The Cell–Surface Interaction - Tissue Engineering: Cell-Surface Interactions for Tissue Culture - page 5

osteoblast differentiation by activating Osf2 (Cbfa1) transcription factor and
induction of osteoblast-specific osteocalcin gene expression [ 13 ]. The a 5 integrin
subunit is responsible for the interactions with fibronectin. Furthermore, the a 5 b 1
integrin has been reported to be necessary for osteoblast proliferation and differ-
entiation. It is responsible for bone-like nodule formation in vitro by osteopro-
genitor cells grown on various synthetic biomaterials. In a study by Petrie et al.
[ 14 ], coatings consisting of defined multimer constructs with monomer, dimer,
tetramer and pentamer recombinant fragments of fibronectin were prepared. The
authors aimed to assess how nanoscale ligand clustering affects integrin binding,
stem cell responses, tissue healing and biomaterial integration. Clinical-grade
titanium was grafted with polymer brushes that presented monomers, dimers,
trimers or pentamers of the a 5 b 1 integrin-specific fibronectin III (7-10) domain.
Their results indicate that coatings with trimer and pentamer modifications
enhanced integrin-mediated adhesion in vitro, osteogenic signalling, and differ-
entiation in human mesenchymal stem cells (MSCs) [ 14 ]. Keselowsky et al. [ 15 ]
showed that the expression of the a 5 integrin subunits regulates alkaline phos-
phatase activity, but this function is not affected by surface microtopography.
However, the microrough surfaces increase the expression of the b 1 integrin
subunit, which is essential in osteoblastic differentiation through the protein kinase
C pathway [ 16 ]. More recent data obtained using molecular beacons targeted to the
b 1 integrin subunit messenger RNA (mRNA) in order to visualize surface-
dependent changes in its expression in individual MG63 cells showed that effects
of the substrate on b 1 mRNA previously observed in confluent cultures were also
evident in preconfluent cultures, supporting the hypothesis that b 1 integrin is
important in proliferation as well as differentiation of osteoblasts [ 17 ].
Many in vitro studies have focused on identifying integrins on osteoblast cell
membranes to determine their role in the mechanism of initial attachment of cells
to the implant surface [ 17 - 19 ]. Gronowicz and McCarthy [ 20 ] have revealed that
the initial attachment to metal materials involves integrins. SaOs-2 cells were
incubated on titanium-4% aluminium-6% vanadium (TAV), polystyrene, glass
and cobalt-chromium-molybdenum (CoCrMo) with antibodies to the fibronectin
receptor a 5 b 1 and the vitronectin receptor (a v b 3 /a v b 5 ). On samples with fibronectin
receptor antibody on TAV and CoCrMo the attachment was reduced by 63 and
49%, respectively. In contrast, no significant effect was seen on samples with the
antibody to the vitronectin receptor. Further analysis of changes in integrin
expression within 24 h on samples without antibodies revealed that the a 5 integrin
subunit has the highest expression level after 24 h of adhesion on TAV compared
with polystyrene-, glass-, CoCrMo- and fibronectin-covered surfaces. The a 2 and
a v subunits were also detected, but their expression was lower on TAV than on
polystyrene, whereas the production of a 1 was inhibited only in cells cultured on
polystyrene, but not in cells cultured on the other surfaces.
Further studies on primary human osteoblasts confirmed these findings. Sinha
and Tuan [ 21 ] reported differences in integrin expression on polished and rough
(R a not given) TAV and CoCrMo after 12 h in culture. For cells on the polished
TAV surface, a 2 , a 3 , a 4 , a 6 , a v , b 1 and b 3 were detected. Results for rough TAV