Date of Award

2019

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Cell & Molecular Biology

First Advisor

Sandra W. Ryeom

Second Advisor

Ellen Puré

Abstract

Fibroblast activation is a crucial step in tumor growth and metastatic progression; activated fibroblasts remodel extracellular matrix (ECM) in primary tumor and metastatic microenvironments, exerting both pro- and anti-tumorigenic effects. However, the intrinsic mechanisms that regulate the activation of fibroblasts are not well defined. The signaling axis comprising the calcium-activated Ser/Thr phosphatase calcineurin (CN), and its downstream target nuclear factor of activated T cells (NFAT), has been shown to play important roles in in endothelial and immune cell activation, but its role in fibroblasts is not known. We have shown that deletion of CN in fibroblasts in vitro results in alterations in fibroblast morphology and function consistent with an activated phenotype relative to wild-type fibroblasts. CN-null fibroblasts have greater migratory capacity, demonstrate increased collagen secretion and remodeling, and promote more robust endothelial cell activation in vitro. ECM generated by CN-null fibroblasts contain more collagen with greater alignment of fibrillar collagen compared to wild-type fibroblast-derived matrix. These differences in matrix composition and organization impose distinct changes in morphology and cytoskeletal architecture of both fibroblasts and tumor cells that may represent changes in their biological function, e.g. activation or epithelial-mesenchymal transition. However, while they are more activated at baseline, Cn-/- fibroblasts cannot functionally respond to TGF-beta stimulation in multiple activation assays. They are also unable to upregulate FAP on soft, fibronectin-coated substrates to the same extent as WT fibroblasts. Consistent with this in vitro phenotype, we show that mice with stromal CN deletion exhibit a greater incidence and larger lung metastases. Our data suggest that CN/NFAT signaling contributes to the maintenance of fibroblast homeostasis and that loss of CN/NFAT signaling is sufficient to promote fibroblast activation, leading to the outgrowth of lung metastases. With a better understanding of how CN affects fibroblast homeostasis, new therapeutic targets may be identified that prevent the priming of the metastatic niche.

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