NANOFIBER REGULATION OF CELLULAR MIGRATORY MECHANISMS
Open Access
- Author:
- Bowers, Daniel Thorn
- Graduate Program:
- Bioengineering
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- September 26, 2017
- Committee Members:
- Justin L. Brown, Ph.D., Dissertation Advisor/Co-Advisor
Justin L. Brown, Ph.D., Committee Chair/Co-Chair
Peter J. Butler, Ph.D., Committee Member
William O. Hancock, Ph.D., Committee Member
Christopher U. Niyibizi, Ph.D., Outside Member - Keywords:
- tissue engineering
scaffold
nanofibers
diameter
migration
focal adhesion - Abstract:
- Cell migration is crucial for a diverse array of physiological and pathological processes from cancer metastases and wound healing to the transformation of tissue engineering constructs into tissue mimicking spaces. Native tissues present a range of fiber diameters from 10’s of nanometers to several microns during homeostasis and natural tissue reconstruction. The purpose of this dissertation was to understand cell migration through mechanosensitive signaling pathways that respond to ‘out-of-plane’ curvature. AIM 1 was to characterize the cellular morphology on nanofibers and to develop direct diameter control. A non-degradable high molecular weight poly(methyl methacrylate), was used to electrospin fibers of 5 diameters and to spin coat a flat surface control. Cell area was significantly smaller on all fibers compared to the flat surface control, arising from a smaller short axis on fibers. Serial block face electron microscopy and nanoComputed Tomography have demonstrated that the cell membrane conforms to the fiber shape. AIM 2 was to measure the cellular velocity dependence on nanofiber diameter. Human mesenchymal stem cells and mouse embryonic fibroblasts displayed increased migration velocity on the middle range fiber diameters (250 to 500 nm). When exploring a larger range there was a relative velocity peak at ~500 nm in diameter, with another at about 1500 nm. Interestingly, the velocity decreased to a value similar to the flat surface control when the diameter reached ~2000 nm. Finally in AIM 3, the mechanotransduction pathways regulated by ‘our-of-plane’ curvature were investigated. The effect of inhibiting ROCK and cdc42 followed the trend of the flat surface when cells were on the ~2000nm fibers. Differential activation states of Focal Adhesion Kinase (FAK) and signaling partner Src followed trends seen in the migration velocity. Since the length of focal adhesions did not match velocity trends, the size of focal adhesions may be decoupled from activation on 3D substrates. We find that nanofiber based migration is a plastic process in which a cell can display lobopodial and mesenchymal migration modes. When considered alongside stem cell differentiation dependence on diameter, we are building a framework from which we can rationally design bioinstructive scaffolds for tissue regeneration.