Thesis (Ph. D.)--University of Rochester. Department of Biomedical Engineering, 2017.
Cell signaling in response to platelet-derived growth factor (PDGF) has been
found to be either up- or downregulated in a variety of pathological conditions, making
PDGF-induced signaling pathways important and attractive therapeutic targets. The
difficulty in translating PDGF signaling to therapeutic interventions arises from the
complexity of the cellular responses to PDGF, wherein broadly up- or downregulating
PDGF signaling often has off-target effects due to the multitude of PDGF-induced
signaling pathways and cellular responses to PDGF. The limited success of PDGF in
tissue engineering and regenerative medicine applications may be overcome with a
clearer understanding of how cells integrate signals from PDGF and the extracellular
matrix (ECM). Fibronectin is a principal component of the ECM that is produced in a
soluble, protomeric form and then assembled by cells into an insoluble, fibrillar matrix.
ECM fibronectin is known to influence ECM composition, as well as cell and tissue
function. Understanding how ECM fibronectin directs cell and tissue responses to PDGF is
critical to designing targeted therapies for pathologies characterized by both aberrant ECM
assembly and PDGF signaling, such as asthma, liver cirrhosis, atherosclerosis, pulmonary
fibrosis, cancer and chronic wounds. In this thesis, the effect of cell-assembled ECM
fibronectin on cellular responsiveness to PDGF was assessed using intracellular calcium
release as a measure of cellular responsiveness to PDGF. Results of this work
demonstrate that ECM fibronectin attenuates the PDGF-PI3K-calcium signaling axis at the
level of PI3K activation. ECM fibronectin did not have an effect on other intracellular
signals activated by PDGF, including activation of PDGF receptor β, AKT, phospholipase
Cγ1, or ERK1/2. The bioactive effects of fibronectin were localized to the α5β1 integrinbinding
FNIII9-10 region. Finally, a cell-binding fibronectin fragment with a truncated FNIII module, FNIII8c-10, attenuated PDGF-induced intracellular calcium release and PI3K activation. Thus, this newly developed protein has the potential to be employed as a therapeutic to specifically attenuate the PDGF-induced intracellular calcium release cascade while leaving intact other PDGF signaling pathways.
