TEMPERATURE-RESPONSIVE CULTURE SURFACES FOR REGENERATIVE MEDICINE - Intelligent Scaffolds for Tissue Engineering and Regenerative Medicine - page 908

interactions of ligands and cell membrane receptors play multiple
important roles in inducing cell spreading, proliferation, differen-
tiation, and signal transduction. 26 − 31 The ECM elaborated by cells
on surfaces constitutes a regulator of the cell adhesion process
and cell differentiation and contributes to the mechanical proper-
ties of tissue. These regulatory effects of ECM are mediated through
transmembraneproteinsspecializedincell-substrateadhesion.The
peptide sequence Arg-Gly-Asp (RGD) found in fibronectin, type I
collagen, and other ECM proteins has been widely studied as an
immobilized cell adhesion ligand specific to integrin-mediated cell
adhesion. 32 , 33 The surface density and chemical presentation of
these synthetic ligands and their effectiveness as surface modi-
fication agents in cell culture are correlated to their ability to
enhance cell adhesion, which is particularly useful for applica-
tions in tissue engineering. 34 − 38 Temperature-responsive PIPAAm-
grafted surfaces are modified with arginine-glycine-aspartic acid-
serine (RGDS) peptides. Surface swelling, wettability, and elastic
modulus of the grafted surface can be changed by temperature con-
trol. At temperatures greater than the LCST of PIPAAm, the grafted
polymer chains collapse, and the immobilized peptide is therefore
exposed to adherent cells. Below the LCST, hydrated polymer chains
soften, expand, and swell, shielding immobilized RGDS from inte-
grin access, limiting cell-surface attachment tension, and mechani-
cally disrupting cell-surface contacts. Adherent cell populations are
firstroundedandthenremovedunderthisthermaltreatmentinthe
absence of serum proteins or trypsin treatment. Because PIPAAm
doesnothavefunctionalgroupssuchascarboxyl,amino,orhydroxyl
groups, these moieties have been introduced by co-polymerization
in order to provide a means for the stable binding of peptides.
Here, we have designed a functional co-monomer, 2-carboxyisopr-
opylacrylamide (CIPAAm), because the use of conventional co-
monomers such as acrylic acid (AAc) shifts the LCST to higher than
37 ◦ C. 8
Poly(IPAAm-co-CIPAAm) was grafted onto a TCPS dish via
electron-beam irradiation polymerization. Peptides were immobi-
lized onto the grafted polymer chains via the carboxyl groups in
CIPAAm using N -hydroxysuccinimide/1-ethyl-3-(3-dimethylam-
inopropyl) carbodiimide hydrochloride (NHS/EDC) chemistry. 39 , 40