The initial stages of the hydration process have been simulated on a single-Ca(I) terminated hydroxyapatite (0001) surface in step-by-step fashion using periodic slab density functional theory (DFT). Adsorption configurations and energetic properties have been described at different H(2)O coverage. At low H(2)O coverage, oxygen prefers to form CaO bonds with surface Ca cations, but as coverage increases, H(2)O tends to loosely float on the already-formed water layer. The height of the first layer H(2)O relative to the surface is found to be 1.6A. The hydration process does not cause the decomposition of surface phosphate groups and hydroxyl channel, but does affect the energetics of subsequent Zn substitution and occupation on Ca(I) and Ca(II) sites. The Ca(II) vacancy site is found to be energetically more favorable for occupation due to less spatial constraint. This suggested mechanism of preferential occupation is different from previous attempts to explain the cation substitution site preference in bulk by ionic radius and electronegativity differences.
The cytocompatibility and adhesion of cells to biomaterials are key to their success in the clinic. Here we report a study of the toxicity, cell-adhesive properties and biocompatibility of a range of alkyl-aminated hydrogels and amphiphilic conetworks comprising 1,2-propandiol-3-methacrylate (GMA) as the hydrophilic component. Previously we had shown that addition of amines containing alkyl spacers of at 3-6 carbons or addition of oligo(butyl methacrylate) sequences to crosslinked polyGMA hydrogels could be used to produce a step change in cell adhesion. In this work we produced two series of polymer networks, based on polyGMA, which contained both of these structural features and we examined the effects that these materials had on A549 epithelial cells and human dermal fibroblasts. No toxicity was observed from either direct contact or from supernatants extracted over 48h. Each of the alkyl-aminated materials provided a good substrate for adhesion of both cell types whereas the non-alkyl-aminated materials were essentially non-cell-adhesive. Peritoneal murine macrophages were present on all of the materials and the activation of these adhered macrophages was investigated by determining the production of the pro-inflammatory cytokines, TNF-alpha, IL1beta and IL-6. All of the materials behaved similarly to a clinically acceptable control, Permacol (a decellularized collagen-based porcine derived material), and each material was much less activating than when the macrophages were in contact with lipopolysaccharide endotoxin. There were no differences in the capacity of the materials to activate TNF-alpha production by macrophages however, there was a trend towards stimulation of lower levels of IL-6 and IL-1beta by the alkyl-aminated materials.
Thiol-modified surfaces are chemically well defined and suited for surface biological model experiments and biomaterials research. 3-Mercapto-1,2-propanediol (mercaptoglycerol, MG), immobilized on gold, spontaneously binds immunoglobulins from human serum and activates the complement system. The surface-bound complement factors were detected by ellipsometry-antibody techniques. The overall complement activation was subsequently corroborated independently with enzyme immunosorbent assay (EIA) and sheep and chicken erythrocyte haemolytic complement techniques. EIA experiments indicated elevated levels of C4d, but no significant increase of factor Bb was evident in the test serum from the MG system. The haemolytic assays show that MG surfaces consume complement factors from both pathways. Ellipsometry revealed that immunoglobulin G (IgG) and complement factor 1q (C1q) are transiently antibody detectable on MG after exposure to whole serum by the use of antibody techniques. Complement factor 3 (C3), C2, C4 and properdin could be detected on the surface, but not factors H and B. The total adsorbed mass and particularly C3 antibody deposition were suppressed by using EGTA-Mg2+ serum. The results suggest that MG surfaces initially activate complement via the classical pathway. Other IgG binding surfaces also appear to behave in a similar manner.
Titanium (Ti) surface roughness affects bone formation in vivo and osteoblast attachment, proliferation and differentiation in vitro. MG63 cells exhibit decreased proliferation and increased differentiation when cultured on rough Ti surfaces (Ra > 2 microm) and response to 1,25-(OH)2D3 is enhanced, resulting in synergistic increases in TGF-beta1 and PGE2. To examine the hypothesis that surface roughness and 1,25-(OH)2D3 exert their effects on local factor production through independent, but convergent, signaling pathways, MG63 cells were cultured on tissue culture plastic or on smooth (PT, Ra = 0.60 microm) and rough (SLA, Ra = 3.97 microm; TPS, Ra = 5.21 microm) Ti disks. At confluence (5 days), cultures were treated for 24h with 10(-8) M 1alpha,25-(OH)2D3 and active and latent TGF-beta1 in the conditioned media measured by ELISA. Cell layers were digested with plasmin and released TGF-beta1 was also measured. 1,25-(OH)2D3 regulated the distribution of TGF-beta1 between the media and the matrix in a surface-dependent manner; the effect was greatest in the matrix of cells cultured on SLA and TPS. Inhibition of PKA with H8 for the last 24 h of culture increased PGE2 on SLA and TPS, but when present throughout the entire culture period H8 caused an increase in PGE2 on all surfaces. 1,25-(OH)2D3 reduced the effect of H8 on PGE2 production in cultures treated for 24 h. H8 had no effect on TGF-beta1 in the media by itself but caused a complete inhibition of the 1,25-(OH)2D3 dependent increase. Inhibition of PKC with chelerythrine increased PGE2 in a surface-dependent manner and 1,25-(OH)2D3 reduced the effect of the PKC inhibitor. Chelerythrine also increased TGF-beta1 but the effect was not surface dependent; however, 1,25-(OH)2D3 reduced the effects of chelerythrine with the greatest effects on the smooth surface. Thus, the distribution of TGF-beta1 between the media and the matrix is regulated by 1,25-(OH)2D3 in a surface-dependent manner. Surface roughness exerts its effects on TGF-beta1 production via PKC but not PKA. The effect of 1,25-(OH)2D3 on TGF-beta1 production is not via PKC. PKA is involved in the surface-dependent regulation of PGE2 but not in the regulation of PGE2 by 1,25-(OH)2D3 on rough surfaces. Regulation of PKC affects PGE2 production but it is not involved in the surface roughness-dependent response to 1,25-(OH)2D3. These results suggest two independent but interconnected pathways are involved.
Surface microroughness increases osteoblast differentiation and enhances responses of osteoblasts to 1,25-dihydroxyvitamin D3 [1alpha,25(OH)2D3]. The observations that beta1 integrin expression is increased in osteoblasts grown on Ti substrates with rough microarchitecture, and that it is regulated by 1alpha,25(OH)2D3 in a surface-dependent manner, suggest that beta1 may play a role in mediating osteoblast response. To test this hypothesis, we silenced beta1 expression in MG63 human osteoblast-like cells using small interfering RNA (siRNA) and examined the responses of the beta1-silenced osteoblasts to surface microtopography and 1alpha,25(OH)2D3. To better understand the role of beta1, MG63 cells were also treated with two different monoclonal antibodies to human beta1 to block ligand binding. beta1-silenced MG63 cells grown on a tissue culture plastic had reduced alkaline phosphatase activity and levels of osteocalcin, transforming growth factor beta1, prostaglandin E2, and osteoprotegerin in comparison with control cells. Moreover, beta1-silencing inhibited the effects of surface roughness on these parameters and partially inhibited effects of 1alpha,25(OH)2D3. Anti beta1 antibody AIIB2 had no significant effect on cell number and osteocalcin, but decreased alkaline phosphatase; MAB2253Z caused dose-dependent decreases in cell number and alkaline phosphatase and an increase in osteocalcin. Effects of 1alpha,25(OH)2D3 on cell number and alkaline phosphatase were reduced and effects on osteocalcin were increased. These findings indicate that beta1 plays a major and complex role in osteoblastic differentiation modulated by either surface microarchitecture or 1alpha,25(OH)2D3. The results also show that beta1 mediates, in part, the synergistic effects of surface roughness and 1alpha,25(OH)2D3.
The monocyte is an essential element in cellular immunity. Consequently, its role in the biocompatibility process is important. A model study of the degradation of bioactive ceramics (calcium phosphate) using human peripheral blood monocytes activated by 1,25 (OH)2 VD3 and interferon (IFN)-gamma was implemented. Activated monocytes cultivated on calcium phosphate tablets led to cellular morphological modifications and to changes in the number of nuclei (from d 2 to 14). IFN-gamma promoted adhesion, the appearance of cytoplasmic extensions and multinuclei. Cavity resorption activity on the material was observed simultaneously.
Poly-N-[(2,2-dimethyl-1,3-dioxolane)methyl]acrylamide (PDMDOMA) is a neutral synthetic water-soluble polymer. In this report, we evaluated the influence of PDMDOMA on blood hemostasis by studying the fibrin polymerization process, the three-dimensional clot structure, and the mechanical properties and fibrinolysis. PDMDOMA altered the normal fibrin polymerization by changing the rate of protofibril aggregation and resulting in a 5-fold increase in the overall turbidity. Fibrin clots formed in presence of PDMDOMA exhibited thinner fibers with less branching which resulted in a more porous and heterogeneous clot structure in scanning electron micrographs. The overall strength and rigidity of the whole blood clot also decreased up to 10-fold. When a combination of plasminogen and tissue-plasminogen activators were included in clotting reactions, fibrin clots formed in the presence of PDMDOMA exhibited highly shortened clot lysis times and was supported by the enhanced clot lysis measured by thromboelastography in whole blood. Further evidence of the altered clot structure and clot cross-linking was obtained from the significant decrease in d-dimer levels measured from degraded plasma clot. Thus, PDMDOMA may play an important role as an antithrombotic agent useful in prophylactic treatments for thrombosis by modulating fibrin clot structure to enhance fibrinolysis.
We have fabricated a robust temperature sensitive hydrogel by photoinitiated copolymerization of 2-(2-methoxyethoxy)ethyl methacrylate (MeO(2)MA), 2-vinyl-4,6-diamino-1,3,5-triazine (VDT) and crosslinker polyethylene glycol diacrylate (PEGDA). It was shown that self-hydrogen bondings of VDT moieties in the bulk considerably strengthened the mechanical properties of gels, which was dependent on the weight ratio of MeO(2)/VDT and the initial monomer concentration; the VDT motifs on the surface could efficiently bind DNA for reverse gene transfection. On this soft-wet platform, gene expression lasted 7 days and the re-treated gels could be reused for new cycle of transfection. The gene modified cells could be detached by thermo-triggered switchable surface hydrophilicity of PMEO(2)MA in hydrogel. MTT assay showed low cytotoxicity of hydrogels. The results suggested that this type of mechanically strong H-bonded and thermoresponsive hydrogels hold a great potential as an integrated functional soft-wet platform for the unharmful harvest of gene modified seed cells for tissue engineering or as implantable scaffold for gene therapy and regenerative medicine applications.
New crosslinked ionic polyurethanes and copolyurethanes were yielded by reaction of telechelic cis-1,4-oligoisoprenes, bearing a variable number of ammonium and hydroxy groups, with isocyanurate of isophorone diisocyanate (I-IPDI). Aiming for a comparative study, polyurethane elastomers based on non-ionic telechelic oligomers were also synthesized. Thermo-mechanical behavior and crosslinking density of these three families of materials were investigated by DMTA and swelling test, respectively. Surface properties were examined by static contact angle measurements and AFM imaging. The bactericidal activity of the polymers was investigated by enumerating living Pseudomonas aeruginosa on material surfaces and on water suspensions. The number of attached living bacteria was found to depend on the chemical structure of the material and on the contact time between the microorganisms and the surface. An exclusive bactericidal activity was obtained with the ionic copolyurethane family. Materials with weak crosslinking density were found to release bactericidal moieties. The abilities of the polymers to prevent bacterial growth were examined through zone of inhibition experiments against P. aeruginosa, which shown a bacteriostatical effect for each synthesized material. These experiments were not sufficiently sensitive to detect the leaching of bactericidal moieties from the materials with weak crosslinking density. When the zone of inhibition experiments was performed on more sensitive bacteria, namely Staphylococcus epidermidis, the leaching of bactericidal moieties as well as bacteriostatic effect was detected. This work demonstrates the potentiality for making functional biomaterials from natural rubber, a renewable resource.
Polyurethanes based on poly(epsilon-caprolactone) (PCL) (750-2800 g/mol) and 1,4-butane diisocyanate (BDI) with different soft segment lengths and constant uniform hard segment length were synthesized in absence of catalysts for the production of a degradable meniscus scaffold. First the polyesterdiols were endcapped with BDI yielding a macrodiisocyanate with a minimal amount of side reactions and a functionality of 2.0. Subsequently, the macrodiisocyanates were extended with 1,4-butanediol in order to obtain the corresponding polyurethane. The polyurethanes had molecular weights between 78 and 160 kg/mol. Above molar masses of 1900 g/mol of the polyesterdiol crystalline PCL was found while the hard segment showed an increase in melting point from 78 to 122 degrees C with increasing hard segment content. It was estimated that the percentage crystallinity of the hard segment varied between 92 and 26%. The Young's modulus varied between 30 and 264 MPa, the strain at break varied between 870 and 1200% and tear strengths varied between 97 and 237 kJ/m2.
N,N'-dimethyl,-N,N'-di(methacryloxy ethyl)-1,6-hexanediamine (NDMH) was synthesized for the purpose of replacing both triethylene glycol dimethacrylate (TEGDMA) and the non-polymerizable amine which is added as a coinitiator in dental resin mixtures, 2,2-bis[4(2-hydroxy-3-methacryloxypropoxy)phenyl] propane (bis-GMA), camphorquinone (CQ) and ethyl-4-dimethylaminobenzoate (EDAB) were used as monomer, photoinitiator and coinitiator, respectively, in these model dental resin systems. Mixtures of bis-GMA/TEGDMA/CQ/EDAB and bis-GMA/TEGDMA/CQ/NDMH were found to reach final conversions of about 45%, slightly higher than his-GMA/NDMH/CQ (40%) under comparable visible light irradiation conditions. In addition, samples cured to these conversions were tested with dynamic mechanical analysis. The bis-GMA/TEGDMA/CQ/EDAB, his-GMA/TEGDMA/CQ/NDMH and bis-GMA/NDMH/CQ mixtures were found to have approximately the same glass transition temperature and modulus. Finally, the water sorption and solubility of bis-GMA/NDMH/CQ were higher than those of the bis-GMA/TEGDMA/CQ/EDAB, and bis-GMA/TEGDMA/CQ/NDMH. However, the values were still within the range of the ISO 9000's standards. These results suggest that NDMH is a viable alternative to conventional photocuring dental resins, serving both as a diluent and coinitiator, since there are no large differences in physical and mechanical properties when using NDMH to replace the amine coinitiator and TEGDMA diluent. The key advantage to this system is that the dimethacrylate NDMH can copolymerize with bis-GMA and TEGDMA, limiting the amount of extractable amine.
The purpose of this study was: (a) to examine the effect of plasma-gas composition on plasma polymer oxygen/carbon (O/C) ratio, functional group composition and stability in water, and then (b) to examine cell attachment to surfaces containing different concentrations of O/C and functional groups. Oxygen-functionalised surfaces were deposited by means of the plasma copolymerisation of acrylic acid/1,7-octadiene. The use of a diluent hydrocarbon allowed the deposition of surfaces with a range of O/C concentrations. Plasma copolymer surfaces were characterised by X-ray photoelectron spectroscopy (XPS). Changes in functional group composition with % acrylic acid monomer and the non-dispersive and dispersive parts of the surface energy of these plasma copolymers were measured. The solubility of the plasma copolymers was assessed by means of XPS. The degree of attachment of ROS 17/2.8 osteoblast-like cells to plasma copolymer surfaces deemed to be 'stable' in aqueous medium was measured. Tissue culture polystyrene (TCPS) was included as a control. Attachment was found to be greatest to the plasma copolymer surface with an O/C of 0.11. This surface had a carboxylic acid concentration of ca. 3%. Attachment did not correlate with increased surface wettability (i.e. the non-dispersive component of the surface energy).
Bladder regeneration studies have yielded inconclusive results possibly due to the use of unfavorable cells and primitive scaffold design. We hypothesized that human mesenchymal stem cells seeded onto poly(1,8-octanediol-co-citrate) elastomeric thin films would provide a suitable milieu for partial bladder regeneration. POCfs were created by reacting citric acid with 1,8-octanediol and seeded on opposing faces with human MSCs and urothelial cells; normal bladder smooth muscle cells and UCs, or unseeded POCfs. Partial cystectomized nude rats were augmented with the aforementioned POCfs, enveloped with omentum and sacrificed at 4 and 10 weeks. Isolated bladders were subjected to Trichrome and anti-human gamma-tubulin, calponin, caldesmon, smooth muscle gamma-actin, and elastin stainings. Mechanical testing of POCfs revealed a Young's modulus of 138 kPa with elongation 137% its initial length without permanent deformation demonstrating its high uniaxial elastic potential. Trichrome and immunofluorescent staining of MSC/UC POCf augmented bladders exhibited typical bladder architecture with muscle bundle formation and the expression and retention of bladder smooth muscle contractile proteins of human derivation. Quantitative morphometry of MSC/UC samples revealed muscle/collagen ratios approximately 1.75x greater than SMC/UC controls at 10 weeks. Data demonstrate MSC seeded POCfs support partial regeneration of bladder tissue in vivo.
The osteoblastic cell-line hFOB 1.19 with the potential to proliferate and differentiate revealed that cellular differentiation is not affected by material and roughness on newly developed zirconia implant materials. Materials under investigation were surfaces machined titanium (Ti-m), modified titanium (TiUnite, machined zirconia (TZP-A-m), modified zirconia (ZiUnitemachined alumina-toughened zirconia (ATZ-m) and modified alumina-toughened zirconia (ATZ-mod). After surface description by scanning electron microscopy (SEM) and atomic force microscopy (AFM), cellular proliferation (EZ4U, Casy1) and differentiation were examined after days 1, 3, 7, 14, 21, and 28. Osteogenic differentiation was visualized by alkaline phosphatase staining, mineralization assay (alizarin red) and by expression analysis (RT-PCR) of bone- and extracellular matrix-related genes. Proliferation on rough surfaces was reduced on both titanium and zirconia. Cell-attachment and cytoskeleton organization documented by confocal laser scanning microscopy (CLSM) elucidated attenuated cell attachment within the first 4h to be the reason for impaired proliferation. A specific up-regulation of m-RNAs in an early event (RUNX2, NELL-1, RUNX3, and BMP7) and a late event (Integrin B3) could be observed on TiUnite and ZiUnite. For titanium an up-regulation of IBSP and Integrin B1 could be described at day 21. In total, differentiation was neither affected by material nor by roughness.
Time-dependent phenotypic response of a model osteoblast cell line (hFOB 1.19, ATCC, and CRL-11372) to substrata with varying surface chemistry and topography is reviewed within the context of extant cell-adhesion theory. Cell-attachment and proliferation kinetics are compared using morphology as a leading indicator of cell phenotype. Expression of (α2, α3, α4, α5, αv, β1, and β3) integrins, vinculin, as well as secretion of osteopontin (OP) and type I collagen (Col I) supplement this visual assessment of hFOB growth. It is concluded that significant cell-adhesion events—contact, attachment, spreading, and proliferation—are similar on all surfaces, independent of substratum surface chemistry/energy. However, this sequence of events is significantly delayed and attenuated on hydrophobic (poorly water-wettable) surfaces exhibiting characteristically low-attachment efficiency and long induction periods before cells engage in an exponential-growth phase. Results suggest that a ‘time–cell–substratum–compatibility–superposition principle’ is at work wherein similar bioadhesive outcomes can be ultimately achieved on all surface types with varying hydrophilicity, but the time required to arrive at this outcome increases with decreasing cell–substratum–compatibility. Genomic and proteomic tools offer unprecedented opportunity to directly measure changes in the cellular machinery that lead to observed cell responses to different materials. But for the purpose of measuring structure–property relationships that can guide biomaterial development, genomic/proteomic tools should be applied early in the adhesion/spreading process before cells have an opportunity to significantly remodel the cell–substratum interface, effectively erasing cause and effect relationships between cell–substratum–compatibility and substratum properties.
Powdered samples (170-230 mesh) of a glass of composition 1.25CaO.SiO2 were soaked in a simulated body fluid (SBF). The powders were submitted to Fourier transform infrared transmission spectroscopy as coarse powders (such as drawn out from the SBF) and as fine powders (soaked and subsequently ground). Soaked samples were submitted to differential thermal analysis (DTA) and the crystalline phases formed during heating in the DTA apparatus were identified by means of X-ray diffraction analysis. The method appears to be useful in studying the mechanism of deposition of the hydroxyapatite layer. It is documented, by using the same method, that the mechanism involves the reactions of hydrolysis and successive condensation and repolymerization of the silicate substrate. These reactions are very fast. Extensive Ca2+ cation depletion occurs, but appears to be slower.
Hoechst RCH 1000 and Hercules Hi-Fax 1900 ultra-high molecular weight (UHMW) polyethylenes have been compared, for the first time, in tests, including wear, fatigue and creep, relevant to artificial human joints. In none of the tests was the behaviour of the Hercules material inferior to that of RCH 1000, and in the wear and creep tests it was superior.
We synthesized vitamin E TPGS (d-α-Tocopheryl-co-poly(ethylene glycol) 1000 succinate) micelles for superparamagnetic iron oxides formulation for nanothermotherapy and magnetic resonance imaging (MRI), which showed better thermal and magnetic properties, and in vitro cellular uptake and lower cytotoxicity as well as better in vivo therapeutic and imaging effects in comparison with the commercial Resovist and the Pluronic F127 micelles reported in the recent literature. The superparamagnetic iron oxides originally coated with oleic acid and oleylamine were formulated in the core of the TPGS micelles using a simple solvent-exchange method. The IOs-loaded TPGS showed greatest colloidal stability due to the critical micelle concentration (CMC) of vitamin E TPGS. Highly monodisperse and water soluble suspension was obtained which were stable in 0.9% normal saline for a period of 12 days. The micelles were characterized for their size and size distribution. Their morphology was examined through transmission electron microscopy (TEM). The enhanced thermal and superparamagnetic properties of the IOs-loaded TPGS micelles were assessed. Cellular uptake and cytotoxicity were investigated in vitro with MCF-7 cancer cells. Relaxivity study showed that the IOs-loaded TPGS micelles can have better effects for T2-weighted imaging using MRI. T2 mapped images of xenograft grown on SCID mice showed that the TPGS micelle formulation of IOs had ∼1.7 times and ∼1.05 times T2 decrease at the tumor site compared to Resovist and the F127 micelle formulation, respectively.
To develop a polymer-anticancer drug conjugate, D-alpha-tocopheryl polyethylene glycol 1000 succinate (TPGS) was employed as a carrier of doxorubicin (DOX) to enhance its therapeutic effects and reduce its side effects. Doxorubicin was chemically conjugated to TPGS. The molecular structure, drug loading efficiency, drug release kinetics and stability of the conjugate were characterized. The cellular uptake, intracellular distribution, and cytotoxicity were accessed by using MCF-7 breast cancer cells and C6 glioma cells as in vitro cell model. The conjugate showed higher cellular uptake efficiency and broader distribution within the cells. Judged by IC(50), the conjugate was found 31.8, 69.6, 84.1% more effective with MCF-7 cells and 43.9, 87.7, 42.2% more effective with C6 cells than the parent drug after 24, 48, 72 h culture, respectively. The in vivo pharmacokinetics and biodistribution were investigated after an i.v. administration at 5 mg DOX/kg body weight in rats. Promisingly, 4.5-fold increase in the half-life and 24-fold increase in the area-under-the-curve (AUC) of DOX were achieved for the TPGS-DOX conjugate compared with the free DOX. The drug level in heart, gastric and intestine was significantly reduced, which is an indication of reduced side effects. Our TPGS-DOX conjugate showed great potential to be a prodrug of higher therapeutic effects and fewer side effects than DOX itself.
We developed a strategy to formulate supraparamagnetic iron oxides (SPIOs) in nanoparticles (NPs) of biodegradable copolymer made up of poly(lactic acid) (PLA) and d-alpha-tocopherol polyethylene glycol 1000 succinate (TPGS) for medical imaging by magnetic resonance imaging (MRI) of high contrast and low side effects. The IOs-loaded PLA-TPGS NPs (IOs-PNPs) were prepared by the single emulsion method and the nanoprecipitation method. Effects of the process parameters such as the emulsifier concentration, IOs loading in the nanoparticles, and the solvent to non-solvent ratio on the IOs distribution within the polymeric matrix were investigated and the formulation was then optimized. The transmission electron microscopy (TEM) showed direct visual evidence for the well dispersed distribution of the IOs within the NPs. We further investigated the biocompatibility and cellular uptake of the IOs-PNPs in vitro with MCF-7 breast cancer cells and NIH-3T3 mouse fibroblast in close comparison with the commercial IOs imaging agent Resovist. MRI imaging was further carried out to investigate the biodistribution of the IOs formulated in the IOs-PNPs, especially in the liver to understand the liver clearance process, which was also made in close comparison with Resovist. We found that the PLA-TPGS NPs formulation at the clinically approved dose of 0.8 mg Fe/kg could be cleared within 24 h in comparison with several weeks for Resovist. Xenograft tumor model MRI confirmed the advantages of the IOs-PNPs formulation versus Resovist through the enhanced permeation and retention (EPR) effect of the tumor vasculature.
This work developed a multimodal imaging system by co-encapsulating superparamagnetic iron oxides (IOs) and quantum dots (QDs) in the nanoparticles of poly (lactic acid) - d-α-tocopheryl polyethylene glycol 1000 succinate (PLA-TPGS) for concurrent imaging of the magnetic resonance imaging (MRI) and the fluorescence imaging to combine their advantages and to overcome their disadvantages as well as to promote a sustained and controlled imaging with passive targeting effects to the diseased cells. The QDs and IOs-loaded PLA-TPGS NPs were prepared by a modified nanoprecipitation method, which were then characterized for their size and size distribution, zeta potential and the imaging agent encapsulation efficiency. The transmission electron microscopy (TEM) images showed direct evidence for the well-dispersed distribution of the QDs and IOs within the PLA-TPGS NPs. The cellular uptake and the cytotoxicity of the PLA-TPGS NPs formulation of QDs and IOs were investigated in vitro with MCF-7 breast cancer cells, which were conducted in close comparison with the free QDs and IOs at the same agent dose. The Xenograft model was also conducted for biodistribution of the QDs and IOs-loaded PLA-TPGS NPs among the various organs, which showed greatly enhanced tumor imaging due to the passively targeting effects of the NPs to the tumor. Images of tumors were acquired in vivo by a 7T MRI scanner. Further ex vivo images of the tumors were obtained by confocal laser scanning microscopy. Such a multimodal imaging system shows great advantages of both contrast agents making the resultant probe highly sensitive with good depth penetration, which confirms the diagnosis obtained from each individual imaging. With therapeutics co-encapsulation and ligand conjugation, such nanoparticles system can realize a multi-functional system for medical diagnosis and treatment.
We recently, developed a simple one-day one-step incubation method to obtain bone-like apatite coating on flexible and biodegradable Polyactive 1000PEGT70PBT30. The present study reports a preliminary biological evaluation on the coated polymer after implantation in rabbit femurs. The porous cylindrical implants were produced from a block fabricated by injection molding and salt leaching. This technique provided the block necessary mechanical integrity to make small cylinders (diameter 3.5 x 5 mm2) that were suitable for implantation in rabbits. The coating continuously covered the surface of the polymer, preserving the porous architecture of outer contour of the cylinders. Two defects with a diameter of 3.5 or 4 mm were drilled in the proximal and distal part of femur diaphysis. The implants were inserted as press-fit or undersized into the cortex as well as in the marrow cavity. The polymer swelled after implantation due to hydration, leading to a tight contact with the surrounding bone in both defects. The adherence of the coating on the polymer proved to be sufficient to endure a steam sterilization process as well as the 15% swelling of the polymer in vivo. The coated Polyactive 1000PEGT70PBT30 has a good osteoconductive property, as manifested by abundant bone growth into marrow cavity along the implant surface during 4-week implantation. A favorable bioactive effect of the coating with an intimate bone contact and extensive bone bonding with this polymer was qualitatively confirmed. Concerning the bone ingrowth into the porous implant in the defect of 4 mm diameter, only marginal bone formation was observed up to 8 weeks with a maximal penetration depth of about 1 mm. The pore interconnectivity is important not only for producing a coating inside the porous structure but also for bone ingrowth into this biodegradable material. This preliminary study provided promising evidence for a further study using a bigger animal model.
Polyactive 1000PEOT70PBT30 (a segmented block copolymer of poly(ethylene oxide terephtalate)/poly(butylene terephtalate) with 70/30 PEOT/PBT ratio) was processed into three different types of samples: injection molded to rods, hot-pressed to films and to composite membranes made by hot-pressing a tubular mesh of poly-L,D-lactide 96/4 between two films of Polyactive. The molecular weight of Polyactive was not influenced by processing, but gamma-sterilization seemed to increase the weight average molecular weight (Mw). Mechanical properties of the rods and films decreased rapidly in hydrolytic conditions due to the hydrogel nature of the polymer, swelling and degradation. Mesh reinforcement increased the mechanical properties, but the components separated during soaking. In vitro the molecular weight of the rods and films started to decrease immediately, but the PEOT (or PEO) proportion remained relatively constant for 26 weeks. Macroscopically, all the wet devices remained intact, but fragmented on drying. Microscopically, topographical formations of polymer were found on the surfaces and small sodium-rich spots were precipitated onto and inside the polymer. Thermal measurements showed that polymer consisted of amorphous PEOT segments and both amorphous and crystalline PBT segments.
The most important features of the degradation and erosion of degradable polymers in vitro are discussed. Parameters of chemical degradation, which is the scission of the polymer backbone, are described such as the type of polymer bond, pH and copolymer composition. Examples are given how these parameters can be used to control degradation rates. Degradation leads finally to polymer erosion, the loss of material from the polymer bulk. The resulting changes in morphology, pH, oligomer and monomer properties as well as crystallinity are illustrated with selected examples. Finally, a brief survey on approaches to polymer degradation and erosion is given.
In this paper, the high-temperature stabilized beta-tricalcium phosphate (betaTCP, beta-Ca3(PO4)2) were prepared by heating the deficient HAP (d-HAP, Ca10-x(HPO4)x(PO4)6-x(OH)2-x) with tetra-sodium diphosphate decahydrate (NP, Na4P2O7 x 10H2O) addition. The betaTCP, d-HAP and d-HAP doped with 2.5, 5, 7.5 and 10 wt % NP were heated to different temperatures and were investigated by X-ray diffraction analysis (XRD) and Fourier-transformed infrared spectroscopy (FTIR). The results demonstrated that the HPO4(2-) of d-HAP condensed into P2O7(4-) occurred before 650 degrees C. The P2O7(4-) ions could be traced in the FTIR spectrum when the d-HAP was heated up to 750 degrees C. The reaction of P2O7(4-) with OH- did not occur instantly but over a wide range of temperatures. The d-HAP doped with NP would decrease the decomposition temperature of d-HAP. NP doped into d-HAP not only induced the d-HAP decomposition at lower temperature but also stabilized the betaTCP crystal structure at higher-temperature. It could also increase the conversion temperature of betaTCP to alphaTCP from 1180 degrees C up to 1300 degrees C. We could successfully prepare high-temperature (up to 1300 C) stabilized ffTCP by heating NP doped d-HAP.
A newly produced bioceramic, beta-Ca2P2O7 with addition of Na4P2O7.10H2O (SDCP), has been implanted into the femoral condyle of rabbits. Within 6 weeks after implantation, most of the bioceramic is replaced by new woven bone. On the contrary, block from hydroxyapatite (HA) and beta-tricalcium phosphate (beta-TCP), which are osteoconductible, do not resorb within a short period of time. We believe that the biodegradable behaviour of SDCP may occur in two steps. The first and most important step is the digestion of particles and migration of the particles by phagocytosis. The object of this study is to examine the change in morphologies, chemical compositions and crystal structure of SDCP after soaking in distilled water for a certain period of time. The SDCP ceramic was also co-cultured with leucocytes to observe how the SDCP particles were digested by the leucocytes, so that the mechanism of biodegradable behaviour of SDCP ceramic in vivo might be clarified. Four types of sintered calcium phosphate ceramics were tested in the experiment: SDCP, pure beta-Ca2P2O7 (DCP), HA and beta-TCP. They wee soaked in distilled water at 37 degrees C for up to 30 days. The microstructure and morphology of crystals deposited on the surface were observed using scanning electron microscopy. Sodium, calcium and phosphorus ion contents in the supernatant solution were detected by atomic absorption analysis and ion coupled plasma. In summary, HA and DCP showed no significant evidence of dissolution in distilled water. In static distilled water, calcium ions may be released from beta-TCP into solution during the initial 7 days and then converted into HA by reprecipitation. The results showed that the SDCP was firstly dissolved into small grains or fragments by the solution. The small fragments should be so small as to be digested by the phagocytes in a physiological environment.
The biodistribution of lanthanide-based upconversion nanophosphors (UCNPs) has attracted increasing attention, and all of the reported UCNPs display metabolism in the liver and spleen mainly. Herein, ∼8 nm poly(ethylene glycol) (PEG)-coated NaYF4 nanoparticles codoped with Yb(3+), Er(3+), and (or) radioactive (153)Sm(3+) ions were synthesized, through a hydrothermal synthetic system assisted by binary cooperative ligands with oleic acid and PEG dicarboxylic acids. The as-prepared PEG-coating NaYF4:Yb,Er and NaYF4:Yb,Er,(153)Sm are denoted as PEG-UCNPs and PEG-UCNPs((153)Sm), respectively. PEG-UCNPs were characterized by transmission electron microscope (TEM), X-ray diffraction (XRD) analysis, and Fourier-transform infrared (FTIR) spectroscopy. The PEG-UCNPs showed excellent water solubility with a hydrodynamic diameter of ∼10 nm and displayed upconversion luminescence (UCL) under continuous-wave excitation at 980 nm. At the same time, the (153)Sm-doped nanoparticles PEG-UCNPs((153)Sm) displayed radioactivity, and time-dependent biodistribution of PEG-UCNPs((153)Sm) was investigated, through single-photon emission computed tomography (SPECT) imaging and γ-counter analysis. Interestingly, PEG-UCNPs((153)Sm) had a long blood retention time and were partly eliminated through urinary pathways in vivo. Therefore, the concept of fabricating PEG-coated, small nanosize (sub-10 nm) nanoparticles with radioactive property is a useful strategy for providing a potential method to monitor lanthanide nanoparticles renal clearable.
The mechanism and kinetics of hydroxyapatite (HAP) precipitation from aqueous solution at the conditions of pH 10-11 and at calcium ion concentrations of more than 0.5 mol/l were investigated. The results show that the reaction undergoes the following process: transferring from octacalcium phosphate (OCP) to amorphous calcium phosphate (ACP) rapidly, and then from ACP to calcium-deficient hydroxyapatite (DAP) and HAP. DAP was the non-stoichiometric material that ACP converts to HAP. Reaction temperature greatly affects the reaction rate of the conversion from ACP to HAP. It takes about 24 h to form pure-phase HAP at 25 degrees C while it takes only 5 min at 60 degrees C. The temperature also has a great deal of influence on the particle size and morphology of precipitated hydroxyapatite. The reaction of transformation from ACP to DAP was second order and the activation energy was 95 kJ/mol (22.7 kCal/mol) and therefore it was inferred that the conversion reaction was a surface control process.
Bone induction is a multistep process that includes chemotaxis and attachment of mesenchymal stem cells, proliferation of progenitor cells and differentiation into cartilage and bone. Endochondral bone formation is the predominant sequence. Bone induction can be operationally divided into the following phases: initiation, promotion and maintenance. The initiation of bone induction is primarily regulated by osteogenin and bone morphogenetic proteins. Recent work has led to the isolation, purification and cloning of osteogenin and other bone morphogenetic proteins. Other growth factors such as platelet derived growth factor (PDGF), transforming growth factor beta (TGF-beta isoforms, insulin like growth factors (IGF 1 and 2), and fibroblast growth factor (FGF) may promote and perhaps maintain the newly induced bone.
In this paper, a new kind of aliphatic biodegradable polyesteramide copolymers P(CL/AU)x/y based on epsilon-caprolactone and 11-aminoundecanoic acid were synthesized by the melt polycondensation method. Hydrolytic degradation behavior of P(CL/AU) copolymers were studied by using FTIR, 1H-NMR and DSC. Chemical compositions, macromolecular weight, thickness of the test sample, and pH of the degradation medium have great effect on degradation rate. The degradation rate decreased with increase in aminoundecanoic acid content, macromolecular weight, and thickness of the test samples, but increased with incubation temperature and pH of the degradation medium. The degradation mechanism was studied according to the mathematical model developed by professor Göpferich.
Current prosthetic small diameter vascular grafts show poor long-term patency rates, leading to the pursuit of a biological alternative. Hyaff-11 is a hyaluronan-based biodegradable polymer developed for tissue-engineering applications. This study aimed to determine whether human vascular endothelial cells attach to Hyaff-11 scaffolds and produce a subendothelial matrix. Two forms of fibrous, non-woven Hyaff-11 scaffolds: unpressed and pressed felts, were analysed. Attachment of human venous endothelial cells was investigated after 1, 5, 10 and 20 days in culture using SEM and confocal microscopy. The deposition of subendothelial matrix components was investigated by immunofluorescent staining. We demonstrate that endothelial cells adhere to the individual fibres of both unpressed and pressed scaffolds: with a seeding density of 1 x 10(6) cells/cm(2), 94% of the cells attached to Hyaff-11 fibres after 24 h. The pressed material provided the best environment for cell growth, allowing the formation of a complete endothelial monolayer after 20 days. Furthermore, endothelial cells on Hyaff-11 pressed felts deposited an organised subendothelial matrix containing laminin, fibronectin, type IV and type VIII collagen. This work indicates Hyaff-11 based biopolymers as suitable scaffolds to promote endothelialisation within the next generation of vascular grafts.
Although cisplatin is still one of the most effective chemotherapy agents for human cancers, its clinical use is limited by serious side effects, especially nephrotoxicity. Oxidative stress is an important mediator of cisplatin-induced nephrotoxicity. In the present study, a simple method for functionalization of selenium nanoparticles by self-assembly of 11-mercapto-1-undecanol (Se@MUN) to achieve enhanced antioxidant activity and antagonis against cisplatin-induced nephrotoxicity has been demonstrated. The chemical structure of the nanoparticles was characterized by various microscopic and spectroscopic methods. The results revealed that the spherical nanoparticles were capped with MUN on the surface through formation of Se-S bond. The in vitro protective effects of Se@MUN on HK-2 proximal tubular cells against cisplatin-induced nephrotoxicity and the underlying mechanisms were also investigated. Se@MUN exhibited free radical scavenging activity and higher cellular uptake in human normal cells by comparing with SeNPs. Se@MUN significantly attenuated cisplatin-induced reduction in cell viability, appearance of Sub-G1 peak, nuclear condensation and DNA fragmentation in HK-2 cells. Activation of caspase-3 in cells exposed to cisplatin was also effectively blocked by Se@MUN. Moreover, Se@MUN significantly prevented the cisplatin-induced overproduction of intracellular ROS. Our findings suggest that Se@MUN is a promising selenium species with potential application in prevention of cisplatin-induced renal injury.
Association of biomaterials with autologous cells can provide a new generation of implantable devices for cartilage repair. Such scaffolds should provide a preformed three-dimensional shape and prevent cells from escaping into the articular cavity. Furthermore, these constructs should have sufficient mechanical strength to facilitate handling in a clinical setting and stimulate the uniform spreading of cells and their phenotype redifferentiation. The aim of this study was to verify the ability of HYAFF 11, a recently developed hyaluronic-acid-based biodegradable polymer, to support the growth of human chondrocytes and to maintain their original phenotype. This capability was assessed by the evaluation of collagen types I, II and aggrecan mRNA expression. Immunohistochemical analyses were also performed to evaluate collagen types I, II and proteoglycans synthesis. A field emission in lens scanning microscopy was utilized to verify the interactions between the cells and the biomaterial. Our data indicate that human chondrocytes seeded on HYAFF 11 express and produce collagen type II and aggrecan and downregulate the production of collagen type I. These results provide an in vitro demonstration for the therapeutic potential of HYAFF 11 as a delivery vehicle in a tissue-engineered approach towards the repair of articular cartilage defects.
Different methods have been used to improve chondrocyte transplantation for the repair of articular cartilage defects. Several groups of biomaterials have been proposed as support for in vitro cell growth and for in vivo implantation. Here. we describe a new approach investigating the healing of rabbit cartilage by means of autologous chondrocytes seeded on a hyaluronan derivative referred to as Hyaff-11. Full thickness defects were created bilaterally in the weight-bearing surface of the medial femoral condyle of both femora of New Zealand male rabbits. The wounds were then repaired using both chondrocytes seeded on the biomaterial and biomaterial alone. Controls were similarly treated but received either no treatment or implants of the delivery substance. Histologic samples from in and around the defect sites were examined 1, 3 and 6 months after surgery and were scored from 0 to 16. Statistically significant differences in the quality of the regenerated tissue were found between the grafts carried out with biomaterial carrying chondrocyte cells compared to the biomaterial alone or controls. This study demonstrates the efficacy of this hyaluronan-based scaffold for autologous chondrocytes transplantation.
Fe(3)O(4)@SiO(2) core-shell nanoparticles were synthesized and used to label human mesenchymal stem cells (hMSCs) for in vitro and in vivo magnetic resonance imaging study. The diameter of the nanoparticles is 24-30 nm with a Fe(3)O(4) core of ∼8 nm and a SiO(2) shell of ∼8 nm. Transverse relaxivity of the nanoparticles dispersed in water is measured to be ∼106 mM(-1) s(-1). After incubation with hMSCs for 12 h at a concentration of 100 μg Fe/mL, cellular uptake of Fe(3)O(4)@SiO(2) is 20-100 pg Fe/cell, which are located predominantly in the cytoplasm of cells. This level of uptake exhibits no significant influence on hMSCs' viability and differentiation. In vitro imaging of Fe(3)O(4)@SiO(2)-labeled hMSCs evenly distributed in agarose gel yields single cell sensitivity at 11.7 T. In vivo imaging of Fe(3)O(4)@SiO(2)-labeled hMSCs injected into the left brain hemisphere of nude mice yields imaging sensitivity of ∼130 hMSCs.
We have identified a number of cell-adhesive peptides from laminins, a major component of basement membranes. Cell-adhesive peptides derived from basement membrane proteins are potential candidates for incorporating cell-binding activities into scaffold materials for tissue engineering. Our goal is development of a chemically synthetic basement membrane using laminin-derived cell-adhesive peptides and polymeric materials. In this study, we used hyaluronic acid (HA) as a scaffold material and laminin-derived cell-adhesive peptides, A99 (AGTFALRGDNPQG, binds to integrin αvβ3), AG73 (RKRLQVQLSIRT, binds to syndecans), and an A99/AG73 mixture (molar ratio = 9:1) conjugated to two-dimensional (2D) HA matrices. As a result, it was found that the 2D A99/AG73-HA matrices have strong biological functions, such as cell attachment, cell spreading, and neurite outgrowth, similar to that of basement membrane extract (BME)-coated plates. Next, we developed three-dimensional (3D) peptide-HA matrices using the A99/AG73 mixture. The 3D A99/AG73-HA matrices promoted cell spreading and improved cell viability and collagen gene expression. Further, PC12 neurite extension was observed in the 3D A99/AG73-HA matrices. These biological activities of the 3D A99/AG73-HA matrices were similar to those of the 3D BME matrices. These results suggest that the peptide-HA matrices are useful as 2D and 3D matrices and can be applied for tissue engineering as a synthetic basement membrane.
The ability of biomaterial surfaces to regulate cell behavior requires control over surface chemistry and microstructure. One of the greatest challenges with silicon-based biomedical microdevices such as those recently developed for neural stimulation, implantable encapsulation, biosensors, and drug delivery, is to improve biocompatibility and tissue integration. This may be achieved by modifying the exposed silicon surface with bioactive peptides. In this study, Arg-Gly-Asp (RGD) peptide conjugated surfaces were prepared and characterized. The effect of these surfaces on fibroblast adhesion and proliferation was examined over 4 days. Silicon surfaces coupled with a synthetic RGD peptide, as characterized with X-ray photoelectron spectroscopy and atomic force microscopy, display enhanced cell proliferation and bioactivity. Results demonstrate an almost three-fold greater cell attachment! proliferation on RGD immobilized surfaces compared to unmodified (control) silicon surfaces. Modulating the biological response of inorganic materials such as silicon will allow us to design more appropriate interfaces for implantable diagnostic and therapeutic silicon-based microdevices.
Laminin-111, a multifunctional matrix protein, has diverse biological functions. Previously, we have identified various biologically active sequences in laminin-111 by a systematic peptide screening. We also demonstrated that peptide-conjugated chitosan matrices enhance the biological functions of the active sequences and are useful as a scaffold. Here, we conjugated sixty biologically active laminin-111 peptides onto chitosan matrices. The twenty-nine peptide-chitosan matrices promoted various biological activities, including cell attachment, spreading, and neurite outgrowth. The biological activities of peptide-chitosan matrices depend on the peptide. These peptide-chitosan matrices are categorized into six groups depending on their biological activities. Next, we conjugated five active peptides, which showed strong cell attachment activity in the each group, onto a single chitosan matrix to mimic the multiple activities of laminin-111. The mixed peptides-chitosan matrix significantly promoted cell attachment and cell spreading over that observed with the individual peptides. We also demonstrated that a mixed peptides-chitosan matrix, using four neurite outgrowth-promoting peptides each from a different group, enhanced the activity. These data suggest that the mixed peptides synergistically induce laminin-like biological activities on a chitosan matrix. The active peptides-chitosan matrices described here have potential for use as biomaterial for tissue engineering and regeneration.
Many studies have been dedicated to the development of scaffolds for improving post-traumatic nerve regeneration. The goal of this study was to develop and test hybrid chitosan membranes to use in peripheral nerve reconstruction, either alone or enriched with N1E-115 neural cells. Hybrid chitosan membranes were tested in vitro, to assess their ability in supporting N1E-115 cell survival and differentiation, and in vivo to assess biocompatibility as well as to evaluate their effects on nerve fiber regeneration and functional recovery after a standardized rat sciatic nerve crush injury. Functional recovery was evaluated using the sciatic functional index (SFI), the static sciatic index (SSI), the extensor postural thrust (EPT), the withdrawal reflex latency (WRL) and ankle kinematics. Nerve fiber regeneration was assessed by quantitative stereological analysis and electron microscopy. All chitosan membranes showed good biocompatibility and proved to be a suitable substrate for plating the N1E-115 cellular system. By contrast, in vivo nerve regeneration assessment after crush injury showed that the freeze-dried chitosan type III, without N1E-115 cell addition, was the only type of membrane that significantly improved posttraumatic axonal regrowth and functional recovery. It can be thus suggested that local enwrapping with this type of chitosan membrane may represent an effective approach for the improvement of the clinical outcome in patients receiving peripheral nerve surgery.
Characterization of synthetic calcium phosphates is reported and compared with previous studies. Barium hydroxyapatite was also synthesized. Shrinkage on sintering, water absorption, tensile strength, porosity and X-ray diffraction patterns were studied. Hydroxyapatite and beta-tricalcium phosphate show compositional and mechanical property variation dependent on sintering temperatures. The property change on introduction of barium removes the practical value. X-ray diffraction analysis is considered essential before clinical use due to sensitivity of composition to sintering conditions.
In chronic wounds, a number of host factors are released which perpetuate the inflammatory process, including polymorphonuclear leukocyte (PMN)-derived reactive oxygen species (ROS), such as superoxide radical (O2−) and hydroxyl radical (OH) species. The glycosaminoglycan, hyaluronan, has been shown to act as an antioxidant towards ROS, although the potential for biomaterials, such as HYAFF®-11p75 (the 75% benzyl ester of hyaluronan) and AQUACEL® (carboxymethylcellulose), to act in this manner has yet to be elucidated. This study compared the antioxidant properties of high and low molecular weight hyaluronan (HMWT HA and LMWT HA), HYAFF®-11p75, AQUACEL® and an AQUACEL®/hyaluronan composite (AQUACEL®/HA) against O2− and OH. The antioxidant capacities of each material were assessed by their ability to inhibit cytochrome C reduction by O2− fluxes, generated via the oxidation of hypoxanthine by xanthine oxidase, and their inhibition of 2-deoxy-d-ribose degradation by OH fluxes, generated by the reaction of hydrogen peroxide (H2O2) and iron (Fe2+).
This study is concerned with the development of an agent for single photon emission computer tomography (SPECT) for imaging inflammation and tumor progression. [(123)I]Iodooctyl fenbufen amide ([(123)I]IOFA) was prepared from the precursor N-octyl-4-oxo-4-(4'-(trimethylstannyl)biphenyl-4-yl)butanamide with a radiochemical yield of 15%, specific activity of 37 GBq/μmol, and radiochemical purity of 95%. Analysis of the binding of [(123)I]IOFA to COX-1 and COX-2 enzymes by using HPLC and a gel filtration column showed a selectivity ratio of 1:1.3. An assay for the competitive inhibition of substrate transfer showed that IOFA exhibited a comparable IC(50) value compared to fenbufen. In the normal rat liver, a lower level and homogeneous pattern of [(123)I]IOFA radioactivity was observed by SPECT. In contrast, in the rat liver with thioacetamide-induced cholangiocarcinoma, a higher uptake and heterogeneous pattern of [(123)I]IOFA radioactivity was seen as hot spots in tumor lesions by SPECT imaging. Importantly, elevated COX-1 and COX-2 expressions from immunostaining were found in the bile ducts of tumor rats but not of normal rats. Therefore, [(123)I]IOFA was found to exhibit the potential for imaging tumors that over-express COX.
Targeting of therapeutics or imaging agents to the endothelium has the potential to improve specificity and effectiveness of treatment for many diseases. One strategy to achieve this goal is the use of nanoparticles (NPs) targeted to the endothelium by ligands of protein determinants present on this tissue, including cell adhesion molecules, peptidases, and cell receptors. However, detachment of the radiolabel probes from NPs poses a significant problem. In this study, we devised polymeric NPs directly labeled with radioiodine isotopes including the positron emission tomography (PET) isotope (124)I, and characterized their targeting to specific endothelial determinants. This approach provided sizable, targetable probes for specific detection of endothelial surface determinants non-invasively in live animals. Direct conjugation of radiolabel to NPs allowed for stable longitudinal tracking of tissue distribution without label detachment even in an aggressive proteolytic environment. Further, this approach permits tracking of NP pharmacokinetics in real-time and non-invasive imaging of the lung in mice using micro-PET imaging. The use of this strategy will considerably improve investigation of NP interactions with target cells and PET imaging in small animals, which ultimately can aid in the optimization of targeted drug delivery.
Diffusivity of 125I-calmodulin (MW congruent to 17,000) through collagen membranes was studied as a model for the release of macromolecules from collagen matrices. The diffusion coefficient of calmodulin through collagen membranes was determined from time-lag experiments conducted in a dialysis cell at 24 degrees C. Based on time-lag experiments, the diffusion coefficient was observed to be a function of source concentration and membrane swelling ratio after denaturation. The dependence of the diffusion coefficient on source concentration was consistent with a model involving calmodulin immobilization by the collagen membrane. At high source concentrations the diffusion coefficient of calmodulin through collagen membranes was observed to vary from about 10(-8) for uncrosslinked membranes to 10(-9) cm2/s for highly crosslinked membranes. Based on theoretical calculations, the release rate from collagen matrices may be altered by a factor of three. It was concluded that the release rate of biologically active molecules from collagen matrices can be controlled by varying the extent of crosslinking and the macromolecular concentration. Further studies are necessary to characterize the release of other macromolecules from collagen matrices.
Diffusion of angiotensin II, albumin and aldolase was studied through collagen membranes with swelling ratios between 4 and 15. The diffusion coefficient was measured from the time-lag for the onset of steady-state flux through the membrane. Binding of macromolecules to collagen was evaluated from the results of sorption studies conducted as a function of macromolecular concentration. Results presented indicate that the diffusion of macromolecules through collagen membrane is slowed by electrostatic and hydrogen bonding between individual macromolecular chains and collagen. The extent of adsorption is increased as the molecular weight of the diffusant increases. Diffusion of water soluble macromolecules through collagen occurs rapidly, suggesting that diffusion occurs through water filled channels as opposed to between collagen molecules. The results of these studies are useful in understanding diffusion through connective tissues and in the design of drug delivery systems based on collagen.
Due to increasing clinical demand for adipose tissue, a suitable scaffold for engineering adipose tissue constructs is needed. In this study, we have developed a three-dimensional (3-D) culture system using bone marrow-derived mesenchymal stem cells (BM-MSC) and a Pluronic F-127 hydrogel scaffold as a step towards the in vitro tissue engineering of fat. BM-MSC were dispersed into a Pluronic F-127 hydrogel with or without type I collagen added. The adipogenic differentiation of the BM-MSC was assessed by cellular morphology and further confirmed by Oil Red O staining. The BM-MSC differentiated into adipocytes in Pluronic F-127 in the presence of adipogenic stimuli over a period of 2 weeks, with some differentiation present even in absence of such stimuli. The addition of type I collagen to the Pluronic F-127 caused the BM-MSC to aggregate into clumps, thereby generating an uneven adipogenic response, which was not desirable.