GROWTH FACTORS IN MAMMALIAN EYE DEVELOPMENT

描述：(候选人摘要)这项提议的最终目的 就是制定一个多方面的计划来指导我的发展 作为一名独立科学家。这一目标是通过一个 基础研究和教育活动的混合体。我的研究将 重点研究神经营养因子的生物学功能 脑源性神经营养因子和神经营养因子-3在脑出血中的表达 哺乳动物眼睛的发育和维持。这些努力 将辅以校外讲习班、眼科指导 病理学，以及参加科学会议。 在过去的几年里，我一直在完成我的博士研究和 专业培训。我现在准备开始一份长期的职业生涯 在学术神经眼科，临床实践与基础相结合 研究。科罗拉多大学将为我提供一个理想的 在这样的环境中，我可以实现眼前和长期的目标。这个 神经内科的综合教员和资源， 眼科学，以及分子、细胞和发育生物学 特别适合为我提供紧张的训练 系统和眼睛发育、小鼠遗传学、组织化学和 转基因和胚胎干细胞技术。 在凯文·琼斯博士的研究指导下，我将调查 脑源性神经营养因子和神经营养因子-3在神经视网膜和前节中的生物学作用 眼睛的。多学科方法将解决三个方面的问题 主要领域。 第一，脑源性神经营养因子和神经营养因子-3的细胞特异性表达及其相互关系 各自的受体。TrkB和TrkC，将在 神经视网膜、晶状体、睫状体和角膜。我们将确定 双标记表达BDNF、NT-3、TrkB和TrkC的特异性细胞 免疫组织化学和共聚焦显微镜检查。这些结果将表明 这些神经营养素在眼睛中的可能功能。 其次，BDNF和NT-3在神经视网膜中的生物学功能将 使用分区域受限的基因敲除技术进行研究。 将产生神经视网膜中缺乏BDNF和NT-3的小鼠。至 要做到这一点，我们将使用Cre/loxP介导的重组，受 Six3或BRN-3b启动子。将表达Cre重组酶 或者作为转基因或作为双顺反子信息使用干预 核糖体进入序列(IRES)。 第三，脑源性神经营养因子和神经营养因子-3在颈椎前段的生物学作用 眼睛将在我们最近发现的受管制的 神经营养因子在睫状体上皮、晶状体上皮、 还有角膜。限制性基因敲除将使用以下方法构建 内源性水通道蛋白-1基因驱动的双顺反子Cre表达。 这些研究应该为分子遗传学提供新的见解。 可能会拓宽我们对眼睛发育的理解 退行性眼病及其治疗的新途径 增长因素。

DESCRIPTION: (Candidate's Abstract) The ultimate aim of this proposal is to formulate a multifaceted program which will guide my development as an independent scientist. This goal is to be accomplished through a mixture of basic research and educational activities. My research will be focused on determining the biologic function of the neurotrophins brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) in the development and maintenance of the mammalian eye. These endeavors will be complemented by extramural workshops, instruction in ocular pathology, and participation at scientific meetings. I have spent the past several years completing my doctoral research and professional training. I am now prepared to embark on a long-term career in academic neuro-ophthalmology, combining clinical practice with basic research. The University of Colorado will provide me with an ideal environment in which to accomplish my immediate and long-term goals. The combined faculties and resources of the Department of Neurology, Ophthalmology, and Molecular, Cellular, and Developmental Biology are particularly well suited for providing me with training in nervous system and eye development, mouse genetics, histochemistry, and transgenic and embryonic stem cell technology. Under the research mentorship of Dr. Kevin Jones, I will investigate the biologic roles of BDNF and NT-3 in the neuroretina and anterior segment of the eye. A multidisciplinary approach will address questions in three main areas. First, the cell specific expression of BDNF and NT-3 and their respective receptors. TrkB and TrkC, will be examined in the neuroretina, lens, ciliary body, and cornea. We will identify the specific cells expressing BDNF, NT-3, TrkB, and TrkC using dual-label immunohistochemistry and confocal microscopy. These results will suggest possible functions for these neurotrophins in the eye. Second, the biologic functions of BDNF and NT-3 in the neuroretina will be investigated using subregion-restricted gene knockout technology. Mice will be generated which lack BDNF and NT-3 in the neuroretina. To achieve this, we will use Cre/loxP-mediated recombination restricted by either the Six3 or Brn-3b promoter. Cre recombinase will be expressed either as a transgene or as a dicistronic message using an intervening ribosome entry sequence (IRES). Third, the biologic role of BDNF and NT-3 in the anterior segment of the eye will be characterized following our recent finding of regulated neurotrophin expression in the ciliary body epithelium, lens epithelium, and cornea. Restricted gene knockouts will be constructed using dicistronic Cre expression driven from the endogenous aquaporin-1 gene. These studies should provide novel insight into the molecular genetics of eye development, potentially broaden our understanding of degenerative eye disorders and open new avenues for their treatment with growth factors.