Integrated Microfluidics for Nerve Regeneration Studies using Laser Nano-Axotomy

使用激光纳米轴切术进行神经再生研究的集成微流体

• 批准号: 7477675
• 负责人: ADELA BEN-YAKAR
• 金额: $ 18.63万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7477675
• 关键词: Ablation; Adverse effects; Affect; Age; Alzheimer' s Disease; Anesthetics; Animals; Area; Automation; Axon; Axotomy; Biological; Caenorhabditis elegans; Cells; Chemicals; Complex; Computer Assisted; Computers; Development; Devices; Dimensions; Disease; Gene Family; Genes; Genetic; Goals; Growth; Heating; Human; Huntington Disease; Image; Immobilization; Individual; Investigation; Lasers; Membrane; Microfluidic Microchips; Microfluidics; Mus; Natural regeneration; Nematoda; Nerve Regeneration; Nervous system structure; Neurodegenerative Disorders; Operative Surgical Procedures; Organism; Parkinson Disease; Performance; Pharmaceutical Preparations; Physiologic pulse; Process; Pulse taking; Recovery; Recovery of Function; Research; Research Project Grants; Resveratrol; Role; Screening procedure; Shapes; Speed; Stress; System; Techniques; Thick; Time; Tissues; Width; Zebrafish; axon regeneration; cost; design; developmental neurobiology; feeding; follow-up; high throughput screening; in vivo; injured; nano; nanosurgery; nervous system disorder; novel; pressure; programs; size; tool

DESCRIPTION (provided by applicant): Understanding the biological mechanisms of nerve regeneration and degeneration is an important step towards the development of novel therapies for human neurodegenerative diseases such as Alzheimer's, Parkinson's, Huntington's, and other neurological disorders affecting millions of people. The regeneration processes can be studied by severing an axon in a controlled manner and then observing its re-growth and functional recovery. To sever an axon in-vivo, a high precision and non-intrusive cutting tool is required. In the absence of precision techniques for severing axons (axotomy), investigations have so far been limited to complex organisms (mouse and zebrafish). Just recently we demonstrated that femtosecond laser pulses can be used for axotomy in the roundworm Caenorhabditis elegans (C. elegans) and those axons functionally can regenerate after the operation. The advantage of these ultrashort laser pulses is their ability to evaporate an extremely small volume of tissue without heating or damaging the surrounding cells. Application of this precise surgical technique now enables nerve regeneration to be studied in-vivo in organisms with simple nervous systems. Since simple organisms such as C. elegans have amenable genetics, application of the femtosecond laser axotomy technique will help in the rapid identification of genes and molecules that affect nerve regeneration and degeneration. The goal of this research project is to develop a high-throughput laser nano-surgery platform for axon regeneration & degeneration studies in C. elegans in-vivo. Specifically, an integrated microfluidic device will be developed to trap the worms. This microfluidic trap will facilitate the appropriate immobilization of the animals for precision nanosurgery of axons using femtosecond laser pulses and for imaging the re-growth of the injured axons following nanosurgery. Development of a high throughput screening platform requires integration of different modules for nanosurgery, feeding, and imaging and their synchronization through computer controlled automation. A platform having 100's of chambers for feeding of individual worms will facilitate automated surgical and screening studies of multiple worms, thus greatly reducing time and cost. For successful development of a high-throughout nanosurgery platform for in-vivo nerve regeneration/degeneration studies, we have assembled a multi-disciplinary team with expertise in three critical areas (1) ultrafast laser ablation of bio-materials (Dr. Ben-Yakar), (2) fabrication and assembly of microfluidic systems (Dr. Chronis), and (3) developmental neurobiology of C. elegans (Dr. Bargmann). Concurrently with the development of a high-throughput microfluidic platform, two important nerve regeneration/degeneration problems will be investigated: (1) time-lapse in-vivo imaging of axon re-growth and degeneration of injured axons and (2) the role of the Sir2 family of genes and the drug resveratrol in regeneration of injured axons. Understanding the biological mechanisms of nerve regeneration and degeneration is an important step towards the development of novel therapies for human neurodegenerative diseases such as Alzheimer's, Parkinson's, Huntington's, and other neurological disorders affecting millions of people. Development of a high-throughput laser nano-surgery platform applicable to simple organism such as C. elegans in-vivo will help in the rapid identification of genes and molecules that affect nerve regeneration and degeneration.

描述(申请人提供)：了解神经再生和退化的生物学机制是为人类神经退行性疾病(如阿尔茨海默氏症、帕金森氏症、亨廷顿氏症和其他影响数百万人的神经疾病)开发新疗法的重要一步。可以通过有控制地切断轴突，然后观察其重新生长和功能恢复来研究再生过程。要在活体内切断轴突，需要一种高精度的非侵入性切割工具。在缺乏切断轴突(轴突切断)的精确技术的情况下，到目前为止，研究仅限于复杂的生物(老鼠和斑马鱼)。就在最近，我们证明了飞秒激光脉冲可以用于线虫的轴突切断，并且这些轴突在功能上可以在手术后再生。这些超短激光脉冲的优势在于，它们能够蒸发极小体积的组织，而不会加热或破坏周围的细胞。应用这种精确的外科技术，现在可以在只有简单神经系统的生物体中研究神经再生。由于线虫等简单生物具有顺从的遗传学，应用飞秒激光轴突切断术将有助于快速识别影响神经再生和退化的基因和分子。本研究项目的目标是开发一个高通量激光纳米手术平台，用于线虫体内轴突再生和变性的研究。具体地说，将开发一种集成的微流控设备来捕捉蠕虫。这种微流体捕捉器将有助于适当地固定动物，以便使用飞秒激光脉冲进行精确的轴突纳米手术，并对纳米手术后受损轴突的重新生长进行成像。高通量筛选平台的开发需要集成用于纳米手术、喂养和成像的不同模块，并通过计算机控制的自动化来实现它们的同步。一个拥有100英尺S单虫饲养室的平台将有助于多条蠕虫的自动化手术和筛查研究，从而大大减少时间和成本。为了成功开发用于体内神经再生/变性研究的高通量纳米外科平台，我们组建了一个拥有三个关键领域专业知识的多学科团队(1)超快激光生物材料消融(Ben-Yakar博士)，(2)微流体系统的制造和组装(Chronis博士)，(3)线虫发育神经生物学(Bargmann博士)。在开发高通量微流控平台的同时，还将研究两个重要的神经再生/变性问题：(1)损伤轴突再生和变性的在体时延成像；(2)Sir2基因家族和药物白藜芦醇在损伤轴突再生中的作用。了解神经再生和退化的生物学机制是开发新的治疗人类神经退行性疾病的重要一步，这些疾病包括阿尔茨海默氏症、帕金森氏症、亨廷顿氏症和其他影响数百万人的神经疾病。开发一个适用于体内线虫等简单生物体的高通量激光纳米手术平台，将有助于快速识别影响神经再生和退化的基因和分子。

项目成果

Active segmentation of 3D axonal images.

3D 轴突图像的主动分割。

• DOI: 10.1109/embc.2012.6346845
• 发表时间: 2012
• 期刊: Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Annual International Conference
• 作者: Muralidhar,GautamS;Gopinath,Ajay;Bovik,AlanC;Ben-Yakar,Adela
• 通讯作者: Ben-Yakar,Adela

A fully automated microfluidic femtosecond laser axotomy platform for nerve regeneration studies in C. elegans.

• DOI: 10.1371/journal.pone.0113917
• 发表时间: 2014
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Gokce SK;Guo SX;Ghorashian N;Everett WN;Jarrell T;Kottek A;Bovik AC;Ben-Yakar A
• 通讯作者: Ben-Yakar A

A multi-trap microfluidic chip enabling longitudinal studies of nerve regeneration in Caenorhabditis elegans.

多陷阱的微流体芯片，实现了秀丽隐杆线虫神经再生的纵向研究。

• DOI: 10.1038/s41598-017-10302-4
• 发表时间: 2017-08-29
• 期刊: Scientific reports
• 影响因子: 4.6
• 作者: Gokce SK;Hegarty EM;Mondal S;Zhao P;Ghorashian N;Hilliard MA;Ben-Yakar A
• 通讯作者: Ben-Yakar A

Femtosecond laser nanoaxotomy properties and their effect on axonal recovery in C. elegans: erratum.

飞秒激光纳米轴切术特性及其对线虫轴突恢复的影响：勘误表。

• 发表时间: 2008
• 期刊: Optics express
• 影响因子: 3.8
• 作者: Bourgeois,Frederic;Ben-Yakar,Adela
• 通讯作者: Ben-Yakar,Adela