Probing the Bioeffects of Cavitation at Single-Cell Level

描述（由申请人提供）：由于其非侵入性和非电离性，治疗性超声和聚焦冲击波已广泛用于临床治疗癌症，靶向药物递送（如打开血脑屏障），结痂崩解，软组织分离，骨折不愈合愈合以及疼痛治疗。尽管在所有这些应用中，空化通常被认为是产生理想治疗效果的关键机制，在某些情况下，产生不期望的副作用，但泡-组织相互作用，特别是泡-细胞相互作用的动态过程在很大程度上是未知的。这主要是由于缺乏可行的实验系统，可以用足够的空间和时间分辨率来研究这种动态相互作用。为了克服这一主要挑战，我们建议开发一种新的实验系统和相关技术，在单细胞水平上研究空化气泡产生的生物效应。该项目将由一个多学科研究团队进行，该团队在治疗超声和空化（钟实验室）和细胞力学（Guilak实验室）方面具有公认的专业知识。提出了两个具体目标：1)开发一种基于微流体的系统，该系统可以精确控制每个通道中生长的单个细胞的位置和方向，以及它们与激光和超声波产生的空化泡的空间对齐；2)研究生物效应(细胞裂解，

DESCRIPTION (provided by applicant): Because of their non-invasiveness and non-ionization nature, therapeutic ultrasound and focused shock waves have been used extensively in clinic for the treatment of cancers, targeted drug delivery (such as opening of blood-brain-barriers), disintegration of concretions, fractionation of soft tissues, and healing of non-union fractures, a well as pain therapy. Although in all these applications cavitation has often been implicated as a key mechanism whereby the desirable therapeutic effects and, in some cases, the undesirable adverse effects are produced, the dynamic processes of bubble(s)-tissue interaction and, in particular, bubble(s)-cell interaction, are largely unknown. This is primarily due to the lack of viable experimental systems that can be used to investigate such dynamic interactions with sufficient spatial and temporal resolutions. To overcome this primary challenge, we propose to develop a new experimental system and associated technologies to investigate the bioeffects produced by cavitation bubbles at the single cell level. This project will be carried out by a multidisciplinary research team with recognized expertise in therapeutic ultrasound and cavitation (Zhong Lab) and cell mechanics (Guilak Lab). Two specific aims are proposed: 1) development of a microfluidic based system that allows for precise control of the location and orientation of individual cells grown in each channel, as well as their spatial alignment with laser- and ultrasound-generated cavitation bubbles, 2) investigation of the bioeffects (cell lysis, membrane poration, Ca2+ influx, cytoskeleton re-arrangement, injury repair, viability, and proliferation) and mechanical deformation of the cell produced by the bubble(s)-cell interaction using representative cell lines (i.e., HeLa cells, MDCK epithelial cells, and chondrocytes) relevant to therapeutic ultrasound and shock wave therapy. With a better understanding of the bubble(s)-cell interaction and the resultant mechanical and biological consequences elicited in different cell types, we hope to gain insights that may be used in the future to improve the design of therapeutic ultrasound and shock wave devices, as well as treatment protocols for safe and more effective medical applications. Furthermore, the new knowledge acquired from this project may be used to develop novel ultrasonic techniques that can be applied to stimulate stem cells under precise mechanical loading conditions to impact their differentiation, growth and phenotypic expression with tissue engineering and regenerative medicine applications. The goal of this R03 application is to demonstrate the feasibility in constructing such a novel experimental system and developing associated technologies for assessing the bioeffects produced by cavitation bubbles at the single-cell level, while the comprehensive utilization of this novel system will be exploited in a follow-up RO1 application.