Development and characterization of reinforced poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactide) scaffolds for bone tissue engineering

Novel reinforced poly(l-lactic acid) (PLLA) scaffolds such as solid shell, porous shell, one beam and two beam reinforced scaffolds were developed to improve the mechanical properties of a standard PLLA scaffold. Experimental results clearly indicated that the compressive mechanical properties such as the strength and the modulus are effectively improved by introducing the reinforcement structures. A linear elastic model consisting of three phases, that is, the reinforcement, the porous matrix and the boundary layer was also introduced in order to predict the compressive moduli of the reinforced scaffolds. The comparative study clearly showed that the simple theoretical model can reasonably predict the moduli of the scaffolds with three phase structures. The failure mechanism of the solid shell and the porous shell reinforced scaffolds under compression were found to be buckling of the solid shell and localized buckling of the struts constructing the pores in the porous shell, respectively. For the beam reinforced scaffolds, on the contrary, the primary failure mechanism was understood to be micro-cracking within the beams and the subsequent formation of the main-crack due to the coalescence of the micro-racks. The biological study was exhibited that osteoblast-like cells, MC3T3-E1, were well adhered and proliferated on the surfaces of the scaffolds after 12 days culturing.     

Electrospun poly(<Emphasis Type="SmallCaps">d</Emphasis>/<Emphasis Type="SmallCaps">l</Emphasis>-lactide-<Emphasis Type="Italic">co</Emphasis>-<Emphasis Type="SmallCaps">l</Emphasis>-lactide) hybrid matrix: a novel scaffold material for soft tissue engineering

Electrospinning is a long-known polymer processing technique that has received more interest and attention in recent years due to its versatility and potential use in the field of biomedical research. The fabrication of three-dimensional (3D) electrospun matrices for drug delivery and tissue engineering is of particular interest. In the present study, we identified optimal conditions to generate novel electrospun polymeric scaffolds composed of poly-d/l-lactide and poly-l-lactide in the ratio 50:50. Scanning electron microscopic analyses revealed that the generated poly(d/l-lactide-co-l-lactide) electrospun hybrid microfibers possessed a unique porous high surface area mimicking native extracellular matrix (ECM). To assess cytocompatibility, we isolated dermal fibroblasts from human skin biopsies. After 5 days of in vitro culture, the fibroblasts adhered, migrated and proliferated on the newly created 3D scaffolds. Our data demonstrate the applicability of electrospun poly(d/l-lactide-co-l-lactide) scaffolds to serve as substrates for regenerative medicine applications with special focus on skin tissue engineering.     

Selective laser sintering of porous tissue engineering scaffolds from poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactide)/carbonated hydroxyapatite nanocomposite microspheres

This study focuses on the use of bio-nanocomposite microspheres, consisting of carbonated hydroxyapatite (CHAp) nanospheres within a poly(L: -lactide) (PLLA) matrix, to produce tissue engineering (TE) scaffolds using a modified selective laser sintering (SLS) machine. PLLA microspheres and PLLA/CHAp nanocomposite microspheres were prepared by emulsion techniques. The resultant microspheres had a size range of 5-30 mum, suitable for the SLS process. Microstructural analyses revealed that the CHAp nanospheres were embedded throughout the PLLA microsphere, forming a nanocomposite structure. A custom-made miniature sintering platform was installed in a commercial Sinterstation((R)) 2000 SLS machine. This platform allowed the use of small quantities of biomaterials for TE scaffold production. The effects of laser power; scan spacing and part bed temperature were investigated and optimized. Finally, porous scaffolds were successfully fabricated from the PLLA microspheres and PLLA/CHAp nanocomposite microspheres. In particular, the PLLA/CHAp nanocomposite microspheres appeared to be promising for porous bone TE scaffold production using the SLS technique.     

Programmed water-induced shape-memory of bioabsorbable poly(<Emphasis Type="SmallCaps">d</Emphasis>,<Emphasis Type="SmallCaps">l</Emphasis>-lactide): activation and properties in physiological temperature

This study reports of the novel water-induced shape-memory of bioabsorbable poly(d,l-lactide). We have developed an orientation-based programming process that generates an ability for poly(d,l-lactide) to transform its shape at 37°C in an aqueous environment without external energy and to adapt to a predefined stress level by stress generation or relaxation. In this orientation-programming process, polymer material is deformed and oriented at an elevated temperature and subsequently cooled down while retaining its deformed shape, tension, and polymer chain entanglements. At body temperature and in an aqueous environment, the shape-memory is activated by the plasticizing effect of water molecules diffused into the polymer matrix causing an entropy-driven directed relaxation of oriented and preloaded polymer chains. This plasticizing effect is clearly seen as a decrease of the onset glass transition temperature by 10–13°C. We found that γ-irradiation used for sterilizing the orientation-programmed materials strongly affected the shape-recovery rate, but not the recovery ratio. Both non-γ-irradiated and γ-irradiated sample materials showed excellent shape-recovery ratios during a ten-week test period: 94 and 97%, respectively. The orientation-programmed materials generated a predefined load in a 37°C aqueous environment when their shape-recovery was restricted, but when external tension was applied to them, they adapted to the predefined level by stress relaxation. Our results show that functionality in terms of shape-memory can be generated in bioabsorbable polymers without tailoring the polymer chain structure thus shortening the time from development of technology to its utilization in medical devices.     

Histological study of surface modified three dimensional poly (<Emphasis Type="SmallCaps">d</Emphasis>,<Emphasis Type="SmallCaps"> l</Emphasis>-lactic acid) scaffolds with chitosan in vivo

Biocompatibility and tissue regenerating capacity are essential for biomaterials that used in tissue engineering. The aim of this study was to histologically assess the tissue reactions and bone conductivities of surface modified three dimensional (3-D) poly (d, l-lactic acid) (PDLLA) scaffolds, which were coated with chitosan via a physical entrapment method. The native PDLLA scaffold was prepared via thermally induced phrase separation technique and was characterized by scanning electron microscopy (SEM) and differential scanning calorimetry (DSC). Osteocalcin assay, a method to evaluate the bone formation potential, has shown that the osteocalcin production in chitosan-modified 3-D PDLLA scaffold group was significantly higher (p < 0.05) than that of in control. The tissue reactions and bone conductivities between surface modified PDLLA and native PDLLA scaffolds were evaluated using a rabbit radialis defect model in vivo and compared at different implantation intervals (2, 4, 8 and 12 weeks). The histological results have shown a higher bone formation potential and better biocompatibility of chitosan-modified 3-D PDLLA scaffolds as compared with the control group scaffolds.     

Body distribution of poly(<Emphasis Type="SmallCaps">d</Emphasis>,<Emphasis Type="SmallCaps">l</Emphasis>-lactide-<Emphasis Type="Italic">co</Emphasis>-glycolide) copolymer degradation products in rats

Poly (d,l-lactide-co-glycolide) (PLGA) copolymers are among the few synthetic polymers approved for human use, but the biocompatibility of PLGA-derived oligomers and particles remains questionable. Here, high molecular weight PLGA (Mw = 32,000) was radiolabeled with ¹²⁵I in chloroform solution, and the body distribution of PLGA copolymer degradation products was examined following subcutaneous implantation of round ¹²⁵I-PLGA films on the back of Sprague Dawley rats. Autoradiographic images of the PLGA implant taken at 2, 4, 6, 8, 10, and 12 weeks revealed that the central portion of the film degraded much more rapidly than the marginal portions. Examination of the body compartment distribution at these time points revealed that over one-half of the radioactivity was recovered from skin. The remaining radioactivity was concentrated in the blood, liver, and kidneys. Radioactivity steadily appeared in the blood and remained elevated up to 12 weeks after implantation, while the liver to kidney distribution began to decrease after 6 weeks. Cumulatively, these results indicate that the clearance of degraded particles and fragments from the implantation site is extremely delayed. Moreover, the degraded particles and fragments were selectively concentrated in the liver and kidneys, following release of degraded products into the bloodstream from the implantation site.     

Titanium dioxide (TiO<Subscript>2</Subscript>) nanoparticles filled poly(<Emphasis Type="SmallCaps">d</Emphasis>,<Emphasis Type="SmallCaps">l</Emphasis> lactid acid) (PDLLA) matrix composites for bone tissue engineering

Titanium dioxide (TiO₂) nanoparticles were investigated for bone tissue engineering applications with regard to bioactivity and particle cytotoxicity. Composite films on the basis of poly(d,l lactid acid) (PDLLA) filled with 0, 5 and 30 wt% TiO₂ nanoparticles were processed by solvent casting. Bioactivity, characterised by formation of hydroxyapatite (HA) on the materials surface, was investigated for both the free TiO₂ nanoparticles and PDLLA/TiO₂ composite films upon immersion in supersaturated simulated body fluid (1.5 SBF) for up to 3 weeks. Non-stoichiometric HA nanocrystals (ns-HA) with an average diameter of 40 nm were formed on the high content (30 wt% TiO₂) composite films after 2 weeks of immersion in 1.5 SBF. For the pure PDLLA film and the low content composite films (5 wt% TiO₂) trace amounts of ns-HA nanocrystals were apparent after 3 weeks. The TiO₂ nanopowder alone showed no bioactivity. The effect of TiO₂ nanoparticles (0.5–10,000 μg/mL) on MG-63 osteoblast-like cell metabolic activity was assessed by the MTT assay. TiO₂ particle concentrations of up to 100 μg/mL had no significant effect on MG-63 cell viability.     

Morphology and rheology of poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactide)/polystyrene blends filled with silica nanoparticles

The successful development of co-continuous structure from poly(l-lactide) (PLLA) blends by melt mixing with lower PLLA content is highly desired in preparing macroporous biomaterials. However, the low viscosity of PLLA makes it difficult to prepare co-continuous PLLA blends at low PLLA concentration. In this study, hydrophilic silica nanoparticle is adopted to control the morphology of co-continuous polystyrene (PS)/PLLA blends. The influence of nanoparticle concentration on the co-continuity intervals and rheological properties of PS/PLLA blends are examined. The morphological stability of blends against melt annealing is also determined and discussed with a conceptual coarsening model for co-continuous structure. The results demonstrate that the incorporation of silica nanoparticles into PS/PLLA blends can be used to prepare macroporous PLLA structure with controllable pore size at lower PLLA content.     

Elastomeric electrospun scaffolds of poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactide-<Emphasis Type="Italic">co</Emphasis>-trimethylene carbonate) for myocardial tissue engineering

In myocardial tissue engineering the use of synthetically bioengineered flexible patches implanted in the infarcted area is considered one of the promising strategy for cardiac repair. In this work the potentialities of a biomimetic electrospun scaffold made of a commercial copolymer of (l)-lactic acid with trimethylene carbonate (P(l)LA-co-TMC) are investigated in comparison to electrospun poly(l)lactic acid. The P(l)LA-co-TMC scaffold used in this work is a glassy rigid material at room temperature while it is a rubbery soft material at 37°C. Mechanical characterization results (tensile stress–strain and creep-recovery measurements) show that at 37°C electrospun P(l)LA-co-TMC displays an elastic modulus of around 20 MPa and the ability to completely recover up to 10% of deformation. Cell culture experiments show that P(l)LA-co-TMC scaffold promotes cardiomyocyte proliferation and efficiently preserve cell morphology, without hampering expression of sarcomeric alpha actinin marker, thus demonstrating its potentialities as synthetic biomaterial for myocardial tissue engineering.     

Study of poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactide) microparticles based on supercritical CO<Subscript>2</Subscript>

Poly(l-lactide) (PLLA) microparticles were prepared in supercritical anti-solvent process. The effects of several key factors on surface morphology, and particle size and particle size distribution were investigated. These factors included initial drops size, saturation ratio of PLLA solution, pressure, temperature, concentration of the organic solution, the flow rate of the solution and molecular weight of PLLA. The results indicated that the saturation ratio of PLLA solution, concentration of the organic solution and flow rate of the solution played important roles on the properties of products. Various microparticles with the mean particle size ranging from 0.64 to 6.64 μm, could be prepared by adjusting the operational parameters. Fine microparticles were obtained in a process namely solution-enhanced dispersion by supercritical fluids (SEDS) process with dichloromethane/acetone mixture as solution.     

In vitro investigation of nanohydroxyapatite/poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactic acid) spindle composites used for bone tissue engineering

Calcium phosphate ceramics such as synthetic hydroxyapatite and tricalcium phosphate are widely used in the clinic, but they stimulate less bone regeneration. In this paper, nano-hydroxyapatite/poly(l-lactic acid) (nano-HA/PLLA) spindle composites with good mechanical performance were fabricated by a modified in situ precipitation method. The HA part of composite, distributing homogenously in PLLA matrix, is spindle shape with size of 10–30 nm in diameter and 60–100 nm in length. The molar ratio of Ca/P in the synthesized nano-HA spindles was deduced as 1.52 from the EDS spectra, which is close to the stoichiometric composition of HA (Ca/P & 1.67). The compress strength is up to 150 MPa when the HA content increase to 20 %. The in vitro tests indicate that HA/PLLA bio-composites have good biodegradability and bioactivity when immersed in simulated body fluid solutions. All the results suggested that HA/PLLA nano-biocomposites are appropriate to be applied as bone substitute in bone tissue engineering.     

The search for novel superhard and incompressible materials on the basis of higher borides of <Emphasis Type="Italic">s</Emphasis>, <Emphasis Type="Italic">p</Emphasis>, <Emphasis Type="Italic">d</Emphasis> metals

The state of the art in the search for novel superhard and (or) incompressible materials on the basis of higher borides of s, p, d metals has been briefly reviewed. The information has been considered about experimental and theoretical studies of the following groups of borides: diborides of 4d, 5d heavy metals (Tc, Ru, Rh, Re, Os, and Ir), hexagonal tetraborides with the WB₄-type structure, and AMB₁₄ borides (where A, M are s, p metals) as well as of a number of related systems.     

Hydroxyapatite nanorod-reinforced biodegradable poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactic acid) composites for bone plate applications

Novel PLLA composite fibers containing hydroxyapatite (HAp) nanorods with or without surface lactic acid grafting were produced by extrusion for use as reinforcements in PLLA-based bone plates. Fibers containing 0–50% (w/w) HAp nanorods, aligned parallel to fiber axis, were extruded. Lactic acid surface grafting of HAp nanorods (lacHAp) improved the tensile properties of composites fibers better than the non-grafted ones (nHAp). Best tensile modulus values of 2.59, 2.49, and 4.12 GPa were obtained for loadings (w/w) with 30% lacHAp, 10% nHAp, and 50% amorphous HAp nanoparticles, respectively. Bone plates reinforced with parallel rows of these composite fibers were molded by melt pressing. The best compressive properties for plates were obtained with nHAp reinforcement (1.31 GPa Young’s Modulus, 110.3 MPa compressive strength). In vitro testing with osteoblasts showed good cellular attachment and spreading on composite fibers. In situ degradation tests revealed faster degradation rates with increasing HAp content. To our knowledge, this is the first study containing calcium phosphate–polymer nanocomposite fibers for reinforcement of a biodegradable bone plate or other such implants and this biomimetic design was concluded to have potential for production of polymer-based biodegradable bone plates even for load bearing applications.     

Osteointegration of poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactic acid)PLLA and poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactic acid)PLLA/poly(ethylene oxide)PEO implants in rat tibiae

Natural or synthetic materials may be used to aid tissue repair of fracture or pathologies where there has been a loss of bone mass. Polymeric materials have been widely studied, aiming at their use in orthopaedics and aesthetic plastic surgery. Polymeric biodegradable blends formed from two or more kinds of polymers could present faster degradation rate than homopolymers. The purpose of this work was to compare the biological response of two biomaterials: poly(L: -lactic acid)PLLA and poly(L: -lactic acid)PLLA/poly(ethylene oxide)PEO blend. Forty four-week-old rats were divided into two groups of 20 animals, of which one group received PLLA and the other PLLA/PEO implants. In each of the animals, one of the biomaterials was implanted in the proximal epiphysis of the right tibia. Each group was divided into subgroups of 5 animals, and sacrificed 2, 4, 8 and 16 weeks after surgery, respectively. Samples were then processed for analysis by light microscopy. Newly formed bone was found around both PLLA and PLLA/PEO implants. PLLA/PEO blends had a porous morphology after immersion in a buffer solution and in vivo implantation. The proportion 50/50 PLLA/PEO blend was adequate to promote this porous morphology, which resulted in gradual bone tissue growth into the implant.     

Porous-conductive chitosan scaffolds for tissue engineering II. <Emphasis Type="Italic">in vitro</Emphasis> and <Emphasis Type="Italic">in vivo</Emphasis> degradation

Porous-conductive chitosan scaffolds were fabricated by blending conductive polypyrrole (PPy) particles with chitosan solution and employing an improved phase separation method. In vitro and in vivo degradation behaviors of these scaffolds were investigated. In the case of in vitro degradation, an enzymatic degradation system was employed and lysozyme was used as a working enzyme. Meanwhile, the degradation products of scaffolds, glucosamine and N-acetyl-glucosamine, were also analyzed with a HPLC method. In vivo degradation of scaffolds was performed by subcutaneously implanting these scaffolds in rat for prescheduled time intervals. In the both cases, the weight-loss of scaffolds was monitored during the whole degradation process for evaluating the degradation of scaffolds. The changes in conductivity of scaffolds afterin vitro or in vivo degradation were also measured using a four-point technique. It was observed that the pore parameters of scaffolds themselves could significantly influence the degradation behaviors of scaffolds but the PPy content in the scaffolds seemed not to impart its effect to the degradation of scaffolds. Degradation dynamics of scaffolds and conductivity measurements indicated that these scaffolds shown fairly different behaviors in their in vitro and in vivo degradation process. According to the results obtained from in vitro and in vivo degradation of scaffolds and based on some requirements of practical tissue engineering application, it was suggested that the PPy content in the scaffold should be slightly higher than 3 wt.% but lower than 6 wt.%.     

The Activation Energy <Emphasis Type="Italic">U</Emphasis>(<Emphasis Type="Italic">T</Emphasis>,<Emphasis Type="Italic">H</Emphasis>) in Y-based Superconductors

Based on the Arrhenius equation, a method to calculate the activation energy from the resistance transition is proposed for high temperature superconductors. This method is applied to the Y-based superconductors. The activation energy is found to be U(T,H)∼(1−T/T _c )^4.8(H/H ₀)^−3.8 of YBCO crystal, and U(T,H)∼(1−T/T _c )^3.3(H/H ₀)^−2.2 of Er doped MTG YBCO crystal, respectively. With the obtained activation energy U(T,H), the lower part of the experimental curve ρ(T,H) and its derivative can be reproduced.      

Effect of ceramic filler content on the mechanical and thermal behaviour of poly-<Emphasis Type="SmallCaps">l</Emphasis>-lactic acid and poly-<Emphasis Type="SmallCaps">l</Emphasis>-lactic-co-glycolic acid composites for medical applications

One main application of resorbable poly-l-lactic acid (PLLA) and poly-l-lactic-co-glycolic acid (PLGA) based materials is in medical implants. In this study composites were made from PLLA and PLGA with hydroxyapatite (HAp) respective β-tricalcium phosphate (β-TCP) fillers. The filler content and particle size were varied, and the thermal properties as well as the mechanical strength of the composites were investigated. The composites were made by an extrusion compounding process giving 2–2.5 mm diameter sized profiles. The results verified that the thermal stability of the composites was reasonable during the optimized compounding conditions. Scanning electron microscopy revealed that the fillers were well dispersed in the polymer matrices. The mechanical properties were improved by the addition of the fillers. The optimum mechanical properties for the extruded profiles were obtained with the HAp fillers. The E-modulus was increased from 3.3 to 4.6 GPa by addition of filler particles (30 wt%) whereas the flexural strength was reduced from 133 to 106 MPa.     

Preparation of poly(3-hydroxybutyrate)/nano-hydroxyapatite composite scaffolds for bone tissue engineering

Poly(3-hydroxybutyrate)/nano-hydroxyapatite (PHB/nHA) composite scaffolds were fabricated via powder mixing, compression moulding, and particle leaching technique. The scaffolds had high porosity with interconnected porous architecture, a favorable structure for cell attachment and new bone tissue ingrowth. A homogeneous dispersion and a uniform distribution of HA nanoparticles in the polymer matrix were obtained. The scaffolds exhibited improved compressive modulus and compressive strength, which were all in the range of compressive modulus and compressive strength of cancellous bone. In addition, the use of toxic organic solvents was eliminated. Thus, the fabricated PHB/nHA composite scaffolds tend to be promising for application in bone tissue engineering.     

Preparation and <Emphasis Type="Italic">in vitro</Emphasis> investigation of chitosan/nano-hydroxyapatite composite used as bone substitute materials

Chitosan/nano-hydroxyapatite composites with different weight ratios were prepared through a co-precipitation method using Ca(OH)₂, H₃PO₄ and chitosan as starting materials. The properties of these composites were characterized by means of TEM, IR, XRD, burn-out test and universal matertial test machine. Additionally, in vitro tests were also conducted to investigate the biodegradability and bioactivity of the composite. The results showed that the HA synthesized here was poorly crystalline carbonated nanometer crystals and dispersed uniformly in chitosan phase and there is no phase-separation between the two phases. Because of the interactions between chitosan and n-HA, the mechanical properties of these composites were improved, and the maximum value of the compressive strength was measured about 120 MPa corresponding to the chitosan/n-HA composite with a weight ratio of 30/70. The specimens made of 30/70 chitosan/n-HA composite exhibit high biodegradability and bioactivity when being immersed in SBF solutions. The composite is appropriate to being used as scaffold materials for bone tissue engineering. © Springer Science + Business Media, Inc.     

In vitro degradation and bioactivity of composite poly-<Emphasis Type="SmallCaps">l</Emphasis>-lactic (PLLA)/bioactive glass (BG) scaffolds: comparison of 45S5 and 1393BG compositions

The objective of this study was to compare the effect of two bioglass (BG) compositions 45S5 and 1393 in poly-l-lactic composite scaffolds in terms of morphology, mechanical properties, biodegradation, water uptake and bioactivity. The scaffolds were produced via thermally induced phase separation starting from a ternary polymer solution (polymer/solvent/non-solvent). Furthermore, different BG to polymer ratios have been selected (1, 2.5, 5% wt/wt) to evaluate the effect of the amount of filler on the composite structure. Results show that the addition of 1393BG does not affect the scaffold morphology, whereas the 45S5BG at the highest amount tends to appreciably modify the scaffold architecture interacting with the phase separation process. Bioactivity tests confirmed the formation of a hydroxycarbonateapatite-layer in both types of BGs (detected via scanning electron microscopy, X-ray diffractometry and Fourier Transform Infrared Spectroscopy). Overall, the results showed that 1393BG composition affects the experimental preparation protocol to a minimal extent thus allowing a better control of the scaffold’s morphology compared to 45S5BG.