RESUMO
Clinically, bone tissue replacements and/or bone repair are challenging. Strategies based on well-defined combinations of osteoconductive materials and osteogenic cells are promising to improve bone regeneration but still require improvement. Herein, we combined polycaprolactone (PCL) fibers, carbon nanotubes (CNT), and hydroxyapatite (nHap) nanoparticles to develop the next generation of bone regeneration material. Fibers formed by rotary jet spinning (RJS) instead of traditional electrospinning (ES) with embedded bone marrow mesenchymal stem cells (BMMSCs) showed the best outcomes to repair rat calvarial defects after 6 weeks. To understand this, it was observed that different morphologies were formed depending on the manufacturing method used. RJS fibers presented a particular topography with rough fibers, which allowed for better cellular growth and cell spreading in vitro around and into a three-dimensional (3D) mesh, while fibers made by ES were more smooth and cellular growth was only measured on the 3D mesh surface. The fibers with incorporated nHap/CNT nanoparticles enhanced in vitro cell performance as indicated by more cellular proliferation, alkaline phosphatase activity, proliferation, and deposition of calcium. Greater bone neoformation occurred by combining three characteristics: the presence of nHap and CNT nanoparticles, the topography of the RJS fibers, and the addition of BMMSCs. RJS fibers with nanoparticles and seeded with BMMSCs showed 10â¯136 mm3 of bone neoformation, meaning a 10-fold increase compared to using RJS only and BMMSCs (0.853 mm3) and a 5-fold increase from using ES only (2054 mm3) after 6 weeks of implantation. Conversely, none of these approaches used individually showed any significant difference for in vivo bone neoformation, suggesting that their combination is essential for optimizing bone formation. In summary, our work generated a potential material composed of well-defined combinations of suitable scaffolds seeded with BMMSCs for enhancing numerous orthopedic tissue engineering applications.
Assuntos
Células-Tronco Mesenquimais , Nanotubos de Carbono , Animais , Osso e Ossos , Durapatita/farmacologia , Poliésteres , Ratos , Alicerces TeciduaisRESUMO
Herein, we report the design of electrospun ultrathin fibers based on the combination of three different polymers polycaprolactone (PCL), polyethylene glycol (PEG), and gelatin methacryloyl (GelMA), and their potential bactericidal activity against three different bacteria Staphylococcus aureus (S. aureus), Pseudomonas aeruginosa (P. aeruginosa), and Methicillin-resistant Staphylococcus aureus (MRSA). We evaluated the morphology, chemical structure and wettability before and after UV photocrosslinking of the produced scaffolds. Results showed that the developed scaffolds presented hydrophilic properties after PEG and GelMA incorporation. Moreover, they were able to significantly reduce gram-positive, negative, and MRSA bacteria mainly after UV photocrosslinking (PCL:PEG:GelMa-UV). Furthermore, we performed a series of study for gaining a better mechanistic understanding of the scaffolds bactericidal activity through protein adsorption study and analysis of the reactive oxygen species (ROS) levels. Furthermore, the in vivo subcutaneous implantation performed in rats confirmed the biocompatibility of our designed scaffolds.
Assuntos
Materiais Biocompatíveis/farmacologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Nanofibras/química , Pseudomonas aeruginosa/efeitos dos fármacos , Alicerces Teciduais/química , Animais , Materiais Biocompatíveis/química , Linhagem Celular , Gelatina/química , Humanos , Masculino , Teste de Materiais/métodos , Metacrilatos/química , Testes de Sensibilidade Microbiana , Modelos Animais , Poliésteres/química , Polietilenoglicóis/química , Ratos , Ratos Wistar , Engenharia Tecidual/métodos , Infecção dos Ferimentos/microbiologia , Infecção dos Ferimentos/prevenção & controleRESUMO
BACKGROUND: Here, electrospun fibers based on a blend of polycaprolactone (PCL), poly(ethylene glycol) (PEG), and gelatin methacryloyl (GelMA) were developed. The careful choice of this polymer combination allowed for the preparation of a biomaterial that preserved the mechanical strength of PCL, while at the same time improving the hydrophilicity of the blended material and human osteoblast maturation. METHODS: The morphology, chemical structure, wettability, and mechanical properties before and after UV photocrosslinking were evaluated. Furthermore, human osteoblasts (hFOB) were cultivated for up to 21 days on the scaffolds, and their potential to upregulate cell proliferation, alkaline phosphatase (ALP) activity, and calcium deposition were investigated. RESULTS: Contact angle measurement results showed that the developed scaffolds presented hydrophilic properties after PEG and GelMA incorporation before (25°) and after UV photocross-linking (69°) compared to pure PCL (149°). PCL:PEG:GelMA-UV displayed a slight increase in mechanical strength (elastic modulus ~37 MPa) over PCL alone (~33 MPa). Normally, an increase in strength of fibers leads to a decrease in elongation at break, due to the material becoming less deformable and stiffer, thus leading to breaks at low strain. This behavior was observed by comparing PCL (elongation at break ~106%) and PCL:PEG:GelMA-UV (~50%). Moreover, increases in ALP activity (10-fold at day 14) and calcium deposition (1.3-fold at day 21) by hFOBs were detected after PEG and GelMA incorporation after UV photocross-linking compared to pure PCL. Ultrathin and hydrophilic fibers were obtained after PEG and GelMA incorporation after UV photocrosslinking, but the strength of PCL was maintained. Interestingly, those ultrathin fiber characteristics improved hFOB functions. CONCLUSION: These findings appear promising for the use of these electrospun scaffolds, based on the combination of polymers used here for numerous orthopedic applications.
Assuntos
Teste de Materiais , Nanofibras/química , Engenharia Tecidual/métodos , Alicerces Teciduais/química , Fosfatase Alcalina/metabolismo , Animais , Módulo de Elasticidade , Gelatina/química , Humanos , Metacrilatos/química , Nanofibras/ultraestrutura , Osteoblastos/citologia , Osteogênese/fisiologia , Poliésteres/química , Polietilenoglicóis/química , Espectroscopia de Infravermelho com Transformada de Fourier , Tensão Superficial , Sus scrofaRESUMO
The treatment of musculoskeletal defects is currently limited by the tissue-regenerative materials available to orthopedic surgeons: autologous bone grafts only have a finite amount of harvestable material within a given patient, while allografts are prone to severe immunological complications and host rejection. With this motivation, the production of poly(ε-caprolactone) (PCL) scaffolds as synthetic, biomimetic biomaterials was investigated, with a specific focus on potential orthopedic translation. PCL scaffolds were produced through three different fabrication techniques: electrospinning (ES), rotary jet spinning (RJS), and airbrush (AB). ES and RJS were observed to produce microfibrillar scaffolds, while all AB products were nanofibrous. Osteoblast viability, within the PCL scaffolds, and the osteogenic phenotype were assessed in vitro through a combination of adherence, metabolic activity, proliferation, gene expression, alkaline phosphatase bioactivity, and calcium deposition assays. While the polymeric scaffolds induced slight reductions in initial osteoblast adhesion and metabolic activity, seeded cells were able to proliferate and demonstrate the bone formation phenotype. AB products demonstrated reduced bacterial surface colonization when inoculated with both Gram-positive (Staphylococcus aureus) and Gram-negative (Pseudomonas aeruginosa) bacterial strains, in comparison to the microfibrous ES and RJS products, without any small-molecule antibiotics, antimicrobial peptides, or reactive nanomaterials included during scaffold synthesis.