Follicular Fluid, Ovarian Cortical Inclusion Cysts, and Substrate Curvature Increase the Cancerous Progression of Fallopian Tube Epithelial Cells

• Main Author: Fleszar, Andrew Joseph
• Format: Dissertation
• Language: English
• Published: ProQuest Dissertations & Theses 01.01.2019
• Subjects: Bioengineering; Biomedical engineering; Oncology
• ISBN: 9781687921116; 1687921113

A growing body of research supports the idea that the fallopian tube epithelium (FTE) is the precursor for most high-grade serous ovarian cancers (HGSOCs). Studies indicate that the ovary plays a critical role in tumor metastasis; however, the mechanism or mechanisms underlying the impact of the ovary on disease progression remain poorly understood. The central hypothesis of this thesis was that follicular fluid secreted during ovulation and the microenvironment of the ovary encourage the cancerous progression of FTE cells. To address this hypothesis, we developed in vitro models to examine the impact that chemical stimuli and physical interactions within the ovarian microenvironment had on FTE cell behavior. This thesis first demonstrated that chemical stimuli released during ovulation enabled FTE cells to evade anoikis by evaluating the impact of follicular fluid on the anchorage-independent survival of FTE cells. Next, this thesis investigated the role that cortical inclusion cysts (CICs) in the ovarian cortex played in HGSOC as these cysts have been hypothesized to create a niche environment for cancer progression. Through histological analysis of pathology samples from human ovaries, we determined that collagen I and III were elevated near CICs and that the collagen fibers in this dense region were oriented parallel to the cyst boundary. Using this information from human samples as design parameters, we engineered an in vitro model that recreated the size, shape, and extracellular matrix properties of CICs. We found that FTE cells within our model underwent robust invasion that was responsive to stimulation with follicular fluid, while ovarian surface epithelial cells, the native cells of the ovary, were non-invasive. We provided experimental evidence to support a role of the extracellular matrix in modulating FTE cell invasion, as a decrease in collagen I concentration or the addition of collagen III to the matrix surrounding the CIC mimic increased FTE cell invasion. During histological analysis of ovarian CICs, we noticed that these structures had a diverse range of curvatures, and we hypothesized that variations in ovarian CIC curvature impact the ability of trapped FTE cells to invade into the surrounding stroma. Using our in vitro model of CICs, we determined that increased curvature resulted in more invasion of FTE cells. To isolate curvature as a system parameter, we developed a novel technique to pattern concave curvatures into collagen gels. When FTE cells were seeded to confluency on curved substrates, increased curvature increased the number of invading FTE cells and the invasion distance. FTE invasion into collagen substrates with higher curvature depended on matrix metalloproteinases (MMPs), but expression of collagen I degrading Mmps was not different on curved and flat regions. A finite element (FE) model predicted that contractility and cell-cell connections were essential for increased invasion on substrates with higher curvature, while cell-substrate interactions had minimal effect. Experiments supported these predictions, with invasion decreased by blebbistatin, EGTA, or N-cadherin blocking antibody, but with no effect from a focal adhesion kinase (FAK) inhibitor. Finally, experimental evidence supported that cell invasion on curved substrates occurred in two phases—a cell-cell dependent initiation phase where individual cells broke away from the monolayer and a MMP dependent phase as cells migrated further into the collagen matrix. Taken together, this thesis showed that in vitro models of anchorage-independent cell survival, ovarian CICs, and substrate curvature can act as important tools for understanding FTE cell interactions with their environment and the cancerous progression of these cells.