Genetic approaches to skin regeneration in zebrafish

斑马鱼皮肤再生的遗传方法

• 批准号: 10112829
• 负责人: David Andrew Brown
• 金额: $ 16.56万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-03-01 至 2025-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10112829
• 关键词: Award; Basal Cell; Behavior; Biological Assay; Biomedical Engineering; Cell Cycle Progression; Cells; Chromatin; Complex; Data; Defect; Development; Development Plans; Developmental Biology; Doctor of Medicine; Doctor of Philosophy; Educational Status; Elements; Epidermis; Epithelial; Equilibrium; Event; Evolution; Exhibits; Faculty; Future; Gene Expression; Gene Transfer Techniques; Genetic; Genetic Enhancer Element; Genetic Transcription; Genomics; Goals; Heart; Human; Image; Imaging technology; Individual; Innovative Therapy; K-Series Research Career Programs; Laboratories; Lead; Learning; Link; Mammals; Maps; Mediating; Mentorship; Modeling; Molecular; Morphology; Natural regeneration; Nucleic Acid Regulatory Sequences; Optics; Organism; Pathway interactions; Pattern; Phylogenetic Analysis; Plastic Surgeon; Positioning Attribute; Process; Regulation; Regulatory Element; Research; Research Personnel; Resolution; Role; Scientist; Shapes; Signal Transduction; Site; Skin; Skin injury; Spinal Cord; Structure; Surgeon; Techniques; Testing; Tissue Engineering; Tissue Model; Tissues; Training; Transcriptional Regulation; Transgenic Organisms; Universities; Work; Wound models; Zebrafish; appendage; base; career; career development; chronic wound; design; epidermal stem cell; epithelial wound; experimental study; genetic approach; genetic manipulation; genetic technology; imaging genetics; improved; in vivo; in vivo imaging; insight; keratinocyte; member; migration; neonatal mice; novel; novel strategies; prevent; programs; quantitative imaging; reconstruction; regenerative; regenerative approach; regenerative biology; regenerative therapy; skin regeneration; skin wound; tissue regeneration; tissue repair; tool; transcriptome sequencing; wound; wound bed; wound healing

PROJECT SUMMARY/ABSTRACT The proposed career development award is designed to support the transition of David A. Brown, M.D., Ph.D., to an independent investigator in the field of wound healing. Dr. Brown is a plastic and reconstructive surgeon at Duke University with a Ph.D. in tissue engineering. His long-term career goal is to utilize principles of bioengineering and regeneration biology to develop novel regenerative therapies for chronic wounds. Advanced training in regeneration biology is proposed, comprised of laboratory work under the mentorship of Kenneth Poss, Ph.D., a leader in the field of zebrafish models for tissue regeneration, as well as structured activities, coursework, and mentorship from faculty in related fields. The project will focus on re- epithelialization, which is mediated by migration of basal epidermal keratinocytes and stands as a central process in wound healing required to prevent progression to chronic wounds. Zebrafish harbor an innate ability to re-epithelialize wounds and regenerate skin more rapidly and completely than mammals, although the underlying mechanisms of this ability remain unclear. Based on prior work that demonstrates the existence of tissue regeneration enhancer elements (TREEs) involved in zebrafish heart and appendage regeneration, the central hypothesis of this project is that re-epithelialization is governed by regeneration-linked regulatory elements that orchestrate coordinated basal keratinocyte behaviors and gene expression. This proposal represents a novel approach to wound healing research and will provide the candidate with high-level training in live imaging, transgenesis, and genomics that will supplement his background in tissue engineering. In Aim 1, live clonal analysis and transgenic zebrafish lines will be used to map dynamic basal keratinocytes behaviors in re-epithelialization. Experiments will track cell fate decisions and morphologic changes during re- epithelialization and identify subpopulations of basal cells and their relative contributions to the neoepidermis. A quantitative imaging practical course will be pursued along with mentorship from experts in wound healing and live imaging. In Aim 2, the candidate will determine the role of the 103runx1 TREE in basal keratinocyte fate decisions, then use transcriptome sequencing, chromatin profiling, and transgenic assays to identify other key cis-acting regulatory sequences involved in skin regeneration. University courses in transcriptional regulation and computational genomics will be completed as well. Altogether, these experiments are expected to link critical cell fate decisions of basal keratinocytes to activation of specific regulatory elements. Future studies will test the hypothesis that genetic manipulation of these elements can enhance re-epithelialization by promoting basal keratinocyte migration and proliferation, thus leading to improved wound healing. This project will position the candidate to submit an R01 application in the final years of the award period, which will be focused on the next translational step in a mammalian model.

项目总结/摘要 拟议的职业发展奖旨在支持大卫A的过渡。布朗，医学博士，哲学博士、 伤口愈合领域的独立调查员布朗医生是一位整形外科医生 在杜克大学获得博士学位在组织工程学中。他的长期职业目标是利用 生物工程和再生生物学为慢性伤口开发新型再生疗法。 再生生物学的高级培训建议，包括实验室工作的指导下， 肯尼斯·波斯博士，斑马鱼组织再生模型领域的领导者，以及结构化的 活动，课程，并从教师在相关领域的指导。该项目将侧重于重新- 上皮形成，其由基底表皮角质形成细胞的迁移介导，并作为中央上皮形成的一部分。 在伤口愈合过程中需要防止进展为慢性伤口。斑马鱼有一种天生的能力 比哺乳动物更迅速和完全地使伤口重新上皮化和再生皮肤，尽管 这种能力的潜在机制仍不清楚。根据先前的工作，证明存在的 组织再生增强因子（TREE）参与斑马鱼心脏和附件再生， 该项目的中心假设是，上皮再生是由再生相关的调节因子控制的。 协调基础角质形成细胞行为和基因表达的元素。这项建议 代表了伤口愈合研究的新方法，并将为候选人提供高水平的培训 在活体成像、转基因和基因组学方面，这将补充他在组织工程方面的背景。在Aim中 1、活克隆分析和转基因斑马鱼品系将用于基底角质形成细胞的动态作图 再上皮化的行为。实验将跟踪细胞命运的决定和形态学变化， 上皮形成和鉴定基底细胞的亚群及其对新表皮的相对贡献。 一个定量成像实践课程将沿着与指导专家在伤口愈合 和实时成像。在目标2中，候选人将确定103 runx 1树在基底角质形成细胞中的作用。 命运的决定，然后使用转录组测序，染色质分析和转基因检测，以确定其他 关键的顺式作用调控序列参与皮肤再生。大学转录课程 调控和计算基因组学也将完成。总的来说，这些实验预计 将基底角质形成细胞的关键细胞命运决定与特定调节元件的激活联系起来。未来 研究将检验这一假设，即对这些元素的遗传操作可以通过以下方式增强上皮再生： 促进基底角质形成细胞迁移和增殖，从而改善伤口愈合。这个项目 将使候选人在奖励期的最后几年提交R 01申请， 专注于哺乳动物模型中的下一个翻译步骤。