Effect of hydroxyapatite materials properties on fibronectin-mediated breast cancer cell adhesion and pro-angiogenic secretion

Abstract

Event Abstract Back to Event Effect of hydroxyapatite materials properties on fibronectin-mediated breast cancer cell adhesion and pro-angiogenic secretion Fei Wu1, Weisi Chen2, Brian Gillis1, Claudia Fischbach2, 3, Lara Estroff1, 3 and Delphine Gourdon1, 2 1 Cornell University, Materials Science and Engineering, United States 2 Cornell University, Biomedical Engineering, United States 3 Kavli Institute at Cornell for Nanoscale Science, United States Introduction: Breast cancer preferentially metastasizes to bone and induces pathological remodeling. There has been evidence suggesting that the materials properties of bone mineral may play an important role in breast cancer metastasis[1],[2]. Hydroxyapatite (HAP, Ca10(PO4)6(OH)2) is a calcium phosphate mineral with structural and mechanical properties closely related to bone apatite. In this study, we seek to understand: (i) whether HAP materials properties, such as surface chemistry and roughness, alter the mineral/extracellular matrix (ECM) interface in bone, and (ii) how the altered ECM impacts breast cancer cell functions. Specifically, we investigated the conformation of fibronectin (Fn, a major ECM protein), breast cancer cell adhesion and pro-angiogenic secretion as a function of underlying HAP materials properties and associated Fn conformation. Materials and Methods: Geologic HAP crystals were bought from Etsy. Fibronectin (Fn) was obtained from Life Technologies, NY, and labeled for intramolecular Fӧrster resonance energy transfer (FRET) as previously described[3]. Smooth HAP facets (100)S and (001)S and rough HAP facets (100)R and (001)R were obtained by polishing HAP slabs to roughnesses either below 1 nm, i.e., smaller than Fn radius of gyration, or around 200 nm. Then, we incubated Fn (10% FRET-labeled) on HAP facets and analyzed Fn conformation via FRET. Finally, MDA-MB-231 breast cancer cells (ATCC, VA) were seeded onto Fn coated HAP facets to investigate cell adhesion and secretion of vascular endothelial growth factors (VEGF). Student’s t-tests were used to determine statistical significance between conditions in GraphPad Prism (GraphPad Software Inc., CA). Results and Discussion: We first quantified Fn conformation adsorbed onto HAP via FRET (Figure 1). FRET ratios of Fn adsorbed onto (100) were higher than those onto (001) for both smooth and rough HAP, indicating that Fn adsorbed in more compact conformations onto (100) facets. Interestingly, Fn FRET ratios were not significantly different between smooth and rough HAP, suggesting that Fn conformation was independent of roughness within the range of interest. We then assessed MDA-MB-231 cells behavior. Our preliminary results showed (i) higher cell adhesion onto (001) than onto (100) HAP facets, (ii) overall higher cell adhesion onto rough HAP than onto smooth HAP, and (iii) lower VEGF secretion associated with higher adhesion. Conclusions: Our FRET analysis indicated that Fn conformation (% of unfolded Fn) depends exclusively on HAP surface chemistry. In contrast, our cells study revealed enhanced adhesion and decreased pro-angiogenic secretion of cancer cells linked with both (001) HAP facets (coated with more unfolded Fn) and rougher HAP. These findings deconvolute the roles of mineral surface chemistry and roughness in interfacial ECM deposition and have important implications for our understanding of breast cancer bone metastasis. Karin Wang; Jin Ho Chang; Young Hye Song; Siyoung Choi; Maura Weathers