Heparin-modified dendrimer crosslinked collagen matrices for the delivery of heparin-binding epidermal growth factor.

A tissue engineered corneal equivalent (TECEs) requires host integration to achieve adequate anchorage and long-term device stability. Corneal integration through epithelialization and stromal integration can be manipulated by growth factors. We investigated the potential of heparin-binding epidermal growth factor (HB-EGF) for mediating interactions with human corneal epithelial cells (HCEC) and compared its efficacy to epidermal growth factor (EGF) in vitro. Furthermore, we utilized heparinized dendrimer crosslinked collagen gels, intended for use as TECE, for delivery of HB-EGF in a sustained manner. HCEC were exposed to HB-EGF at varying concentrations between 0.1 and 1000 ng/mL. Cell proliferation increased with growth factor concentration up to a concentration of 50 ng/mL, suggesting growth factor receptor down-regulation at higher HB-EGF concentrations. Response to HB-EGF was comparable to EGF at low concentrations of 0.1 and 1 ng/mL but at a concentration of 10 ng/mL, HB-EGF induced significantly better proliferation than EGF. Proliferation was found to be dependent on the initial seeding density. Heparinized dendrimer crosslinked collagen (CHG) gels were capable of HB-EGF uptake, which was influenced by heparin concentration within the gel, growth factor concentration and exposure time to the growth factor. HB-EGF release followed first order kinetics, with ∼90% of the growth factor released after 2 weeks. Growth factor stability was verified with in vitro HCEC culture studies. Bioavailability was maintained in the gels through heparin interaction. Overall, HB-EGF induced proliferation of HCEC in vitro and can be released from heparinized collagen gels making it potentially suitable for promoting epithelialization of TECEs.

Modified dendrimer cross-linked collagen-based matrices.

Dendrimer cross-linking has been achieved with pepsin digested over 80% type-I bovine collagen to create strong hydrogels with good cell compatibility. Herein we investigate the use of commercially available collagen-based products with the dendrimer cross-linking technology. Specifically PureCol(®) (PC), a 97% bovine type-I collagen, human collagen (HC) and human extracellular matrix (hECM) were concentrated, and then cross-linked with polypropyleneimine octaamine generation two dendrimers using 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) chemistry. PC gels with 30 and 20 mg/ml bovine collagen were fabricated, and despite similar concentrations to >80% type-I bovine collagen dendrimer cross-linked gels (CG), PC gels demonstrated increased swelling and decreased stability, as determined with collagenase digestion. The highly purified bovine (PC) and human sourced-collagen (HC) gels were similar in performance, but not as stable as the CG gels, which may correlate to the manufacturer's collagen purification and storage. Finally, the addition of hECM components to PC to create PC-hECM gels, resulted in a looser gel network, compared to heparinized dendrimer cross- linked bovine >80% type-I collagen gels (CHG). However, all collagen-based gels supported 3T3 fibroblast cell growth over 4 days, indicating these gels may be suitable for tissue-engineering applications.

Photosensitive controlled release with polyethylene glycol-anthracene modified alginate.

Covalent modification of alginate with polyethylene glycol-conjugated anthracene molecules has the potential to both stabilize the alginate and act as a photosensitive crosslinker. Release studies with Coomassie Blue show lengthy release times from the alginate photogels that extend past 70 days with, for example, 17% versus 27% release at 1750 h (73 days) for photogels with and without 365-nm UV light treatment for 30 min at 10 mW/cm(2) in the initial release period. Photocrosslinking of the photogels after loading effectively "locks" in drug compounds to control their release. Effective crosslinking densities and controls of polyethylene glycol-crosslinked alginate and physically crosslinked calcium alginate gels suggest strong interactions between Coomassie Blue and both alginate and anthracene. Photogels containing anthracene-capped star-polyethylene glycol show increased photosensitivity with modified release profiles. Ultimately, the covalent modification of alginate with photoactive crosslinkers has the potential to produce a long-term, photosensitive, controlled release system.

Corneal epithelial cell biocompatibility to silicone hydrogel and conventional hydrogel contact lens packaging solutions.

PURPOSE: Although all contact lenses (CLs) are applied initially to the eye directly from a packaging solution, little is known about the effects of these solutions on human corneal epithelial cells (HCECs). Due to the porous nature of CL materials, they have the potential to sorb components of the packaging solution during storage, which could then be subsequently released upon insertion of the CL on the eye. The purpose of this study was to investigate the effect of various packaging solutions on HCECs, using an in vitro model. METHODS: An in vitro assay was developed whereby various silicone hydrogels and conventional, poly-2-hydroxyethylmethacrylate (polyHEMA)-based lens materials were removed directly from their packaging and then incubated for up to 24 h with HCECs. The effect of the retained and released packaging solution components on HCECs was assessed by measuring cell viability, adhesion phenotype, and apoptosis. RESULTS: Incubation of HCECs with CLs stored in borate-buffered packaging solutions resulted in a significant reduction in cell viability. Adherent cells incubated with these CLs also exhibited reduced levels of beta(1) and alpha(3) integrin. Soaking borate-buffered packaged CLs in PBS before cell incubation resolved viability and integrin expression in all cases, with the exception of galyfilcon A and balafilcon A, from which a 20% reduction in cell viability was still observed. In comparison, CLs stored in phosphate-buffered packaging solutions had cellular viability and expression of integrins similar to control cells (cells incubated in the absence of a lens). When incubated with cells at a 10% concentration in serum-free medium, borate-buffered packaging solutions and borate-containing saline (Unisol 4) significantly reduced cell viability and integrin expression. Neither caspase activation nor annexin V binding was observed on cells following exposure to borate buffer solution. However, a significant decrease in reactive oxygen species was observed at 24 h. These latter results suggest that in vitro exposure to low concentration of borate/boric acid results in cell dysfunction, leading to necrosis rather than apoptosis. CONCLUSIONS: Borate-buffered packaging solutions were shown to adversely affect the viability and integrin expression of HCECs in vitro. When used in ophthalmic packaging solutions, the antimicrobial properties of borate buffer may be outweighed by its relatively cytotoxic effects on cells.

A high-density PEG interfacial layer alters the response to an EGF tethered polydimethylsiloxane surface.

Previously, epidermal growth factor (EGF)-modified surfaces have shown promise in supporting cellular growth and adhesion on synthetic polymeric substrates. Surfaces prepared using a novel modification technique were investigated in the current work for their ability to support corneal epithelialization, important to the integration of a synthetic artificial cornea. EGF could be tethered to PDMS surfaces via a high-density, hetero-bifunctional PEG-NSC linking layer with a tunable surface concentration of up to 300 ng/cm(2). Only a small fraction of the EGF on these surfaces could be removed with SDS rinsing, indicative of covalent tethering. Studies with human corneal epithelial cells suggest a relatively linear increase in the number of corneal epithelial cells with increasing EGF concentration at all times. However, confluence was not achieved at any time point. It is believed that the presence of the non-adsorbent PEG layer, useful for preventing non-specific adsorption of proteins, may limit the cellular response by minimizing the adsorption of adhesion molecules. The effects of the EGF alone are clearly not sufficient to result in epithelialization of an artificial cornea surface. Altering both the adhesion and growth of corneal epithelial cells in a controlled manner may be necessary for epithelialization of an artificial cornea.

Heparin-modified dendrimer cross-linked collagen matrices for the delivery of basic fibroblast growth factor (FGF-2).

Tissue integration between a tissue-engineered corneal equivalent and the host eye is of critical importance in ensuring long-term implant success. A novel dendrimer cross-linked collagen scaffold has previously shown good corneal epithelial cell compatibility in vitro particularly when the highly functional dendrimer cross-linkers were functionalized to introduce additional biological groups. Herein we investigated heparinization of these materials and their potential to facilitate the delivery of basic fibroblast growth factor (FGF-2) in an active form, ultimately for use as a corneal tissue scaffold. Collagen gels cross-linked with 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) chemistry, and varying amounts of polypropyleneimine octaamine generation 2 (G2) dendimer and heparin were synthesized. Swelling studies and differential scanning calorimetry characterization indicated higher gel stability with the introduction of dendrimer cross-linking, which was not compromised by heparin integration. Dendrimer cross-linked gels with or without heparin gave multiple denaturation peaks, as did the heparinized EDC gels. This is thought to be the result of the heterogeneous cross-linking possible between collagen, the dendrimer and heparin. Release of FGF-2 from collagen gels showed typical first-order kinetics, with an initial burst followed by a prolonged gradual release. Heparinized dendrimer cross-linked gels released approx. 40% of the growth factor over a 2-week period, with significance maintenance of growth factor activity. Incorporation of heparin resulted in somewhat prolonged release from these systems.

Dendrimer-grafted cell adhesion peptide-modified PDMS.

Surface concentration of cell adhesion peptides is thought to play a role in the interactions between biomaterials and cells. The high density of functional groups at the periphery of dendrimers has been exploited in various applications, but their full potential for generating surfaces with high functional group concentrations has not yet been realized. Poly(dimethylsiloxane) elastomers were surface modified with both polyethylene oxide (PEO) and generation 3 diaminobutane dendrimers. PEO and the dendrimers were subsequently used as linker molecules for surface grafting of cell adhesion peptides. ATR-FTIR, X-ray photoelectron spectroscopy, and water contact angle results confirmed the successful attachment of the polymer linkers and peptides. Peptide grafting density was quantified by means of (125)I radiolabeling. Maximum surface peptide grafting density on dendrimer-modified surfaces was twofold greater than the maximum peptide grafting density achieved via the PEO linker. However, vascular endothelial cell adhesion was significantly greater on surfaces modified with the PEO linker, presumably due to the highly flexible PEO spacer making the peptide more accessible for binding with the cell surface receptors. These results suggest that, although peptide surface density may be important, optimizing surface density may not be sufficient for improving biological interactions.

Incorporation of cell-adhesion peptides into collagen scaffolds promotes corneal epithelial stratification.

The objective of this study was to examine the effects of cell-adhesion peptides incorporated into collagen scaffolds on corneal epithelial cell stratification. Peptides (YIGSR, YIGSRIKVAV, IKVAVYIGSR and negative control YISGR) were first chemically attached to dendrimers. The peptide-modified dendrimers were then used as collagen cross-linkers. This permitted the incorporation of the peptides into the bulk structure of the collagen gels. The amount of peptide incorporated into the collagen gels was determined by 125I radiolabelling to be between 0.064 and 6.4 microg/mg collagen for YIGSR, and between 0.1187 to 11.87 microg/mg collagen for YIGSRIKVAV and IKVAVYIGSR. Corneal epithelial cell monolayers were grown on the surface of the collagen scaffolds and then exposed to conditions that promoted stratification as a stratified epithelial layer is desired in a tissue-engineered cornea. It was found that all of the incorporated peptides promoted stratification of the cells with the exception of the negative control YISGR. A synergistic effect of the combined sequences from laminin was observed, with the orientation of the peptide sequences having a great impact on the ability of the materials to promote cell stratification.

Extended release of high pI proteins from alginate microspheres via a novel encapsulation technique.

Alginate has potential as a matrix for controlled delivery of protein-based drugs that require site-specific long-term delivery. In the current work albumin, lysozyme and chymotrypsin were encapsulated into alginate microspheres using a novel method that involved soaking the microspheres in a protein-containing NaCl solution. This was followed by recrosslinking with calcium chloride. High pI proteins also appeared to physically crosslink the sodium alginate which resulted in more sustained release. Release was affected by the nature of the releasate solution. In TRIS buffered saline, the high pI proteins chymotrypsin and lysozyme showed sustained release lasting over 150 h. Release into 0.15% NaCl led to relatively constant release of lysozyme and chymotrypsin over more than 2000 h; reduction of the releasate volume lengthened the lysozyme release to greater than 8 months. Released lysozyme was shown to remain active for at least 16 days, in some cases with activity greater than 100% of the active control. This encapsulation technique can therefore be used to rapidly load alginate microspheres with proteins, with high isoelectric point proteins showing particular promise. Furthermore, the interactions between the high pI proteins and the alginate gel could potentially be exploited to generate new protein delivery systems.

Dendrimer crosslinked collagen as a corneal tissue engineering scaffold: mechanical properties and corneal epithelial cell interactions.

Generation 2 polypropyleneimine octaamine dendrimers were used to generate highly crosslinked collagen with mechanical properties that would make it appropriate for use as a corneal tissue-engineering scaffold. Crosslinking of a highly concentrated collagen solution (2-4%) was effected using the water-soluble carbodiimide 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC). The multifunctional dendrimers were introduced as novel multifunctional crosslinkers after the activation of the carboxylic acid groups of glutamic and aspartic acid residues in collagen. Glutaraldehyde, a common collagen crosslinker, was used as comparison, as was EDC, itself an alternative crosslinker, which forms "zero-length or self-crosslinking". The mechanical properties resultant gels were determined. Young's modulus of the dendrimer crosslinked gels was significantly higher than that observed with the other crosslinkers, increasing to 5 MPa compared with 0.1 MPa for the EDC crosslinked gels. Transmission electron microscopy (TEM) analysis of the gels demonstrated the presence of fibrils in the thermally gelled collagen controls; no fibrils were observed in the dendrimer crosslinked gels. As a result, the optical transparency of the dendrimer crosslinked collagen was significantly better than that of the collagen thermal gels. The EDC and glutaraldehyde crosslinked gels were generally less transparent than those crosslinked with the dendrimers. Glucose permeation results demonstrated that the dendrimer crosslinked collagen had higher glucose permeability than natural human cornea. Dendrimer crosslinked collagen gels supported human corneal epithelial cell growth and adhesion, with no cell toxicity. In comparison, some potentially cytotoxic effects were observed with glutaraldehyde crosslinked collagen. Overall, the dendrimer crosslinked collagen gels showed promising properties that suggest that these might be suitable scaffolds for corneal tissue engineering and potentially other tissue engineering applications.

2-methacryloyloxyethyl N-butylcarbamate: a new co-monomer for synthesis of polyurethane hydrogels with improved mechanical properties for biomedical applications.

Hydrogels with tunable hydrophilic and mechanical properties were synthesized by the free radical polymerization of 2-hydroxyethyl methacrylate (HEMA) and 2-methacryloyloxyethyl N-butylcarbamate. The resulting hydrogels were investigated for their equilibrium water content, sessile drop water contact angles, gel fraction, mechanical properties and protein adsorption. Results indicated that co-polymer hydrogels have good hydrophilicity and that, with the incorporation of the 2-methacryloyloxyethyl N-butylcarbamate, mechanical properties could be improved significantly. without affecting other important properties. Lysozyme and albumin adsorption experiments demonstrated that, similar to most hydrogel materials, the co-polymer hydrogels adsorb more lysozyme than albumin and that the adsorption was dependent on hydrophilicity. The control poly(HEMA) hydrogels were found to adsorb more protein than the co-polymer hydrogels; this is thought to be primarily a consequence of protein absorption rather than protein adsorption.

Crosslinking of collagen with dendrimers.

Polypropyleneimine octaamine dendrimers were studied as an alternative means of generating highly crosslinked collagen. Crosslinking was effected by using the water-soluble carbodiimide 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC). The multifunctional dendrimers were introduced as novel crosslinkers after the activation of the carboxylic acid groups of glutamic and aspartic acid residues in collagen. The conventional crosslinker glutaraldehyde was used as a control. EDC, itself an alternative crosslinker, which forms zero-length crosslinks by directly covalently binding collagen molecules, as well as a low molecular weight diamine and a low molecular weight triamine, were also studied. All of the resultant gels were freeze-dried to obtain sponges for characterization. Water uptake of the gels decreased from 90% to 60% after dendrimer crosslinking compared with EDC crosslinking. DSC results showed an increase of denaturation temperature of collagen after crosslinking with the various methods. The generation 2 and 3 dendrimer-crosslinked collagen samples had the highest denaturation temperature, at up to 90 degrees C compared with 50 degrees C in the uncrosslinked collagen control. The dendrimer-crosslinked collagen also showed unique thermal characteristics, with multiple denaturation temperature peaks in contrast to the single peak noted with the other crosslinked collagens. This is thought to be due to the heterogeneous nature of dendrimer crosslinking. Collagenase results revealed that the dendrimer-crosslinked collagen had a comparative resistance to proteolysis to glutaraldehyde-crosslinked collagen. Measurement of activated carboxylic acid groups before and after crosslinking indicated that 40-70% of the activated carboxylic acid was consumed during crosslinking with dendrimers. The results suggest that dendrimer crosslinking of collagen produces stable gels. The presence of a large number of excess amine groups in the dendrimers may also be useful for subsequent modification with biologically relevant groups.

EGF-grafted PDMS surfaces in artificial cornea applications.

Lack of epithelial cell coverage has remained a persistent problem in the design of an artificial cornea. In this work, polydimethylsiloxane (PDMS) surfaces were modified with epidermal growth factor (EGF) to improve the growth of corneal epithelial cells. The EGF was covalently tethered to PDMS substrates aminated by plasma polymerization of allylamine via a homobifunctional polyethylene glycol (PEG) spacer. Surface modification was confirmed by contact angle and X-ray photoelectron spectroscopy measurements. By varying the ratio of EGF to PEG from 1:50 to 1:5, EGF amounts from 40 to 90 ng/cm2 could be bound, as determined by surface plasmon resonance (SPR) and 125I radiolabelling. Human corneal epithelial cells on the various modified surfaces were cultured both in the presence and absence of EGF in the culture medium to determine the effect of covalently bound EGF on the cells. The results demonstrated that covalently bound EGF on the surfaces is active with respect to promoting epithelial cell coverage. This was significant when compared to unmodified controls.

Glucose permeable poly (dimethyl siloxane) poly (N-isopropyl acrylamide) interpenetrating networks as ophthalmic biomaterials.

Poly (dimethyl siloxane) (PDMS) has been widely used as a biomaterial in ophthalmic and other applications due to its good compatibility, high mechanical strength, excellent oxygen permeability and transparency. However, for use as an artificial cornea, contact lens and in other applications, modifications with hydrophilic functional groups or polymers are necessary to improve wettability for tear protein and mucin interactions and to improve glucose permeability for cellular health. Poly (N-isopropyl acrylamide) (PNIPAAM) is a biocompatible and hydrophilic polymer that has been extensively studied on controlled drug release applications due to its lower critical solution temperature (LCST) phenomenon. In the current work, a composite interpenetrating network (IPN) of PDMS and PNIPAAM was formed to generate polymers with oxygen and glucose permeability as well as improved wettability compared to PDMS homopolymers and greater mechanical strength than PNIPAAM homopolymers. Transparent vinyl and hydroxyl terminated PDMS/PNIPAAM IPNs (PDMS-V and PDMS-OH IPNs, respectively) were successfully synthesized. Transmission electron microscopy images verified the structure of the IPNs. Surface analysis suggested that PNIPAAM was present on the surface as well as in the bulk material. PDMS-OH IPNs generated from a PDMS-OH matrix cured in the presence of solvent had the highest glucose permeability at 10(-7)cm2/s, comparable to that of the native cornea. The LCST phenomenon remained in these materials, although changes were not as abrupt as with pure PNIPAAM. These results suggest that these materials may be further developed as ophthalmic biomaterials or for controlled drug-release applications.

Recruitment of multiple cell lines by collagen-synthetic copolymer matrices in corneal regeneration.

Collagen hydrogel matrices with high optical clarity have been developed from collagen I, cross-linked with a copolymer based on N-isopropylacrylamide, acrylic acid and acryloxysuccinimide. The controlled reaction of collagen amine groups with this copolymer under neutral pH and aqueous conditions gave robust, optically clear hydrogels and prevented the excessive collagen fibrillogenesis that can lead to collagen opacity. These sterile, non-cytotoxic hydrogels allowed epithelial cell overgrowth and both stromal cell and nerve neurite ingrowth from the host tissue. This regenerative ability appeared to result from the high glucose permeability, nanoporosity and the presence of cell adhesion factors, RGD in collagen and the laminin pentapeptide, YIGSR, grafted onto the copolymer. Under physiological conditions, optical clarity superior to the human cornea and tensile performance adequate for suturing were obtained from some formulations.

Partitioning of model toxins to hydrophobically terminated DAB dendrimers.

Dendrimers are attractive in biological and biomedical applications due to the similarity in their molecular size to biologically relevant molecules and the large number of chain ends available functionalization. In the current work, we examined the potential of diamino butane (DAB) dendrimers functionalized with long alkyl chains as partitioning agents for hydrophobic toxins for use as a prefiltering stage in a bioartiticial liver. DAB dendrimers of various generations that had been previously fully modified with palmitoyl chloride were obtained. A study of the kinetics of partitioning of acetylsalicylic acid (ASA) suggested that while significant toxin removal occurred in 30 s, although a slight time dependent increase in removal was noted up to 60 minutes. The partitioning of 6 hydrophobic toxins from aqueous solution to the modified dendrimers in 30 minutes was examined. The results demonstrated that a number of factors, including the pKa of the toxin, its octanol water partitioning coefficient and molecular size contributed to the level of toxin removal. Toxin removal on a molar basis increased with increasing dendrimer generation for all toxins, with the modified G5 dendrimers partitioning 50-100 toxin molecules in most cases. Dendrimer modification with C4 alkyl chains rather than Cl5 chains significantly decreased toxin removal, although chains longer than C10 seemed to partition equal amounts of toxins. The results of the study demonstrate that water-soluble dendrimers modified with hydrophobic end groups may be useful for the removal of toxins from the blood in a prereaction step for a bioartificial liver, but that a better understanding of the molecular mechanisms of removal may be necessary before it is possible to predict the levels of toxin removal.

Interactions of corneal cells with transforming growth factor beta 2-modified poly dimethyl siloxane surfaces.

The downgrowth of corneal epithelial cells at the interface of an artificial cornea and the host eye tissue poses a significant problem to be overcome in developing a successful implant. As a means of inhibiting the proliferation of corneal epithelial cells on the stromal surface of the implant, we examined the immobilization of transforming growth factor beta-2 (TGF-beta2) via a bifunctional poly ethylene glycol (PEG) spacer to poly dimethyl siloxane (PDMS) surfaces. Growth factor immobilization was confirmed by modification with (125)I-labeled TGF-beta 2. The modified surfaces were also characterized by advancing water contact angles, X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM). Although the amount of growth factor covalently bound to the surface was difficult to quantify apparently due to strong interactions between the growth factor and the PEG layer and high levels of adsorption, differences in the modified surfaces, suggestive of the presence of a significant amount of TGF-beta 2, were found. In vitro interactions of the modified surfaces with human corneal epithelial and stromal cells were examined. Growth factor surface concentrations as well as culture in the absence and presence of serum and other adhesive proteins were examined. Corneal stromal and epithelial cells cultured on the TGF-beta 2-modified surfaces consistently gave results opposite to those expected. Likely, the most notable and surprising result was the almost complete lack of adhesion of the stromal cells, with coverage averaging between 3 and 5%. In comparison, corneal epithelial cell growth appeared to be promoted by the presence of the immobilized growth factor, with cell coverage averaging 50-60% at 7 days of culture. A TGF-beta 2 concentration effect was noted with both cell types in the absence of serum, with increases in the coverage at higher TGF-beta 2 concentrations. The observed cell growth appeared to be the result of interactions between the cells and active growth factor, because the addition of anti-TGF-beta 2 to the culture medium reduced cell coverage to levels similar to those noted on control surfaces. Therefore, although TGF-beta 2-modified surfaces may not be suitable as corneal epithelial cell inhibiting surfaces, interactions of surface immobilized growth factor and corneal cells are complex and should be further examined.

Surface analysis methods for characterizing polymeric biomaterials.

Surface properties have an enormous effect on the success or failure of a biomaterial device, thus signifying the considerable importance of and the need for adequate characterization of the biomaterial surface. Microscopy techniques used in the analysis of biomaterial surfaces include scanning electron microscopy, transmission electron microscopy, atomic force microscopy, and confocal microscopy. Spectroscopic techniques include X-ray photoelectron spectroscopy, Fourier Transform infrared attenuated total reflection and secondary ion mass spectrometry. The measurement of contact angles, although one of the earlier techniques developed remains a very useful tool in the evaluation of surface hydrophobicity/hydrophilicity. This paper provides a brief, easy to understand synopsis of these and other techniques including emerging techniques, which are proving useful in the analysis of the surface properties of polymeric biomaterials. Cautionary statements have been made, numerous authors referenced and examples used to show the specific type of information that can be acquired from the different techniques used in the characterization of polymeric biomaterials surfaces.

Novel dendrimer based polyurethanes for PEO incorporation.

A series of segmented polyurethanes based on methylene diisocyanate/poly (tetramethylene oxide) and chain extended with either ethylene diamine or butane diol in combination with a generation 2 polypropylenimine octaamine dendrimer were synthesized. For polymer synthesis, the dendrimers were protected with either t-boc or Fmoc groups and were incorporated into the polyurethane microstructure to permit further functionalization with biologically active groups. Following deprotection, the dendrimers were reacted with succinimidyl propionate polyethylene oxide (SPA-PEO) to improve the protein resistance of the polymers and to examine the potential of this technique for polymer functionalization. Different synthesis techniques were examined to optimize the incorporation of the PEO into the polymer microstructure. Incorporation of the dendrimers and the PEO were confirmed by NMR and FTIR. Gel permeation chromatography was used to examine the molecular weights of the various polyurethanes. The dendrimer incorporated polymers had significantly lower molecular weights than the ED or BDO chain extended controls, likely due to lower reactivity of the dendrimers as a result of steric factors. Following PEO reaction, the molecular weights of the resultant polymers were consistent with the levels of PEO incorporation noted by comparison of peak intensities in the NMR spectra. Due to the highly hydrophilic nature of the PEO, some migration to the polymer surface was expected. Water contact angles and XPS, used to characterize the surfaces, suggest that there was some PEO enrichment at the surface of the polymers. Adsorption of radiolabeled fibrinogen to the polymer surfaces was decreased by a factor of approximately 40% in some of the PEO incorporated polymers. There were also differences in the patterns of plasma protein adsorption on the various surfaces as evaluated by SDS PAGE and immunoblotting. Therefore, the use of dendrimers in biomaterials for incorporation of a large number of functional groups seems to be promising.