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Attachment and growth of cultured fibroblast cells on PVA/chitosan-blended hydrogels.

Hydrogels were prepared from poly(vinyl alcohol) and chitosan in various blend ratios. The water contents of the hydrogels were in the range of 65 to 75 wt %. The attachment and growth of fibroblast cells (L-929) on the hydrogels were studied with a cell culture method. On the hydrogels with more than 15 wt % chitosan content, the attached cells were able not only to remain viable but also to proliferate. The relative cell attachment after incubation for 30 h increased with increasing chitosan content in the hydrogels. Cell attachment and growth on the hydrogel with 40 wt % chitosan content exceeded those on collagen, a widely-used mammalian cell culture substrate. The morphology of the cells attached onto the hydrogels with a lower chitosan content was spherical, but in hydrogels with more than 15 wt % chitosan content, the number of spindle-shaped cells increased with increasing chitosan content.

Biocompatible Materials↗

Implantable controlled delivery systems for proteins based on collagen--pHEMA hydrogels.

Co-polymeric hydrogels of collagen and poly(hydroxyethyl methacrylate) (pHEMA) were prepared using an ammonium persulphate-sodium metabisulphite redox initiation technique. Fluorescein isothiocyanate (FITC)-labelled bovine serum albumin (BSA) was entrapped into these hydrogels. These gels were modified differently by external cross-linking and made highly porous by incorporation of ethylene glycol. The morphological characteristics of the gels containing FITC-BSA before and after release were studied. The in vitro release of FITC-labelled BSA was studied in phosphate buffer, pH 7.4, at 37 degrees C. The results clearly indicated that collagen-pHEMA hydrogels release FITC-BSA in a zero-order fashion. In the case of uncross-linked hydrogels, the release of FITC-BSA was about 81% within 8 d, compared to 49% for the same period in the case of cross-linked hydrogels. It was also observed that in the case of porous hydrogels the release was very fast (19% in the first hour) and lasted for about 12 h, releasing 76% of FITC-BSA. These results clearly indicated that collagen-pHEMA hydrogels released BSA in a zero-order fashion and that the release rate can be programmed by appropriate modification of the hydrogel matrices.

Animals↗

Comparison of covalently and physically cross-linked polyethylene glycol-based hydrogels for the prevention of postoperative adhesions in a rat model.

A covalently and a physicochemically cross-linked hydrogel, both based primarily on polyethylene glycol and both formed in situ, were compared side by side in a rat uterine horn devascularization and serosal injury model for efficacy in adhesion prevention. The primary difference between the two materials was the nature of their cross-linking. The covalently cross-linked hydrogel was a photopolymerized polyethylene glycol-co-lactic acid diacrylate, and the physically cross-linked hydrogel was a polyethylene glycol-co-polypropylene glycol, Poloxamer 407. In the surgical model employed, application of the covalently cross-linked hydrogel reduced the extent of adhesion formation from 75 +/- 10% in the control group to 16 +/- 6% (mean +/- s.d., P < 0.001). Application of the physically cross-linked hydrogel reduced adhesion formation to 38 +/- 19% (P < 0.01). Retention of the two hydrogels upon the site of application was also evaluated. The covalently cross-linked hydrogel formed a continuous barrier upon the uterine horns for more than 4 d, while the physicochemically cross-linked hydrogel was present upon the uterine horns for less than 2 d. This difference in retention was probably the cause of the difference in efficacy and may be attributed to the nature of the cross-linking.

Animals↗

Macroporous poly(sucrose acrylate) hydrogel for controlled release of macromolecules.

We have studied the controlled release of proteins from poly(sucrose acrylate) hydrogels. The hydrogels were prepared by a two-step procedure in which sucrose was first acylated to sucrose-1'-acrylate followed by free radical polymerization. By adjusting the cross-link ratio and initial monomer concentration, the swelling ratio of the hydrogel was varied from five to 28. The mechanical strength of these hydrogels was comparable to that of the hydrogels with approximately the same swelling ratio. Scanning electron micrographs and mesh size calculations indicate that the hydrogel is macroporous, suggesting it may be suitable for a variety of biomedical applications. The release kinetics of beta-lactoglobulin, bovine serum albumin and gamma-globulin were studied as a function of initial monomer concentrations for the sucrose-based hydrogel. All of the release profiles were characterized by an initial burst of protein in the first 25 h followed by a long period of sustained release (> 500 h). The magnitude of the initial burst was reduced by increasing the initial monomer concentration and by increasing the molecular weight of the protein. A quantitative model based on the heterogeneous nature of hydrogel was developed to explain the observed release kinetics.

Acrylic Resins↗

Molding of hydrogel microstructures to create multiphenotype cell microarrays.

The fabrication of mammalian cell-containing poly(ethylene glycol) (PEG) hydrogel microstructures on glass and silicon substrates is described. Using photoreaction injection molding in poly(dimethylsiloxane) microfluidic channels, three-dimensional hydrogel microstructures encapsulating cells (fibroblasts, hepatocytes, macrophage) were fabricated with cells uniformly distributed to each hydrogel microstructure, and the number of cells in each hydrogel microstructure was controlled by changing the cell density of the precursor solution. PEG hydrogels were modified using an Arg-Gly-Asp (RGD) peptide sequence, with the incorporation of RGD into the hydrogel matrix promoting the spreading of encapsulated fibroblasts over a 24-h period in culture. Cells remained viable encapsulated in these hydrogel microstructures for a period in excess of 1 week in culture. Arrays of hydrogel microstructures encapsulating two or more phenotypes on a single substrate were successfully fabricated using multimicrofluidic channels, creating the potential for multiphenotype cell-based biosensors.

Journal Article↗

Non-destructive characterization of hydrogels.

Hydrogels have been prepared by a freezing-thawing procedure and investigations made of the effect of both number of freezing-thawing cycles and different content of hyaluronic acid (HA) on the mechanical properties of the PVA-HA hydrogels using non-destructive testing. The bulk elastic modulus K of hydrogels has been determined by pulse-echo measurements. It is noted that hydrogel elastic properties improve with the number of the cycles in PVA-HA 100/0; on the other hand samples with a high HA (1,000,000 molecular weight) content, beyond the third cycle, seem to be unaffected by the number of cycles. A bulk elastic modulus fall-off is then observed in samples submitted to an additional overnight freezing between two subsequent cycles. K increases in hydrogels with the highest HA content, when samples undergo pulse-echo measurements soon after their preparation. When hydrogels reach equilibrium, after having been kept in deionized water for 12 h, K values are lower, showing a nearly constant behaviour with different PVA-HA ratios and cycles. Furthermore, by means of scanning laser acoustic microscopy (SLAM) defects have been detected in the hydrogels. In samples which have reached equilibrium, SLAM images show that these defects disappear in PVA-HA hydrogels.

Journal Article↗

Thermoresponsive hydrogel with rapid response dynamics.

Intelligent hydrogels, particularly poly(N-isopropylacrylamide) (PNIPAAm)-based hydrogels, have attracted extensive interest because the soft wet hydrogels can change their shapes in response to the small changes of environmental factors like temperature. In order to fully make use of this unique property of PNIPAAm-based hydrogels, the response rates of the PNIPAAm hydrogels have to be improved since the dynamics property is critical to certain applications of this material. In this paper, the thermo-sensitive PNIPAAm hydrogels were successfully synthesized by carrying out the polymerization of N-isopropylacrylamide monomer in vacuum (-100 kPa) at room temperature (22 degrees C). The resultant hydrogel has tremendously improved shrinking rate as well as the large volume changes upon temperature stimulation when comparing with the normal PNIPAAm hydrogel. The SEM micrographs revealed that the improved properties were attributed to the macroporous network structure generated during the synthesis under vacuum.

Journal Article↗

Transformation of hydrogel-based inverse opal photonic sensors from FCC to L1(1) during swelling.

The structural evolution of Bragg diffracting inverse opal hydrogel sensors during swelling is directly observed by two-photon laser scanning fluorescence microscopy and compared to predictions from finite element analysis. A fluorescently labeled pH-sensitive hydrogel is UV-polymerized in a dried polystyrene colloidal crystal template, which is etched to yield an inverse opal. Fluorescence imaging of the hydrogel at different pH values reveals an inhomogeneous deformation of the FCC array of aqueous pores. The pores elongate along the sample normal direction and collapse along the sample parallel directions, consistent with the Bragg response, which indicates a 1-D increase in the interlayer distance. Interconnects between the pores serve as anchor points during hydrogel expansion into the pores. Pinning of the hydrogel to the substrate causes a change of the hydrogel lattice symmetry during deformation, from FCC (ABC stacking) to L1(1) (ABCA'B'C' stacking). Reconstructed cross-sections confirm that a 1-D increase in the interlayer distance along the substrate normal direction is responsible for the diffraction response of an inverse opal hydrogel sensor. Comparison with predictions from finite element analysis shows qualitative agreement, although the experimental mesostructure is significantly more deformed than the calculated data, due to buckling in the experimental system that is not captured by the model.

Biosensing Techniques↗

Synthesis and characterization of dextran-maleic acid based hydrogel.

A new class of hydrogel precursor, dextran-maleic acid (Dex-MA), was synthesized by the reaction of dextran with maleic anhydride in the presence of the catalyst triethylamine. The effects of temperature, time, catalyst amount, and reactant concentration on the degree of substitution (DS) by MA was studied to establish an optimum reaction condition. The new hydrogel precursor had excellent solubility in various common organic solvents. The hydrogels based on Dex-MA precursor were made by the irradiation of Dex-MA with a long wave UV lamp. The Dex-MA hydrogels showed a very high swelling ratio in water, and the magnitude of swelling depended on the pH of the medium and the DS by MA. The Dex-MA hydrogels exhibited the highest swelling ratio in neutral pH, followed by acidic (pH 3) and alkaline pH (10). The most distinctive characteristic of Dex-MA hydrogels was that a carboxylic acid group was generated by the reaction of dextran with maleic anhydride. As a result, the swelling ratio increased with an increase of the DS of the MA segment (ionizable moiety that affects swelling ratio) in the Dex-MA hydrogel.

Adsorption↗

Synthesis and characterization of sol-gel phase-reversible hydrogels sensitive to glucose.

A new type of hydrogel capable of sol-gel phase-reversible transition by changes in the environmental glucose concentration has been synthesized. The hydrogel consists of vinylpyrrolidinone-allylglucose (VP/AG) copolymer and concanavalin A (Con A). The formation of the hydrogel is based on the specific interaction between glucose on the copolymer and glucose receptor sites on Con A. Hydrogels were formed immediately after mixing the copolymer and Con A. The critical factors in the formation of hydrogel were the concentrations of the copolymer, of glucose on the copolymer and of Con A. In general, the formation of a hydrogel became more efficient as the concentration of glucose on the copolymer decreased and/or as the Con A concentration increased. The hydrogel formed became a sol in the presence of glucose in the environment. The environmental glucose concentration necessary to dissolve the gel to sol was approximately four times higher than the concentration of glucose on the copolymer. Upon removal of the environmental glucose by dialysis, the sol became a gel again and the sol-gel phase transition could be repeated.

Concanavalin A↗

Biodegradable dextran-polylactide hydrogel network and its controlled release of albumin.

The objective of this paper was to study the release of bovine serum albumin (BSA) from a series of biodegradable hydrogels having a wide range of hydrophilicity to hydrophobicity, swelling, and biodegradation properties. BSA was incorporated into a series of biodegradable hydrogels made from a dextran derivative of allyl isocyanate (dex-AI, as the hydrophilic constituent) and poly(DL-lactic acid) diacrylate macromer (PDLLAM, as the hydrophobic constituent). The release kinetics of BSA from these dex-AI/PDLLAM hydrogels was studied. Laser confocal scanning microscopy was used to investigate the morphological change of the hydrogels, as well as BSA distribution in the hydrogels, as a function of dex-AI to PDLLAM composition ratio and incubation time. We found that the incorporation of PDLLAM into dex-AI reduced the initial burst release of BSA due to its more homogeneous distribution in the hydrogels. As the PDLLAM component increased, the rate of formation of a loose three-dimensional (3D) network structure increased; consequently, the sustained rate and extent of BSA release increased. Both release index and diffusion coefficient (from release kinetics data) increased as the PDLLAM component increased in the hydrogels. The data suggest that the release of BSA was controlled by both diffusion of BSA through swelling of the hydrophilic phase during an early stage, and degradation of the hydrophobic phase during a late stage, and also that the magnitude of diffusion versus degradation controlled release is dependent on composition ratio and immersion time.

Biocompatible Materials↗

Attachment of fibronectin to poly(vinyl alcohol) hydrogels promotes NIH3T3 cell adhesion, proliferation, and migration.

Hydrogels have been used in biology and medicine for many years, and they possess many properties that make them advantageous for tissue engineering applications. Their high water content and tissue-like elasticity are similar to the native extracellular matrix of many tissues. In this work, we investigated the potential of a modified poly(vinyl alcohol) (PVA) hydrogel as a biomaterial for tissue engineering applications. First, the ability of NIH3T3 fibroblast cells to attach to PVA hydrogels was evaluated. Because of PVA's extremely hydrophilic nature, important cell adhesion proteins do not adsorb to PVA hydrogels, and consequently, cells are unable to adhere to the hydrogel. By covalently attaching the important cell adhesion protein fibronectin onto the PVA hydrogel surface, the rate of fibroblast attachment and proliferation was dramatically improved, and promoted two-dimensional cell migration. These studies illustrate that a fibronectin-modified PVA hydrogel is a potential biomaterial for tissue engineering applications.

3T3 Cells↗

Effect of chitosan-polyvinyl pyrrolidone hydrogel on proliferation and cytokine expression of endothelial cells: implications in islet immunoisolation.

Earlier we have shown the suitability of chitosan-polyvinyl pyrrolidone (PVP) hydrogel for islet immunoisolation and its inability to activate macrophages. Biomaterials that support vascularization without activating immune competent endothelial cells are desirous in islet immunoisolation. The aim of the present study was to evaluate effect of chitosan-PVP hydrogel on proliferation and activation of endothelial cells. Hydrogel did not allow the majority of cells to adhere well but maintained their viability. Hydrogel leachouts were nontoxic to the cells, as confirmed by tetrazolium reduction (MTT) and Neutral red uptake assays. Exposure to leachouts also did not alter their functionality as seen from normal expression of von Willebrand factor. 3H-thymidine incorporation revealed that hydrogel leachouts did not induce endothelial cell proliferation. Cells cultured on hydrogel and polystyrene control showed comparable expression of interleukin (IL) 6, IL-10, and transforming growth factor beta, with higher expression of tumor necrosis factor alpha as determined by reverse transcription-polymerase chain reaction. Taken together these results point out that hydrogel is compatible with endothelial cells and maintains their nonactivated status and hence is suitable as immunoisolation matrix.

Biocompatible Materials↗

Modeling ionic hydrogels swelling: characterization of the non-steady state.

Ionic hydrogels can be used as controlled release systems that respond to an external substrate or trigger by swelling or de-swelling. One example is a glucose-sensitive system for insulin-controlled release based on pH-sensitive hydrogel. To enhance understanding of non-steady state swelling, and to facilitate design of specifications (e.g., glucose-sensitivity) of the pH-sensitive ionic hydrogel based on the copolymer poly (2-hydroxyethyl methacrylate-co-N, N-dimethylaminoethyl methacrylate) (poly (HEMA-co-DMAEMA)), we developed a mathematical compartmental model using the software SAAM II. Current analytical and computational methods focus on equilibrium swelling of hydrogels; although for many stimuli-responsive hydrogel applications, the dynamic process is significant. We now report, using a combination of experimental data and kinetic analysis that in the poly (HEMA-co-DMAEMA) the rate of proton entry is governed by a different rate coefficient than water entry rate. The transport coefficient governing water uptake is dependent upon three variables: pH of external media, amine groups incorporated into the polymer, and crosslinking density of the polymer. An additional result is that swelling equilibrium is reached when all the amine groups are protonated. In this study we also demonstrate the predictive capability of the model for both interpolated and extrapolated data, and its use in design of future bench experiments. Uncovering these fundamental properties of pH-sensitive hydrogels with the aid of a kinetic model suggests that the complexities of hydrogel research and development can be overcome by combining experimental and computational approaches.

Computer Simulation↗

The influence of hydrogel functional groups on cell behavior.

This study provides information on the behavior of macrophages on the surface of the subcutaneously implanted hydrogel strips. Hydrogel containing -OH, -CO-NH-, and (CH3)2N- groups induced a spreading of macrophages on the implants. The materials containing -SO3H groups slightly, and materials containing -COOH groups more intensively, inhibited spreading of the macrophages. The fusion of macrophages into multinucleate cells was inhibited on the surface of materials containing acidic groups (-SO3H, -COOH) and increased on the hydrogels containing 30 mol% of alkaline (CH3)2N- groups in comparison with hydrogels containing -OH or -CO -NH- groups. The differences of the behavior of macrophages on the surface of individual types of hydrogels are probably independent on the adsorption of plasma fibronectin onto the hydrogels. The correlation between the macrophages spreading and fusion and surface charge of the hydrogel implant can be hypothetically explained by electrostatic interaction between macrophages cell membrane and implant.

Animals↗

Hydrogels based on poly(ethylene oxide) and poly(tetramethylene oxide) or poly(dimethyl siloxane). III. In vivo biocompatibility and biostability.

To investigate the effects of polymer chemistry and topology (linear or graft copolymer) on in vivo biocompatibility and biostability based on cage implant system, various hydrogels, composed of short hydrophilic [polyethylene oxide (PEO)] and hydrophobic block, were prepared by polycondensation reaction. Poly(tetramethylene oxide) (PTMO) or poly(dimethyl siloxane) (PDMS) was chosen as a hydrophobic block because of their wide utilization as a biomaterial. By using the specimens retrieved from rats killed after 1, 2, 3, 5, and 7 weeks' implantation, cellular and material responses were assessed. Most hydrogels showed a comparable value of macrophage density to Pellethane(R), control polymer, whereas they did significantly lower foreign body giant cell (FBGC) density and coverage because of the presence of PEO. However, PEO block length and polymer topology did not affect macrophage adhesion and FBGC formation in our polymer composition. The hydrogel based on PDMS alone showed significantly lower macrophage density and FBGC density than Pellethane(R), indicating that PDMS plays a role in inhibiting cellular adhesion. The results obtained from gel permeation chromatography curve and Fourier transform infrared spectra exhibited that all the polymers were susceptible to oxidative degradation in vivo. Although Pellethane(R) revealed surface degradation by 5 weeks in vivo, hydrogels showed rapid degradation in the bulk within 2 weeks because of the penetration of oxidative chemicals released from phagocytic cells into PEO domain of phase-separated hydrogels. The more significant degradation was observed in the hydrogels with longer PEO block and PTMO as a hydrophobic block instead of PDMS. It was evident that the minor degradation could be achieved by grafting PEO and adopting PDMS as a hydrophobic block in the hydrogel.

Animals↗

Attachment and spreading of fibroblasts on an RGD peptide-modified injectable hyaluronan hydrogel.

Hyaluronan (HA) hydrogels resist attachment and spreading of fibroblasts and most other mammalian cell types. A thiol-modified HA (3,3'-dithiobis(propanoic dihydrazide) [HA-DTPH]) was modified with peptides containing the Arg-Gly-Asp (RGD) sequence and then crosslinked with polyethylene glycol (PEG) diacrylate (PEGDA) to create a biomaterial that supported cell attachment, spreading, and proliferation. The hydrogels were evaluated in vitro and in vivo in three assay systems. First, the behavior of human and murine fibroblasts on the surface of the hydrogels was evaluated. The concentration and structure of the RGD peptides and the length of the PEG spacer influenced cell attachment and spreading. Second, murine fibroblasts were seeded into HA-DTPH solutions and encapsulated via in situ crosslinking with or without bound RGD peptides. Cells remained viable and proliferated within the hydrogel for 15 days in vitro. Although the RGD peptides significantly enhanced cell proliferation on the hydrogel surface, the cell proliferation inside the hydrogel in vitro was increased only modestly. Third, HA-DTPH/PEGDA/peptide hydrogels were evaluated as injectable tissue engineering materials in vivo. A suspension of murine fibroblasts in HA-DTPH was crosslinked using PEGDA plus PEGDA peptide, and the viscous, gelling mixture was injected subcutaneously into the flanks of nude mice; gels formed in vivo following injection. After 4 weeks, growth of new fibrous tissue had been accelerated by the sense RGD peptides. Thus, attachment, spreading, and proliferation of cells is dramatically enhanced on RGD-modified surfaces but only modestly accelerated in vivo tissue formation.

Animals↗

Biocompatibility of chemoenzymatically derived dextran-acrylate hydrogels.

The biocompatibility of chemoenzymatically generated dextran-acrylate hydrogels has been evaluated in vitro, using human foreskin fibroblasts, and in vivo, by subcutaneous and intramuscular implantation in Wistar rats for up to 40 days. In vitro tests show that hydrogel extracts only minimally reduced (<10%) the mitochondrial metabolic activity of fibroblasts. Direct contact of the hydrogels with cells induced a cellular proliferation inhibition index (CPII) of 50-80%, compared with a control, whereas through indirect contact, the CPII values were <16%, suggesting that the high CPII values achieved in the direct assay test were likely due to mechanical stress or limitations in oxygen diffusion. Hence, the hydrogels were noncytotoxic. Moreover, cell-material interaction studies show that these hydrogels were nonadhesive. Finally, histologic evaluation of tissue response to subcutaneous and intramuscular implants showed acceptable levels of biocompatibility, as characterized by a normal cellular response and the absence of necrosis of the surrounding tissues of the implant. In the first 10 days, the foreign-body reaction in the intramuscular implantation was more severe than in subcutaneous implantation, becoming identical after 30 days. In both cases, dextran hydrogels did not show signs of degradation 6 weeks postimplantation and were surrounded by a thin fibrous capsule and some macrophages and giant cells. This response is typical with a number of nondegradable biocompatible materials. These results indicate that dextran hydrogels are biocompatible, and may have suitable applications as implantable long-term peptide/protein delivery systems or scaffolds for tissue engineering.

Acrylates↗