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Investigation of oral cancer development by using apoptosis and its application for clinical evaluation

利用细胞凋亡研究口腔癌发生发展及其在临床评价中的应用

基本信息

批准号：
12671836
负责人：
OHBA Takeshi
金额：
$ 2.11万
依托单位：
Kyushu Dental College
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12671836/
关键词：
Anti-canccer drug Apoptosis Egr-1 NORs Oral carcinoma Protein phosphatase Protein phosphatase Fas antigen nucleolin anti-cancerdrug

项目摘要

Oral squamous carcinoma cell line SSCKN cells were shown to be highly sensitive to bleomycin, whereas SCCKN cells were minimally sensitive to this reagent. Peplomycin caused apoptosis SCCKN and SCCTF cells in a dose-dependent fashion with the maximal effect at concentrations of 5 μM and 50 μM, respecnvely, as determined by phase-contrast microscopy, WST-1 cell viability assay, Hoechst 33342 staining, and DNA ladder formation. These results indicate that peplomycin-induced apoptosis in oral squamous carcinoma cell lines depends on the sensitivity of these cells to bleomycin.We examined the expression of Egr-1 in human squamous carcinoma cell line SCCKN cells treated with okada1C acid. Western blot analysis revealed that Egr-1 was strongly expressed in SCCKN cells and that okadaic acid decreased the expression of Egr-1 protein in these cells. Suppression of Egr-1 protein expression okadaic acid-treated cells stemmed from the suppression of the Egr-1 mRNA level, as determined by the RT-RCR method. A gel mobility shift assay showed that treatment of SCCKN cells with okadaic acid suppressed Egr-1 binding to the DIG-labeled Egr-1 consensus oligonucleotide probe. Treatment of SCCKN cells with okadaic acid suppressed the cell viability of SCCKN cells and induced apoptosis in these cells.The expression and cytolocalization of protein phosphatase type 1 δ isoform and nucleolin in human osteoblastic MG63 and Saos-2 cells. Anti-nucleolin antibody interacted with the 100- and 95-kDa proteins present in the whole cell lysate. The 100-kDa protein was detected in nuclear and nucleolar fractions. The 95-kDa protein was detected in cytosolic and nucleoplasmic fractions. Protein phosphatase 1δ and nucleolin were co-localized and interact with each other in nucleolus in MG63 and Saos-2 cells revealed bv immunofluorescent and immunoprecipitation method.

口腔鳞状细胞癌细胞系SSCKN细胞对博来霉素高度敏感，而SCCKN细胞对该试剂的敏感性最低。通过相差显微镜、WST-1细胞活力测定、Hoechst 33342染色和DNA梯状条带形成测定，发现5 μM和50 μM的培洛霉素可诱导SCCKN和SCCTF细胞凋亡，且呈剂量依赖性。这些结果表明，在口腔鳞状细胞癌细胞系中，培洛霉素诱导的凋亡依赖于这些细胞的敏感性，以博莱霉素。我们研究了Egr-1在人鳞状细胞癌细胞系SCCKN细胞与冈田酸处理的表达。Western blot分析显示Egr-1在SCCKN细胞中强烈表达，冈田酸降低了Egr-1蛋白在这些细胞中的表达。通过RT-RCR方法测定，冈田酸处理的细胞对Egr-1蛋白表达的抑制源于对Egr-1 mRNA水平的抑制。凝胶迁移率变动分析显示，用冈田酸处理SCCKN细胞抑制了Egr-1与DIG标记的Egr-1共有寡核苷酸探针的结合。冈田酸可抑制SCCKN细胞的生长，诱导其凋亡。蛋白磷酸酶1 δ亚型和核仁素在人成骨细胞MG 63和Saos-2中的表达和定位抗核仁素抗体与全细胞裂解物中存在的100-和95-kDa蛋白质相互作用。100-kDa的蛋白质中检测到的核和核仁组分。在胞质和核质组分中检测到95-kDa蛋白。用免疫荧光和免疫沉淀方法研究了MG 63和Saos-2细胞核仁中蛋白磷酸酶1δ和核仁素的共定位和相互作用。