Biomedical Engineering Reference
In-Depth Information
46 CHAPTER 3.
IN VITRO
TISSUE ENGINEERING
Growth factors are typically delivered as soluble components in culture media cocktails. While
this is acceptable for
in vitro
experiments, delivery becomes more complicated once the construct
is implanted. An alternative approach is to include polymeric carriers, such as microspheres, in the
construct so that growth factors are released over time [
334
]. Early approaches found little success
since most polymer release times lasted only a few days. Once freed, growth factors can degrade
within a week, so long-term treatments using these carriers would be infeasible [
413
]. However,
current research has indicated that alternative polymers, such as elastin-like polypeptides, have the
capability to extend the release time of drugs to weeks or months [
414
]. These carriers would allow
long-term stimulation of the implant with a local source of growth factors, further stimulating matrix
growth and possibly helping integration with the surrounding tissue.
Another alternative to long-term growth factor stimulation is to modify the gene expression
of implanted cells using either transfection or other forms of genetic modification [
415
,
416
]. In
this case, growth factors are secreted by cells within the defect site. Local stimulation means that
implant growth and integration do not rely on externally provided drugs. Many different approaches
can be used to apply this technique. For example, all implanted cells could be modified or only a
fraction. Alternatively, the modified gene could be conditionally active, which would be advantageous
if stimulation is only desired for certain periods during regeneration. While this approach seems
attractive, the practicality of genetic modification creates a large barrier to its current success. Total
control of how the gene is expressed is not currently feasible, which creates a safety issue. Furthermore,
the long-term effects of elevated growth factor levels are not known, especially on neighboring tissues
that are not involved with the cartilage repair process.
Growth factors do not necessarily need to be available as unbound molecules to induce a
response from resident cells. Modifying the scaffold material itself with growth factors is a possible
means to stimulate cells growing in the construct. Proteins such as TGF-
β
, IGF, PDGF, HGF,
and FGF have all been used in such modifications. The benefit of scaffold-bound growth factors
for
in vivo
experiments is clear, but even
in vitro
experiments could be enhanced since bioactive
molecules would be distributed evenly throughout the scaffold, affecting all regions equally. With-
out restriction of the growth factors, diffusion could result in loss of bioactive molecules to the
surrounding environment.
There are two primary methods for including growth factors in engineered constructs. The
first, encapsulation, was mentioned previously and involves sequestering stimulatory proteins in
polymeric materials that have controllable release characteristics. Alternatively, a growth factor can
be encapsulated in the bulk scaffold material, assuming that the scaffold has characteristics that can
restrict the diffusion of certain molecules out of the construct. For example, hydrogels can restrict the
diffusion of growth factors for short periods of time in comparison to meshes or felts. Incorporating
multiple types of materials in a scaffold is another means to allow drug release over time. A two-
phase PLGA implant loaded with TGF-
β
showed good results when implanted into osteochondral
defects [
346
]. Microparticles are the most common methodology for including growth factors in
