Abstract
Background: Corneal stromal cells (keratocytes) are responsible for developing and maintaining normal corneal structure and transparency, and for repairing the tissue after injury. Corneal keratocytes reside between highly aligned collagen lamellae in vivo. In addition to growth factors and other soluble biochemical factors, feedback from the extracellular matrix (ECM) itself has been shown to modulate corneal keratocyte behavior. Methods: In this study, we fabricate aligned collagen substrates using a microfluidics approach and assess their impact on corneal keratocyte morphology, cytoskeletal organization, and patterning after stimulation with platelet derived growth factor (PDGF) or transforming growth factor beta 1 (TGFβ). We also use time-lapse imaging to visualize the dynamic interactions between cells and fibrillar collagen during wound repopulation following an in vitro freeze injury. Results: Significant co-alignment between keratocytes and aligned collagen fibrils was detected, and the degree of cell/ECM co-alignment further increased in the presence of PDGF or TGFβ. Freeze injury produced an area of cell death without disrupting the collagen. High magnification, time-lapse differential interference contrast (DIC) imaging allowed cell movement and subcellular interactions with the underlying collagen fibrils to be directly visualized. Conclusions: With continued development, this experimental model could be an important tool for accessing how the integration of multiple biophysical and biochemical signals regulate corneal keratocyte differentiation.
Highlights
The corneal stroma is a highly ordered structure consisting of approximately 200 collagen lamellae [1]
Following injury or refractive surgery in the adult cornea, wound contraction and tissue remodeling depend on mechanical interactions between corneal keratocytes and extracellular matrix (ECM) fibrils [9,10,11]
We evaluate for the first time the effects of aligned fibrillar collagen substrates created using a microfluidics approach on corneal keratocyte morphology, cytoskeletal organization, and alignment after stimulation with platelet derived growth factor (PDGF) or TGFβ
Summary
The corneal stroma is a highly ordered structure consisting of approximately 200 collagen lamellae [1]. Corneal stromal cells (keratocytes) reside between these lamellae and are responsible for secreting the extracellular matrix (ECM) components required to develop and maintain normal corneal structure and function [2,3,4] Biophysical interactions between these cells and their surrounding ECM play a central role in corneal morphogenesis and wound healing. Following injury or refractive surgery in the adult cornea, wound contraction and tissue remodeling depend on mechanical interactions between corneal keratocytes and ECM fibrils [9,10,11] Understanding these behaviors is important to developing new advances in the field of tissue engineering, where it is necessary to modulate cell and ECM patterning to direct the formation of specific tissue geometries [12], or to prevent cell-induced matrix disruption of pre-fabricated 3-D constructs [13]. Conclusions: With continued development, this experimental model could be an important tool for accessing how the integration of multiple biophysical and biochemical signals regulate corneal keratocyte differentiation
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have