Immunological and Molecular Biological Study on Oncogenes In Human Oral Cancer.

1)应用抗p21单抗免疫组织化学方法检测ras p21在口腔癌组织中的表达。不仅恶性肿瘤阳性，良性病变也呈阳性，且染色强度呈相对强弱。因此，对临床恶性程度进行评估似乎是不可能的。此外，我们还研究了7个口腔癌细胞系中Rb基因的表达，发现所有细胞系中Rb基因的表达都是正常的。2)EJ c-H-ras-1的2.9kb的Sac I片段有4个编码外显子，但没有自己的5‘端启动子区域，3’端的VTR与肿瘤病毒的启动子/增强子或仅在3‘端的增强子区域反向连接。将pUC18中的这些构建物导入NIH3T3细胞中，发现其诱导病灶的效率约为启动子/增强子片段与片段同向连接的相应构建体的10%，而截短的c-H-ras-1单独的转化效率不到1%。结果表明，病毒增强剂提高了截短的c-H-ras-1的转化活性。为了进一步研究这种增强作用，在与Moloney LTR反向连接的截短的e-H-ras-1中引入上游缺失，并将缺失质粒导入NIH3T3细胞。起始密码子上游-615位至-150位的缺失并未显著降低转化活性，而-150位至-50位缺失突变体的转化活性较低，提示控制元件可能位于该区域内。我们还检测了截短的c-H-ras-1与MoMSV LTR连接的c-myc在3Y1细胞中的转化活性。诱导灶数目增加数倍，表明c-myc增强了截短的c-H-ras-1的转化活性。

1) The ras p21 expression in human oral carcinoma tissue specimens was studied by immunohistochemistry with monoclonal antibodies against p2l. Not only malignant tumors, but also benigns were stained positively, and the staining intensities were relative. Therefore, it seemed to be impossible to evaluate the clinical malignancy. In addition, the expressions of Rb gene mRNA was investigated in seven human oral carcinoma cell lines and found to be normal in all of the cell lines.2) The 2.9kb Sac I fragment of EJ c-H-ras-1 with 4 coding exons but without its own 5' promoter region and the VTR at 3' side was linked to tumor viruses's promoter/enhancer or only enhancer region at the 3' end in the opposite direction. These constructions in pUC18 plasmid were transfected onto NIH3T3 cells and found to induce foci with efficiencies of about 10% of those of the corresponding constructs in which the promoter/enhancer fragments were linked to the fragment in the same direction, while the transforming efficiency of the truncated c-H-ras-1 alone was less than 1%. The results suggests that viral enhancers increase transforming activity of the truncated c-H-ras-1. To further study the enhancement, upstream deletions were introduced into the truncated e-H-ras-1 linked to Moloney LTR in the opposite direction, and deletion plasmids were transfected onto NIH3T3 cells. Deletions from the upstream SacI site (-615) to around -150 from the initiation codon did not significantly lower the transforming activities, while the transforming activities of deletion mutants from -15O to -50 were low, suggesting that the controlling element may reside within this region. We examined also transforming activity of the truncated c-H-ras-1 by cotransfecting with c-myc linked to MoMSV LTR, in 3Y1 cells. The number of induced foci increased several fold, suggesting that c-myc enhances transforming activity of the truncated c-H-ras-1.