Background: Controlled release of transforming
growth factor-&betabeta;1 (TGF-&betabeta;1) to a bone
defect may be beneficial for the induction of a bone regeneration cascade.
The objectives of this work were to assess the feasibility of using
biodegradable polymer microparticles as carriers for controlled
TGF-&betabeta;1 delivery and the effects of released TGF-&betabeta;1
on the proliferation and differentiation of marrow stromal cells in
Methods: Recombinant human TGF-&betabeta;1
was incorporated into microparticles of blends of poly(DL-lactic-co-glycolic
acid) (PLGA) and poly(ethylene glycol) (PEG). Fluorescein isothiocynate-labeled
bovine serum albumin (FITC-BSA) was co-encapsulated as a porogen.
The effects of PEG content (0, 1, or 5% by weight [wt%])
and buffer pH (3, 5, or 7.4) on the protein release kinetics and
the degradation of PLGA were determined in vitro for
as long as 28 days. Rat marrow stromal cells were seeded on a biodegradable
poly(propylene fumarate) (PPF) substrate. The dose response and biological
activity of released TGF-&betabeta;1 was determined after 3
days in culture. The effects of TGF-&betabeta;1 released from
PLGA/PEG microparticles on marrow stromal cell proliferation
and osteoblastic differentiation were assessed during a 21-day period.
Results: TGF-&betabeta;1 was encapsulated along
with FITC-BSA into PLGA/PEG blend microparticles and released
in a multiphasic fashion including an initial burst for as long
as 28 days in vitro. Increasing the initial PEG
content resulted in a decreased cumulative mass of released proteins. Aggregation
of FITC-BSA occurred at lower buffer pH, which led to decreased
release rates of both proteins. The degradation of PLGA was increased at
higher PEG content and significantly accelerated at acidic pH conditions.
Rat marrow stromal cells cultured on PPF substrates showed a dose
response to TGF-&betabeta;1 released from the microparticles similar
to that of added TGF-&betabeta;1, indicating that the activity
of TGF-&betabeta;1 was retained during microparticle fabrication
and after growth factor release. At an optimal TGF-&betabeta;1
dosage of 1.0 ng/ml after 3 days, the released TGF-&betabeta;1 enhanced
the proliferation and osteoblastic differentiation of marrow stromal
cells over 21 days of culture, with increased total cell number,
alkaline phosphatase activity, and osteocalcin production.
Conclusions: PLGA/PEG blend microparticles
can serve as delivery vehicles for controlled release of TGF-&betabeta;1,
and the released growth factor enhances marrow stromal cell proliferation
and osteoblastic differentiation in vitro.
Clinical Relevance: Controlled release of TGF-&betabeta;1
from PLGA/PEG microparticles is representative of emerging tissue
engineering technologies that may modulate cellular responses to
encourage bone regeneration at a skeletal defect site.