A carboxymethyl chitosan/oxidized hyaluronic acid composite hydrogel dressing loading with stem cell exosome for chronic inflammation wounds healing.

Stem cell exosomes (Exo) play an important role in the transformation of macrophages, but the rapid clearance of Exo in vivo limits their therapeutic effects for chronic inflammation wounds healing. Here, stem cell Exo was isolated and introduced to a composite hydrogel including carboxymethyl chitosan (CMCS) and oxidized hyaluronic acid (OHA) through chemical cross-linking, which formed an Exo-loaded (CMCS/OHA/Exo) hydrogel. The CMCS/OHA/Exo hydrogel exhibited a function of Exo sustained release and an Exo protection within 6 days. This CMCS/OHA/Exo hydrogel was much better than CMCS/OHA hydrogel or Exo solution in macrophage cell phagocytosis, proliferation and migration in vitro, especially, played an obviously positive role in the transformation of macrophages compared with the reference groups. For the treatment of the chronic inflammation wounds in vivo, the CMCS/OHA/Exo hydrogel had the best results at wound heal rate and inhibiting the secretion of inflammatory factors, and it was far superior to reference groups in wound re-epithelization and collagen production. CMCS/OHA/Exo hydrogels can promote Exo release based on hydrogel degradation to regulate macrophages transformation and accelerate chronic wound healing. The study offers a method for preparing Exo-loaded hydrogels that effectively promote the transformation of macrophages and accelerate chronic inflammatory wound healing.

Functional Oxidized Hyaluronic Acid Cross-Linked Decellularized Heart Valves for Improved Immunomodulation, Anti-Calcification, and Recellularization.

Tissue engineering heart valves (TEHVs) are expected to address the limitations of mechanical and bioprosthetic valves used in clinical practice. Decellularized heart valve (DHV) is an important scaffold of TEHVs due to its natural three-dimensional structure and bioactive extracellular matrix, but its mechanical properties and hemocompatibility are impaired. In this study, DHV is cross-linked with three different molecular weights of oxidized hyaluronic acid (OHA) by a Schiff base reaction and presented enhanced stability and hemocompatibility, which could be mediated by the molecular weight of OHA. Notably, DHV cross-linked with middle- and high-molecular-weight OHA could drive the macrophage polarization toward the M2 phenotype in vitro. Moreover, DHV cross-linked with middle-molecular-weight OHA scaffolds are further modified with RGD-PHSRN peptide (RPF-OHA/DHV) to block the residual aldehyde groups of the unreacted OHA. The results show that RPF-OHA/DHV not only exhibits anti-calcification properties, but also facilitates endothelial cell adhesion and proliferation in vitro. Furthermore, RPF-OHA/DHV shows excellent performance under an in vivo hemodynamic environment with favorable recellularization and immune regulation without calcification. The optimistic results demonstrate that OHA with different molecular weights has different cross-linking effects on DHV and that RPF-OHA/DHV scaffold with enhanced immune regulation, anti-calcification, and recellularization properties for clinical transformation.

Protamine-grafted carboxymethyl chitosan based hydrogel with adhesive and long-term antibacterial properties for hemostasis and skin wound healing.

In this study, we developed a tissue-adhesive and long-term antibacterial hydrogel consisting of protamine (PRTM) grafted carboxymethyl chitosan (CMC) (PCMC), catechol groups modified CMC (DCMC), and oxidized hyaluronic acid (OHA), named DCMC-OHA-PCMC. According to the antibacterial experiments, the PCMC-treated groups showed obvious and long-lasting inhibition zones against E. coli (and S. aureus), and the corresponding diameters varied from 10.1 mm (and 15.3 mm) on day 1 to 9.8 mm (and 15.3 mm) on day 7. The DCMC-OHA-PCMC hydrogel treated groups also exhibited durable antibacterial ability against E. coli (and S. aureus), and the antibacterial rates changed from 99.3 ± 0.21 % (and 99.6 ± 0.36 %) on day 1 to 76.2 ± 1.74 % (and 84.2 ± 1.11 %) on day 5. Apart from good mechanical and tissue adhesion properties, the hydrogel had excellent hemostatic ability mainly because of the grafted positive-charged PRTM. As the animal assay results showed, the hydrogel was conducive to promoting the deposition of new collagen (0.84 ± 0.03), the regeneration of epidermis (98.91 ± 6.99 µm) and wound closure in the process of wound repairing. In conclusion, the presented outcomes underline the prospective potential of the multifunctional CMC-based hydrogel for applications in wound dressings.

What Is Wrong with Hyaluronic Acid Chemistry? A 15N/13C Solid-State NMR Re-Evaluation of Its Dopamine Conjugates.

In this work, a systematic 15N/13C solid-state NMR investigation is performed on three dopamine (DA) conjugates of hyaluronic acid, considered in both its native (HA) and NaIO4-oxidized (HAOx) forms. Two of them, here named HAEDC-DA and HAOx-DA, have been previously introduced as covalent conjugates involving DA amine nitrogen: the former by EDC-mediated amide bond formation, and the latter by reaction of the Schiff base with the aldehyde moieties presumed to exist in HAOx. The third conjugate, HA-DA, is reported here for the first time; it is obtained by simply mixing hyaluronan with DA∙HCl at pH 5. The 15N ss-NMR spectra were found to be consistent in all the systems, and the DA molecules were found to be in their charged -NH3+ form, which contradicts the HAEDC-DA/HAOx-DA covalent bonding schemes proposed in the literature. The 13C ss-NMR results add useful new insights into the structure and interaction patterns of the conjugates. All of our findings are relevant for future practical applications, for instance in developing novel HA-based hydrogels. In addition, the present study demonstrates the importance of using the most appropriate analytical tools when investigating composite systems due to the complexity of hyaluronic acid conjugates. Solid-state NMR proved essential to answering the question in the title: actually, there is nothing wrong with hyaluronic acid chemistry; the claimed covalent bonds between DA and the HA(HAOx) chain do not exist in these systems, because the conditions for their formation do not hold in practice.

An injectable and pH-responsive hyaluronic acid hydrogel as metformin carrier for prevention of breast cancer recurrence.

To achieve the pH-responsive release of metformin in tumor acidic microenvironment, we prepared OHA-Met by covalently grafting metformin (Met) onto oxidized hyaluronic acid (OHA) through imine bonds, and then prepared carboxymethyl chitosan (CMCS)/OHA-Met drug loaded hydrogels. The CMCS/OHA-Met hydrogels showed the in-situ injection performance. At pH = 7.4, the cumulative release rate of metformin from CMCS/OHA-Met20 hydrogel was 42.7 ± 2.6 % in 6 h, and the release tended to balance after 72 h. At pH = 5.5, the release kept constant and the cumulative release rate was 79.3 ± 4.7 % at 6 h, showing good pH-responsive behavior. Metformin induced apoptosis of MCF-7 cells through the caspase 3/PARP pathway. CMCS/OHA-Met20 hydrogel could effectively kill MCF-7 cells, while reducing the cytotoxicity of free metformin to L929 cells. In vivo breast cancer recurrence experiments showed CMCS/OHA-Met20 hydrogel could achieve local injection and pH-responsive smart drug delivery at the tumor resection site, inhibiting breast cancer recurrence. Compared with direct administration, CMCS/OHA-Met20 hydrogel reduced the metformin dosage, frequency of administration and systemic side effects.

New Smart Bioactive and Biomimetic Chitosan-Based Hydrogels for Wounds Care Management.

Wound management represents a continuous challenge for health systems worldwide, considering the growing incidence of wound-related comorbidities, such as diabetes, high blood pressure, obesity, and autoimmune diseases. In this context, hydrogels are considered viable options since they mimic the skin structure and promote autolysis and growth factor synthesis. Unfortunately, hydrogels are associated with several drawbacks, such as low mechanical strength and the potential toxicity of byproducts released after crosslinking reactions. To overcome these aspects, in this study new smart chitosan (CS)-based hydrogels were developed, using oxidized chitosan (oxCS) and hyaluronic acid (oxHA) as nontoxic crosslinkers. Three active product ingredients (APIs) (fusidic acid, allantoin, and coenzyme Q10), with proven biological effects, were considered for inclusion in the 3D polymer matrix. Therefore, six API-CS-oxCS/oxHA hydrogels were obtained. The presence of dynamic imino bonds in the hydrogels' structure, which supports their self-healing and self-adapting properties, was confirmed by spectral methods. The hydrogels were characterized by SEM, swelling degree, pH, and the internal organization of the 3D matrix was studied by rheological behavior. Moreover, the cytotoxicity degree and the antimicrobial effects were also investigated. In conclusion, the developed API-CS-oxCS/oxHA hydrogels have real potential as smart materials in wound management, based on their self-healing and self-adapting properties, as well as on the benefits of APIs.

Ultrasound-triggered piezocatalytic composite hydrogels for promoting bacterial-infected wound healing.

Wound healing has become one of the basic issues faced by the medical community because of the susceptibility of skin wounds to bacterial infection. As such, it is highly desired to design a nanocomposite hydrogel with excellent antibacterial activity to achieve high wound closure effectiveness. Here, based on ultrasound-triggered piezocatalytic therapy, a multifunctional hydrogel is designed to promote bacteria-infected wound healing. Under ultrasonic vibration, the surface of barium titanate (BaTiO3, BT) nanoparticles embedded in the hydrogel rapidly generate reactive oxygen species (ROS) owing to the established strong built-in electric field, endowing the hydrogel with superior antibacterial efficacy. This modality shows intriguing advantages over conventional photodynamic therapy, such as prominent soft tissue penetration ability and the avoidance of serious skin phototoxicity after systemic administration of photosensitizers. Moreover, the hydrogel based on N-[tris(hydroxymethyl)methyl]acrylamide (THM), N-(3-aminopropyl)methacrylamide hydrochloride (APMH) and oxidized hyaluronic acid (OHA) exhibits outstanding self-healing and bioadhesive properties able to accelerate full-thickness skin wound healing. Notably, compared with the widely reported mussel-inspired adhesive hydrogels, OHA/THM-APMH hydrogel due to the multiple hydrogen bonds from unique tri-hydroxyl structure overcomes the shortage that catechol groups are easily oxidized, giving it long-term and repeatable adhesion performance. Importantly, this hybrid hydrogel confines BT nanoparticles to wound area and locally induced piezoelectric catalysis under ultrasound to eradicate bacteria, markedly improving the therapeutic biosafety and exhibits great potential for harmless treatment of bacteria-infected tissues.

Extracellular vesicle-loaded hydrogels for tissue repair and regeneration.

Extracellular vesicles (EVs) are a collective term for nanoscale or microscale vesicles secreted by cells that play important biological roles. Mesenchymal stem cells are a class of cells with the potential for self-healing and multidirectional differentiation. In recent years, numerous studies have shown that EVs, especially those secreted by mesenchymal stem cells, can promote the repair and regeneration of various tissues and, thus, have significant potential in regenerative medicine. However, due to the rapid clearance capacity of the circulatory system, EVs are barely able to act persistently at specific sites for repair of target tissues. Hydrogels have good biocompatibility and loose and porous structural properties that allow them to serve as EV carriers, thereby prolonging the retention in certain specific areas and slowing the release of EVs. When EVs are needed to function at specific sites, the EV-loaded hydrogels can stand as an excellent approach. In this review, we first introduce the sources, roles, and extraction and characterization methods of EVs and describe their current application status. We then review the different types of hydrogels and discuss factors influencing their abilities to carry and release EVs. We summarize several strategies for loading EVs into hydrogels and characterizing EV-loaded hydrogels. Furthermore, we discuss application strategies for EV-loaded hydrogels and review their specific applications in tissue regeneration and repair. This article concludes with a summary of the current state of research on EV-loaded hydrogels and an outlook on future research directions, which we hope will provide promising ideas for researchers.

Oxidized Hyaluronic Acid-Gelatin-Based Hydrogels for Tissue Engineering and Soft Tissue Mimicking.

Hydrogels are ideal materials for mimicking and engineering soft tissue. Hyaluronic acid is a linear polysaccharide native to the human extracellular matrix. In this study, we first develop and characterize two hydrogel compositions built from oxidized HA and gelatin with and without alginate-di-aldehyde (ADA) crosslinked by ionic and enzymatic agents with potential applications in soft tissue engineering and tissue mimicking structures. The stability under incubation conditions was improved by adjusting crosslinking times. Through large-strain mechanical measurements, the hydrogels' properties were compared to human brain tissue and the samples containing ADA revealed similar mechanical properties to the native tissue specimens in cyclic compression-tension. In vitro characterization demonstrated a high viability of encapsulated mouse embryonic fibroblasts and a spreading of the cells in case of ADA-free samples. Impact statement Brain mimicking materials are required in several medical and industrial fields for the development of safety gear, testing of medical imaging techniques, surgical training, tissue engineering, and modeling of the mechanical behavior of tissues. The materials must resemble the microstructure, chemistry, and mechanical properties of the native tissue extracellular matrix while being adjustable in degradation to suit the various applications. In this article, different methods are used to evaluate a novel hydrogel material and its suitability as brain mimicking matrix.

Construction of Konjac Glucomannan/Oxidized Hyaluronic Acid Hydrogels for Controlled Drug Release.

Konjac glucomannan (KGM) hydrogel has favorable gel-forming abilities, but its insufficient swelling capacity and poor control release characteristics limit its application. Therefore, in this study, oxidized hyaluronic acid (OHA) was used to improve the properties of KGM hydrogel. The influence of OHA on the structure and properties of KGM hydrogels was evaluated. The results show that the swelling capacity and rheological properties of the composite hydrogels increased with OHA concentration, which might be attributed to the hydrogen bond between the KGM and OHA, resulting in a compact three-dimensional gel network structure. Furthermore, epigallocatechin gallate (EGCG) was efficiently loaded into the KGM/OHA composite hydrogels and liberated in a sustained pattern. The cumulative EGCG release rate of the KGM/OHA hydrogels was enhanced by the increasing addition of OHA. The results show that the release rate of composite hydrogel can be controlled by the content of OHA. These results suggest that OHA has the potential to improve the properties and control release characteristics of KGM hydrogels.

New injectable chitosan-hyaluronic acid based hydrogels for hemostasis and wound healing.

It is a challenge to develop hemostatic and wound dressings that are used for irregular shape and deep wound. Herein, a series of novel N-succinyl chitosan-oxidized hyaluronic acid based (NSC-OHA-based) hydrogels were fabricated, while calcium ions (Ca2+) and/or four-armed amine-terminated poly(ethylene glycol) (4-arm-PEG-NH2, labeled as PEG1) were introduced to regulate the mechanical behavior and bioactivities. We found all NSC-OHA-based hydrogels displayed self-healing and injectable performances. Besides, the addition of Ca2+ or PEG1 exhibited a positive effect on the adjustable mechanical behavior of hydrogels, providing the possibility to meet different mechanical requirements. Furthermore, Ca2+ or PEG1 significantly improved the biocompatibility, hemostasis and wound healing abilities of NSC-OHA hydrogel. Notably, compared with the commercial hemostatic agent (Arista™), hydrogels containing Ca2+ showed comparable hemostatic effects and significantly accelerated wound healing. Overall, the calcium-containing NSC-OHA hydrogels are promising for hemostasis and accelerating wound healing.

Self-healing/pH-responsive/inherently antibacterial polysaccharide-based hydrogel for a photothermal strengthened wound dressing.

Hydrogels have been used widely as wound dressings for maintaining a moist environment to rapid wound healing. However, the lack of antibacterial effect limits their further applications. Herein, we developed a polysaccharide-based hydrogel that can achieve photothermal-assisted bacterial inactivation. The hydrogel has inherent antibacterial activity due to introduction of quaternised chitosan (QCS) with a protonated amine group-modified hydrophilic polycationic structure. Under near infrared (NIR) irradiation, the antibacterial effect of the hydrogel was significantly improved because thermal ablation could also combat bacteria. The hydrogel showed self-healing property through reversible Schiff base bonds between QCS and oxidized hyaluronic acid (OHA). The hydrogel is also pH-sensitive to release drugs in acidic wound. The full-thickness skin defect model showed it had a promoting effect on wound healing. Overall, we provide a theoretical basis for a promising wound dressing based on a photothermally improved polysaccharide-based hydrogel with self-healing/pH-responsive/inherently antibacterial capacity.

Feasibility study of oxidized hyaluronic acid cross-linking acellular bovine pericardium with potential application for abdominal wall repair.

Bovine pericardium(BP)is one of the biological membranes with extensive application in tissue engineering. To fully investigate the potential clinical applications of this natural biological material, a suitable cross-linking reagent is hopefully adopted for modification. Glutaraldehyde (GA) is a clinically most common synthetic cross-linking reagent. In the study, oxidized hyaluronic acid (AHA) was developed to substitute GA to fix acellular bovine pericardium (ABP) for lower cytotoxicity, aiming to evaluate the feasibility of AHA as a cross-linking reagent and develop AHA-fixed ABP as a biological patch for abdominal wall repair. The AHA with the feeding ratio (1.8:1.0) has an appropriate molecular weight and oxidation degree, almost no cytotoxicity and good cross-linking effect. The critical cross-linking characteristics and cytocompatibility of AHA-fixed ABP were also investigated. The results demonstrated that 2.0% AHA-fixed ABP had the most suitable mechanical properties, thermal stability, resistance to enzymatic degradation and hydrophilicity. Moreover, 2.0% AHA-fixed samples exhibited an excellent cytocompatibility with human peritoneal mesothelial cells (HPMC) and low antigenicity. It also showed a prominent anti-calcification ability required for abdominal wall repair. Our data provided experimental basis for future research on AHA as a new cross-linking reagent and AHA-fixed ABP for abdominal wall repair.

3D Printing of Collagen/Oligomeric Proanthocyanidin/Oxidized Hyaluronic Acid Composite Scaffolds for Articular Cartilage Repair.

Articular cartilage defects affect millions of people worldwide, including children, adolescents, and adults. Progressive wear and tear of articular cartilage can lead to progressive tissue loss, further exposing the bony ends and leaving them unprotected, which may ultimately cause osteoarthritis (degenerative joint disease). Unlike other self-repairing tissues, cartilage has a low regenerative capacity; once injured, the cartilage is much more difficult to heal. Consequently, developing methods to repair this defect remains a challenge in clinical practice. In recent years, tissue engineering applications have employed the use of three-dimensional (3D) porous scaffolds for growing cells to regenerate damaged cartilage. However, these scaffolds are mainly chemically synthesized polymers or are crosslinked using organic solvents. Utilizing 3D printing technologies to prepare biodegradable natural composite scaffolds could replace chemically synthesized polymers with more natural polymers or low-toxicity crosslinkers. In this study, collagen/oligomeric proanthocyanidin/oxidized hyaluronic acid composite scaffolds showing high biocompatibility and excellent mechanical properties were prepared. The compressive strengths of the scaffolds were between 0.25-0.55 MPa. Cell viability of the 3D scaffolds reached up to 90%, which indicates that they are favorable surfaces for the deposition of apatite. An in vivo test was performed using the Sprague Dawley (SD) rat skull model. Histological images revealed signs of angiogenesis and new bone formation. Therefore, 3D collagen-based scaffolds can be used as potential candidates for articular cartilage repair.

Glycol chitosan/oxidized hyaluronic acid hydrogel film for topical ocular delivery of dexamethasone and levofloxacin.

In the present study, we fabricated a glycol chitosan/oxidized hyaluronic acid hydrogel film with promising potential for the dual ophthalmic delivery of dexamethasone (Dex) and levofloxacin (Lev). Utilizing different oxidation degrees of oxidized hyaluronic acid (OHA), several blank hydrogel films and Lev-loaded hydrogel films were successfully fabricated. With an increase in the oxidation degree of OHA, the swelling ratio of the hydrogel films decreased accordingly. The hydrogel films displayed a stepwise release of Lev and Dex, with Lev rapidly released from the hydrogel film, followed by a sustained release of Dex. Lev-loaded hydrogel films revealed a potent capacity to inhibit bacterial growth in different bacterial strains. In lipopolysaccharide-activated RAW264.7 macrophages, the formulated hydrogel films displayed potent in vitro anti-inflammatory activity by significantly downregulating various inflammatory cytokines. Overall, the fabricated hydrogel film acting as a dual drug delivery system might be a promising vehicle for the treatment of postoperative endophthalmitis.

Study of injectable Blueberry anthocyanins-loaded hydrogel for promoting full-thickness wound healing.

Injectable hydrogels with high anti-inflammatory and wound-healing properties are highly desirable for clinical application. In the present study, injectable hydrogels were prepared based on carboxymethyl chitosan and oxidized hyaluronic acid. Blueberry anthocyanins (BA), which are known for their antioxidant and antiinflammatory properties, were successfully loaded into the hydrogels. The gelation kinetics and mechanical properties of the hydrogels were investigated. Oxidized hyaluronic acid with an oxidation degree of 38.1% conferred a suitable gelation time (~70 s) and mechanical properties (76.0 kPa compression stress at strain of 80%) of the hydrogel. The injectable BA-loaded hydrogel significantly accelerated the wound healing process in a full-thickness skin wound model in rats, promoted epithelial and tissue regeneration, exerted antiinflammatory effects, and promoted collagen deposition and angiogenesis. Besides, the hydrogel could upregulate the expression of VEGF and IL-10 proteins, downregulate the NF-κB level, and promote macrophage transformation from M1 phenotype to M2. The promotion of the BA-loaded hydrogel on wound healing were mainly realized by its biological effects, including antioxidant and anti-inflammatory effects, and regulation of various wound healing related factors. The results suggested that BA and the hydrogels exert synergistic effects in promoting wound healing. Injectable BA-loaded hydrogels appear to be promising candidates for wound healing application.

Hyaluronic acid targeted and pH-responsive nanocarriers based on hollow mesoporous silica nanoparticles for chemo-photodynamic combination therapy.

In this work, a pH-responsive and tumor targeted multifunctional drug delivery system (RB-DOX@HMSNs-N = C-HA) was designed to realize chemo-photodynamic combination therapy. Hollow mesoporous silica nanoparticles (HMSNs) was served as the host material to encapsulate doxorubicin (DOX) and photosensitizer rose bengal (RB). Hyaluronic acid (HA) was modified on the surface of HMSNs via pH-sensitive Schiff base bonds as gatekeeper as well as targeted agent. Characterization results indicated the successful preparation of HMSNs-N = C-HA with appropriate diameter of 170 nm around and the nanocarriers displayed superior drug loading capacity (15.30 % for DOX and 12.78 % for RB). Notably, the results of in vitro drug release experiments confirmed that the system possessed good pH-sensitivity, which made it possible to release cargoes in slight acid tumor micro-environments. Significantly, the in vitro cell uptake and cytotoxicity assay results fully proved that RB-DOX@HMSNs-N = C-HA could precisely target murine mammary carcinoma (4T1) cells and effectively inhibit tumor cells viability with chemo-photodynamic synergistic therapy. Overall, our work (RB-DOX@HMSNs-N = C-HA) provides an efficient approach for the development of chemo-photodynamic combination therapy.

Ocular biocompatibility of gelatin microcarriers functionalized with oxidized hyaluronic acid.

Given that the presence of aldehyde groups on the oxidized sugar residues may pose toxicity concerns, it is necessary to examine the safety of gelatin microcarriers (GMC) functionalized with oxidized hyaluronic acid (oHA) for potential ophthalmic applications. In this study, the ocular biocompatibility of biopolymer microcarriers was investigated in vitro using primary rabbit corneal cell cultures and in vivo using the anterior chamber of the rabbit eye model. Our results showed that different types of corneal cells including epithelial, stromal, and endothelial cells remain viable and actively proliferate following 2 and 4days of exposure to test materials. In addition, similar interleukin-6 gene expression levels and comet tail lengths were seen in the presence and absence of biopolymer microcarriers, suggesting no cellular inflammation and genotoxicity. After 7 and 14days of intracameral injection in the rabbit eyes, both the GMC samples and their counterparts functionalized with oHA were well tolerated in the ocular anterior chamber as demonstrated by slit-lamp biomicroscopy. Clinical observations including specular microscopic examinations, corneal topography, and corneal thickness measurements also showed that the rabbits bearing biopolymer microcarriers exhibit no signs of corneal edema and astigmatism as well as endothelial damage, indicating the absence of tissue response. It is concluded that the GMC materials functionalized with oHA (oxidation level: 10.4±0.9%) are compatible toward corneal cells and ocular anterior segment tissues at a concentration of 10mg/ml. The information about the effect of coupling of aldehyde-functionalized HA to gelatin on in vitro and in vivo biocompatibility of biopolymer composites can be used as further development of corneal stromal cell microcarriers for tissue engineering applications.

Biofunctionalization of gelatin microcarrier with oxidized hyaluronic acid for corneal keratocyte cultivation.

Development of microcarriers as alternative architecture models to traditional three-dimensional porous scaffolds can potentially eliminate the concern of extensive geographic necrosis during keratocyte cultivation. For the first time, this work was performed to evaluate the in vitro expansion of rabbit corneal keratocytes (RCKs) on the gelatin microcarrier (GMC) modified with hyaluronic acid (HA). In order to confirm the successful synthesis, the oxidized HA (oHA) samples following treatment of HA with sodium periodate were investigated by chemical characterization, including Fourier transform infrared and nuclear magnetic resonance spectroscopy. Biofunctionalization of GMC with oHA was then achieved by the formation of aldimine linkage between free amino groups in protein and aldehyde groups in oxidized polysaccharide. The grafting yield of oHA onto GMC determined by alcian blue staining assay was 26.9±0.1%. Tissue culture polystyrene (TCPS) plates and the microcarriers from either the GMC or GMC-oHA group were used for suspension culture of RCKs. Results of WST-1 assays, scanning electron microscopy, extracellular matrix production assays, and quantitative real-time reverse transcription polymerase chain reaction and Western blot analysis showed that among all the groups studied, the GMC-oHA samples could most effectively support a large scale growth of RCKs while enhancing their total collagen and glycosaminoglycan contents (i.e., indicative of biosynthetic capacity) and keratocan and ALDH1 gene and protein expressions (i.e., indicative of cell phenotype and function). It is concluded that the aforementioned keratocyte behaviors are found to be strongly dependent on the type of culture substrate (i.e., TCPS plate versus biopolymer microcarrier) and on the biomaterial functionalization (i.e., modification of gelatin with oHA).

Using casein and oxidized hyaluronic acid to form biocompatible composite hydrogels for controlled drug release.

To develop biocompatible polymeric hydrogels for the in-situ encapsulation and controlled release of hydrophilic drugs, the oxidized hyaluronic acid containing aldehyde groups was prepared by the reaction between hyaluronic acid and sodium periodate, and then used for the first time to crosslink casein protein in aqueous system. By changing its aldehyde group content or amount, we found that the gelation kinetics and the properties of resultant composite hydrogel could be modulated. Particularly, an increase of its aldehyde group content or amount was found to result in a shorten gelation time, an enhanced gel strength, a reduced swelling ratio and a prolonged drug release. In addition, the as obtained composite hydrogel was also evaluated for its in vitro cytotoxicity on L929 mouse fibroblast cells and was confirmed to have a good biocompatibility.