Catechol-functionalized chitosan/pluronic hydrogels for tissue adhesives and hemostatic materials

Bioinspired from adhesion behaviors of mussels, injectable and thermosensitive chitosan/Pluronic composite hydrogels were synthesized for tissue adhesives and hemostatic materials. Chitosan conjugated with multiple catechol groups in the backbone was cross-linked with terminally thiolated Pluronic F-127 triblock copolymer to produce temperature-sensitive and adhesive sol-gel transition hydrogels. A blend mixture of the catechol-conjugated chitosan and the thiolated Pluronic F-127 was a viscous solution state at room temperature but became a cross-linked gel state with instantaneous solidification at the body temperature and physiological pH. The adhesive chitosan/Pluronic injectable hydrogels with remnant catechol groups showed strong adhesiveness to soft tissues and mucous layers and also demonstrated superior hemostatic properties. These chitosan/Pluronic hydrogels are expected to be usefully exploited for injectable drug delivery depots, tissue engineering hydrogels, tissue adhesives, and antibleeding materials.     

Biocompatible scaffolds based on collagen and oxidized dextran for endothelial cell survival and function in tissue engineering

Angiogenesis is a vital step in tissue regeneration. Hence, the current study aimed to prepare oxidized dextran (Odex)/collagen (Col)-hydrogels with laminin (LMN), as an angiogenic extracellular matrix (ECM) component, for promoting human umbilical vein endothelial cell (HUVEC) proliferation and function. Odex/Col scaffolds were constructed at various concentrations and temperatures. Using oscillatory rheometry, scanning electron microscopy (SEM), and cell viability testing, the scaffolds were characterized, and then HUVEC proliferation and function was compared with or without LMN. The gelation time could be modified by altering the Odex/Col mass ratio as well as the temperature. SEM showed that Odex/Col hydrogels had a more regular three-dimensional (3D) porous structure than the Col hydrogels. Moreover, HUVECs grew faster in the Col scaffold (12 mg/mL), whereas the Odex (30 mg/mL)/Col (6 mg/mL) scaffold exhibited the lowest apoptosis index. Furthermore, the expression level of vascular endothelial growth factor (VEGF) mRNA in the group without LMN was higher than that with LMN, and the Odex (30 mg/mL)/Col (6 mg/mL) scaffold without LMN had the highest VEGF protein secretion, allowing the cells to survive and function effectively. Odex/Col scaffolds, with or without LMN, are proposed as a tissue engineering construct to improve HUVEC survival and function for angiogenesis.     

One-step synthesis of interpenetrating network hydrogels: Environment sensitivities and drug delivery properties

A novel interpenetrating network hydrogel for drug controlled release, composed of modified poly(aspartic acid) (KPAsp) and carboxymethyl chitosan (CMCTS), was prepared in aqueous system. The surface morphology and composition of hydrogels were characterized by SEM and FTIR. The swelling properties of KPAsp, KPAsp/CMCTS semi-IPN and KPAsp/CMCTS IPN hydrogels were investigated and the swelling dynamics of the hydrogels was analyzed based on the Fickian equation. The pH, temperature and salt sensitivities of hydrogels were further studied, and the prepared hydrogels showed extremely sensitive properties to pH, temperature, the ionic salts kinds and concentration. The results of controlled drug release behaviors of the hydrogels revealed that the introduction of IPN observably improved the drug release properties of hydrogels, the release rate of drug from hydrogels can be controlled by the structure of the hydrogels and pH value of the external environment, a relative large amount of drug released was preferred under simulated intestinal fluid. These results illustrated high potential of the KPAsp/CMCTS IPN hydrogels for application as drug carriers.     

Rheological characterization of in situ crosslinkable hydrogels formulated from oxidized dextran and N-carboxyethyl chitosan

The gelation kinetics of an in situ gelable hydrogel formulated from oxidized dextran (Odex) and N-carboxyethyl chitosan (CEC) was investigated rheologically. Both Schiff base mediated chemical and physical crosslinking account for its rapid gelation (30-600 s) between 5 and 37 degrees C. The correlation between gelation kinetics and hydrogel properties with Odex/CEC concentration, their feed ratio, and temperature were elucidated. The gelation time determined from crossing over of storage moduli (G') and loss moduli (G' ') was in good agreement with that deduced from frequency sweeping tests according to the Winter-Chambon power law. The power law exponents for a 2% (w/v) Odex/CEC solution (ratio 5:5) at the gel point was 0.61, which is in excellent agreement with the value predicted from percolation theory (2/3). Temperature dependence of gelation time for the same hydrogel formulation is well-described by an Arrhenius plot with its apparent activation energy calculated at 51.9 kJ/mol.     

Cross-linking and degradation of step-growth hydrogels formed by thiol-ene photoclick chemistry

Thiol-ene photoclick hydrogels have been used for a variety of tissue engineering and controlled release applications. In this step-growth photopolymerization scheme, four-arm poly(ethylene glycol) norbornene (PEG4NB) was cross-linked with dithiol containing cross-linkers to form chemically cross-linked hydrogels. While the mechanism of thiol-ene gelation was well described in the literature, its network ideality and degradation behaviors are not well-characterized. Here, we compared the network cross-linking of thiol-ene hydrogels to Michael-type addition hydrogels and found thiol-ene hydrogels formed with faster gel points and higher degree of cross-linking. However, thiol-ene hydrogels still contained significant network nonideality, demonstrated by a high dependency of hydrogel swelling on macromer contents. In addition, the presence of ester bonds within the PEG-norbornene macromer rendered thiol-ene hydrogels hydrolytically degradable. Through validating model predictions with experimental results, we found that the hydrolytic degradation of thiol-ene hydrogels was not only governed by ester bond hydrolysis, but also affected by the degree of network cross-linking. In an attempt to manipulate network cross-linking and degradation of thiol-ene hydrogels, we incorporated peptide cross-linkers with different sequences and characterized the hydrolytic degradation of these PEG-peptide hydrogels. In addition, we incorporated a chymotrypsin-sensitive peptide as part of the cross-linkers to tune the mode of gel degradation from bulk degradation to surface erosion.     

Thermosensitive hybrid hyaluronan/p(HPMAm-lac)-PEG hydrogels enhance cartilage regeneration in a mouse model of osteoarthritis

Osteoarthritis (OA), due to cartilage degeneration, is one of the leading causes of disability worldwide. Currently, there are not efficacious therapies to reverse cartilage degeneration. In this study we evaluated the potential of hybrid hydrogels, composed of a biodegradable and thermosensitive triblock copolymer cross-linked via Michael addition to thiolated hyaluronic acid, in contrasting inflammatory processes underlying OA. Hydrogels composed of different w/w % concentrations of hyaluronan were investigated for their degradation behavior and capacity to release the polysaccharide in a sustained fashion. It was found that hyaluronic acid was controllably released during network degradation with a zero-order release kinetics, and the release rate depended on cross-link density and degradation kinetics of the hydrogels. When locally administered in vivo in an OA mouse model, the hydrogels demonstrated the ability to restore, to some extent, bone remineralization, proteoglycan production, levels of Sox-9 and Runx-2. Furthermore, the downregulation of proinflammatory mediators, such as TNF-α, NFkB, and RANKL and proinflammatory cytokines was observed. In summary, the investigated hydrogel technology represents an ideal candidate for the potential encapsulation and release of drugs relevant in the field of OA. In this context, the hydrogel matrix could act in synergy with the drug, in reversing phenomena of inflammation, cartilage disruption, and bone demineralization associated with OA.     

Photopolymerized water-soluble chitosan-based hydrogel as potential use in tissue engineering

Novel biodegradable hydrogels by photocrosslinking macromers based on chitosan derivative are reported. Photocrosslinkable macromers, a water-soluble (methacryloyloxy) ethyl carboxyethyl chitosan were prepared by Michael-addition reaction between chitosan and ethylene glycol acrylate methacrylate. The macromers were characterized by Fourier transform infrared spectroscopy, (1)H NMR and (13)C NMR. Hydrogels were fabricated by exposing aqueous solutions of macromers with 0.1% (w/v) photoinitiator to UV light irradiation, and their swelling kinetics as well as degradation behaviors was evaluated. The results demonstrated that the degradation rates were affected strongly by crosslinking density. The hydrogel was compatible to Vero cells, not exhibiting significant cytotoxicity. Cell culture assay also demonstrated that the hydrogels were good in promoting the cell attachment and proliferation, showing their potential as tissue engineering scaffolds.     

Synthesis and degradation test of hyaluronic acid hydrogels

Hyaluronic acid (HA) hydrogels prepared with three different crosslinking reagents were assessed by in vitro and in vivo degradation tests for various tissue engineering applications. Adipic acid dihydrazide grafted HA (HA-ADH) was synthesized and used for the preparation of methacrylated HA (HA-MA) with methacrylic anhydride and thiolated HA (HA-SH) with Traut's reagent (imminothiolane). (1)H NMR analysis showed that the degrees of HA-ADH, HA-MA, and HA-SH modification were 69, 29, and 56 mol%, respectively. HA-ADH hydrogel was prepared by the crosslinking with bis(sulfosuccinimidyl) suberate (BS(3)), HA-MA hydrogel with dithiothreitol (DTT) by Michael addition, and HA-SH hydrogel with sodium tetrathionate by disulfide bond formation. According to in vitro degradation tests, HA-SH hydrogel was degraded very fast, compared to HA-ADH and HA-MA hydrogels. HA-ADH hydrogel was degraded slightly faster than HA-MA hydrogel. Based on these results, HA-MA hydrogels and HA-SH hydrogels were implanted in the back of SD rats and their degradation was assessed according to the pre-determined time schedule. As expected from the in vitro degradation test results, HA-SH hydrogel was in vivo degraded completely only in 2 weeks, whereas HA-MA hydrogels were degraded only partially even in 29 days. The degradation rate of HA hydrogels were thought to be controlled by changing the crosslinking reagents and the functional group of HA derivatives. In addition, the state of HA hydrogel was another factor in controlling the degradation rate. Dried HA hydrogel at 37 degrees C for a day resulted in relatively slow degradation compared to the bulk HA hydrogel. There was no adverse effect during the in vivo tests.     

Preparation and characterization of konjac glucomannan–poly(acrylic acid) IPN hydrogels for controlled release

In this paper, we reported the synthesis and properties of interpenetrating polymer network (IPN) hydrogel systems designed for colon targeted drug delivery. The gels were composed of konjac glucomannan (KGM) and cross-linked poly(acrylic acid) (PAA) by N,N-methylene-bis-(acrylamide) (MBAAm). It was possible to modulate the swelling degree of the gels. And the swelling ratio has sensitive respondence to the environmental pH value variation. The degradation tests show that the hydrogels retain the enzymatic degradation character of KGM. In vitro release of model drug VB₁₂ was studied in the presence of Cellulase E0240 in pH 7.4 phosphate buffer at 37 °C. The accumulative release percent of the model drug reached 85.6% after 48 h and the drug release was controlled by the swelling and the degradation of the hydrogels. The results indicated that the IPN hydrogels can be exploited as potential carriers for colon-specific drug delivery.     

New aspects of the formation of physical hydrogels of chitosan in a hydroalcoholic medium

New aspects concerning the mechanism of formation of chitosan physical hydrogels without any cross-linking agent were studied. The gelation took place during the evaporation of a hydroalcoholic solution of chitosan. We first demonstrated that it was possible to form a physical hydrogel from a hydrochloride form of chitosan. Chromatographic methods showed that during the gel formation, when the initial concentration is over C, the critical concentration of chain entanglement, the water and acid used for the solubilization of the polymer were both eliminated. This particular situation contributed to decrease the dielectric constant of the medium and the apparent charge density of chitosan chains, thus inducing the formation of a three-dimensional network through hydrophobic interactions and hydrogen bonding. In the gelation process, this step was kinetically determining. The speed of evaporation of water and acid were determined and different initial conditions were compared. Thus, we investigated the influence of: the initial polymer concentration, the nature of the counterion and the alcohol, the temperature and the geometry of the reactor. Our results allowed us to confirm the existence of a second critical initial concentration C, from which the evaporation of water became more difficult. We suggested that C corresponded to a reorganization of the solution involving the presence of gel precursors. Then, a mechanism of formation of physical hydrogels of chitosan in a hydroalcoholic medium could be proposed. For the first time, we demonstrated that it was possible to generate physical hydrogels in the presence of various diols, which size of the carbonated chain appeared as a limiting factor for the gelation process. These physical hydrogels of chitosan are currently used in our laboratory for tissue engineering in the treatment of third degree burns with the possibility to adapt their mechanical properties from the choice of both the acid or the alcohol used.     

IPNs based on chitosan with NVP and NVP/HEMA synthesised through photoinitiator-free photopolymerisation technique for biomedical applications

Biocompatible interpenetration polymeric network (IPN) hydrogels based on chitosan with N-vinylpyrrolidinone (NVP) as well as its copolymer with 2-hydroxymethyl methacrylate (HEMA) were synthesised using the photopolymerisation technique without the inclusion of any photoinitiator or crosslinking agent. These hydrogels were characterised using the Fourier-transform infrared spectroscopy (FTIR) technique. Equilibrium swelling of these hydrogels was performed in Milli-Q water and drug release studies were carried out using theophylline as the model drug. These tests showed that the IPN comprised of chitosan and NVP with a very small amount of N-hydroxymethyl maleimide (HMMI) included exhibited higher swelling abilities and fast drug release rates than the IPN which contained chitosan, NVP and HEMA. Kinetic studies of water diffusion into these hydrogels and drug release revealed that with the exception of the IPN with HEMA incorporated, the other hydrogels did not adhere to the Fickian diffusion model. These hydrogels were tested for their biocompatibility with human epidermal keratinocyte cells (HaCaT). A positive cell growth as evidenced by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) cell proliferation assay indicated that these hydrogels are non-toxic to human keratinocytes and can be potentially used as biomaterials for biomedical applications.     

The cytoprotection of chitosan based hydrogels in xenogeneic islet transplantation: An in vivo study in streptozotocin-induced diabetic mouse

Immune rejection and scarcity of donor tissues are the restrictions of islets transplantation. In this study, the cytoprotection of chitosan hydrogels in xenogeneic islet transplantation was demonstrated. Wistar rat islets encapsulated in chitosan hydrogels were performed glucose challenge test and live/dead cell staining in vitro. Islets/chitosan hydrogels were transplanted into the renal subcapsular space of diabetic C57BL/6 mice. Non-fasting blood glucose level (NFBG), body weight, intraperitoneal glucose tolerance test (IPGTT), and glucose disappearance rate were determined perioperatively. The serum insulin level was analyzed, and the kidney transplanted with islets/chitosan hydrogels were retrieved for histological examination after sacrifice. The present results showed that islets encapsulated in chitosan hydrogels secreted insulin in response to the glucose stimulation as naked islets with higher cell survival. The NFBG of diabetic mice transplanted with islets/chitosan hydrogels decreased from 487 ± 46 to 148 ± 32 at one day postoperation and maintained in the range of 201 ± 36 mg/dl for four weeks with an increase in body weight. IPGTT showed the glucose disappearance rate of mice transplanted with islets/chitosan hydrogels was significant faster than that of mice transplanted with naked islets; the serum insulin level increased from 0.29 ± 0.06 to 1.69 ± 0.65 μg/dl postoperatively. Histological examination revealed that the islets successfully engrafted at renal subcapsular space with positive insulin staining. The immunostain was negative for neither the T-cell lineages nor the monocyte/macrophages. This study indicates that the chitosan hydrogels deliver and protect encapsulated islets successfully in xenotransplantation.     

壳聚糖基温敏水凝胶的研究进展

壳聚糖是一种由甲壳素脱乙酰化得到的氨基多糖,具有生物相容性、低细胞毒性和可生物降解性等特点。壳聚糖/β-甘油磷酸钠溶液温敏水凝胶在组织工程、药物缓释等领域多有报道,其成胶性能取决于凝胶的组分和浓度。针对单纯壳聚糖水凝胶强度较低、降解较快、药物突释等缺陷,通常对壳聚糖进行改性或引入新材料共混,获得更符合实际需要的壳聚糖基温敏水凝胶。对近年来壳聚糖基水凝胶的研究进展进行综述,包括改性壳聚糖、共混体系等,概述了其在组织工程(软骨、血管、神经修复)、药物缓释(癌症药物缓释、糖尿病治疗)领域中研究和应用的新进展,以期为后续温敏水凝胶的进一步研究提供参考。     

Preparation and characteristics of novel porous hydrogel films based on chitosan and glycerophosphate

In this study, we prepared thermosensitive hydrogels by adding α-β-glycerophosphate (α-β-GP) to chitosan (CS) solutions. Then the hydrogels were dried to form films at room temperature. Scanning electron microscope (SEM) revealed that the hydrogel films had rough surfaces and porous cross-sections. Compared with pure chitosan films, the CS/GP hydrogel films showed better elasticity and lower tensile strength. Contact angle studies indicated that all these materials have good hydrophilicity. The CS/GP hydrogel films exhibited higher protein adsorption against both negatively charged protein (bovine serum albumin) and positively charged protein (lysozyme) than pure chitosan films. The results of MTT assay performed with the extracts of the CS/GP hydrogel films revealed the films had nontoxicity. The mouse embryonic fibroblast cells cultured on the CS/GP hydrogel films had good spreading and no apparent impairment of cell morphology. The results indicated that the CS/GP hydrogel film could be a promising candidate biomaterial for biomedicine applications.     

Synthesis and properties of thermo- and pH-sensitive poly(N-isopropylacrylamide)/polyaspartic acid IPN hydrogels

Various interpenetrating polymer network (IPN) hydrogels with sensitivity to temperature and pH were prepared by introducing the pH-sensitive polymer polyaspartic acid (PASP) hydrogel, into the poly(N-isopropylacrylamide) (PNIPAAm) hydrogel system for the purpose of improving its response rate to temperature. The morphologies and thermal behavior of the prepared IPN hydrogels were studied by both scanning electron microscopy (SEM) and differential scanning calorimetry (DSC). The IPN hydrogels showed a large and uneven porous network structure, without showing the common PNIPAAm hydrogel structure. The paper moreover studied their swelling properties, such as temperature dependence of equilibrium swelling ratio, shrinking kinetics, re-swelling kinetics and oscillatory swelling behavior in water. The swelling experiment results revealed that IPN hydrogels exhibited much faster shrinking and re-swelling in function of the composition ratio of the two network components. These fast responsive hydrogels foster potential applications in biomedical and biotechnology fields.     

The Impact of Vehicles on the Mucoadhesive Properties of Orally Administrated Nanoparticles: a Case Study with Chitosan-4-Thiobutylamidine Conjugate

The aim of this study was to evaluate the impact of various vehicles on mucoadhesive properties of thiolated chitosan nanoparticles both in vitro and in vivo. Nanoparticles (NPs) were prepared by in situ gelation technique followed by labeling with fluorescein diacetate. Comparative studies on mucoadhesion were done with these thiolated chitosan NPs and unmodified chitosan NPs (control). The obtained nanoparticles displayed a mean diameter of 164.2 ± 6.9 nm and a zeta potential of 21.5 ± 5 mV. In an in vitro adhesion study, unhydrated thiolated NPs adhered strongly to freshly excised porcine small intestine, which was more than threefold increase compared to the control. In contrast, in the presence of various vehicles (PEG 300, miglyol 840, PEG 6000, cremophor EL, and caprylic triglyceride), the mucoadhesive properties of thiolated NPs were comparatively weak. Thiolated NPs suspended in caprylic triglyceride, for example, had a percent mucoadhesion of 22.50 ± 5.35% on the mucosa. Furthermore, results from in vivo mucoadhesion studies revealed that the dry form of nanoparticles exhibits the strongest mucoadhesion, followed by nanoparticles suspended in PEG 300, miglyol, and 100 mM phosphate buffer, in that order. Three hours after administration, the gastrointestinal residence time of the dry form of thiolated NPs was up to 3.6-fold prolonged. These findings should contribute to the design of highly effective oral mucoadhesive nanoparticulate drug delivery systems.     

Chitosan cross-linking with a water-soluble,blocked diisocyanate. 2. Solvates and hydrogels

A water-soluble, blocked diisocyante was used to cross-link chitosan under various degrees of solvation, including hydration to form hydrogels. Thermal cross-linking of films cast from various amounts of organic cosolvents was found to increase with increased level of cosolvent up to a solvation level of 17% (w/w) and to be more efficient than for films prepared without cosolvent. Rheological studies revealed that gel modulus increased and gel time decreased with increasing cross-linker content and that gelation kinetics were consistent with a process having an activation energy of 103 kJ/mol. Swelling of hydrogels indicated that, even at high levels of hydration, the increased molecular mobility of reactants allowed for efficient network formation in a concentration-dependent manner. The extent of solvation via equilibrium swelling correlated well with degradative properties of chitosan networks in the presence of Chitinase (E. C. 3.2.1.14) from Streptomyces griseus with stability increasing with decreasing swelling (i.e., increased cross-linking).     

Photo-cross-linked PLA-PEO-PLA hydrogels from self-assembled physical networks: mechanical properties and influence of assumed constitutive relationships

Poly(lactide)-block-poly(ethylene oxide)-block-poly(lactide) (PLA-PEO-PLA) triblock copolymers are known to form physical hydrogels in water as a result of the polymer's amphiphilicity. Their mechanical properties, biocompatibility, and biodegradability have made them attractive for use as soft tissue scaffolds. However, the network junction points are not covalently cross-linked, and in a highly aqueous environment these hydrogels adsorb more water, transform from gel to sol, and lose the designed mechanical properties. In this article, a hydrogel was formed by the use of a novel two-step approach. In the first step, the end-functionalized PLA-PEO-PLA triblock was self-assembled into a physical hydrogel through hydrophobic micelle network junctions, and in the second step, this self-assembled physical network structure was locked into place by photo-cross-linking the terminal acrylate groups. In contrast with physical hydrogels, the photo-cross-linked gels remained intact in phosphate-buffered solution at body temperature. The swelling, degradation, and mechanical properties were characterized, and they demonstrated an extended degradation time (approximately 65 days), an exponential decrease in modulus with degradation time, and a tunable shear modulus (1.6-133 kPa). We also discuss the various constitutive relationships (Hookean, neo-Hookean, and Mooney-Rivlin) that can be used to describe the stress-strain behavior of these hydrogels. The chosen model and assumptions used for data fitting influenced the obtained modulus values by as much as a factor of 3.5, which demonstrates the importance of clearly stating one's data fitting parameters so that accurate comparisons can be made within the literature.     

Evaluation of Physical and Mechanical Properties of Porous Poly (Ethylene Glycol)-co-(L-Lactic Acid) Hydrogels during Degradation

Porous hydrogels of poly(ethylene glycol) (PEG) have been shown to facilitate vascularized tissue formation. However, PEG hydrogels exhibit limited degradation under physiological conditions which hinders their ultimate applicability for tissue engineering therapies. Introduction of poly(_L-lactic acid) (PLLA) chains into the PEG backbone results in copolymers that exhibit degradation via hydrolysis that can be controlled, in part, by the copolymer conditions. In this study, porous, PEG-PLLA hydrogels were generated by solvent casting/particulate leaching and photopolymerization. The influence of polymer conditions on hydrogel architecture, degradation and mechanical properties was investigated. Autofluorescence exhibited by the hydrogels allowed for three-dimensional, non-destructive monitoring of hydrogel structure under fully swelled conditions. The initial pore size depended on particulate size but not polymer concentration, while degradation time was dependent on polymer concentration. Compressive modulus was a function of polymer concentration and decreased as the hydrogels degraded. Interestingly, pore size did not vary during degradation contrary to what has been observed in other polymer systems. These results provide a technique for generating porous, degradable PEG-PLLA hydrogels and insight into how the degradation, structure, and mechanical properties depend on synthesis conditions.     

Synthesis of interpenetrating network hydrogel from poly(acrylic acid-co-hydroxyethyl methacrylate) and sodium alginate: Modeling and kinetics study for removal of synthetic dyes from water

Several interpenetrating network (IPN) hydrogels were made by free radical in situ crosslink copolymerization of acrylic acid (AA) and hydroxy ethyl methacrylate in aqueous solution of sodium alginate. N,N′-methylenebisacrylamide (MBA) was used as comonomer crosslinker for making these crosslink hydrogels. All of these hydrogels were characterized by carboxylic content, FTIR, SEM, XRD, DTA–TGA and mechanical properties. Swelling, diffusion and network parameters of the hydrogels were studied. These hydrogels were used for adsorption of two important synthetic dyes, i.e. Congo red and methyl violet from water. Isotherms, kinetics and thermodynamics of dye adsorption by these hydrogels were also studied.