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Guiding Cell Behavior in 2D, 3D, and Dynamic Environments

  • Author(s): Mercurio, Kevin
  • Advisor(s): Leppert, Valerie
  • et al.
Abstract

Cell culture substrates for contact guidance studies can be developed through polymer processing to produce various topographic dimensions. Surface topography has a large influence on cell fate and can ultimately be used to help understand and direct cell behaviors. Fabrication techniques for nano- and micro-patterning are typically costly and require large, laboratory-grade equipment. However, in the past decade, researchers have discovered shrink-based techniques to produce nano-scale topography at a fraction of the cost. The fabrication methods are simple, fast, and cost effective, when compared to lithography-based patterning methods. 2D topography is developed using a lab-on-a-chip approach, in which pre-strained polystyrene sheets are coated with metal, constrained from the edges, and heat-treated to develop 320nm and 510nm surface wrinkles. An increase in metal coating thickness from 15nm to 60nm resulted in an increase in wrinkle width and height. To build upon this phenomenon, the polystyrene sheets were further manipulated to generate 3D topography. Acetone solvent was used to craze the polymer sheet and develop surface topography on the micro-scale. Depending on sheet orientation and treatment procedure, wrinkles, channels, and voids ranging in width from 1 μm to 210 μm can be produced. These patterned sheets were then used to mold polydimethylsiloxane (PDMS) cell culture platforms. Biological analysis showed good cellular adhesion of human umbilical vein endothelial cells (HUVECs) on the structures surface. Cellular orientation and actin fiber elongation were analyzed on flat controls, nano-scale wrinkles, crazed topography, and shrunk-crazed PDMS substrates after 24 hours of culture using optical microscopy. HUVECs on the acetone crazed topographies extended the actin filaments roughly 23 μm in length as compared to the flat control having an average length of 51 μm. The shrunk-crazed topography produced better cellular alignment than the crazed topography. Longitudinal and transverse shrunk samples produced cell alignment of about 10 ± 9°and 8 ± 7° from the underlying pattern direction, respectively. In addition to cell analysis on open faced surfaces, cells were also cultured in 3D microchanneled structures to demonstrate the guidance of cells in a dynamically rotating environment. Cells were loaded into PDMS tubular networks and rotated at approximately 1 rotation per hour. As a result, human umbilical vein endothelial cells (HUVECs) and rat aortic smooth muscle cells (RAOSMCs) were able to adhere to the inner surfaces of the channel and confluent layers were formed circumferentially. The use and application for the structures and devices discussed herein are analyzed and evaluated in three separate projects for their potential in bioengineering and tissue engineering applications.

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