NITRIC OXIDE AND FERRITIN MEDIATED TUMOR CELL RESISTANCE

1994年8月，我在哈佛大学医学系担任教职， 威斯康星州医学院（MCW）开始学术生涯 在Owen W. Griilith博士，董事长兼 生物化学系教授。我完成了一个奖学金， 明尼苏达大学（U of M），其中包括2年。 哈利·S博士的实验室Jacob和Greg M.维切洛蒂研究金 是明确的学术导向;我参加了课程和研讨会， 在科学会议（3次国家会议，3次 区域），并产生了3份手稿（1份已发表，1份已接受，1份已发表， 提交）。因为我在实验室的经历很短暂， 在MCW的职位取决于建立一个赞助商。格里菲斯博士 培训了许多博士后，并同意赞助早期阶段的 我的职业生涯我他为获得场地、设备和 为我的实验室提供技术支持，审查并批评了我的提案 广泛地，并帮助建立了与个人的联系， 我需要进一步培训的领域的专业知识。我的实验室毗邻 博士Chris Chitamber是我所在部门的一名教员，他发表了 在铁代谢领域有广泛的研究， 日报该提案是一个令人兴奋的机会，因为有机会 在专家指导下发展新技能。 该提案的目标是确定铁蛋白和一氧化氮的作用 (NO)肿瘤细胞对化疗的抵抗力。研究表明，这些 这些物质可以保护细胞免受氧化损伤。在密歇根大学的时候， 我们发现，内皮细胞暴露于铁化合物， 允许铁蛋白合成，对活化的中性粒细胞，H2O2， 和氧化的LDL用铁化合物类似地处理的肿瘤细胞变得 对博来霉素耐药。 我们检测了NO调节 铁蛋白的合成，并发现NO本身，诱导抵抗 氧化剂，特别是在氯化血红素暴露的细胞中。作为这个的延伸， 工作，我们将研究铁蛋白是否保护肿瘤细胞免受氧化 化疗造成的损伤。在詹妮弗·莫里斯博士的帮助下， 将进行研究，以解决铁蛋白保护的范围， 铁蛋白对损伤细胞中铁的影响，以及铁蛋白的特点， 铁蛋白是必要的保护。Albert Girotti博士和Dr. Chitamber将协助研究铁动力学和细胞损伤。我们 将研究化疗是否诱导肿瘤铁蛋白合成 细胞研究表明，放线菌素D和紫外线照射诱导 铁蛋白的合成，有理由推测化疗， 药物也可以。玛丽克莱尔·肯尼迪博士在 铁硫簇和铁蛋白调节领域，并将提供 化疗诱导铁蛋白改变的研究指南 新陈代谢.将对耐药肿瘤细胞进行研究，以确定 铁蛋白调节的改变可能有助于抗性。最后， 我们将研究淋巴瘤细胞产生的NO对 对化疗的敏感性。在威廉·安东尼博士的指导下， 我们将使用ESR来确定血红素加氧酶（HO）是否催化形成 作为一种保护机制。该提案是一个延伸 我作为一个研究员所做的工作，但包括方法， 超出了我的训练。我对这些研究的援助请求一直是 我满怀热情，希望能发展出许多新技能。

In August 1994, I took a faculty position in the Department of Medicine at the Medical College of Wisconsin (MCW) to begin a career in academic medicine under the sponsorship of Owen W. Griilith Ph.D., Chairman and Professor of the Department of Biochemistry. I completed a fellowship at the University of Minnesota (U of M) which included 2 yrs. in the laboratory of Drs. Harry S. Jacob and Greg M. Vercellotti. The fellowship is unequivically academically oriented; I attended classes and seminars in the basic sciences, presented at scientific meetings (3 national, 3 regional), and generated 3 manuscripts (1 published, 1 accepted, 1 submitted). Because my experience in the lab has been brief, accepting the position at MCW was contingent on establishing a sponsor. Dr. Griffith has trained numerous post-doc's, and has agreed to sponsor the early phase of my career. He has facilitated the acquisition of space, equipment and technical support for my laboratory, reviewed and critiqued my proposal extensively, and has helped establish contacts with individuals that have expertise in areas in which I need further training. My laboratory adjoins Dr. Chris Chitamber, a faculty member within my division, who has published extensively in the field of iron metabolism, and with whom I interact daily. The proposal is an exciting opportunity because of the chance to develop new skills under expert guidance. The goal of the proposal is to define the role of ferritin and nitric oxide (NO) in tumor cell resistance to chemotherapy. Studies suggest that these substances may defend cells against oxidative injury. While at the U of M, we found that endothelial cells exposed to iron compounds and incubated to allow ferritin synthesis, became resistant to activated neutrophils, H202, and oxidized LDL. Tumor cells similarly treated with iron compounds became resistance to bleomycin. We examined the ability of NO to modulate ferritin synthesis and found that NO, itself, induced resistance to oxidants, particularly in hemin exposed cells. As an extension of this work, we will study whether ferritin protects tumor cells against oxidative injury from chemotherapy. With help from Dr. Jennifer Morris, transfection studies will be conducted to address the scope of ferritin protection, the effect of ferritin on iron in the injured cell, and the features of ferritin that are necessary for protection. Dr. Albert Girotti and Dr. Chitamber will assist with studies of iron kinetics and cellular injury. We will investigate whether chemotherapy induces ferritin synthesis in tumor cells. Studies have shown that actinomycin D and UV irradiation induce ferritin synthesis, and there is reason to speculate that chemotherapeutic drugs may also. Dr. Mary Claire Kennedy has published extensively in the field of iron-sulfur clusters and ferritin regulation and will provide guidance in our studies of chemotherapy induced alterations of ferritin metabolism. Drug resistant tumor cells will be studied to determine whether alterations in ferritin regulation may contribute to resistance. Finally, we will study the effects of NO production by lymphoma cells on susceptibility to chemotherapy. With the guidance of Dr. William Antholine, we will use ESR to determine if heme-oxygenase (HO) catalyzes the formation of NO-heme adducts as a protective mechanism. The proposal is an extension of work that I performed as a fellow, but includes methodology that is beyond my training. My requests for assistance with these studies have been granted with enthusiasm and I expect to develop many new skills.