ISSN:
0021-9304
Keywords:
cytokine delivery via covalent binding
;
tissue repair
;
biomaterials
;
collagen
;
transforming growth factor β
;
polyethylene glycol
;
Chemistry
;
Polymer and Materials Science
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Medicine
,
Technology
Notes:
To overcome rapid diffusion and clearance from the implant site and to increase stability, recombinant transforming growth factor β2 (TGF-β2) was covalently bound to injectable bovine dermal fibrillar collagen (FC) and its activity compared to admixed TGF-β2. Covalent binding was achieved in a two-step procedure: First, TGF-β2 was reacted with the difunctional polyethylene glycol (PEG) linker, and then the PEG-attached TGF-β2 (PEG-TGF-β2) was bound to the fibrillar collagen (FC-PEG-TGF-β2). Initial binding of TGF-β2 to difunctional succinimidyl glutarate (D-SG-PEG) or succinimidyl propionate polyethylene glycol (D-SE-PEG) linkers was completed after reacting for 8 or 10 min as monitored by reverse-phase high-performance liquid chromatography. After reaction with injectable fibrillar collagen, extraction of unbound PEG-TGF-β2 and Western blot analysis, using a TGF-β specific antibody, demonstrated that at least 85% of the TGF-β2 was bound to the fibrillar collagen. The activity of PEG-TGF-β2 was fully stable in phosphate-buffered saline at 4°C and 37°C for at least up to 4 weeks. Unmodified TGF-β2 mixed with fibrillar collagen was completely inactivated after 1 week of incubation, as measured by the mink lung epithelial cell (Mv1Lu) growth inhibition assay. Formulations of FC-PEG-TGF-β2 containing 40 μg/mL TGF-β2 were implanted subcutaneously into rats and analyzed after days 7, 21, and 42. All TGF-β2-containing formulations showed the TGF-β typical fibroblastic response at the day 7 time point. Covalent binding of TGF-β2 to collagen with both difunctional PEG crosslinkers resulted in a significantly stronger and longer-lasting TGF-β2 response than that observed with admixed formulations of collagen and TGF-β. The TGF-β response with FC-PEG-TGF-β2 lasted up to day 42 but was not seen after day 7 for TGF-β2 admixed to FC. These findings clearly demonstrate that TGF-β2 remains fully active after being covalently bound to collagen via difunctional PEG. In addition, covalent binding potentiates and prolongs in vivo TGF-β responses and stabilizes the TGF-β in vitro. Results suggest that this method of formulation could be useful to stabilize and deliver similar peptide growth factors or biologically active agents. © 1998 John Wiley & Sons, Inc. J Biomed Mater Res, 39, 539-548, 1998.
Additional Material:
10 Ill.
Type of Medium:
Electronic Resource
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