Influence of radiation crosslinked carboxymethyl-chitosan/gelatin hydrogel on cutaneous wound healing

A series of carboxymethyl chitosan (CM-chitosan) and gelatin hydrogels were prepared by radiation crosslinking. A pre-clinical study was performed by implantation model and full-thickness cutaneous wound model in Sprague–Dawley rats to preliminarily evaluate the biocompatibility, biodegradability and effects on healing. In the implantation test, as a component of the hydrogels, CM-chitosan showed a positive effect on promoting cell proliferation and neovascularization, while gelatin was efficient to stabilize the structure and prolong the degradation time. To evaluate the function on wound healing, the hydrogels were applied to the relatively large full-thickness cutaneous wounds (Φ3.0 cm). Compared with the control groups, the hydrogel group showed significantly higher percentage of wound closure on days 9, 12 and 15 postoperatively, which was consistent with the significantly thicker granulation tissue on days 3 and 6. All results apparently revealed that the radiation crosslinked CM-chitosan/Gelatin hydrogels could induce granulation tissue formation and accelerate the wound healing.     

Electrospun polyvinylidene pyrolidone/gelatin membrane impregnated with silver sulfadiazine as wound dressing for burn treatment

Nanofibrous membranes used for burn treatment have become widely popular due to their large surface area and high porous structure. In this study, electrospinning was used to fabricate a blended nanofibrous membrane of polyvinylidene pyrolidone (PVP) and gelatin, to use as wound dressing. The physical and mechanical properties of this novel membrane were investigated using SEM, FTIR and tensile tests. Results showed that poor mechanical properties of gelatin, which are preferred in medical applications for curing burns as they allow for antigen activity and skin repair, can be enhanced by adding PVP in the solution. Silver sulfadiazine (AgSD), an antibacterial agent, was also impregnated into the PVP/gelatin nanofibrous structure during electrospinning. The membrane thus fabricated showed antibacterial activities against both the Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus. AgSD release behaviour of fabricated samples indicated short-term drug delivery. It was concluded that the proposed drug-loaded membrane can be used as wound dressing, specifically, in treating skin burns.     

Hyaluronic acid and silver sulfadiazine-impregnated polyurethane foams for wound dressing application

Five different kinds of PU foam wound dressings were prepared to investigate their wound healing capability. They include (i) PU+silver sulfadiazine (AgSD), (ii) PU+alginate (Al), (iii) PU+Al+AgSD, (iv) PU+hyaluronic acid (HA), and (v) PU+HA+AgSD. Physical properties and in vitro behaviors of AgSD release and fibroblast adhesion on those dressings were evaluated. From the drug release and fibroblast adhesion studies, it was observed that PU foam impregnated with both HA and AgSD shows good drug release behavior and low adhesion of the cells. Furthermore, the HA and AgSD-containing PU foam showed excellent wound healing effect without any inflammation or yellow cluster. The wound size decreased around 77% after 1 week application of that foam dressing onto a rat skin defect.     

Click chemistry approach for fabricating PVA/gelatin nanofibers for the differentiation of ADSCs to keratinocytes

Every year, millions of people suffer from dermal wounds caused by heat, fire, chemicals, electricity, ultraviolet radiation or disease. Tissue engineering and nanotechnology have enabled the engineering of nanostructured materials to meet the current challenges in skin treatments owing to such rising occurrences of accidental damages, skin diseases and defects. The abundance and accessibility of adipose derived stem cells (ADSCs) may prove to be novel cell therapeutics for skin regeneration. The nanofibrous PVA/gelatin/azide scaffolds were then fabricated by electrospinning using water as solvent and allowed to undergo click reaction. The scaffolds were characterized by SEM, contact angle and FTIR. The cell–scaffold interactions were analyzed by cell proliferation and the results observed that the rate of cell proliferation was significantly increased (P ≤ 0.05) on PVA/gelatin/azide scaffolds compared to PVA/gelatin nanofibers. In the present study, manipulating the biochemical cues by the addition of an induction medium, in combination with environmental and physical factors of the culture substrate by functionalizing with click moieties, we were able to drive ADSCs into epidermal lineage with the development of epidermis-like structures, was further confirmed by the expression of early and intermediate epidermal differentiation markers like keratin and filaggrin. This study not only provides an insight into the design of a site-specific niche-like microenvironment for stem cell lineage commitment, but also sheds light on the therapeutic application of an alternative cell source—ADSCs, for wound healing and skin tissue reconstitution.     

Implantable biodegradable sponges: effect of interpolymer complex formation of chitosan with gelatin on the release behavior of tramadol hydrochloride

The effect of interpolymer complex formation between positively charged chitosan and negatively charged gelatin (Type B) on the release behavior of tramadol hydrochloride from biodegradable chitosan-gelatin sponges was studied. Mixed sponges were prepared by freeze-drying the cross-linked homogenous stable foams produced from chitosan and gelatin solutions where gelatin acts as a foam builder. Generation of stable foams was optimized where concentration, pH of gelatin solution, temperature, speed and duration of whipping process, and, chitosan-gelatin ratio drastically affect the properties and the stability of the produced foams. The prepared sponges were evaluated for their morphology, drug content, and microstructure using scanning electron microscopy, mechanical properties, uptake capacity, drug release profile, and their pharmacodynamic activity in terms of the analgesic effect after implantation in Wistar rats.

It was revealed that whipping 7% (w/w) gelatin solution, of pH 5.5, for 15 min at 25°C with a stirring speed of 1000 rpm was the optimum conditions for stable gelatin foam generation. Moreover, homogenous, uniform chitosan-gelatin foam with small air bubbles were produced by mixing 2.5% w/w chitosan solution with 7% w/w gelatinsolution in 1:5 ratio. Indeed, polyionic complexation between chitosan and gelatin overcame the drawbacks of chitosan sponge mechanical properties where, pliable, soft, and compressible sponge with high fluid uptake capacity was produced at 25°Cand 65% relative humidity without any added plasticizer. Drugreleasestudies showed a successful retardation of the incorporated drug where the t_50% values of the dissolution profiles were 0.55, 3.03, and 4.73 hr for cross-linked gelatin, un-cross-linked chitosan-gelatin, and cross-linked chitosan-gelatin sponges, respectively. All the release experiments followed Higuchi's diffusion mechanism over 12 hr. The achieved drug prolongation was a result of a combined effect of both cross-linking and polyelectrolyte complexation between chitosan and gelatin. The analgesic activity of the implanted tramadol hydrochloride mixed chitosan-gelatin sponge showed reasonable analgesic effect that was maintained for more than 8 hr. Therefore, the use of chitosan and gelatin together appears to allow the formulator to manipulate both the drug release profiles and the mechanical properties of the sponge that could be effectively implanted.     

Effect of pore size and cross-linking of a novel collagen-elastin dermal substitute on wound healing

Collagen-elastin (CE) scaffolds are frequently used for dermal replacement in the treatment of full-thickness skin defects such as burn wounds. But little is known about the optimal pore size and level of cross-linking. Different formulations of dermal substitutes with unidirectional pores were tested in porcine full-thickness wounds in combination with autologous split skin mesh grafts (SSG). Effect on wound healing was evaluated both macro- and microscopically. CE scaffolds with a pore size of 80 or 100 μm resulted in good wound healing after one-stage grafting. Application of scaffolds with a larger average pore size (120 μm) resulted in more myofibroblasts and more foreign body giant cells (FBGC). Moderate crosslinking impaired wound healing as it resulted in more wound contraction, more FBGC and increased epidermal thickness compared to no cross-linking. In addition, take rate and redness were negatively affected compared to SSG only. Vascularization and the number of myofibroblasts were not affected by cross-linking. Surprisingly, stability of cross-linked scaffolds was not increased in the wound environment, in contrast to in vitro results. Cross-linking reduced the proliferation of fibroblasts in vitro, which might explain the reduced clinical outcome. The non-cross-linked CE substitute with unidirectional pores allowed one-stage grafting of SSG, resulting in good wound healing. In addition, only a very mild foreign body reaction was observed. Cross-linking of CE scaffolds negatively affected wound healing on several important parameters. The optimal non-cross-linked CE substitute is a promising candidate for future clinical evaluation.     

Efficacy of chitin-PAA-GTMAC gel in promoting wound healing: animal study

Acrylic grafted chitin (chitin-PAA) was modified with glycidyltrimethylammonium chloride (GTMAC) with the aim of promoting wound healing. The chitin-PAA-GTMAC gels with different GTMAC contents were compared with the original chitin-PAA gel and Intrasite gel for their efficacy in deep wound healing of Wistar rats. Four full-thickness wounds were made on the dorsal skin of rats and then each was treated with 4 materials; chitin-PAA, chitin-PAA-GTMAC(1:4), chitin-PAA-GTMAC(1:10) and Intrasite gel. During 18 days of treatment, the wounds were visually observed and calculated for wound size using image analysis program. Skin wound tissues of sacrificed rats were processed for routine histological observation and immunohistochemistry of proliferating cell nuclear antigen (PCNA). The wounds covered with the chitin derivatives either with or without GTMAC showed a significant reduction in wound size in day 9 in comparison with day 12 for those covered with Intrasite gel. The faster rate and the better pattern of epidermal development observed in histological study as well as the higher dermal cell proliferation (PCNA expression) also demonstrated the better efficiency in wound healing of the chitin derivatives than Intrasite. The earliest epidermal development of the wounds treated with chitin-PAA-GTMAC (1:4) among the tested materials suggested the most promising of this material for the treatment of full-thickness open wound.     

Electrospun gelatin fiber mats containing a herbal-Centella asiatica-extract and release characteristic of asiaticoside

Ultra-fine gelatin (type A, porcine skin, ～180 Bloom) fiber mats containing a methanolic crude extract of Centella asiatica (L.) Urban, a medicinal plant widely known for its traditional medical applications including its wound healing ability, were fabricated, for the first time, from the neat gelatin solution (22% w/v in 70?vol% acetic acid) containing the crude extract (mCA) in various amounts (i.e. 5-30?wt% based on the weight of gelatin powder) by electrospinning. Incorporation of mCA in the neat gelatin solution did not affect both the morphology and the size of the mCA-loaded gelatin fibers, as both of the neat and the mCA-loaded gelatin fibers were smooth and the average diameters of these fibers ranged between 226 and 232?nm. The cross-linked mCA-loaded e-spun gelatin fiber mat from the neat gelatin solution containing 30?wt% of mCA was further investigated for the release characteristic of asiaticoside, identified as the most active compound associated with the healing of wounds, in two different types of releasing medium, i.e. acetate buffer and the buffer containing 10?vol% of methanol, based on the thin-layer chromatography (TLC)-densitometry technique. Based on the unit weight of the actual amount of asiaticoside present in the specimens, the total amount of asiaticoside released from the fiber mat specimens was lower than that from the film counterparts while, based on the unit weight of the specimens, an opposite trend was observed.     

Electrospinning of gelatin and gelatin/poly(l-lactide) blend and its characteristics for wound dressing

Gelatin nanofibres were electrospun from its aqueous acetic acid solution. Electrospinning parameters, such as concentration of aqueous acetic acid and gelatin solutions, electric field and spinning distance, were examined to investigate the electrospinnability of gelatin solutions and the morphology of gelatin nanofibrous mats. Nanofibrous mats from poly(l-lactide) (PLLA) and gelatin/PLLA solutions were obtained. The electrospun mats showed controlled evaporative water loss, promoted fluid drainage ability, and excellent biocompatibility, and therefore have a potential application as wound dressing.     

Nanofibers prepared by needleless electrospinning technology as scaffolds for wound healing

Electrospun gelatin and poly-ε-caprolactone (PCL) nanofibers were prepared using needleless technology and their biocompatibility and therapeutic efficacy have been characterized in vitro in cell cultures and in an experimental model of a skin wound. Human dermal fibroblasts, keratinocytes and mesenchymal stem cells seeded on the nanofibers revealed that both nanofibers promoted cell adhesion and proliferation. The effect of nanofibers on wound healing was examined using a full thickness wound model in rats and compared with a standard control treatment with gauze. Significantly faster wound closure was found with gelatin after 5 and 10 days of treatment, but no enhancement with PCL nanofibers was observed. Histological analysis revealed enhanced epithelialisation, increased depth of granulation tissue and increased density of myofibroblasts in the wound area with gelatin nanofibers. The results show that gelatin nanofibers produced by needleless technology accelerate wound healing and may be suitable as a scaffold for cell transfer and skin regeneration.     

Promotion of fibrovascular tissue ingrowth into porous sponges by basic fibroblast growth factor

Fibrovascular tissue ingrowth into poly(vinyl alcohol) (PVA) sponges of different pore sizes was investigated by incorporating basic fibroblast growth factor (bFGF) into the sponges. The average pore size of PVA sponges used in this study was 30, 60, 110, 250, 350, and 700 m and gelatin microspheres were employed as release carrier of bFGF. The sponges were subcutaneously implanted into the back of mice after incorporating free bFGF or gelatin microspheres containing bFGF into the sponges. Fibrovascular tissue infiltrated with time into the sponge pores and the extent of fibrous tissue ingrowth showed a maximum at a pore size around 250 m 1 and 6 weeks after implantation. Significant promotion of the growth of fibrous tissue by bFGF was observed only at 3 weeks post-implantation (p < 0.05). New capillaries were formed in the tissue at any time, as long as bFGF was given to the sponges. Both empty gelatin microspheres and phosphate buffered solution neither promoted tissue ingrowth nor induced capillary formation in the sponges. It was concluded that bFGF was essential to induce the fibrovascular tissue ingrowth into the pores of PVA sponges. ©2000 Kluwer Academic Publishers     

Silver sulfadiazine loaded chitosan/chondroitin sulfate films for a potential wound dressing application

Silver sulfadiazine (AgSD) loaded chitosan/chondroitin sulfate (CHI/CS) films were formed to be applied as a potential wound dressing material. The liquid uptake capacity of both, CHI/CS and CHI/CS/AgSD, films exhibited a pH-dependent behavior. Tensile tests showed that the amount of CS used to form the films and the further incorporation of AgSD affect the mechanical properties of the films. In vitro AgSD-release assays showed that the CHI/CS mass ratio influences the AgSD release rate. All the investigated CHI/CS/AgSD films sustain the AgSD release up to 96 h at physiological pH. Antibacterial activity and cell viability assays showed that all the CHI/CS/AgSD films have activity against Pseudomonas aeruginosa and Staphylococcus aureus but they were not toxic to Vero cells. The results presented in this work indicate that the CHI/CS/AgSD exhibits potential to be applied as a wound dressing material.     

Moisture-Adaptive Contractile Biopolymer-Derived Fibers for Wound Healing Promotion

The advancement in thermosensitive active hydrogels has opened promising opportunities to dynamic full-thickness skin wound healing. However, conventional hydrogels lack breathability to avoid wound infection and cannot adapt to wounds with different shapes due to the isotropic contraction. Herein, a moisture-adaptive fiber that rapidly absorbs wound tissue fluid and produces a large lengthwise contractile force during the drying process is reported. The incorporation of hydroxyl-rich silica nanoparticles in the sodium alginate/gelatin composite fiber greatly improves the hydrophilicity, toughness, and axial contraction performance of the fiber. This fiber exhibits a dynamic contractile behavior as a function of humidity, generating ≈15% maximum contraction strain or ≈24 MPa maximum isometric contractile stress. The textile knitted by the fibers features excellent breathability and generates adaptive contraction in the target direction during the natural desorption of tissue fluid from the wounds. In vivo animal experiments further demonstrate the advantages of the textiles over traditional dressings in accelerating wound healing.     

Polycaprolactone/gelatin electrospun nanofibres containing biologically produced tellurium nanoparticles as a potential wound dressing scaffold: Physicochemical,mechanical, and biological characterisation

The biologically synthesised tellurium nanoparticles (Te NPs) were applied in the fabrication of Te NP‐embedded polycaprolactone/gelatin (PCL/GEL) electrospun nanofibres and their antioxidant and in vivo wound healing properties were determined. The as‐synthesised nanofibres were characterised using scanning electron microscopy (SEM), energy‐dispersive X‐ray (EDX) spectroscopy and elemental mapping, thermogravimetric analysis (TGA), and Fourier‐transform infrared (FTIR) spectroscopy. The mechanical properties and surface hydrophobicity of scaffolds were investigated using tensile analysis and contact angle tests, respectively. The biocompatibility of the produced scaffolds on mouse embryonic fibroblast cells (3T3) was evaluated using MTT assay. The highest wound healing activity (score 15/19) was achieved for scaffolds containing Te NPs. The wounds treated with PCL/GEL/Te NPs had inflammation state equal to the positive control. Also, the mentioned scaffold represented positive effects on collagen formation and collagen fibre''s horizontalisation in a dose‐dependent manner. The antioxidative potency of Te NP‐containing scaffolds was demonstrated with lower levels of malondialdehyde (MDA) and catalase (∼3 times) and a higher level of glutathione (GSH) (∼2 times) in PCL/GEL/Te NP‐treated samples than the negative control. The obtained results strongly demonstrated the healing activity of the produced nanofibres, and it can be inferred that scaffolds containing Te NPs are suitable for wound dressing.     

Effects of PVA sponge containing chitooligosaccharide in the early stage of wound healing

Poly(vinyl alcohol) (PVA) sponges with different chitooligosaccharide (COS) content were prepared for wound-dressing application. The morphological structure of PVA sponges was observed by scanning electron microscopy. As the concentration of COS-loaded PVA sponge increased, the average pore size of sponge decreased and the release rate of COS from the sponge also slightly decreased. The accelerating effect of the COS-loaded PVA sponges on open wound healing in rats was investigated by macroscopic examination and measurement of wound area. The COS-loaded sponges were found to be very effective as a wound-healing accelerator in the early stage of wound healing. The wound treated with the COS-loaded PVA sponge was almost reepithelialized, granulation tissues in the wound were considerably replaced by fibrosis at 8 days after initial wounding. The COS-loaded PVA sponge was considered to be a suitable wound-healing formulation due to its easy preparation and high effectiveness.     

Preparation of Adrenochrome Hydrogel Patch, Gel, Ointment, and the Comparison of Their Blood Coagulating and Wound Healing Capability

The aim of this study is to make an effective blood coagulant and wound healing agent, which on its topical application on ruptured skin would help in instant coagulation of blood and ongoing healing of wound. The hydrogel has been prepared by mixing 28% w/v gelatin and 21% w/v PVA in distilled water, and heated to 40°C followed by addition of a blood coagulant at a lower temperature. Beeswax, alcohol, liquid paraffin, and adrenochrome were mixed, triturated, and heated accordingly to prepare adrenochrome ointment. Polyvinyl alcohol and glycerin were mixed and heated and the drug was added at a lower temperature, and stored at 4-5°C to form adrenochrome gel. Gelatin alone has cell adhesion property. Adrenochrome is a blood coagulant. Therefore, gelatin with adrenochrome in hydrogel has a synergistic effect in wound healing. To evaluate the efficacy of these three different formulations, incisions were made on the backs of three mice and simultaneously adrenochrome containing hydrogel patch, gel, and ointment were applied on the wound and observed at regular intervals for half an hour to examine the rate of blood coagulation and kept under observation for 2 days to study the rate of wound healing. The efficacy of all these three formulations was compared to appraise the most effective blood coagulating and wound healing agent.     

Evaluation of proanthocyanidin-crosslinked electrospun gelatin nanofibers for drug delivering system

Electrospun nanofibers are excellent candidates for various biomedical applications. We successfully fabricated proanthocyanidin‐crosslinked gelatin electrospun nanofibers. Proanthocyanidin, a low cytotoxic collagen crosslinking reagent, increased the gelatin crosslinking percentage in the nanofibers from 53% to 64%. The addition of proanthocyanidin kept the nanofibers from swelling, and, thus, made the fibers more stable in the aqueous state. The compatibility and the release behavior of the drug in the nanofibers were examined using magnesium ascorbyl phosphate as the model drug. Proanthocyanidin also promoted drug loading and kept the drug release rate constant. These properties make the proanthocyanidin‐crosslinked gelatin nanofibers an excellent material for drug delivery. In the cell culture study, L929 fibroblast cells had a significantly higher proliferation rate when cultured with the gelatin/proanthocyanidin blended nanofibers. This characteristic showed that proanthocyanidin‐crosslinked gelatin electrospun nanofibers could potentially be employed as a wound healing material by increasing cell spreading and proliferation.     

Hydroxyapatite coating of cellulose sponges attracts bone-marrow-derived stem cells in rat subcutaneous tissue

The presence of bone-marrow-derived stem cells was investigated in a wound-healing model where subcutaneously implanted cellulose sponges were used to induce granulation tissue formation. When cellulose was coated with hydroxyapatite (HA), the sponges attracted circulating haemopoietic and mesenchymal progenitor cells more efficiently than uncoated cellulose. We hypothesized that the giant cells/macrophages of HA-coated sponges recognize HA as foreign material, phagocyte or hydrolyse it and release calcium ions, which are recognized by the calcium-sensing receptors (CaRs) expressed on many cells including haemopoietic progenitors. Our results showed, indeed, that the HA-coated sponges contained more CaR-positive cells than untreated sponges. The stem cells are, most probably, responsible for the richly vascularized granulation tissue formed in HA-coated sponges. This cell-guiding property of HA-coated cellulose might be useful in clinical situations involving impaired wound repair.     

Probing the stability of biocompatible sodium hyaluronate/chitosan nanocoatings against changes in salinity and pH

The stability of polyelectrolyte multilayer assemblies was investigated with emphasis on the effects of solution ionic strength, pH, and polymer molecular weight on the film thickness and surface topography. The multilayers consisting of two polysaccharides, the polyanion sodium hyaluronate (HA) and the polycation chitosan (CH) were studied using surface plasmon resonance (SPR) spectroscopy, impedance quartz crystal microbalance (QCM), and atomic force microscopy (AFM). SPR/QCM experiments show that coatings consisting of four HA/CH bilayers assembled at pH 4.5 in the presence of 0.15 M NaCl are stable in NaCl solutions of concentration less than 0.8 M. These multilayers are stable when placed in contact with aqueous solutions ranging in pH from 3.5 to 9. The molecular weight of the polysaccharides has only a marginal effect on the stability of the films in the range explored here (HA: Mn = 360,000 or 31,000 g/mol; CH: Mn = 160,000 or 30,000 g/mol). AFM imaging reveals that different mechanisms may account for the multilayers stability versus salt and pH treatments. While increasing the ionic strength induces reorganization of the surface topography from isolated spherical islets to elongated worm-like features, changes in pH have no appreciable effects on the coating topography prior to complete disintegration.     

Experimental wound dressings of degradable PHA for skin defect repair

The present study reports construction of wound dressing materials from degradable natural polymers such as hydroxy derivatives of carboxylic acids (PHAs) and 3-hydroxybutyrate/4-hydroxybutyrate [P(3HB/4HB)] as copolymer. The developed polymer films and electrospun membranes were evaluated for its wound healing properties with Grafts—elastic nonwoven membranes carrying fibroblast cells derived from adipose tissue multipotent mesenchymal stem cells. The efficacy of nonwoven membranes of P(3HB/4HB) carrying the culture of allogenic fibroblasts was assessed against model skin defects in Wistar rats. The morphological, histological and molecular studies revealed the presence of fibroblasts on dressing materials which facilitated wound healing, vascularization and regeneration. Further it was also observed that cells secreted extracellular matrix proteins which formed a layer on the surface of membranes and promoted the migration of epidermal cells from the neighboring tissues surrounding the wound. The wounds under the P(3HB/4HB) membrane carrying cells healed 1.4 times faster than the wounds under the cell-free membrane and 3.5 times faster than the wounds healing under the eschar (control).The complete wound healing process was achieved at Day 14. Thus the study highlights the importance of nonwoven membranes developed from degradable P(3HB/4HB) polymers in reducing inflammation, enhancing angiogenic properties of skin and facilitating better wound healing process.