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J Vis Exp. 2010 Dec 22;(46). pii: 2413. doi: 10.3791/2413.

Micropatterned surfaces to study hyaluronic acid interactions with cancer cells.

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1
Department of Chemical and Biomolecular Engineering, Johns Hopkins Physical Sciences Oncology Center and Institute for NanoBioTechnology, Johns Hopkins University, USA.

Abstract

Cancer invasion and progression involves a motile cell phenotype, which is under complex regulation by growth factors/cytokines and extracellular matrix (ECM) components within the tumor microenvironment. Hyaluronic acid (HA) is one stromal ECM component that is known to facilitate tumor progression by enhancing invasion, growth, and angiogenesis(1). Interaction of HA with its cell surface receptor CD44 induces signaling events that promote tumor cell growth, survival, and migration, thereby increasing metastatic spread(2-3). HA is an anionic, nonsulfated glycosaminoglycan composed of repeating units of D-glucuronic acid and D-N-acetylglucosamine. Due to the presence of carboxyl and hydroxyl groups on repeating disaccharide units, native HA is largely hydrophilic and amenable to chemical modifications that introduce sulfate groups for photoreative immobilization (4-5). Previous studies involving the immobilizations of HA onto surfaces utilize the bioresistant behavior of HA and its sulfated derivative to control cell adhesion onto surfaces(6-7). In these studies cell adhesion preferentially occurs on non-HA patterned regions. To analyze cellular interactions with exogenous HA, we have developed patterned functionalized surfaces that enable a controllable study and high-resolution visualization of cancer cell interactions with HA. We utilized microcontact printing (uCP) to define discrete patterned regions of HA on glass surfaces. A "tethering" approach that applies carbodiimide linking chemistry to immobilize HA was used (8). Glass surfaces were microcontact printed with an aminosilane and reacted with a HA solution of optimized ratios of EDC and NHS to enable HA immobilization in patterned arrays. Incorporating carbodiimide chemistry with mCP enabled the immobilization of HA to defined regions, creating surfaces suitable for in vitro applications. Both colon cancer cells and breast cancer cells implicitly interacted with the HA micropatterned surfaces. Cancer cell adhesion occurred within 24 hours with proliferation by 48 hours. Using HA micropatterned surfaces, we demonstrated that cancer cell adhesion occurs through the HA receptor CD44. Furthermore, HA patterned surfaces were compatible with scanning electron microscopy (SEM) and allowed high resolution imaging of cancer cell adhesive protrusions and spreading on HA patterns to analyze cancer cell motility on exogenous HA.

PMID:
21206473
PMCID:
PMC3159670
DOI:
10.3791/2413
[Indexed for MEDLINE]
Free PMC Article
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