Multilayered microspheres for the controlled release of growth factors in tissue engineering.

Tissue regeneration may be stimulated by growth factors but to be effective, this delivery must be sustained and requires delivery vehicles that overcome the short half-life of these molecules in vivo. One promising approach is to couple growth factors to the biomaterial surface so that they are readily bioavailable. Here the layer-by-layer process was used to construct a multilayered polyelectrolyte delivery system on the surface of poly(lactic-co-glycolic) acid constructs. The system was first optimized on a planar surface before translation to a 3D microsphere system. The layers incorporated heparin to facilitate the loading of basic fibroblast growth factor and increase growth factor stability. Cross-linked capping layers also reduced any burst release. The model growth factor was released in a sustained manner and stimulated significantly higher cell proliferation in vitro on release compared with the addition of the growth factor heparin complex free in solution, demonstrating the promise of this approach.

Porous PLGA microspheres tailored for dual delivery of biomolecules via layer-by-layer assembly.

Tissue engineering is a complex and dynamic process that requires varied biomolecular cues to promote optimal tissue growth. Consequently, the development of delivery systems capable of sequestering more than one biomolecule with controllable release profiles is a key step in the advancement of this field. This study develops multilayered polyelectrolyte films incorporating alpha-melanocyte stimulating hormone (α-MSH), an anti-inflammatory molecule, and basic fibroblast growth factor (bFGF). The layers were successfully formed on macroporous poly lactic-co-glycolic acid microspheres produced using a combined inkjet and thermally induced phase separation technique. Release profiles could be varied by altering layer properties including the number of layers and concentrations of layering molecules. α-MSH and bFGF were released in a sustained manner and the bioactivity of α-MSH was shown to be preserved using an activated macrophage cell assay in vitro. The system performance was also tested in vivo subcutaneously in rats. The multilayered microspheres reduced the inflammatory response induced by a carrageenan stimulus 6 weeks after implantation compared to the non-layered microspheres without the anti-inflammatory and growth factors, demonstrating the potential of such multilayered constructs for the controlled delivery of bioactive molecules.

Use of a short peptide as a building block in the layer-by-layer assembly of biomolecules on polymeric surfaces.

The incorporation of low molecular weight drugs and therapeutic peptides into multilayer films assembled via the layer-by-layer technique can potentially provide means to deliver small molecules to target sites and to tune their release. This study describes the use of both hydrophobic and electrostatic interactions to incorporate a tridecapeptide antiinflammatory hormone, α-melanocyte stimulating hormone (α-MSH), as a building block at the base of a multilayer assembly of hyaluronic acid (HA) and chitosan (CS) on poly(lactic-co-glycolic acid) (PLGA) surfaces. A range of switching layers, including a neutral lipid, dioleylphosphatidylcholine (DOPC), a negatively charged lipid mixture DOPC/dioleylphosphatidylserine (DOPS) and a negatively charged polysaccharide, HA, were investigated for their ability to support subsequent HA and CS layers. Molecular dynamics simulations were performed to examine the structure and surface chemistry of α-MSH in solution and on surfaces to provide insights into the conditions most likely to support multilayer assembly. The multilayer assembly was stable at physiological pH and was successfully applied to particulate systems.

Coating and release of an anti-inflammatory hormone from PLGA microspheres for tissue engineering.

Many biomaterials used in tissue engineering cause a foreign body response in vivo, which left untreated can severely reduce the effectiveness of tissue regeneration. In this study, an anti-inflammatory hormone α-melanocyte stimulating hormone (α-MSH) was physically adsorbed to the surface of biodegradable poly (lactic-co-glycolic) acid (PLGA) microspheres to reduce inflammatory responses to this material. The stability and adsorption isotherm of peptide binding were characterized. The peptide secondary structure was not perturbed by the adsorption and subsequent desorption process. The α-MSH payload was released over 72 h and reduced the expression of the inflammatory cytokine, Tumor necrosis factor-α (TNF-α) in lipopolysaccharide activated RAW 264.7 macrophage cells, indicating that the biological activity of α-MSH was preserved. α-MSH coated PLGA microspheres also appeared to reduce the influx of inflammatory cells in a subcutaneous implantation model in rats. This study demonstrates the potential of α-MSH coatings for anti-inflammatory delivery and this approach may be applied to other tissue engineering applications.