Characterization of Poly(ε-caprolactone)/Polyfumarate Blends as Scaffolds for Bone Tissue Engineering

There is considerable interest in the design of polymeric biomaterials that can be used for the repair of bone defects. In this study, we used ultrasound to prepare a compatibilized blend of poly(ε-caprolactone) (PCL) and poly(diisopropyl fumarate) (PDIPF). The formation of post-sonication inter-polymer coupling products was verified by SEC analysis of a blend with azo-labeled PDIPF. We also analyzed the physicochemical and mechanical properties of the compatibilized blend. When compared to PCL alone, the PCL/PDIPF blend showed no difference in its resistance as evaluated by the elastic modulus, although it did show a 50% decrease in ultimate tensile stress (P < 0.05) and an 84% decrease in elongation-at-break (P < 0.05). However, the mechanical properties of this blend were comparable to those of trabecular bone. We next evaluated biocompatibility of the PCL/PDIPF blend, and of homo-polymeric PCL and PDIPF films for comparison, with UMR106 and MC3T3E1 osteoblastic cells. Osteoblasts plated on the compatibilized blend adhered and proliferated more than on either homo-polymer, showed a greater number of cellular processes with a better organized actin cytoskeleton and expressed more type-I collagen and mineral, both markers of osteoblast phenotype. These results support the hypothesis that this new compatibilized blend could be useful in future applications for bone regeneration.     

Design of peptides for thin films,coatings and microcapsules for applications in biotechnology

A highly-interdisciplinary approach has been developed for minimizing the immunogenicity of films, coatings, microcapsules and other nano-structured materials fabricated from designed polypeptide chains. It is to base the amino-acid sequences on solvent-exposed regions in the folded states of proteins from the same organism. Each such region that meets defined criteria with respect to charge is called a sequence motif. The approach becomes more specifically tailored for intravenous applications by requiring an employed sequence motif to correspond to a known blood protein. An algorithm has been developed to identify sequence motifs in protein-encoding regions of a genome. This work is focused on sequence motifs of charge per unit length >0.5 at neutral pH. It has been found that the number of unique sequence motifs meeting this criterion in available human genome data is maximal for motifs of approx. 7 residues in length. We have designed polypeptides on the basis of computational analysis and shown that they can be used to fabricate nano-structured thin films by electrostatic layer-by-layer assembly (ELBL). The results of this work are discussed with a view to possible applications in biotechnology, notably development of biocompatible coatings and microcapsules.     

Modification of porous aminopropyl-silicate microcapsule membrane by electrically-bonded external anionic polymers

Biocompatibility and permeability of a microcapsule membrane governs the function of a microcapsule-shaped bioartificial pancreas. We have previously developed an alginate/sol-gel-synthesized aminopropyl-silicate/alginate microcapsule (Alg/AS/Alg), which had insufficient biocompatibility. The purpose of this study was to investigate whether the biocompatibility could improve by modifying the external surface with other anionic polymers and to investigate an influence of the modification on the permeability of the membrane. Four kinds of anionic polymers, poly(oxyethylene)diglycolic acid (3 kDa), heparin (15 kDa), Alg (54 kDa) and carboxymethylcellulose (CMC, 60 kDa) were used as the external anionic polymers. The heparin-bonded gel bead had the largest resistance to the diffusion of small molecules. The molecular mass cut-off point of 150 kDa required for immunoisolation was maintained for all anionic polymers. Cellular overgrowth to the implanted islet-enclosing microcapsule, a sign of insufficient biocompatibility, was suppressed by altering the external surface material from Alg to CMC. These results suggest that the biocompatibility of the Alg/AS/anionic polymer membrane can be improved by using a biocompatible anionic polymer. At the same time, it is suggested the influence on the permeability has to be investigated to develop an optimal microcapsule for bioartificial pancreas.     

In vitro toxicity of spiroorthocarbonate monomers designed for non-shrinking dental restoratives

In development of photopolymerized expanding monomers with epoxy resin systems, there is a need for reactive expanding monomers that exert a good biocompatibility profile. The objective of this study was to evaluate the in vitro toxicology of new spiroorthocarbonates designed to be expanding monomers. The expanding monomers investigated were: trans/trans-2,3,8,9- di(tetramethylene)-1,5,7,11-tetraoxaspiro[5,5]undecane (DTM-TOSU), 5,5-diethyl-19-oxadispiro[1,3-dioxane-2,2′-1,3-dioxane-5′,4-bi (DECHE-TOSU); 3,9-diethyl-3,9-dipropionyloxy methyl-1,5,7,11-tetraoxaspiro[5.5]undecane (DEDPM-TOSU); and 3,9-diethyl-3,9- diacetoxy methyl-1,5,7,11-tetraoxaspiro[5.5]undecane (DAMDE-TOSU). The in vitro toxicology of these monomers measured their cytotoxicity and mutagenicity potential. Succinic dehydrogenase (SDH) activity in the MTT assay was used to assess the toxic dose that kills 50% of cells (TC₅₀) for all the monomers. Their mutagenic potential was measured in the Ames Salmonella assay with and without metabolic activation. Two solvents, DMSO and acetone, were used to validate effects. Appropriate controls included the solvents alone. All the expanding monomers in this study were less cytotoxic than BISGMA (p < 0.01), a commercial component of dental restoratives. The relative cytotoxicity of the expanding monomers in DMSO was defined in the following order: DTM-TOSU (more toxic) > DECHE-TOSU > DEDPM-TOSU > DAMDE-TOSU. Each was significantly different from the other (p < 0.05). Overall, the TC₅₀ values of all expanding monomers were significantly greater in DMSO than in acetone (p < 0.05). However, for BISGMA this trend was opposite. For mutagenicity results, the expanding monomers were non-mutagenic and there was no solvent effect on this outcome. The non-mutagenicity and low cytotoxicity profile of these expanding monomers suggests their potential for development of biocompatible non-shrinking composites.     

Polyamide 6 composite membranes: Properties and in vitro biocompatibility evaluation

The aim of the present study was to develop polyamide 6 membrane blended with gelatin and chondroitin sulfate using the phase precipitation method and evaluate its in vitro biocompatibility. Morphology of membranes was studied by laser scanning confocal microscopy which allowed the nondestructive visualization of internal bulk morphology of membranes. Membranes exhibited porous morphology with pores spanning across the membrane width with interconnections at various depths. Membranes showed adequate mechanical properties with tensile strengths of 20.10 ± 0.64 MPa, % strain of 3.01±0.07, and modulus of 1082.50±23.50 MPa. In vitro biocompatibility of membranes by direct contact test did not show degenerative effects on NIH3T3 cells and also its leach-out products (LOP), as determined by tetrazolium (MTT) and neutral red uptake (NRU) assay. Mouse peritoneal macrophage cultured in contact with membranes and PTFE control showed comparable expression of activation markers such as CD11b/CD18, CD45, CD14, and CD86 suggesting the membranes' non-activating nature. Membrane LOP did not induce excessive proliferation of mouse splenocytes suggesting its non-antigenic nature. Preliminary blood compatibility of membranes was observed with no detectable hemolysis in static incubation assay. Taken collectively, the present data demonstrate that polyamide 6 composite membranes are biocompatible and prospective candidates for tissue engineering applications.     

Tissue responses to thermally-responsive hydrogel nanoparticles

—Thermally-responsive hydrogel nanoparticles of poly(N-isopropylacrylamide) (PNIPAM) and hydroxypropyl cellulose (HPC) have been synthesized. The particle size has been correlated to surfactant concentration and polymer concentration using dynamic laser light scattering techniques. The tissue compatibility of these hydrogel nanoparticles has been evaluated by comparing with poly-L-lactic acid and polystyrene nanoparticles using a mouse implantation model. Our results suggested that both PNIPAM and HPC nanoparticles triggered lesser inflammatory and fibrotic responses among all nanoparticles tested. It is likely that these hydrogel nanoparticles may be suitable for tissue augmentation or drug-delivery devices.     

Biocompatibility of Electrospun Halloysite Nanotube-Doped Poly(Lactic-co-Glycolic Acid) Composite Nanofibers

Organic/inorganic hybrid nanofiber systems have generated great interest in the area of tissue engineering and drug delivery. In this study, halloysite nanotube (HNT)-doped poly(lactic-co-glycolic acid) (PLGA) composite nanofibers were fabricated via electrospinning and the influence of the incorporation of HNTs within PLGA nanofibers on their in vitro biocompatibility was investigated. The morphology, mechanical and thermal properties of the composite nanofibers were characterized by scanning electron microscopy (SEM), tensile test, differential scanning calorimetry and thermogravimetric analysis. The adhesion and proliferation of mouse fibroblast cells cultured on both PLGA and HNT-doped PLGA fibrous scaffolds were compared through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay of cell viability and SEM observation of cell morphology. We show that the morphology of the PLGA nanofibers does not appreciably change with the incorporation of HNTs, except that the mean diameter of the fibers increased with the increase of HNT incorporation in the composite. More importantly, the mechanical properties of the nanofibers were greatly improved. Similar to electrospun PLGA nanofibers, HNT-doped PLGA nanofibers were able to promote cell attachment and proliferation, suggesting that the incorporation of HNTs within PLGA nanofibers does not compromise the biocompatibility of the PLGA nanofibers. In addition, we show that HNT-doped PLGA scaffolds allow more protein adsorption than those without HNTs, which may provide sufficient nutrition for cell growth and proliferation. The developed electrospun HNT-doped composite fibrous scaffold may find applications in tissue engineering and pharmaceutical sciences.     

In Vitro Biocompatibility of New PVA-Based Hydrogels as Vitreous Body Substitutes

In order to synthesize injectable hydrogels suitable as vitreous body substitutes, a new method based on the use of trisodium trimetaphosphate (STMP) to cross-link PVA was recently proposed. Hydrogels with different molar ratios between STMP and PVA were realised. The aim of the present study was the evaluation of the biocompatibility of the different STMP/PVA hydrogels synthesised by analysing the effects of their in vitro interaction with cultures of mouse fibroblasts NIH3T3, primary human microvascular endothelial cells adult (HMVECad) and human lens cells. Cytotoxicity of hydrogels was first evaluated by analysing cell density and proliferation. Morphological and morphometric analysis of cell in contact with hydrogels was then performed using light microscopy and scanning electron microscopy, respectively. Moreover, cell adhesion and growth onto the hydrogels surface was evaluated and correlated to the amount of adsorbed proteins. At last, the biocompatibility of the sheared STMP/PVA 1:8 hydrogel was tested. The in vitro data of all the STMP/PVA hydrogels demonstrated their good biocompatibility, and indicated that the 1:8 sample was the most promising as vitreous body substitute.     

The Effects of One-Step Self-Etch Adhesives on the Induction of Oxidative Stress and Production of TGF-β1 and BMP-2 by Human Gingival Fibroblasts

The aim of this study was to evaluate and compare the effects of two self-etch adhesive materials on the induction of oxidative stress and production of transforming growth factor-β1 (TGF-β1) and bone morphogenetic protein-2 (BMP-2) by cultured human gingival fibroblasts (HGF). Inflammation-free attached gingiva was obtained from healthy donors under informed consent. Following 24- and 72-h exposure of HGF to two different elutes of the test materials, cell viability was determined using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Lipid peroxidation, a major indicator of oxidative stress, was measured by the thiobarbituric acid reactive substance (TBARS) assay. TGF-β1 and BMP-2 levels in cell-free culture media were determined by enzyme-linked immunosorbent assay (ELISA). Cell viability of the test groups was significantly lower than those of control at 24 and 72 h (P < 0.001), but showed an increase at 72 h (P < 0.001). The TBARS levels of both test groups were significantly greater than that of control (P < 0.05), and displayed similar values at 72 h (P > 0.05). For both materials, the levels of TGF-β1 and BMP-2 were significantly greater than that of control (P < 0.05). Both test groups showed increased TGF-β1 levels. These results indicate that the tested self-etch adhesives might be capable of inducing production of TGF-β1 and BMP-2 in cultured HGF, despite their cytotoxic and oxidative stress-producing potential.     

Retinal prostheses: current challenges and future outlook

Blindness from retinal diseases, including age-related macular degeneration (AMD) and retinitis pigmentosa (RP), usually causes a significant decline in quality of life for affected patients. Currently there is no cure for these conditions. However, over the last decade, several groups have been developing retinal prostheses which hopefully will provide some degree of improved visual function to these patients. Several such devices are now in clinical trials. Unfortunately, the possibility of electrode or tissue damage limits excitation schemes to those that may be employed with electrodes that have relatively low charge densities. Further, the excitation thresholds that have been required to achieve vision to date, in general, are relatively high. This may result in part from poor apposition between neurons and the stimulating electrodes and is confounded by the effects of the photoreceptor loss, which initiates other pathology in the surviving retinal tissue. The combination of these and other factors imposes a restriction on the pixel density that can be used for devices that actively deliver electrical stimulation to the retina. The resultant use of devices with relatively low pixel densities presumably will limit the degree of visual resolution that can be obtained with these devices. Further increases in pixel density, and therefore increased visual acuity, will necessitate either improved electrode-tissue biocompatibility or lower stimulation thresholds. To meet this challenge, innovations in materials and devices have been proposed. Here, we review the types of retinal prostheses investigated, the extent of their current biocompatibility and future improvements designed to surmount these limitations.