GENE EXPRESSION IN IMMORTAL HUMAN CELLS

我提出了一个分两个阶段的培训计划，为毕业前的 胃肠道肿瘤学研究生涯。在第一阶段，我将 参加分子生物学、分子遗传学、病毒学和癌症方面的课程 生物医学科学研究生院生物学/遗传学。L将 培养细胞和分子生物学的实验室技能，并发起 将成为第二阶段的主要重点的研究计划 将在临床胃肠病学和肿瘤学方面投入1%的精力 保持我的临床技能。 在体内，癌细胞逃脱了正常的生长调节并表现出 自主地。培养中的恶性细胞的行为类似，并且 不朽的。很少(如果有的话)正常的人类细胞显示出永生。因此 永生化似乎是人类致癌过程中的重要一步。 我们将用一个独特的系列来研究长生不老的转变。 相合的永生前和永生SV40转化的人成纤维细胞 (SV1HF)细胞株。从一名未长生不老的人身上制备了一个cdna文库。 SV/HF(SVtsA/HF-C)。一系列 已经确定了检测差异表达的mRNA的cDNA 在永生细胞系中表达。具有代表性的cDNA将进一步 其特点是评估它们在不朽中的作用。MRNA低表达 在永生细胞中将进行生长抑制功能的测试。mrna 在永生细胞系中过度表达的细胞将进行生长测试 刺激。全长cdna将从适当的 表达式库。它对正常增殖期和增殖期生长的影响 衰老细胞、永生前细胞和永生细胞将用 瞬时和稳定表达分析。几个不朽的细胞系将 被研究以验证一致性。确定的次级效应器 永生可能导致癌症预防、检测的新方法 和治疗。这些方法稍后将应用于特定的 胃肠道恶性肿瘤的研究。

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.