For healthy individuals, wound healing mainly occurs without medical intervention, yet for the growing elderly, diabetic, or obese populations, as well as for those recovering from surgery, disregulated wound healing poses a serious health risk. Therefore, understanding the cellular processes regulating wound healing and correcting them when they go awry is essential for meeting these population’s healthcare needs. Wound healing is a complex process consisting of a suite of injury repair programs executed by cells in the injured tissue. While several of these programs have been previously described, there are many possible cellular signalling pathways that can mediate a given repair program, and its unclear which pathway mediates a specific process. In this work, we aimed to identify the key cellular signaling pathway that regulates the injury contraction process in a dermal microtissue on a chip model. We found that a balance of tissue forces generated via RhoA activation is critical for injury contraction, and that spatially localized RhoA activation can recruit new cells to participate in injury contraction. During our experiments, we also discovered and characterized a novel actin cytoskeleton-plasma membrane topology present in human dermal fibroblasts at the extreme end of cellular contractility. We also developed several technical advances: the real-time imaging and manipulation of calcium in 3D microtissues, the development of a reporter for smooth muscle actin and a labeled cellular fibronectin fusion protein, and the optimization of Forster Resonance Energy Transfer sensors. Taken together, our experimental results demonstrate the importance of RhoA-mediated force balance during injury contraction, which also has implications for scarring wound pathologies, while the tools we developed provide support for future investigations into the cellular signaling mediating injury repair programs.
| Identifer | oai:union.ndltd.org:bu.edu/oai:open.bu.edu:2144/44766 |
| Date | 23 May 2022 |
| Creators | Ghilardi, Samuel J. |
| Contributors | Sgro, Allyson E. |
| Source Sets | Boston University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
| Rights | Attribution-NonCommercial-ShareAlike 4.0 International, http://creativecommons.org/licenses/by-nc-sa/4.0/ |
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