Moist-Retaining, Self-Recoverable, Bioadhesive, and Transparent in Situ Forming Hydrogels To Accelerate Wound Healing.

In the management of accelerating wound healing, moist environments play an important role. Compared with other scaffolds of various forms, hydrogels can maintain a moist environment in the wound area. They are cross-linked hydrophilic polymeric networks that resemble natural soft tissues and extracellular matrices. Among them, injectable hydrogels have attracted great attention in wound repair, as they can be injected into irregular-shaped skin defects and formed in situ to shape the contour of different dimensions. The excellent compliance makes hydrogels easy to adapt to the wound under different conditions of skin movement. Here, we oxidized hydroxyethyl starch (O-HES) and modified carboxymethyl chitosan (M-CMCS) to fabricate an in situ forming hydrogel with excellent self-recoverable extensibility-compressibility, biocompatibility, biodegradability, and transparency for accelerating wound healing. The oxidation degree of O-HES was 74%. The amino modification degree of M-CMCS was 63%. M-CMCS/O-HES hydrogels were formed through the Schiff base reaction. The physicochemical properties of M-CMCS/O-HES hydrogels with various ratios were investigated, and M-CMCS/O-HES hydrogel with a volume ratio of 5:5 exhibited appropriate gelation time, notable water-retaining capacity, self-recoverable conformal deformation, suitable biodegradability, and good biocompatibility for wound-healing application. Then, skin wound-healing experimental studies were carried out in Sprague-Dawley rats with full-thickness skin defects. Significant outcomes were achieved in the M-CMCS/O-HES hydrogel-treated group including higher wound closure percentage, more granulation tissue formation, faster epithelialization, and decreased collagen deposition. These findings demonstrate that using the obtained M-CMCS/O-HES hydrogels is a promising therapeutic strategy for wound healing.

In situ forming hydrogel of natural polysaccharides through Schiff base reaction for soft tissue adhesive and hemostasis.

In this study, a novel injectable hydrogel with biocompatibility and biodegradability through Schiff base reaction was prepared for soft tissue adhesive and hemostasis. Aldehyde hydroxyethyl starch (AHES) was prepared by oxidizing hydroxyethyl starch to get aldehyde groups. Amino carboxymethyl chitosan (ACC) was prepared by grafting ethylenediamine onto carboxymethyl chitosan to get more amino groups. Two-component AHES/ACC hydrogel was formed through Schiff base reaction between aldehyde and amino groups. By changing the reaction conditions various contents of aldehyde and amino group were achieved. The properties of AHES/ACC hydrogel were tunable including gelation time, swelling ratio, degradation and mechanical tensile by varying the content of aldehyde and amino groups. Then biocompatibility measurements showed that AHES/ACC hydrogels supported cell viability and proliferation in vitro and exhibited good biodegradability and biocompatibility in vivo. AHES/ACC hydrogel also had effective hemostatic ability. Thus, this study provides a strategy for the design and fabrication of fast in situ forming hydrogels. Through Schiff base reaction in situ forming hydrogel derived from natural polysaccharides can be modulated and prepared for soft tissue adhesive, hemostasis or other biomedical applications in future.

An in situ forming tissue adhesive based on poly(ethylene glycol)-dimethacrylate and thiolated chitosan through the Michael reaction.

In this paper, a novel biocompatible and biodegradable tissue adhesive composed of poly(ethylene glycol)-methacrylate (PEGDMA) and thiolated chitosan (CSS) was prepared. PEGDMA and CSS cross-linked rapidly under physiological conditions through the Michael addition reaction via UV lamp irradiation. The chemical structures of PEGDMA and CSS were confirmed via FTIR and 1H NMR. The equilibrium swelling ratio and biodegradation of the hydrogels were tunable by varying the component ratios of the hydrogels. The compression strength and adhesive strength of the resulting hydrogels were measured with a tensile tester, and the adhesion strength of the hydrogel was higher than the fibrin glues. Moreover, the cytotoxicity of the PEGDMA/CSS hydrogels for L929 cells was evaluated by the MTT assay, and the results indicate that the photocured hydrogels are biocompatible and less cytotoxic towards the growth of L929 cells. These findings imply that the obtained hydrogel adhesives are a potential bioadhesive for clinical application in the future.

Evaluation of synovium-derived mesenchymal stem cells and 3D printed nanocomposite scaffolds for tissue engineering.

Stem cells and scaffolds play a very important role in tissue engineering. Here, we isolated synovium-derived mesenchymal stem cells (SMSCs) from synovial membrane tissue and characterized stem-cell properties. Gelatin nanoparticles (NP) were prepared using a two-step desolvation method and then pre-mixed into different host matrix (silk fibroin (SF), gelatin (Gel), or SF-Gel mixture) to generate various 3D printed nanocomposite scaffolds (NP/SF, NP/SF-Gel, NP/Gel-1, and NP/Gel-2). The microstructure was examined by scanning electron microscopy. Biocompatibility assessment was performed through CCK-8 assay by coculturing with SMSCs at 1, 3, 7 and 14 days. According to the results, SMSCs are similar to other MSCs in their surface epitope expression, which are negative for CD45 and positive for CD44, CD90, and CD105. After incubation in lineage-specific medium, SMSCs could differentiate into chondrocytes, osteocytes and adipocytes. 3D printed nanocomposite scaffolds exhibited a good biocompatibility in the process of coculturing with SMSCs and had no negative effect on cell behavior. The study provides a strategy to obtain SMSCs and fabricate 3D printed nanocomposite scaffolds, the combination of which could be used for practical applications in tissue engineering.

Fabrication of modified dextran-gelatin in situ forming hydrogel and application in cartilage tissue engineering.

Hydrogels play a very important role in cartilage tissue engineering. Here, we oxidized dextran (Odex) and modified gelatin (Mgel) to fabricate a fast forming hydrogel without the addition of a chemical crosslinking agent. The dynamic gelling process was measured through rheological measurements. The microstructure was examined by lyophilizing to get porous scaffolds. Biological assessment was performed through CCK-8 assays by using synovium-derived mesenchymal cells (SMSCs) at 1, 3, 7 and 14 days. In vivo evaluation for application in cartilage tissue engineering was performed 8 weeks after subcutaneous injection of SMSC-loaded Odex/Mgel hydrogels combined with TGF-ß3 in the dorsa of nude mice. According to the results, a fast forming hydrogel was obtained by simply modifying dextran and gelatin. Moreover, the Odex/Mgel hydrogel exhibited good biocompatibility in cultures of SMSCs and a homogeneous distribution of live cells was achieved inside the hydrogels. After 8 weeks, newly formed cartilage was achieved in the dorsa of nude mice; no inflammatory reaction was observed and high production of GAGs was shown. The method provides a strategy for the design and fabrication of fast in situ forming hydrogels. The Odex/Mgel hydrogel could be used for the regeneration of cartilage in tissue engineering.

Coaxial electrospinning of P(LLA-CL)/heparin biodegradable polymer nanofibers: potential vascular graft for substitution of femoral artery.

Electrospinning is one of the most simple and effective methods to prepare polymer fibers with the diameters ranging from nanometer to several micrometers. Poly(L-lactide)-co-poly (ɛ-caprolactone) (P(LLA-CL)) fibers and P(LLA-CL)/heparin coaxial composite fibers herein were successfully prepared by single electrospinning and coaxial electrospinning, respectively. The prepared endothelialized P(LLA-CL) and P(LLA-CL)/heparin vascular grafts were used in the Beagle dogs experiment to evaluate the feasibility of thus made different scaffolds for substitution of dog femoral artery in early period, medium term, and long term, meanwhile the pure P(LLA-CL) vascular graft was used as the control group during all the experiments. The animal model was established by using the graft materials to anastomose both femoral arteries of dogs. The vascular grafts patency rates (i.e., the unobstructed capacity of blood vessel) were detected by color Doppler flow imaging technology and digital subtraction angiography. To observe the histological morphology at different periods, the vascular grafts were removed after 7, 14, and 30 days, and the corresponding histological changes were evaluated by hematoxylin and eosin staining. The experimental results show that in the early period, the patency rates of pure P(LLA-CL) graft, endothelial P(LLA-CL) graft, and P(LLA-CL)/heparin graft were 75%, 75%, and 100%, respectively; in the medium term, the patency rates of pure P(LLA-CL) graft and endothelial P(LLA-CL) graft were 25%, whereas that of P(LLA-CL)/heparin graft was 50%; the patency rates of pure P(LLA-CL) graft and endothelial P(LLA-CL) graft were down to 0%, whereas the patency rate of P(LLA-CL)/heparin graft was 25% in the long term. This preliminary study has demonstrated that P(LLA-CL)/heparin coaxial composite fiber maybe a reliable artificial graft for the replacement of femoral artery. © 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2013.

The effect of aligned core-shell nanofibres delivering NGF on the promotion of sciatic nerve regeneration.

Recent bioengineering strategies for peripheral nerve regeneration have been focusing on the development of alternative treatments for nerve repair. In this study, we incorporated nerve growth factor (NGF) into aligned core-shell nanofibres by coaxial electrospinning, and reeled the scaffold into aligned fibrous nerve guidance conduits (NGCs) for nerve regeneration study. This aligned PLGA/NGF NGC combined physical guidance cues and biomolecular signals to closely mimic the native extracellular matrix (ECM). The effect of this aligned PLGA/NGF NGC on the promotion of nerve regeneration was evaluated in a 13-mm rat sciatic nerve defect using functional and morphological analysis. After 12 weeks implantation, the results of electrophysiological and muscle weight examination demonstrated that the functional recovery of the regenerated nerve in the PLGA/NGF NGC group was significantly better than that in the PLGA group, yet had no significant difference compared with the autograft group. The toluidine blue staining study showed that more nerve fibres were regenerated in the PLGA/NGF group, while the electron microscopy study indicated that the regenerated nerve in the PLGA/NGF group was more mature than that in the PLGA group. This study demonstrated that the aligned PLGA/NGF could greatly promote peripheral nerve regeneration and have a potential application in nerve regeneration.

Fabrication and intermolecular interactions of silk fibroin/hydroxybutyl chitosan blended nanofibers.

The native extracellular matrix (ECM) is composed of a cross-linked porous network of multifibril collagens and glycosaminoglycans. Nanofibrous scaffolds of silk fibroin (SF) and hydroxybutyl chitosan (HBC) blends were fabricated using 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP) and trifluoroacetic acid (TFA) as solvents to biomimic the native ECM via electrospinning. Scanning electronic microscope (SEM) showed that relatively uniform nanofibers could be obtained when 12% SF was blended with 6% HBC at the weight ratio of 50:50. Meanwhile, the average nanofibrous diameter increased when the content of HBC in SF/HBC blends was raised from 20% to 100%. Fourier transform infrared spectra (FTIR) and (13)C nuclear magnetic resonance (NMR) showed SF and HBC molecules existed in hydrogen bonding interactions but HBC did not induce conformation of SF transforming from random coil form to ß-sheet structure. X-ray diffraction (XRD) confirmed the different structure of SF/HBC blended nanofibers from both SF and HBC. Thermogravimetry-Differential thermogravimetry (TG-DTG) results demonstrated that the thermal stability of SF/HBC blend nanofibrous scaffolds was improved. The results indicated that the rearrangement of HBC and SF molecular chain formed a new structure due to stronger hydrogen bonding between SF and HBC. These electrospun SF/HBC blended nanofibers may provide an ideal tissue engineering scaffold and wound dressing.

Europium-doped amorphous calcium phosphate porous nanospheres: preparation and application as luminescent drug carriers.

Calcium phosphate is the most important inorganic constituent of biological tissues, and synthetic calcium phosphate has been widely used as biomaterials. In this study, a facile method has been developed for the fabrication of amorphous calcium phosphate (ACP)/polylactide-block-monomethoxy(polyethyleneglycol) hybrid nanoparticles and ACP porous nanospheres. Europium-doping is performed to enable photoluminescence (PL) function of ACP porous nanospheres. A high specific surface area of the europium-doped ACP (Eu3+:ACP) porous nanospheres is achieved (126.7 m2/g). PL properties of Eu3+:ACP porous nanospheres are investigated, and the most intense peak at 612 nm is observed at 5 mol% Eu3+ doping. In vitro cytotoxicity experiments indicate that the as-prepared Eu3+:ACP porous nanospheres are biocompatible. In vitro drug release experiments indicate that the ibuprofen-loaded Eu3+:ACP porous nanospheres show a slow and sustained drug release in simulated body fluid. We have found that the cumulative amount of released drug has a linear relationship with the natural logarithm of release time (ln(t)). The Eu3+:ACP porous nanospheres are bioactive, and can transform to hydroxyapatite during drug release. The PL properties of drug-loaded nanocarriers before and after drug release are also investigated.

Cross-linking of gelatin and chitosan complex nanofibers for tissue-engineering scaffolds.

The aim of this study is to investigate cross-linked gelatin-chitosan nanofibers produced by means of electrospinning. Gelatin and chitosan nanofibers were electrospun and then cross-linked by glutaraldehyde (GTA) vapor at room temperature. Scanning electron microscopy (SEM) images showed that the cross-linked mats could keep their nanofibrous structure after being soaked in deionized water at 37° C. The cross-linking mechanism was discussed based on FT-IR results. The two main mechanisms of cross-linking for chitosan and gelatin-chitosan complex are Schiff base reaction and acetalization reaction. For gelatin, the mechanism of cross-linking was Schiff base reaction. The mechanical properties of nanofibrous mats were improved after cross-linking. The biocompatibility of electrospun nanofibrous mats after cross-linking was investigated by the viability of porcine iliac endothelial cells (PIECs). The morphologies of PIECs on the cross-linked nanofibrous mats were observed by SEM. In addition, proliferation of PIECs was tested with the method of methylthiazol tetrazolium (MTT) assay. The results indicate that gelatin-chitosan nanofibrous mats could be a promising candidate for tissue-engineering scaffolds.

Aligned natural-synthetic polyblend nanofibers for peripheral nerve regeneration.

Peripheral nerve regeneration remains a significant clinical challenge to researchers. Progress in the design of tissue engineering scaffolds provides an alternative approach for neural regeneration. In this study aligned silk fibroin (SF) blended poly(L-lactic acid-co-ε-caprolactone) (P(LLA-CL)) nanofibrous scaffolds were fabricated by electrospinning methods and then reeled into aligned nerve guidance conduits (NGC) to promote nerve regeneration. The aligned SF/P(LLA-CL) NGC was used as a bridge implanted across a 10mm defect in the sciatic nerve of rats and the outcome in terms of of regenerated nerve at 4 and 8 weeks was evaluated by a combination of electrophysiological assessment and histological and immunohistological analysis, as well as electron microscopy. The electrophysiological examination showed that functional recovery of the regenerated nerve in the SF/P(LLA-CL) NGC group was superior to that in the P(LLA-CL) NGC group. The morphological analysis also indicated that the regenerated nerve in the SF/P(LLA-CL) NGC was more mature. All the results demonstrated that the aligned SF/P(LLA-CL) NGC promoted peripheral nerve regeneration significantly better in comparison with the aligned P(LLA-CL) NGC, thus suggesting a potential application in nerve regeneration.

Fabrication of chitosan/silk fibroin composite nanofibers for wound-dressing applications.

Chitosan, a naturally occurring polysaccharide with abundant resources, has been extensively exploited for various biomedical applications, typically as wound dressings owing to its unique biocompatibility, good biodegradability and excellent antibacterial properties. In this work, composite nanofibrous membranes of chitosan (CS) and silk fibroin (SF) were successfully fabricated by electrospinning. The morphology of electrospun blend nanofibers was observed by scanning electron microscopy (SEM) and the fiber diameters decreased with the increasing percentage of chitosan. Further, the mechanical test illustrated that the addition of silk fibroin enhanced the mechanical properties of CS/SF nanofibers. The antibacterial activities against Escherichia coli (Gram negative) and Staphylococcus aureus (Gram positive) were evaluated by the turbidity measurement method; and results suggest that the antibacterial effect of composite nanofibers varied on the type of bacteria. Furthermore, the biocompatibility of murine fibroblast on as-prepared nanofibrous membranes was investigated by hematoxylin and eosin (H&E) staining and MTT assays in vitro, and the membranes were found to promote the cell attachment and proliferation. These results suggest that as-prepared chitosan/silk fibroin (CS/SF) composite nanofibrous membranes could be a promising candidate for wound healing applications.

Preparation of core-shell biodegradable microfibers for long-term drug delivery.

A coaxial electrospun technique to fabricate core-shell microfibers (MFs) for drug delivery application is described. In one-step, Paclitaxel (PTX)-loaded poly(L-lactic acid-co-epsilon-caprolactone) (75:25) (P(LLA-CL)(core/shell)) was electrospun into MFs using 2,2,2-trifluoroethanol as the solvent. The physical and chemical properties of electrospun fibers were characterized by various techniques, such as scanning electron microscopy, transmission electron microscopy, X-ray diffractometry, and Fourier-transform infrared. The fiber diameter depended on both the polymer concentration and the flow ratio of PTX to P(LLA-CL). The encapsulation efficiency and in vitro release profile were measured using high performance liquid chromatography methods. PTX released from the MFs in a short burst over 24 h followed by very slow release over the following 60 days. In addition, the cytotoxicity of PTX-loaded P(LLA-CL) MFs was evaluated using 3-[4,5-dimehyl-2-thiazolyl]-2, 5-diphenyl-2H-tetrazolium bromide assay on HeLa cell lines. These results indicate that PTX could be released from P(LLA-CL) fibers in a steady manner and effectively inhibit the activity of HeLa cells.