GENE EXPRESSION IN IMMORTAL HUMAN CELLS

我提出了一个两阶段的培训计划， 胃肠道肿瘤学的研究生涯在第一阶段，我将 学习分子生物学、分子遗传学、病毒学和癌症课程 生物医学科学研究生院的生物学/遗传学。我会 发展细胞和分子生物学的实验室技能， 研究计划，这将成为第二阶段的主要重点。 二十 %的努力将用于临床胃肠病学和肿瘤学， 保持我的临床技能 在体内，癌细胞逃避正常的生长调节， 自主地。培养中的恶性细胞表现相似， 仙 正常的人体细胞很少有不死的。因此 永生化似乎是人类致癌作用中的重要步骤。 将通过使用一个独特的系列来研究向不朽的转化 匹配的永生前和永生SV 40转化的人成纤维细胞 （SV 1HF）细胞系。 一个cDNA文库已经从一个永生的 SV/HF（SVtsA/HF-C）的杂交，并通过差减杂交进行分析。一系列 已经鉴定了检测mRNA的cDNA， 在永生细胞系中表达。 代表性cDNA将进一步 以评估它们在永生化中的作用。mRNA低表达 将测试永生细胞的生长抑制功能。mRNA 将测试在永生细胞系中过表达的细胞的生长 刺激. 全长cDNA将从合适的细胞中分离。 表达式库。它对正常增殖细胞生长的影响 衰老细胞、永生前细胞和永生细胞将使用 瞬时和稳定表达测定。 几种永生细胞系将 进行研究以验证一致性。确定次级效应器 永生可能会导致癌症预防，检测 和治疗。这些方法将在以后应用于具体的 胃肠道恶性肿瘤研究

I propose a two-phase plan for training in preparation for an academic research career in gastrointestinal oncology. In the first phase, I will take courses in molecular biology, molecular genetics, virology and cancer biology/genetics in the Graduate School of Biomedical Sciences. l will develop laboratory skills in cell and molecular biology and initiate a research plan which will become the major focus of phase II. Twenty percent effort will be spent in clinical gastroenterology and oncology to maintain my clinical skills. In vivo, cancer cells escape normal growth regulation and behave autonomously. Malignant cells in culture behave similarly and are immortal. Few, if any, normal human cells show immortality. Therefore immortalization appears to be an important step in human carcinogenesis. The transformation to immortality will be studied by using a unique series of matched preimmortal and immortal SV40 transformed human fibroblast (SV1HF) cell lines. A cDNA library has been prepared from a preimmortal SV/HF (SVtsA/HF-C) and analyzed by subtraction hybridization. A series of cDNAs have been identified which detect mRNAs which are differentially expressed in immortal cell lines. Representative cDNAs will be further characterized to assess their role in immortalization. mRNA underexpressed in immortal cells will be tested for growth suppressor function. mRNA overexpressed in the immortal cell line will be tested for growth stimulation. The full length cDNA will- be isolated from the appropriate expression library. Its effect on growth in normal proliferating and senescent cells, preimmortal and immortal cells will be determined using transient and stable expression assays. Several immortal cell lines will be studied to verify consistency. Determining the secondary effectors of immortality may lead to novel approaches in cancer prevention, detection and treatment. These approaches will later be applied to the specific study of gastrointestinal malignancy.