Acrylonitrile Butadiene Styrene/Thermoplastic Polyurethane Blends for Material Extrusion Three-Dimensional Printing: Effects of Blend Composition on Printability and Properties.

Blend filaments of acrylonitrile butadiene styrene (ABS) and thermoplastic polyurethane (TPU) were prepared at different weight ratios, i.e., 100:0, 70:30, 50:50, 30:70, and 0:100, for FDM printing; the prepared filaments, with an average diameter of 2.77 ± 0.19 mm, were encoded as A100, A70T30, A50T50, A30T70, and T100, respectively. The properties and printability of the filaments were thoroughly investigated. The blend composition, as well as the printing parameters, were optimized to obtain the FDM-printed objects with a well-defined surface structure and minimized warpages. The glass transition temperatures of ABS and TPU in the blends were not much altered from those of the parent filaments, whereas the thermal degradation characteristics of the blend filaments still fell between those of the neat filaments. The fractured surfaces of the filaments, observed by SEM, appeared smoother when higher amounts of TPU integrated; the smoothest surface of the ABS-based filament was found in A30T70, indicating the well-compatible blend characteristic. This was also confirmed by its rheological behavior examined by a parallel plate rheometer at 225 °C. Not only was the printability of the filaments improved, but also the warpages of the 3D-printed specimens were decreased when increasing amount of TPU was incorporated into the filaments. Among the printed objects, the A30T70 specimen exhibited the evenest surface morphology with the lowest surface roughness value of 32.9 ± 13.2 nm and the most uniform and consistent linear printing structure when being fabricated at the nozzle temperature of 225 °C and the printing bed temperature of 60 °C. However, the incorporation of TPU into the filaments markedly cut down both strength and modulus values of the fabricated materials up to about half but assisted the printed articles to absorb more energy, demonstrating that this polymer served as a good and effective toughener for ABS.

Chondrogenic Differentiation of Human Mesenchymal Stem Cells and Macrophage Polarization on 3D-Printed Poly(ε-caprolactone)/Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) Blended Scaffolds with Different Secondary Porous Structures.

This study was aimed to evaluate the chondrogenic differentiation of human mesenchymal stem cells (hMSCs) and polarization of THP-1-derived macrophages cultured on poly(ε-caprolactone) (PC)/poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PH) blended scaffolds with dual primary (PP) and secondary (SP) pores, which were fabricated via a 3D printing technique, i.e., fused deposition modeling, followed by a salt-leaching process at 50 °C for varied times, i.e., 15, 30, and 60 min. Sodium chloride (SC), a porogen, was initially incorporated in the blend at varied weight percentages, i.e., 0, 25, and 50%, whereas 1 M NaOH solution and deionized water were used as salt-leaching agents. To elucidate the surface properties of the developed scaffolds, directly governed by the amount of the salt originally mixed and the salt-leaching efficiency, several characterization techniques, e.g., scanning electron microscopy, X-ray microcomputed tomography, mercury intrusion porosimetry, atomic force microscopy, and contact angle measurement, were used. Meanwhile, the salt-leaching efficiency was determined by means of weight loss measurement and thermogravimetric analysis. It was found that the alkaline solution could satisfactorily leach out the salt particles in 60 min with a mild etching of the polymer framework. The most immensely and homogeneously pitted filament surface was observed in the NaOH-treated scaffold initially integrated with 50% salt, i.e., 60B_PC/PH/50SC; the SP structure was mostly open and interconnected. The size of most of micropores was about 0.14 µm. With its suitable microsurface roughness and hydrophilicity, 60B_PC/PH/50SC could properly support the initial attachment and lamellipodia formation of hMSCs, which was favorable for chondrogenesis. Consequently, a significantly increased ratio of glycosaminoglycans/deoxyribonucleic acid and a superior expression of the COL2A1 gene were detected when cells were grown on this material. Although 60B_PC/PH/50SC induced the macrophages to secrete a slightly high level of IL-1ß during the first few days of culture, the polarized M1 cells could return to a nearly normal stage at Day7, suggesting no unfavorable chronic inflammation caused by the material.

Hyaluronic acid-coated gold nanorods enhancing BMP-2 peptide delivery for chondrogenesis.

Bone morphogenic protein-2 (BMP-2) knuckle epitope peptide has been recently discovered and known to activate chondrogenesis. However, the applications of this soluble peptide remain very limited due to rapid diffusion resulting in poor cellular uptake into target cells. We herein designed nanoparticles made from hyaluronic acid functionalized gold nanorods (GNRs) to conjugate with thiolated BMP-2 knuckle epitope peptide via a two-step reaction. Hyaluronic acid was modified to have thiol functional groups to replace the cetyl trimethylammonium bromide ligands on the surface of GNRs. The thiolated peptides were subsequently reacted with hyaluronic acid on the surface on GNRs via a maleimide-hydrazide crosslinker. The conjugation was confirmed by the change of surface charge of GNRs and the plasmon shift. A colorimetric peptide assay suggested more than 69% of the thiolated peptides were conjugated with the hyaluronic acid coated gold nanorods. Moreover, in vitro cell viability showed that BMP-2 conjugated hyaluronic acid functionalized gold nanorods (B2HGR) were cytocompatible and did not cause cytotoxicity to fibroblast cells. The B2HGRs also significantly promote cellular uptake of the BMP-2 peptides in both human mesenchymal stem cells and porcine chondrocytes due to multivalent ligand binding to the BMP receptors on the cell surface resulting in receptor-mediated endocytosis. The enhanced cellular uptake was clearly observed under a confocal microscope resulting in the significant activation of type II collagen gene expression and glucosaminoglycan secretion in those cells. Furthermore, our delivery system is a proof-of-concept of using scaffolds in combination with nanodelivery platform to enhance cartilaginous repair. The peptide loading capacity and the release is not limited by the scaffolds. Therefore, our delivery platform has potential applications for cartilage regeneration in a preclinical and clinical setting in the future.

Chondrogenic phenotype in responses to poly(ɛ-caprolactone) scaffolds catalyzed by bioenzymes: effects of surface topography and chemistry.

Biodegradable poly(ε-caprolactone) (PCL) has been increasingly investigated as a promising scaffolding material for articular cartilage tissue repair. However, its use can be limited due to its surface hydrophobicity and topography. In this study, 3D porous PCL scaffolds fabricated by a fused deposition modeling (FDM) machine were enzymatically hydrolyzed using two different biocatalysts, namely Novozyme®435 and Amano lipase PS, at varied treatment conditions in a pH 8.0 phosphate buffer solution. The improved surface topography and chemistry of the PCL scaffolds were anticipated to ultimately boost the growth of porcine articular chondrocytes and promote the chondrogenic phenotype during cell culture. Alterations in surface roughness, wettability, and chemistry of the PCL scaffolds after enzymatic treatment were thoroughly investigated using several techniques, e.g., SEM, AFM, contact angle and surface energy measurement, and XPS. With increasing enzyme content, incubation time, and incubation temperature, the surfaces of the PCL scaffolds became rougher and more hydrophilic. In addition, Novozyme®435 was found to have a higher enzyme activity than Amano lipase PS when both were used in the same enzymatic treatment condition. Interestingly, the enzymatic degradation process rarely induced the deterioration of compressive strength of the bulk porous PCL material and slightly reduced the molecular weight of the material at the filament surface. After 28 days of culture, both porous PCL scaffolds catalyzed by Novozyme®435 and Amano lipase PS could facilitate the chondrocytes to not only proliferate properly, but also function more effectively, compared with the non-modified porous PCL scaffold. Furthermore, the enzymatic treatments with 50 mg of Novozyme®435 at 25 °C from 10 min to 60 min were evidently proven to provide the optimally enhanced surface roughness and hydrophilicity most significantly favorable for induction of chondrogenic phenotype, indicated by the greatest expression level of cartilage-specific gene and the largest production of total glycosaminoglycans.

Bisphosphonate activation of crystallized bioglass scaffolds for enhanced bone formation.

The interplay between bone formation by osteoblasts and bone resorption by osteoclasts has a critical effect on bone remodelling processes, and resultant bone quality. Bone scaffolds combined with anti-resorptive bisphosphonate drugs are a promising approach to achieving bone regeneration. Here, we have examined the synergistic effects of the bisphosphonate alendronate (ALD) coated onto calcium phosphate (CaP) modified, sintered bioactive glass 45S5 (BG) scaffolds, on osteoblast stimulation and osteoclast inhibition. After BG pre-treatment with ALD (10-8 M) for 5 days, human MG-63 osteoblasts displayed increased cellular proliferation and significantly enhanced alkaline phosphatase activity (ALP), in comparison with a non-ALD control BG. In contrast, human THP-1-derived osteoclasts cultured with 10-8 M ALD pretreated BG scaffolds showed a significant decrease in tartrate-resistant acid phosphatase (TRAcP) activity, and morphological changes indicative of functional inhibition, including reduced cell size and disruption of the osteoclast sealing zone (F-actin rings). These findings indicate that ALD-coated BG scaffolds promote osteoblast activity and inhibit osteoclast function to enhance bone formation.

PCL/PHBV blended three dimensional scaffolds fabricated by fused deposition modeling and responses of chondrocytes to the scaffolds.

In this study, poly(ε-caprolactone)/poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PCL/PHBV) blended porous scaffolds were fabricated by fused deposition modeling (FDM). PCL/PHBV filaments, initially prepared at different weight ratios, that is, 100/0, 75/25, 50/50, and 25/75, were fabricated by the lay-down pattern of 0/90/45/135° to obtain scaffolds with dimension of 6.0 × 6.0 × 2.5 mm3 and average filament diameters and channel sizes in the ranges of 370-390 µm and 190-210 µm, respectively. To enhance the surface hydrophilicity of the materials, the scaffolds were subsequently subjected to a low pressure oxygen plasma treatment. The untreated and plasma-treated scaffolds were comparatively characterized, in terms of surface properties, mechanical strength, and biological properties. From SEM, AFM, water contact angle, and XPS results, the surface roughness, wettability, and hydrophilicity of the blended scaffolds were found to be enhanced after plasma treatment, while the compressive strength of the scaffolds was scarcely changed. It was, however, found to increase with an increasing content of PHBV incorporated. The porcine chondrocytes exhibited higher proliferative capacity and chondrogenic potential when being cultured on the scaffolds with greater PHBV contents, especially when they were plasma-treated. The PCL/PHBV scaffolds were proven to possess good physical, mechanical, and biological properties that could be appropriately used in articular cartilage regeneration. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 1141-1150, 2017.

Enhancement of chondrocyte proliferation, distribution, and functions within polycaprolactone scaffolds by surface treatments.

Enhancement of porcine chondrocyte growth, distribution and functions within polycaprolactone (PCL) scaffolds was attempted using alkaline hydrolysis and oxygen plasma treatment. The hydrolysis of PCL was performed either before or after scaffold fabrication in the preparations of pre-hydrolyzed PCL (pre-HPCL) or post-HPCL scaffolds, respectively. The PCL, pre-HPCL, and post-HPCL scaffolds were subsequently plasma-treated to yield plasma-treated PCL, plasma-treated pre-HPCL, and plasma-treated post-HPCL scaffolds, respectively. All scaffolds were comparatively characterized, in terms of surface morphology, hydrophilicity, and atomic composition using scanning electron microscopy, contact angle measurement and X-ray photoelectron spectroscopy, respectively. The interactions of chondrocytes with individual scaffolds were assessed, in terms of cartilage-gene expression and cartilaginous matrix production using reverse transcription polymerase chain reaction analysis and glycosaminoglycans (GAGs) assay, respectively. The cell infiltration and cartilaginous matrix distribution were investigated by histological and immunofluorescence analysis. The results revealed that the plasma treatment exhibited a more prominent effect on the enhancement of surface roughness and hydrophilicity of the scaffolds than the alkaline hydrolysis. The scaffolds subjected to both surface treatments stimulated the cells to secret more GAGs and type II collagen. The sequence of hydrolysis of PCL also evidently played a crucial role in the hydrophilicity of the materials and the cartilage-gene expression and cartilaginous matrix production of the cultured chondrocytes. The hydrolysis of PCL prior to the fabrication, followed by the oxygen plasma treatment of the resulting fabricated scaffold, yielded plasma-treated pre-HPCL scaffold with homogeneous hydrophilic characteristics all over the material. Consequently, the cells could proliferate well, infiltrate most deeply and ultimately produce the highest amounts of the cartilage-specific substances throughout this scaffold.