Mechanoregulation of Basal Keratinocyte Migration in Wounded Tissue

项目摘要 上皮内环境的稳定是由作用于整个组织细胞上的机械力的平衡来维持的， 规模损伤破坏了这种机械平衡，目前还不清楚如何改变稳态机械信号 影响伤口修复所需的细胞行为。不能有效修复可导致纤维化瘢痕形成、慢性炎症和慢性炎症。 不愈合的伤口，并有助于病理学。上皮创伤修复依赖于基底膜的迁移， 将角质形成细胞转移到损伤部位。虽然已知基底角质形成细胞对机械力敏感， 我们缺乏对上皮损伤如何改变体内组织力学以及这些损伤如何诱导 生物物理变化随后协调伤口修复所需的基础角质形成细胞行为。本研究 旨在解决这些问题，通过使用幼斑马鱼，这是适合实时，活体成像， 它们的光学透明度。初步的活体成像实验表明，上皮损伤导致快速的基础 角质形成细胞迁移到伤口部位，这是有效修复所需的。基底角化细胞迁移是 依赖于机械信号，例如由于细胞肿胀引起的膜张力，并且与 伤口边缘上皮组织结构的短暂和局部破坏。基底角化细胞迁移 可以通过阻断Arp 2/3复合物激活或通过Talin 1敲低来抑制，这表明可能的联系 机械信号和F-肌动蛋白或粘着斑复合物在体内重塑之间的关系。此外，短暂地 减弱细胞与细胞外基质的粘附改变了基底角质形成细胞的迁移，导致伤口不良 愈合，并导致上皮结构的慢性破坏。这种表型模拟了 Kindler综合征是一种皮肤病，患者在受伤后表现出伤口愈合缺陷。这些 初步观察表明，斑马鱼幼鱼的基底角质形成细胞对机械信号有反应 通过启动有效伤口愈合所需的迁移反应而在损伤后的上皮组织中。他们 也表明缺陷的基底角质形成细胞迁移可能有助于伤口愈合病理学。的 拟议的研究将调查如何张力传感的机械传感器压电1和塔林1调节 分别通过F-actin和粘着斑重塑观察伤口诱导的基底角质形成细胞行为。这些 研究结果随后将被翻译成研究金德勒综合征的斑马鱼模型，以确定 失调的基础角质形成细胞行为对伤口愈合病理生理学的贡献。确保 该项目的成功，一个量身定制的培训计划已经制定，利用优秀的研究 威斯康星州-麦迪逊大学的环境。专门培训使用斑马鱼模型 用于伤口愈合研究的生物体和用于量化上皮组织的先进体内成像技术 机械师将有助于完成既定目标。这一培训将有助于成功过渡到 研究独立性。

Project Summary Epithelial homeostasis is maintained by the balance of mechanical forces acting upon cells across the tissue- scale. Injury disrupts this mechanical balance and it is unclear how changing homeostatic mechanical signals impacts cell behavior needed for wound repair. Failure to efficiently repair can lead to fibrotic scarring, chronic non-healing wounds, and contribute to pathology. Epithelial wound repair relies on the migration of basal keratinocytes to the site of damage. While it is known that basal keratinocytes are sensitive to mechanical forces, we lack an understanding of how epithelial injury alters tissue mechanics in vivo and how these wound-induced biophysical changes subsequently coordinate basal keratinocyte behavior needed for wound repair. This study aims to address these issues by using larval zebrafish, which are amenable to real-time, intravital imaging due to their optical transparency. Preliminary live-imaging experiments show that epithelial injury causes rapid basal keratinocyte migration to the wound site, which is needed for efficient repair. Basal keratinocyte migration is dependent on mechanical signals, such as membrane tension due to cell swelling, and is associated with a transient and localized disruption of epithelial tissue architecture at the wound edge. Basal keratinocyte migration can be inhibited by blocking Arp2/3 complex activation or through Talin1 knockdown, suggesting a potential link between mechanical signaling and F-actin or focal adhesion complex remodeling in vivo. Further, transiently weakening cell adhesion to the extracellular matrix alters basal keratinocyte migration, causing poor wound healing, and resulting in chronic disruption of epithelial architecture. This phenotype mimics pathology associated with Kindler Syndrome, a skin disease in which patients show wound healing defects in response to injury. These preliminary observations demonstrate that basal keratinocytes of larval zebrafish respond to mechanical signals in epithelial tissue after injury by initiating a migratory response that is required for efficient wound healing. They also suggest that defective basal keratinocyte migration may contribute to wound healing pathology. The proposed study will investigate how tension sensing by the mechanotransducers Piezo1 and Talin1 regulate wound-induced basal keratinocyte behavior by F-actin and focal adhesion remodeling, respectively. These findings will subsequently be translated to investigate a zebrafish model of Kindler Syndrome to determine the contribution of dysregulated basal keratinocyte behavior to wound healing pathophysiology. To ensure the success of this project, a tailored training plan has been developed that takes advantage of the excellent research environment at the University of Wisconsin – Madison. Dedicated training in the use of the zebrafish model organism for wound healing studies and advanced in vivo imaging techniques for quantifying epithelial tissue mechanics will aid in the completion of the stated aims. This training will facilitate a successful transition to research independence.