Collagen/Chitosan Electrospun Nanofibrous Membrane for Wound Dressing

Collagen(Col)/chitosan(CS)nanofibrous membrane has great potential to be used as wound dressing.However,current Col/CS nanofibrous membrane produced from electrospinning can not offer sufficient mechanical strength for practical applications.Herein,a novel mixed solvent was used to prepare next-generation high-strength Col/CS nanofibrous membrane.Meanwhile,the optimal Col to CS weight ratio was investigated as well.The asproduced membrane was examined by scanning electron microscopy(SEM),attenuated total reflectance Fourier transform infrared spectroscopy(ATR-FTIR),differential scanning calorimetry(DSC),and XF-1A tester to study its morphological,chemical,thermal and mechanical properties.The preliminary results demonstrated that the mechanical properties of Col/CS nanofibrous membranes were enhanced substantially with the increase of CS weight ratios from 0 to 90%and the optimal Col to CS weight ratio was determined to be 1∶1.A promising way was presented to fabricate Col/CS electrospun nanofibrous membrane with sufficient mechanical strength for practical wound dressing applications.

Low-temperature 3D-printed collagen/chitosan scaffolds loaded with exosomes derived from neural stem cells pretreated with insulin growth factor-1 enhance neural regeneration after traumatic brain injury

There are various clinical treatments for traumatic brain injury,including surgery,drug therapy,and rehabilitation therapy;howeve r,the therapeutic effects are limited.Scaffolds combined with exosomes represent a promising but challenging method for improving the repair of traumatic brain injury.In this study,we determined the ability of a novel 3D-printed collagen/chitosan scaffold loaded with exosomes derived from neural stem cells pretreated with insulin-like growth factor-1(3D-CC-INEXOS) to improve traumatic brain injury repair and functional recove ry after traumatic brain injury in rats.Composite scaffolds comprising collagen,chitosan,and exosomes derived from neural stem cells pretreated with insulin-like growth fa ctor-1(INEXOS) continuously released exosomes for 2weeks.Transplantation of 3D-CC-INExos scaffolds significantly improved motor and cognitive functions in a rat traumatic brain injury model,as assessed by the Morris water maze test and modified neurological seve rity scores.In addition,immunofluorescence staining and transmission electron microscopy showed that3D-CC-INExos implantation significantly improved the recove ry of damaged nerve tissue in the injured area.In conclusion,this study suggests that transplanted3D-CC-INExos scaffolds might provide a potential strategy for the treatment of traumatic brain injury and lay a solid foundation for clinical translation.

Optimization of the Process Parameters in the Electrospinning of Collagen/Chitosan/Polyethylene Oxide Biological Dressings

Biological activity dressings on wound healing consist of Prionace glauca cartilage type Ⅱ collagen( CII) cartilage,chitosan and polyethylene oxide( PEO). The dressing fibers are successfully obtained by electrospinning from the acetic acid aqueous solution( 80%, volume fraction). An effective quadric polynomial regression model,including the feed speed( m L/h),the applied voltage( kV) and the nozzle-collector distance( cm),was set up by response surface methodology. By using the optimized process parameters( applied voltage 20 kV,nozzle-collector distance 16. 6 cm and feed speed 0. 70 m L/h),the mean diameter( 230. 34 nm) was reduced by about 10% compared with the original one.

Activation of adult endogenous neurogenesis by a hyaluronic acid collagen gel containing basic fibroblast growth factor promotes remodeling and functional recovery of the injured cerebral cortex

The presence of endogenous neural stem/progenitor cells in the adult mammalian brain suggests that the central nervous system can be repaired and regenerated after injury.However,whether it is possible to stimulate neurogenesis and reconstruct cortical layers II to VI in non-neurogenic regions,such as the cortex,remains unknown.In this study,we implanted a hyaluronic acid collagen gel loaded with basic fibroblast growth factor into the motor cortex immediately following traumatic injury.Our findings reveal that this gel effectively stimulated the proliferation and migration of endogenous neural stem/progenitor cells,as well as their differentiation into mature and functionally integrated neurons.Importantly,these new neurons reconstructed the architecture of cortical layers II to VI,integrated into the existing neural circuitry,and ultimately led to improved brain function.These findings offer novel insight into potential clinical treatments for traumatic cerebral cortex injuries.

A proteomics study to explore the role of adsorbed serum proteins for PC12 cell adhesion and growth on chitosan and collagen/chitosan surfaces

The aim of this article is to apply proteomics in the comparison of the molecular mechanisms of PC12 cell adhesion and growth mediated by the adsorbed serum proteins on the surfaces of chitosan and collagen/chitosan films.First,the chitosan and the collagen/chitosan films were prepared by spin coating;and their surface morphologies were characterized by scanning electron microscopy,X-ray energy dispersive spectroscopy,contact angle measurement and Fourier transform infrared spectroscopy.Subsequently,cell proliferation experiments on two materials were performed and the dynamic curves of protein adsorption on their surfaces were measured.Then,proteomics and bioinformatics were used to analyze and compare the adsorbed serum proteins on the surfaces of two biomaterials;and their effects on cell adhesion were discussed.The results showed that the optimum concentration of chitosan film was 2%w/v.When compared with chitosan film,collagen/chitosan film promoted the growth and proliferation of PC12 cells more significantly.Although the dynamic curves showed no significant difference in the total amount of the adsorbed proteins on both surfaces,proteomics and bioinformatics analyses revealed a difference in protein types:the chitosan surface adsorbed more vitronectin whereas collagen/chitosan surface adsorbed more fibronectin 1 and contained more cell surface receptor binding sites and more Leu-Asp-Val sequences in its surface structure;the collagen/chitosan surface were more conducive to promoting cell adhesion and growth.

Integrated transcriptomic and proteomic study on the different molecular mechanisms of PC12 cell growth on chitosan and collagen/chitosan films

The purpose of this article is to integrate the transcriptomic analysis and the proteomic profiles and to reveal and compare the different molecular mechanisms of PC12 cell growth on the surface of chitosan films and collagen/chitosan films.First,the chitosan films and the collagen/chitosan films were prepared.Subsequently,the cell viability assay was performed;the cell viability of the PC12 cells cultured on the collagen/chitosan films for 24 h was significantly higher than that on the chitosan films.Then,with cDNA microarray,the numbers of differentially expressed genes of PC12 cells on the surface of chitosan and collagen/chitosan films were 13349 and 5165,respectively.Next,the biological pathway analysis indicated that the differentially expressed genes were involved in 40 pathways directly related to cell adhesion and growth.The integrated transcriptomic and our previous proteomic analysis revealed that three biological pathways-extracellular matrix-receptor interaction,focal adhesion and regulation of actin cytoskeleton-were regulated in the processes of protein adsorption,cell adhesion and growth.The adsorbed proteins on the material surfaces further influenced the expression of important downstream genes by regulating the expression of related receptor genes in these three pathways.In comparison,chitosan films had a strong inhibitory effect on PC12 cell adhesion and growth,resulting in the significantly lower cell viability on its surface;on the contrary,collagen/chitosan films were more conducive to promoting PC12 cell adhesion and growth,resulting in higher cell viability.

Collagen-chitosan scaffold impregnated with bone marrow mesenchymal stem cells for treatment of traumatic brain injury

Combinations of biomaterials and cells can effectively target delivery of cells or other therapeutic factors to the brain to rebuild damaged nerve pathways after brain injury.Porous collagen-chitosan scaffolds were prepared by a freeze-drying method based on brain tissue engineering.The scaffolds were impregnated with rat bone marrow mesenchymal stem cells.A traumatic brain injury rat model was established using the 300 g weight free fall impact method.Bone marrow mesenchymal stem cells/collagen-chitosan scaffolds were implanted into the injured brain.Modified neurological severity scores were used to assess the recovery of neurological function.The Morris water maze was employed to determine spatial learning and memory abilities.Hematoxylin-eosin staining was performed to measure pathological changes in brain tissue.Immunohistochemistry was performed for vascular endothelial growth factor and for 5-bromo-2-deoxyuridine(BrdU)/neuron specific enolase and BrdU/glial fibrillary acidic protein.Our results demonstrated that the transplantation of bone marrow mesenchymal stem cells and collagen-chitosan scaffolds to traumatic brain injury rats remarkably reduced modified neurological severity scores,shortened the average latency of the Morris water maze,increased the number of platform crossings,diminished the degeneration of damaged brain tissue,and increased the positive reaction of vascular endothelial growth factor in the transplantation and surrounding areas.At 14 days after transplantation,increased BrdU/glial fibrillary acidic protein expression and decreased BrdU/neuron specific enolase expression were observed in bone marrow mesenchymal stem cells in the injured area.The therapeutic effect of bone marrow mesenchymal stem cells and collagen-chitosan scaffolds was superior to stereotactic injection of bone marrow mesenchymal stem cells alone.To test the biocompatibility and immunogenicity of bone marrow mesenchymal stem cells and collagen-chitosan scaffolds,immunosuppressive cyclosporine was intravenously injected 12 hours before transplantation and 1-5 days after transplantation.The above indicators were similar to those of rats treated with bone marrow mesenchymal stem cells and collagen-chitosan scaffolds only.These findings indicate that transplantation of bone marrow mesenchymal stem cells in a collagen-chitosan scaffold can promote the recovery of neuropathological injury in rats with traumatic brain injury.This approach has the potential to be developed as a treatment for traumatic brain injury in humans.All experimental procedures were approved by the Institutional Animal Investigation Committee of Capital Medical University,China(approval No.AEEI-2015-035)in December 2015.

Comparative study of chitosan/fibroin–hydroxyapatite and collagen membranes for guided bone regeneration in rat calvarial defects: micro-computed tomography analysis

This study aimed to utilize micro-computed tomography （micro-CT） analysis to compare new bone formation in rat calvarial defects using chitosan/fibroin-hydroxyapatite （CFB-HAP） or collagen （Bio-Gide） membranes. Fifty-four （54） rats were studied. A circular bony defect （8 mm diameter） was formed in the centre of the calvaria using a trephine bur. The CFB-HAP membrane was prepared by thermally induced phase separation. In the experimental group （n= 18）, the CFB-HAP membrane was used to cover the bony defect, and in the control group （n= 18）, a resorbable collagen membrane （Bio-Gide） was used. In the negative control group （n= 18）, no membrane was used. In each group, six animals were euthanized at 2, 4 and 8 weeks after surgery. The specimens were then analysed using micro-CT. There were significant differences in bone volume （BV） and bone mineral density （BMD） （P〈O.05） between the negative control group and the membrane groups. However, there were no significant differences between the CFB-HAP group and the collagen group. We concluded that the CFB-HAP membrane has significant potential as a guided bone regeneration （GBR） membrane.

Chitosan-collagen porous scaffold and bone marrow mesenchymal stem cell transplantation for ischemic stroke

In this study, we successfully constructed a composite of bone marrow mesenchymal stem cells and a chitosan-collagen scaffold in vitro, transplanted either the composite or bone marrow mesenchymal stem cells alone into the ischemic area in animal models, and compared their effects. At 14 days after co-transplantation of bone marrow mesenchymal stem cells and the hi- tosan-collagen scaffold, neurological function recovered noticeably. Vascular endothelial growth factor expression and nestin-labeled neural precursor cells were detected in the iscbemic area, surrounding tissue, hippocampal dentate gyrus and subventricular zone. Simultaneously, a high level of expression of glial fibrillary acidic protein and a low level of expression of neuron-spe- cific enolase were visible in BrdU-labeled bone marrow mesenchymal stem cells. These findings suggest that transplantation of a composite of bone marrow mesenchymal stem cells and a chi- tosan-collagen scaffold has a neuroprotective effect following ischemic stroke.

Integrated printed BDNF-stimulated HUCMSCs-derived exosomes/collagen/chitosan biological scaffolds with 3D printing technology promoted the remodelling of neural networks after traumatic brain injury

The restoration of nerve dysfunction after traumatic brain injury(TBI)faces huge challenges due to the limited self-regenerative abilities of nerve tissues.In situ inductive recovery can be achieved utilizing biological scaffolds combined with endogenous human umbilical cord mesenchymal stem cells(HUCMSCs)-derived exosomes(MExos).In this study,brain-derived neurotrophic factor-stimulated HUCMSCs-derived exosomes(BMExos)were composited with collagen/chitosan by 3D printing technology.3D-printed collagen/chitosan/BMExos(3D-CC-BMExos)scaffolds have excellent mechanical properties and biocompatibility.Subsequently,in vivo experiments showed that 3D-CC-BMExos therapy could improve the recovery of neuromotor function and cognitive function in a TBI model in rats.Consistent with the behavioural recovery,the results of histomorphological tests showed that 3D-CC-BMExos therapy could facilitate the remodelling of neural networks,such as improving the regeneration of nerve fibres,synaptic connections and myelin sheaths,in lesions after TBI.

Effect of chitosan/type I collagen/gelatin composites in biocompatibility and nerve repair

Chitosan, collagen I and gelatin were mixed in appropriate quantities to develop a new nerve repair material, with good arrangement and structure, as well as even aperture size. The composite material was sterilized by 60Co irradiation for 24 hours prior to implantation in the right thigh of rats following sciatic nerve damage. Results showed that the material was nontoxic to the kidneys and the liver, and did not induce an inflammatory response in the muscles. The composite material enhanced the recovery of sciatic nerve damage in rats. These experimental findings indicate that the composite material offers good biocompatibility and has a positive effect on injured nerve rehabilitation.

Integrated printed BDNF/collagen/chitosan scaffolds with low temperature extrusion 3D printer accelerated neural regeneration after spinal cord injury

Recent studies have shown that 3D printed scaffolds integrated with growth factors can guide the growth of neurites and promote axon regeneration at the injury site.However,heat,organic solvents or cross-linking agents used in conventional 3D printing reduce the biological activity of growth factors.Low temperature 3D printing can incorporate growth factors into the scaffold and maintain their biological activity.In this study,we developed a collagen/chitosan scaffold integrated with brain-derived neurotrophic factor(3D-CC-BDNF)by low temperature extrusion 3D printing as a new type of artificial controlled release system,which could prolong the release of BDNF for the treatment of spinal cord injury(SCI).Eight weeks after the implantation of scaffolds in the transected lesion of T10 of the spinal cord,3D-CC-BDNF significantly ameliorate locomotor function of the rats.Consistent with the recovery of locomotor function,3D-CC-BDNF treatment could fill the gap,facilitate nerve fiber regeneration,accelerate the establishment of synaptic connections and enhance remyelination at the injury site.

Three-dimensional-printed collagen/chitosan/secretome derived from HUCMSCs scaffolds for efficient neural network reconstruction in canines with traumatic brain injury

The secretome secreted by stem cells and bioactive material has emerged as a promising therapeutic choice for traumatic brain injury(TBI).We aimed to determine the effect of 3D-printed collagen/chitosan/secretome derived from human umbilical cord blood mesenchymal stem cells scaffolds(3D-CC-ST)on the injured tissue regeneration process.3D-CC-ST was performed using 3D printing technology at a low temperature(20C),and the physical properties and degeneration rate were measured.The utilization of low temperature contributed to a higher cytocompatibility of fabricating porous 3D architectures that provide a homogeneous distribution of cells.Immediately after the establishment of the canine TBI model,3D-CC-ST and 3D-CC(3D-printed collagen/chitosan scaffolds)were implanted into the cavity of TBI.Following implantation of scaffolds,neurological examination and motor evoked potential detection were performed to analyze locomotor function recovery.Histological and immunofluorescence staining were performed to evaluate neuro-regeneration.The group treated with 3D-CC-ST had good performance of behavior functions.Implanting 3D-CC-ST significantly reduced the cavity area,facilitated the regeneration of nerve fibers and vessel reconstruction,and promoted endogenous neuronal differentiation and synapse formation after TBI.The implantation of 3D-CC-ST also markedly reduced cell apoptosis and regulated the level of systemic inflammatory factors after TBI.

Chitosan conduits enriched with fibrin-collagen hydrogel with or without adipose-derived mesenchymal stem cells for the repair of 15-mm-long sciatic nerve defect

Hollow conduits of natural or synthetic origins have shown acceptable regeneration results in short nerve gap repair;however,results are still not comparable with the current gold standard technique“autografts”.Hollow conduits do not provide a successful regeneration outcome when it comes to critical nerve gap repair.Enriching the lumen of conduits with different extracellular materials and cells could provide a better biomimicry of the natural nerve regenerating environment and is expected to ameliorate the conduit performance.In this study,we evaluated nerve regeneration in vivo using hollow chitosan conduits or conduits enriched with fibrin-collagen hydrogels alone or with the further addition of adipose-derived mesenchymal stem cells in a 15 mm rat sciatic nerve transection model.Unexpected changes in the hydrogel consistency and structural stability in vivo led to a failure of nerve regeneration after 15 weeks.Nevertheless,the molecular assessment in the early regeneration phase(7,14,and 28 days)has shown an upregulation of useful regenerative genes in hydrogel enriched conduits compared with the hollow ones.Hydrogels composed of fibrin-collagen were able to upregulate the expression of soluble NRG1,a growth factor that plays an important role in Schwann cell transdifferentiation.The further enrichment with adipose-derived mesenchymal stem cells has led to the upregulation of other important genes such as ErbB2,VEGF-A,BDNF,c-Jun,and ATF3.

Biological Feature of Collagen-Chitosan Membrane with Basic Fibroblast Growth Factor for the Culture of Human Fibroblast

The effect of collagen-chitosan membrane with different proportionate collagen and bFGF were investigated for culture human fibroblast. The optimum weight ratio of collagen/chitoson and bFGF were selected. Using culture human fibroblast technologies and cytotoxicity evaluated in vitro, Cell morphology was observed. Experimental results show that collagen-chitosan with bFGF promoted human fibroblast adhesion and supported cell proliferation for a long time. Furthermore collagen-chitosan membrane obviously degrade after 18d when human fibroblast was exhibited fusion spreading, compacting and stabilize. Cytotoxic to human fibroblast was revealed very low . Collagen- chitosan with bFGF should be useful as a tissue engineering biomaterial scaffold for cell culture.

Mechanical Properties of Collagen Biomimetic Films Formed in the Presence of Calcium,Silica and Chitosan

Using eucollagen solutions from ox hide, we cast collagen films to assess the influence of calcium and silica on the reconstitution of the fibrous structure of collagen. The tensile strength and the breaking elongation of the reconstituted collagen films were measured and analysed. Significant differences were observed between reconstituted collagen films with and without calcium and silica. The breaking elongation of the films obtained in the presence of silica was significantly greater, and the degradation was lower than other films of reconstituted collagen. Collagen and chitosan do not exist together as blends in nature, but the specific properties of each may be used to produce in biomimetic way man-made blends with biomedical applications, that confer unique structural, mechanical （detail） and in vivo properties.

Electrospun Nanofibers of Hydroxyapatite / Collagen / Chitosan Promote Osteogenic Differentiation of BMSCs

Bone tissue engineering, aiming at developing bone substitutes for repair and regeneration of bone defects instead of using autologous bone grafts,has attracted wide attention in the field of tissue engineering and regenerative medicine.Developing biomimetic biomaterial scaffolds able to regulate osteogenic differentiation of stem cells could be a promising strategy to improve the therapeutic efficacy.In this study, electrospun composite nanofibers of hydroxyapatite / collagen / chitosan( HAp / Col / CTS)resembling the fibrous nanostructure and constituents of the hierarchically organized natural bone,were prepared to investigate their capacity for promoting bone mesenchymal stem cells( BMSCs)to differentiate into the osteogenic lineage in the absence and presence of the osteogenic supplementation, respectively.Cell morphology,proliferation and quantified specific osteogenic protein expression on the electrospun HAp / Col / CTS scaffolds were evaluated in comparison with different controls including electrospun nanofibrous CTS,HAp / CTS and tissue culture plate.Our results showed that the nanofibrous HAp / Col / CTS scaffolds supported better spreading and proliferation of the BMSCs than other substrates( P < 0.01).Expressions of osteogenesis protein markers,alkaline phosphatase( ALP) and Col,were significantly upregulated on the HAp / Col / CTS than those on the CTS( P < 0.01) and HAp /CTS( P < 0.05) scaffolds in the absence of the osteogenic supplementation.Moreover,presence of osteogenic supplementation also proved to enhance osteogenic differentiation of BMSCs on HAp /Col / CTS scaffolds, indicative of a synergistic effect.This study highlights the potential of BMSCs / HAp / Col / CTS cell-scaffold system for functional bone repair and regeneration applications.

Elaboration of Materials with Functionality Gradients by Assembly of Chitosan-Collagen Microspheres Produced by Microfluidics

Biopolymers extracted from renewable resources like chitosan and collagen exhibit interesting properties for the elaboration of materials designed for tissue engineering applications,among which are their hydrophilicity,biocompatibility and biodegradability.In many cases,functional recovery of an injured tissue or organ requires oriented cell outgrowth,which is particularly critical for nerve regeneration.Therefore,there is a growing interest for the elaboration of materials exhibiting functionalization gradients able to guide cells.Here,we explore an original way of elaborating such gradients by assembling particles from a library of functionalized microspheres.We propose a simple process to prepare chitosan-collagen hybrid microspheres by micro-and milli-fluidics,with adaptable dimensions and narrow size distributions.The adhesion and survival rate of PC12 cells on hybrid microspheres were compared to those on pure chitosan ones.Finally,functionalized microspheres were assembled into membranes exhibiting a functionalization gradient.

Activation of endogenous neurogenesis and angiogenesis by basic fibroblast growth factor-chitosan gel in an adult rat model of ischemic stroke

Attempts have been made to use cell transplantation and biomaterials to promote cell proliferation,differentiation,migration,and survival,as well as angiogenesis,in the context of brain injury.However,whether bioactive materials can repair the damage caused by ischemic stroke by activating endogenous neurogenesis and angiogenesis is still unknown.In this study,we applied chitosan gel loaded with basic fibroblast growth factor to the stroke cavity 7 days after ischemic stroke in rats.The gel slowly released basic fibroblast growth factor,which improved the local microenvironment,activated endogenous neural stem/progenitor cells,and recruited these cells to migrate toward the penumbra and stroke cavity and subsequently differentiate into neurons,while enhancing angiogenesis in the penumbra and stroke cavity and ultimately leading to partial functional recovery.This study revealed the mechanism by which bioactive materials repair ischemic strokes,thus providing a new strategy for the clinical application of bioactive materials in the treatment of ischemic stroke.

Chitosan-based triboelectric materials for self-powered sensing at high temperatures

Although biopolymers have been widely utilized as triboelectric materials for the construction of self-powered sensing systems,the annihilation of triboelectric charges at high temperatures restricts the output signals and sensitivity of the assembled sensors.Herein,a novel chitosan/montmorillonite/lignin(CML)composite film was designed and employed as a tribopositive layer in the assembly of a self-powered sensing system for use under hot conditions(25-70℃).The dense contact surface resulting from the strong intermolecular interaction between biopolymers and nanofillers restrained the volatilization of induced electrons.The optimized CML-TENG delivered the highest open-circuit voltage(V_(oc))of 262 V and maximum instantaneous output power of 429 mW/m^(2).Pristine CH-TENG retained only 39%of its initial Voc at 70℃,whereas the optimized CM_(5)L_(3)-TENG retained 66%of its initial Voc.Our work provides a new strategy for suppressing the annihilation of triboelectric charges at high temperatures,thus boosting the development of self-powered sensing devices for application under hot conditions.