TGF-beta1 release from biodegradable polymer microparticles: its effects on marrow stromal osteoblast function.
BACKGROUND
Controlled release of transforming growth factor-beta1 (TGF-beta1) 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-beta1 delivery and the effects of released TGF-beta1 on the proliferation and differentiation of marrow stromal cells in vitro.


METHODS
Recombinant human TGF-beta1 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-beta1 was determined after 3 days in culture. The effects of TGF-beta1 released from PLGA/PEG microparticles on marrow stromal cell proliferation and osteoblastic differentiation were assessed during a 21-day period.


RESULTS
TGF-beta1 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-beta1 released from the microparticles similar to that of added TGF-beta1, indicating that the activity of TGF-beta1 was retained during microparticle fabrication and after growth factor release. At an optimal TGF-beta1 dosage of 1.0 ng/ml after 3 days, the released TGF-beta1 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-beta1, and the released growth factor enhances marrow stromal cell proliferation and osteoblastic differentiation in vitro.


CONCLUSIONS
Controlled release of TGF-beta1 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.
Lu
L
L
Department of Bioengineering, Rice University, Houston, Texas, USA.
Yaszemski
M J
MJ
Mikos
A G
AG
D000818:Animals; D001672:Biocompatible Materials; D001673:Biodegradation, Environmental; D001854:Bone Marrow Cells / Q000187:drug effects / Q000502:physiology*; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004337:Drug Carriers; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives; D008297:Male; D008855:Microscopy, Electron, Scanning; D008863:Microspheres; D010006:Osteoblasts / Q000187:drug effects / Q000502:physiology*; D011092:Polyethylene Glycols; D011098:Polyglactin 910; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000493:pharmacokinetics / Q000494:pharmacology; D012710:Serum Albumin, Bovine; D017154:Stromal Cells / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D016212:Transforming Growth Factor beta / Q000008:administration & dosage / Q000493:pharmacokinetics* / Q000494:pharmacology; D053773:Transforming Growth Factor beta1
D001672:Biocompatible Materials; D004337:Drug Carriers; D011994:Recombinant Proteins; C508960:TGFB1 protein, human; C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C039217:fluorescein isothiocyanate bovine serum albumin; D012710:Serum Albumin, Bovine; D011098:Polyglactin 910; D011092:Polyethylene Glycols; D016650:Fluorescein-5-isothiocyanate