Biophysical Cues and Corneal Wound Healing

生物物理线索和角膜伤口愈合

基本信息

批准号：
9185333
负责人：
CHRISTOPHER John MURPHY
金额：
$ 39.25万
依托单位：
UNIVERSITY OF CALIFORNIA AT DAVIS
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-12-01 至 2020-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9185333
关键词：
Actins Address Adhesions Atomic Force Microscopy Behavior Biochemical Biomechanics Biophysics Blindness Cells Chemistry Coculture Techniques Communication Complex Cornea Corneal Injury Corneal Opacity Corneal Stroma Cues Cytoskeleton Data Drug usage Engineering Environment Epithelial Epithelial Cells Evolution Extracellular Matrix Fiber Fibroblasts Fibrosis Funding Goals Homeostasis Hydrogels In Vitro Infection Investigation Knowledge Laboratories Laser In Situ Keratomileusis Maintenance Measures Mechanics Methods Microfluidic Microchips Microfluidics Modeling Myofibroblast Operative Surgical Procedures Oryctolagus cuniculus Outcome Patient-Focused Outcomes Phenotype Process Production Research Research Personnel Signal Transduction Signaling Molecule Stimulus Stromal Cells System Testing Therapeutic Therapeutic Effect Time Tissues Traction Transforming Growth Factor beta Trauma Visual Acuity Visual impairment Wound Healing cell behavior cell type clinically relevant corneal epithelium crosslink design experimental study improved in vivo intervention effect mechanical properties migration novel public health relevance repaired response to injury wound

项目摘要

DESCRIPTION (provided by applicant): Corneal opacities, a leading cause of blindness worldwide, are commonly a result of stromal fibrosis or haze from dysregulated wound healing. Corneal wound healing is an enormously complex process that requires the simultaneous cellular integration of multiple soluble biochemical as well as biophysical cues associated with the wound space. Research from our laboratory and others have demonstrated biophysical attributes of the extracellular matrix profoundly modulate a host of fundamental cellular behaviors essential to the maintenance of homeostasis and wound repair including adhesion, migration, proliferation and differentiation. We have demonstrated that the intrinsic mechanical properties of the corneal stroma are significantly altered in a time dependent manner throughout wound healing. The matrix associated with the wound space was stiffest at 7 days with the greatest myofibroblast numbers and degree of stromal haze occurring at a later time point. This temporal relationship, combined with in vitro data documenting stiffer substrates to promote myofibroblast transformation, suggests a causal relationship. In Aim 1 we propose to directly modulate matrix stiffness in vivo using clinically relevant approaches and establish the impact of stiffening and softening of the wound matrix on wound healing outcomes using a well-defined rabbit PTK model. The complex inter-relationship between cytoskeletal dynamics and matrix remodeling and stiffness is central to determining wound healing outcomes but remains little investigated and poorly understood and are the central focus of Aim 2. Here, the impact of altering cytoskeletal dynamics in vitro on cell derived extracellular matrix elaboration, remodeling and mechanics, and the impact of altering cell derived matrix stiffness on keratocytes to myofibroblast transformation will be determined. Preliminary data demonstrate that the mechanical properties of ECM derived from cell in vitro can be modulated by applying diverse clinically relevant crosslinking methods. Preliminary data also document that modulating the cellular stiffness using drugs that alter the cytoskeleton can dramatically influence myofibroblast transformation. Stiffening of the cell promoted myofibroblast transformation. While softening of the cell inhibited transformation even in the presence of TGFβ signaling. Additionally, the complex signaling dynamic that exist between stromal and epithelial cells will be investigated using a novel microfluidic co-culture platform in Aim 3. This novel system allows for the simultaneous presentation of multiple yet distinct biophysical and biochemical stimuli in addition to controlled exchange of soluble signaling molecules between epithelial and stromal cells. We will determine if soluble signals from the epithelial cells have a differential effect on KFM transformation under various biophysical stimuli. In aggregate the experiments detailed in this proposal are aimed at testing our central concept that optimal outcomes in corneal wound repair can be facilitated by engineering the intrinsic biophysical attributes of cells and/or matrices.

描述（由申请人提供）：角膜混浊是全世界失明的主要原因，通常是由于伤口愈合失调引起的基质纤维化或混浊造成的。角膜伤口愈合是一个极其复杂的过程，需要多种可溶性生化物质同时进行细胞整合。我们实验室和其他实验室的研究表明，细胞外基质的生物物理属性深刻地调节了许多对于维持体内平衡和维持至关重要的基本细胞行为。伤口修复，包括粘附、迁移、增殖和分化，在整个伤口愈合过程中，角膜基质的内在机械特性以时间依赖性方式显着改变，与伤口空间相关的基质在7天时最硬。在稍后的时间点发生的肌成纤维细胞数量和基质混浊程度，与记录较硬基质促进肌成纤维细胞转化的体外数据相结合，表明了因果关系。在目标 1 中，我们建议直接调节基质。使用临床相关方法测定体内硬度，并使用明确的兔 PTK 模型确定伤口基质的硬化和软化对伤口愈合结果的影响，细胞骨架动力学与基质重塑和硬度之间复杂的相互关系是确定伤口的核心。愈合结果，但仍然很少进行研究和了解，这是目标 2 的中心焦点。这里，体外改变细胞骨架动力学对细胞衍生的细胞外基质的加工、重塑和力学的影响，以及改变细胞骨架动力学的影响初步数据表明，体外细胞衍生的 ECM 的机械特性可以通过应用临床上不同的相关交联方法来调节。初步数据还证明，使用药物调节细胞硬度。改变细胞骨架可以显着影响肌成纤维细胞的转化，而细胞的软化甚至在存在 TGFβ 的情况下也会抑制转化。此外，在目标 3 中，将使用新型微流体共培养平台来研究基质细胞和上皮细胞之间存在的复杂信号动态。除了受控刺激之外，该新型系统还允许同时呈现多种但不同的生物物理和生化刺激。我们将确定上皮细胞的可溶性信号是否对上皮细胞和基质细胞之间的可溶性信号分子有不同的影响。总的来说，本提案中详述的实验旨在测试我们的中心概念，即通过设计细胞和/或基质的内在生物物理属性可以促进角膜伤口修复的最佳结果。