Engineering Fibrous Tissues and Their Interfaces with Bone - Structural Interfaces and Attachments in Biology - page 329

did not change over time in the co-culture and collagen type II and aggrecan
expression (measured by RT-PCR) were elevated after 28 days [ 136 ]. Mineraliza-
tion was also observed to remain relatively low levels in the co-culture samples.
The results of the study indicate that co-culture may be an important aspect to
include for creation of the fibrocartilaginous tissue that is found at the insertion
point. However, further experiments examining additional cell types and/or soluble
factors to improve the amount and integration of this fibrocartilaginous tissue with
juxtaposing bone and fibrous tissues are needed before engineering of a more
complete fibrous tissue-bone interface can be achieved.
15.5.2 Scaffolds
The scaffold design for an interface region needs to recapitulate the complex
structure of the native tendon/ligament to bone insertion site. The scaffold should
exhibit a gradient of structural and mechanical properties that mimics the native
tissue [ 137 ]. A stratified or multi-phased scaffold is able to achieve this characteris-
tic. Scaffold phases must also have the ability to integrate into the native tissue after
implantation. This can be achieved by using gradually degradable biomaterials.
A study has been performed to develop a triphasic scaffold with co-culture for the
ligament-bone interface [ 138 ]. This scaffold is modeled after the native structure, in
which “Phase A” was designed with a PLGA mesh for fibrous tissue formation and
seeded with human fibroblasts, “Phase B” contained PLGA microspheres for
fibrocartilage culture, and “Phase C” utilized sintered PLGA and bioactive glass
with human osteoblasts for bone formation. After 42 days, fibroblast and osteoblast
cells migrated to “Phase B” and increased matrix production similar to the interface
region, while cell specific matrices were secreted for “Phase A” and “Phase C”
[ 138 ]. When a triphasic system with tri-culture of bovine fibroblasts in “Phase A,”
chondrocytes in “Phase B,” and osteoblasts in “Phase C” was implanted in vivo in a
subcutaneous rat model, matrix production compensated for the decrease inmechan-
ical properties of a degrading scaffold and phase specific controlled matrix hetero-
geneity was maintained 2 months after implantation [ 139 ]. These results show that
stratified scaffolds have the potential to promote multiple zones of tissue regenera-
tion in a single scaffold system [ 137 ].
As an alternative to a construct containing discrete phases, a graded scaffold has
also been examined for fibrous tissue-bone interface regeneration [ 140 ]. This
gradient was achieved by controlling the location of a retrovirus for osteogenic
transcription factor Runx2/Cbfa1 on a collagen I scaffold. Dermal rat fibroblasts
were seeded on these scaffolds and osteogenic and fibroblastic cell activity was
studied. After 42 days of in vitro culture, microCT imaging revealed zonal organi-
zation of mineral deposition and fibroblastic ECM on the graded scaffold. Quanti-
tative RT-PCR confirmed upregulated expression of osteogenic specific markers in
fibroblasts at regions of dense retroviral coating. These constructs were then
implanted into ectopic subcutaneous sites of syngeneic rats. After 14 days in vivo ,