am6b08548_si_006.avi (1017.87 kB)
The Effect of Size and Geometry of Poly(acrylamide) Brush-Based Micropatterns on the Behavior of Cells
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posted on 2016-08-19, 00:00 authored by Inga Lilge, Siyu Jiang, Daniel Wesner, Holger SchönherrIn
this study, the fabrication, detailed characterization, and application
of long-term stable micropatterned bio-interfaces of passivating poly(acrylamide)
(PAAm) brushes on transparent gold for application in the study of
cell-surface interactions is reported. The micropatterns were fabricated
by microcontact printing of an initiator for surface-initiated atom
transfer radical polymerization (SI-ATRP), SI-ATRP of acrylamide,
and subsequently backfilling of the unfunctionalized areas of 400–2500
μm2 size and systematically altered number of corners
with octadecanethiol. As verified by surface plasmon resonance spectroscopy,
the physisorption of fibronectin (FN) was restricted to the adhesive
areas. Exploiting this platform, the effect of micropattern geometry
and size of cell-adhesive FN areas surrounded by passivating PAAm
brushes on transparent gold substrates on the attachment of cells
and cytoskeleton alignment was investigated at the single-cell level.
Exceptional long-term stability of the patterned PAAm brushes and
arrays of adhesive areas, in which human pancreatic tumor cells (Patu
8988T) and fibroblast cells (NIH 3T3) were confined for more than
one week, was observed. Adhesive areas of 1600 μm2 or less constrained the cell shape and caused focal adhesions to
accumulate in the corners of the pattern. These changes were most
obvious for the PatuT cells in adhesive areas of ∼900 μm2, in which the actin filaments were aligned, following the
boundary of the pattern, and merged in the focal adhesions concentrated
in the corners of the pattern. NIH 3T3 cells possessed a larger cell
area, which led to an optimal cytoskeleton alignment in adhesive patterns
of ∼1600 μm2. The alignment of the cytoskeleton
was found to be less pronounced in cells on larger adhesive areas,
where the PatuT cells spread similarly to cells on unpatterned substrates.
By contrast, the NIH 3T3 cells were found to stretch even on larger
adhesive areas, spanning from one corner to the other. The long-term
stability under cell culture conditions of the patterns introduced
here will also be useful for long-term studies of single and multiple
cells, cell motility in toxicity assays, and stem cell differentiation.