatment with CDDP compared with the tTeatment for 0 h.(3)The cdc2 mRNA was stable after the CDDP treatmentTo clarify if the regulation of p34^<cdc2> by treatment with CDDP occurred in post- or pre-transcription, the level of cdc2 mRNA was estimated by Northern blotting analysis. By densitometric analysis, the level of cdc2 mRNA was stable for 36 h after treatment with CDDP.(4)Turnover of p34^<cdc2>To examine the half-life of p34^<cdc2> after treatment with CDDP, metabolic labeling and immunoprecipitation were carried out. The half-life of p34^<cdc2> in F9 cells treated with CDDP for 12 and 24 h was increased, but was decreased by treatment with CDDP for 36 h compared with control samples.

(1)F9用CDDP阻滞在G21 M的细胞由于用CDDP处理导致F9细胞生长的完全抑制，因此使用流式细胞术分析CDDP对细胞周期进程的影响。在用5 × 10 - 6 M CDDP处理后的不同时间，取出细胞等分试样进行流式细胞术分析，以确定细胞周期阶段。从这些发现，细胞周期被阻滞在晚期S + G2/M期。（2）用CDDP处理降低p34 β dc 2通过<cdc2>使用抗p34 β抗体的Western印迹分析来评估CDDP处理的F9细胞中p34 β的水平<cdc2>。从用CDDP处理后不同时间收集的细胞制备总细胞裂解物。这些裂解物以相等的细胞数或相等体积的总蛋白进行测定。用CDDP处理36 h后，<cdc2>在使用等细胞数的检测分析中，p34 β的水平降低。在使用等量总蛋白的测定中，在36 h后观察到类似的结果，但<cdc2>与0 h的tTreatment相比，用CDDP处理24 h的p34 β水平增加。（3）CDDP处理后cdc 2 mRNA水平稳定。为了阐明CDDP对p34 ~+的调控是<cdc2>发生在转录后还是转录前，我们用北方印迹法检测了cdc 2 mRNA水平。光密度分析显示，CDDP处理后36 h内cdc 2 mRNA水平稳定。（4）p34的转换。<cdc2>为了检测<cdc2>用CDDP处理后p34的半衰期，进行代谢标记和免疫沉淀。与<cdc2>对照样品相比，用CDDP处理12和24 h的F9细胞中p34 α的半衰期增加，但用CDDP处理36 h的F9细胞中p34 α的半衰期降低。