Electrospun PLLA Nanofiber Scaffolds and Their Use in Combination with BMP-2 for Reconstruction of Bone Defects

Introduction Adequate migration and differentiation of mesenchymal stem cells is essential for regeneration of large bone defects. To achieve this, modern graft materials are becoming increasingly important. Among them, electrospun nanofiber scaffolds are a promising approach, because of their high physical porosity and potential to mimic the extracellular matrix (ECM). Materials and Methods The objective of the present study was to examine the impact of electrospun PLLA nanofiber scaffolds on bone formation in vivo, using a critical size rat calvarial defect model. In addition we analyzed whether direct incorporation of bone morphogenetic protein 2 (BMP-2) into nanofibers could enhance the osteoinductivity of the scaffolds. Two critical size calvarial defects (5 mm) were created in the parietal bones of adult male Sprague-Dawley rats. Defects were either (1) left unfilled, or treated with (2) bovine spongiosa, (3) PLLA scaffolds alone or (4) PLLA/BMP-2 scaffolds. Cranial CT-scans were taken at fixed intervals in vivo. Specimens obtained after euthanasia were processed for histology, histomorphometry and immunostaining (Osteocalcin, BMP-2 and Smad5). Results PLLA scaffolds were well colonized with cells after implantation, but only showed marginal ossification. PLLA/BMP-2 scaffolds showed much better bone regeneration and several ossification foci were observed throughout the defect. PLLA/BMP-2 scaffolds also stimulated significantly faster bone regeneration during the first eight weeks compared to bovine spongiosa. However, no significant differences between these two scaffolds could be observed after twelve weeks. Expression of osteogenic marker proteins in PLLA/BMP-2 scaffolds continuously increased throughout the observation period. After twelve weeks osteocalcin, BMP-2 and Smad5 were all significantly higher in the PLLA/BMP-2 group than in all other groups. Conclusion Electrospun PLLA nanofibers facilitate colonization of bone defects, while their use in combination with BMP-2 also increases bone regeneration in vivo and thus combines osteoconductivity of the scaffold with the ability to maintain an adequate osteogenic stimulus

PEG-Chitosan Hydrogel with Tunable Stiffness for Study of Drug Response of Breast Cancer Cells

Mechanical properties of the extracellular matrix have a profound effect on the behavior of anchorage-dependent cells. However, the mechanisms that define the effects of matrix stiffness on cell behavior remains unclear. Therefore, the development and fabrication of synthetic matrices with well-defined stiffness is invaluable for studying the interactions of cells with their biophysical microenvironment in vitro. We demonstrate a methoxypolyethylene glycol (mPEG)-modified chitosan hydrogel network where hydrogel stiffness can be easily modulated under physiological conditions by adjusting the degree of mPEG grafting onto chitosan (PEGylation). We show that the storage modulus of the hydrogel increases as PEGylation decreases and the gels exhibit instant self-recovery after deformation. Breast cancer cells cultured on the stiffest hydrogels adopt a more malignant phenotype with increased resistance to doxorubicin as compared with cells cultured on tissue culture polystyrene or Matrigel. This work demonstrates the utility of mPEG-modified chitosan hydrogel, with tunable mechanical properties, as an improved replacement of conventional culture system for in vitro characterization of breast cancer cell phenotype and evaluation of cancer therapies

High Affinity Binding of an Engineered, Modular Peptide to Bone Tissue

Bone grafting procedures have become common due in part to a global trend of population aging. Native bone graft is a popular choice when compared to various synthetic bone graft substitutes, owing to superior biological activity. Nonetheless, the insufficient ability of bone allograft to induce new bone formation and the insufficient remodeling of native bone grafts call for osteoinductive factors during bone repair, exemplified by recombinant human bone morphogenetic protein 2 (rhBMP2). We previously developed a modular bone morphogenetic peptide (mBMP) to address complications associated with the clinical use of rhBMP2 as a bone graft substitute. The mBMP is designed to strongly bind to hydroxyapatite, the main inorganic component of bone and teeth, and to provide pro-osteogenic properties analogous to rhBMP2. Our previous <i>in vivo</i> animal studies showed that mBMP bound to hydroxyapatite-coated orthopedic implants with high affinity and stimulated new bone formation. In this study, we demonstrate specific binding of mBMP to native bone grafts. The results show that mBMP binds with high affinity to both cortical and trabecular bones, and that the binding is dependent on the mBMP concentration and incubation time. Importantly, efficient mBMP binding is also achieved in an <i>ex vivo</i> bone bioreactor where bone tissue is maintained viable for several weeks. In addition, mBMP binding can be localized with spatial control on native bone tissue via simple methods, such as dip-coating, spotting, and direct writing. Taken together with the pro-osteogenic activity of mBMP established in previous bone repair models, these results suggest that mBMP may promote bone healing when coated on native bone grafts in a clinically compatible manner

Extending Foldamer Design beyond α-Helix Mimicry: α/β-Peptide Inhibitors of Vascular Endothelial Growth Factor Signaling

Diverse strategies have been explored to mimic the surface displayed by an α-helical segment of a protein, with the goal of creating inhibitors of helix-mediated protein–protein interactions. Many recognition surfaces on proteins, however, are topologically more complex and less regular than a single α-helix. We describe efforts to develop peptidic foldamers that bind to the irregular receptor-recognition surface of vascular endothelial growth factor (VEGF). Our approach begins with a 19-residue α-peptide previously reported by Fairbrother et al. (<i>Biochemistry</i> <b>1998</b>, <i>37</i>, 17754) to bind to this surface on VEGF. Systematic evaluation of α→β replacements throughout this 19-mer sequence enabled us to identify homologues that contain up to ∼30% β residues, retain significant affinity for VEGF, and display substantial resistance to proteolysis. These α/β-peptides can block VEGF-stimulated proliferation of human umbilical vein endothelial cells