TEMPERATURE-RESPONSIVE CULTURE SURFACES FOR REGENERATIVE MEDICINE - Intelligent Scaffolds for Tissue Engineering and Regenerative Medicine - page 910

To control the dissociation of specific interactions between cul-
tured cell integrins and surface RGDS, spread HUVECs (1-day cul-
ture without serum at 37 ◦ C) were exposed to a lower temperature
treatment at 20 ◦ C (Fig. 44.4). The detachment rates depended on
the immobilized cell adhesive peptide densities represented by the
monomercarboxylategroupcompositionineachfeed.Moreover,the
detachmentratesincreaseddramaticallywiththeadditionofhigher
concentrations of RGDS. Low temperature treatment at 20 ◦ C also
promotedcelldetachmentfromtheRGDS-modifiedpoly(IPAAm-co-
CIPAAm)surface,forwhichthedetachmentratealsodecreasedwith
increasing celladhesive peptide surface content.
Recently, more systematic chemical compositions achieved
by using RGDS-modified poly(IPAAm-co-CIPAAm) systems have
advanced cell attachment anddetachment. 41
By co-immobilizing a Pro-His-Ser-Arg-Asn (PHSRN) sequence
on intelligent surfaces that enables the stable binding of RGDS to
α 5 β 1 integrin, the synergistic roles of the PHSRN sequence have
beeninvestigatedbystudyingtime-dependentcelldetachmentfrom
these surfaces. Notably, PHSRN has been found in FN and is thought
to synergistically enhance the cell adhesive activity of the RGD
sequence. 42 − 44 The synergistic site is located approximately 3.5 nm
from the RGD loop in native FN, and its precise spatial positioning
is thought to play a critical role in the observed synergistic activ-
ity, especially for downstream cell adhesion events involving focal
adhesion kinase (FAK) phosphorylation. 44 − 46 In contrast, very little
isknownaboutthesynergisticeffectsofPHSRNonbindingstrength.
Therefore, here, we have developed a novel but simple protocol for
the comparative study of cell adhesive strengths using RGDS- and
PHSRN-immobilized intelligent surfaces. This approach provides a
noninvasive method for examining time-dependent a nity changes
between cells and peptides.
RGDS-immobilized, temperature-responsive polymer-grafted
surfaces promote HUVEC adhesion and spreading by their biospe-
cificactivityinserum-freemedia.Attemperaturesbelowthegrafted
polymer's LCST, integrin-RGDS association decreases due to a loss
of cell tension and surface anchoring, prompting cells to round
and then detach (Fig. 44.5). Observed “on-off” control of specific
integrin-RGDS binding achieved only by temperature regulation is