Polysaccharide nanofibrous scaffolds as a model for in vitro skin tissue regeneration

Tissue engineering and nanotechnology have advanced a general strategy combining the cellular elements of living tissue with sophisticated functional biocomposites to produce living structures of sufficient size and function at a low cost for clinical relevance. Xylan, a natural polysaccharide was electrospun along with polyvinyl alcohol (PVA) to produce Xylan/PVA nanofibers for skin tissue engineering. The Xylan/PVA glutaraldehyde (Glu) vapor cross-linked nanofibers were characterized by SEM, FT-IR, tensile testing and water contact angle measurements to analyze the morphology, functional groups, mechanical properties and wettability of the fibers for skin tissue regeneration. The cell-biomaterial interactions were studied by culturing human foreskin fibroblasts on Xylan/PVA Glu vapor cross-linked and Xylan/PVA/Glu blend nanofibrous scaffolds. The observed results showed that the mechanical properties (72 %) and fibroblast proliferation significantly increased up to 23 % (P < 0.05) in 48 h Glu vapor cross-linked nanofibers compared to 24 h Glu vapor cross-linked Xylan/PVA nanofibers. The present study may prove that the natural biodegradable Xylan/PVA nanofibrous scaffolds have good potential for fibroblast adhesion, proliferation and cell matrix interactions relevant for skin tissue regeneration.     

Isolation of nanofibers from soybean source and their reinforcing capability on synthetic polymers

The focus of this work is to study nanofibers in three different polymers: polyvinyl alcohol (PVA), polypropylene (PP) and polyethylene (PE). The nanofibers were isolated from a soybean source by combining chemical and mechanical treatments. Isolated nanofibers were shown to have diameter between 50 and 100 nm and the length in micrometer scale which results in very high aspect ratio. The mechanical properties demonstrated an increase in tensile strength from 21 MPa of PVA/UNF5 (untreated-fiber (5 wt%) reinforced PVA) and 65 MPa of pure PVA to 103 MPa of PVA/SBN5 (nanofiber (5 wt%) reinforced PVA). The increased stiffness of PVA/SBN5 nanocomposites was also very promising; it was 6.2 GPa compared to 2.3 GPa of pure PVA and 1.5 GPa of PVA/UNF5. In solid phase melt-mixing, nanofiber was directly incorporated into the polymer matrix using a Brabender. The nanofiber addition significantly changed the stress–strain behavior of the composites: modulus and stress were increased with coated nanofibers by ethylene–acrylic oligomer emulsion as a dispersant; however, elongation was reduced. The dynamic mechanical analysis showed the addition of the soybean nanofiber (SBN) improved the thermal properties for PVA and how the addition of different contents of SBN influenced the tan δ peak and storage modulus of PVA.     

Preparation and properties of biomimetic porous nanofibrous poly(l-lactide) scaffold with chitosan nanofiber network by a dual thermally induced phase separation technique

A biomimetic nanofibrous poly(l-lactide) scaffold decorated by chitosan nanofiber network inside the macropores was fabricated using a dual thermally induced phase separation technique. The first phase separation was used to build a nanofibrous poly(l-lactide) scaffold with interconnected macropores, where chitosan nanofibers about 500 nm in diameter were incorporated via the second phase separation. The content of nanofibrous chitosan was determined to be 5.76 in weight percentage by elemental analysis. The composite scaffold showed the highest protein adsorption of 7225 ± 116 μg/cm³ and the most hydroxyapatite crystal deposition in the mineralization. Compared with non-nanofibrous poly(l-lactide) scaffold, nanofibrous poly(l-lactide) scaffold exhibited a much faster degradation, but it could be restrained by the introduced chitosan nanofibers. The bone mesenchymal stem cell culture results indicated that the cells would rather attach and stretch along the chitosan nanofibers in the composite scaffold that showed the highest viability and the best cytocompatibility may be attributed to the biomimetic nanofibrous network and good cell affinity of chitosan nanofibers.     

Template assisted synthesis of porous nanofibrous cellulose membranes for tissue engineering

Porous, nanofibrous bacterial cellulose (BC) membranes were produced by the bacterium Acetobacter xylinum. The bacterium was cultivated in an appropriate culture medium under static conditions. In situ pore formation was attained through the use of pin templates with diameters varying from 60 to 300 μm composed of polyestirene (ϕ = 300 μm) or optical fibers (ϕ = 60 μm), which were placed on culture medium with the pins immersed in the liquid. Cellulose biosynthesis occurred around the pins leaving tiny pores on the cellulose membrane. After removal of the template the biofilm was dried at 50 °C/24 h. Physico-chemical properties of BC membranes, like degree of crystallinity, swelling and tensile strength were not significantly altered after pore formation. Microstructure evaluation revealed that the film matrix is composed of long nanofibers isotropically distributed on its surface. Round-shaped pores with diameters varying between 60 and 300 μm, depending on the pin template used, were formed in the cellulose membranes. These pores exhibited no border failures that could start crack propagation along the film surface. Microporous membranes could be useful for applications in repairing tissues, which require high oxygenation rates or wound contracture delay.     

Effect of carbonization temperature on properties of aligned electrospun polyacrylonitrile carbon nanofibers

Aligned electrospun nanofibrous bundle was used as the raw material for pretreatment, preoxidation and carbonization processes to prepare carbon nanofibers in a procedure temperature-controlled sintering furnace. Effect of carbonization temperature on the morphology and structural performance of nanofibers was investigated in present study. Results showed that R_I (the relative intensity radio between Disordered peak and Graphite peak) of nanofibers carbonized at 1000 °C is 0.90, carbon content is up to 85.67%, conductivity is 105.44 S·cm^− 1, Young's modulus is 68.8 ± 0.42 GPa, and fiber strength is 306.0 ± 9.0 MPa, all of which endow the fibers with a superior comprehensive property.     

Fabrication of WO3 nanofibers by high voltage electrospinning

Mixtures of 0.1, 0.3, and 0.5 mmol ammonium metatungstate hydrate (AMH), and poly (vinyl alcohol) (PVA) were electrospun by a + 20 kV direct voltage to synthesize fibers. Those of 0.5 mmol AMH were further calcined to have PVA removed and crystalline degree improved. At 500 °C and 2 h calcination, WO₃ nanofibers, including two main stretching modes, 3.24 eV direct energy gap, and 378 nm wavelength violet emission were detected. A possible formation mechanism of WO₃ nanofibers was proposed according to the experimental results.     

Functional shape memory composite nanofibers with graphene oxide filler

In the present study, we prepared a series of graphene oxide (GO) filled shape memory polyurethane (SMPU) nanofibers and systematically investigated the morphological, thermal and mechanical properties, surface wettability, and the shape memory effect (SME) followed by the proposed programming model. The results show that GO can be well dispersed within the SMPU matrix, and the introduction of GO significantly improves the mechanical strength, surface wettability, and thermal stability of the SMPU. Compared with pristine SMPU nanofibrous mats, the prepared SMPU/GO nanofibrous mats have better SME and lower thermal shrinkage. When the loading amount of GO increased to 4.0 wt%, the thermal shrinkage ratio (R_ts) of composite nanofibrous mats could be as low as 4.7 ± 0.3%, while the average fixation ratio (R_f) and recovery ratio (R_r) could be as high as 92.1% and 96.5%, respectively. The study indicates that GO is a desirable reinforcing filler for preparing shape memory nanofibers with improved properties.     

Stem cell differentiation on electrospun nanofibrous substrates for vascular tissue engineering

Nanotechnology has enabled the engineering of a variety of materials to meet the current challenges and requirements in vascular tissue regeneration. In our study, poly-l-lactide (PLLA) and hybrid PLLA/collagen (PLLA/Coll) nanofibers (3:1 and 1:1) with fiber diameters of 210 to 430 nm were fabricated by electrospinning. Their morphological, chemical and mechanical characterizations were carried out using scanning electron microscopy (SEM), attenuated total reflectance Fourier transform infrared (ATR-FTIR), and tensile instrument, respectively. Bone marrow derived mesenchymal stem cells (MSCs) seeded on electrospun nanofibers that are capable of differentiating into vascular cells have great potential for repair of the vascular system. We investigated the potential of MSCs for vascular cell differentiation in vitro on electrospun PLLA/Coll nanofibrous scaffolds using endothelial differentiation media. After 20 days of culture, MSC proliferation on PLLA/Coll(1:1) scaffolds was found 256% higher than the cell proliferation on PLLA scaffolds. SEM images showed that the MSC differentiated endothelial cells on PLLA/Coll scaffolds showed cobblestone morphology in comparison to the fibroblastic type of undifferentiated MSCs. The functionality of the cells in the presence of ‘endothelial induction media’, was further demonstrated from the immunocytochemical analysis, where the MSCs on PLLA/Coll (1:1) scaffolds differentiated to endothelial cells and expressed the endothelial cell specific proteins such as platelet endothelial cell adhesion molecule-1 (PECAM-1 or CD31) and Von Willebrand factor (vWF). From the results of the SEM analysis and protein expression studies, we concluded that the electrospun PLLA/Coll nanofibers could mimic the native vascular ECM environment and might be promising substrates for potential application towards vascular regeneration.     

Physico-chemical/biological properties of tripolyphosphate cross-linked chitosan based nanofibers

In this study, chitosan-PEO blend, prepared in a 15 M acetic acid, was electrospun into nanofibers (~ 78 nm diameter) with bead free morphology. While investigating physico-chemical parameters of blend solutions, effect of yield stress on chitosan based nanofiber fabrication was clearly evidenced. Architectural stability of nanofiber mat in aqueous medium was achieved by ionotropic cross-linking of chitosan by tripolyphosphate (TPP) ions. The TPP cross-linked nanofiber mat showed swelling up to ~ 300% in 1 h and ~ 40% degradation during 30 day study period. 3T3 fibroblast cells showed good attachment, proliferation and viability on TPP treated chitosan based nanofiber mats. The results indicate non-toxic nature of TPP cross-linked chitosan based nanofibers and their potential to be explored as a tissue engineering matrix.     

Gold nanoparticle/polymer nanofibrous composites by laser ablation and electrospinning

Poly(vinylpyrolidone) (PVP) nanofibers incorporating gold nanoparticles (Au-NPs) were produced in combination with laser ablation and electrospinning techniques. The Au-NPs were directly synthesized in PVP solution by laser ablation and then, the electrospinning of PVP/Au-NPs solution was carried out for obtaining nanofibrous composites. The presence of Au-NPs in the PVP nanofibers was confirmed by SEM, TEM and EDX analyses. The SEM imaging elucidated that the electrospun PVP/Au-NPs nanofibers were bead-free having average fiber diameter of 810 ± 480 nm. The TEM imaging indicated that the Au-NPs were in spherical shape having diameters in the range of 5 to 20 nm and the Au-NPs were more or less dispersed homogeneously in the PVP nanofiber matrix. The FTIR study suggested the presence of molecular interactions between PVP matrix and the Au-NPs in the nanofibrous composites. The UV–Vis measurement confirmed the enhancement of the optical properties of the PVP/Au-NPs nanofibers in the solid state due to the surface plasma resonance effect of Au-NPs.     

Strong and optically transparent biocomposites reinforced with cellulose nanofibers isolated from peanut shell

Cellulose nanofibers–reinforced PVA biocomposites were prepared from peanut shell by chemical–mechanical treatments and impregnation method. The composite films were optically transparent and flexible, showed high mechanical and thermal properties. FE-SEM images showed that the isolated fibrous fragments had highly uniform diameters in the range of 15–50 nm and formed fine network structure, which is a guarantee of the transparency of biocomposites. Compared to that of pure PVA resin, the modulus and tensile strength of prepared nanocomposites increased from 0.6 GPa to 6.0 GPa and from 31 MPa to 125 MPa respectively with the fiber content as high as 80 wt%, while the light transmission of the composite only decreased 7% at a 600 nm wavelength. Furthermore, the composites exhibited excellent thermal properties with CTE as low as 19.1 ppm/K. These favorable properties indicated the high reinforcing efficiency of the cellulose nanofibers isolated from peanut shell in PVA composites.     

Bio-functionalized PCL nanofibrous scaffolds for nerve tissue engineering

Surface properties of scaffolds such as hydrophilicity and the presence of functional groups on the surface of scaffolds play a key role in cell adhesion, proliferation and migration. Different modification methods for hydrophilicity improvement and introduction of functional groups on the surface of scaffolds have been carried out on synthetic biodegradable polymers, for tissue engineering applications. In this study, alkaline hydrolysis of poly (ε-caprolactone) (PCL) nanofibrous scaffolds was carried out for different time periods (1 h, 4 h and 12 h) to increase the hydrophilicity of the scaffolds. The formation of reactive groups resulting from alkaline hydrolysis provides opportunities for further surface functionalization of PCL nanofibrous scaffolds. Matrigel was attached covalently on the surface of an optimized 4 h hydrolyzed PCL nanofibrous scaffolds and additionally the fabrication of blended PCL/matrigel nanofibrous scaffolds was carried out. Chemical and mechanical characterization of nanofibrous scaffolds were evaluated using attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, contact angle, scanning electron microscopy (SEM) and tensile measurement. In vitro cell adhesion and proliferation study was carried out after seeding nerve precursor cells (NPCs) on different scaffolds. Results of cell proliferation assay and SEM studies showed that the covalently functionalized PCL/matrigel nanofibrous scaffolds promote the proliferation and neurite outgrowth of NPCs compared to PCL and hydrolyzed PCL nanofibrous scaffolds, providing suitable substrates for nerve tissue engineering.     

Biological evaluation of human hair keratin scaffolds for skin wound repair and regeneration

The cytocompatibility, in vivo biodegradation and wound healing of keratin biomaterials were investigated. For the purposes, three groups of keratin scaffolds were fabricated by freeze-drying reduced solutions at 2 wt.%, 4 wt.% and 8 wt.% keratins extracted from human hairs. These scaffolds exhibited evenly distributed high porous structures with pore size of 120–220 μm and the porosity > 90%. NIH3T3 cells proliferated well on these scaffolds in culture lasting up to 22 days. Confocal micrographs stained with AO visually revealed cell attachment and infiltration as well as scaffold architectural stability. In vivo animal experiments were conducted with 4 wt.% keratin scaffolds. Early degradation of subcutaneously implanted scaffolds occurred at 3 weeks in the outermost surface, in concomitant with inflammatory response. At 5 weeks, the overall porous structure of scaffolds severely deteriorated while the early inflammatory response in the outermost surface obviously subsided. A faster keratin biodegradation was observed in repairing full-thickness skin defects. Compared with the blank control, keratin scaffolds gave rise to more blood vessels at 2 weeks and better complete wound repair at 3 weeks with a thicker epidermis, less contraction and newly formed hair follicles. These preliminary results suggest that human hair keratin scaffolds are promising dermal substitutes for skin regeneration.     

Thermal-crosslinked porous chitosan scaffolds for soft tissue engineering applications

The aim of this study was to demonstrate the feasibility of using a steam autoclave process for sterilization and simultaneously thermal-crosslinking of lyophilized chitosan scaffolds. This process is of great interest in biomaterial development due to its simplicity and low toxicity. The steam autoclave process had no significant effect on the average pore diameter (~ 70 μm) and overall porosity (> 80%) of the resultant chitosan scaffolds, while the sterilized scaffolds possessed more homogenous pore size distribution. The sterilized chitosan scaffolds exhibited an enhanced compressive modulus (109.8 kPa) and comparable equilibrium swelling ratio (23.3). The resultant chitosan scaffolds could be used directly for in vitro cell culture without extra sterilization. The data of in vitro studies demonstrated that the scaffolds facilitated cell attachment and proliferation, indicating great potential for soft tissue engineering applications.     

Fabrication of carbon-core/TiO2-sheath nanofibers by carbonization of poly(vinyl alcohol)/TiO2 composite nanofibers prepared via electrospinning and an interfacial sol–gel reaction

Carbon-core/TiO₂-sheath nanofibers have been fabricated from poly(vinyl alcohol) (PVA)/TiO₂ composite nanofibers that were prepared by electrospinning an aqueous solution of PVA and introducing the thread-like droplets directly into a titanium tetraisopropoxide (TTIP)/heptane solution. The PVA/TiO₂ composite nanofibers were transformed into carbon/TiO₂ nanofibers by iodine vapor treatment at 353 K, which induced dehydration of PVA, followed by carbonization in N₂ at 873 K. The carbon/TiO₂ nanofibers had diameters of a few hundred nanometers and sheathes of tens of nanometers. The nanofibers were composed of carbon cores with a low degree of graphitization and TiO₂ sheaths with an anatase crystal structure. The electrical conductivity of the carbon/TiO₂ composite nanofibers was enhanced by 4 orders of magnitude compared with TiO₂ hollow nanofibers.     

Cross-linked nanocomposite hydrogels based on cellulose nanocrystals and PVA: Mechanical properties and creep recovery

Cellulose nanocrystal (CNC) reinforced poly(vinyl alcohol) (PVA) hydrogels with a water content of ∼92% were successfully prepared with glutaraldehyde (GA) as a cross-linker. The effects of the CNC content on the thermal stability, swelling ratio and mechanical and viscoelastic properties of the cross-linked hydrogels were investigated. The compressive strength at 60% strain for the hydrogels with 1 wt% CNCs increased by 303%, from 17.5 kPa to 53 kPa. The creep results showed that the addition of CNCs decreased the creep elasticity due to molecular chain restriction. The almost complete strain recovery (∼97%) after fixed load removal for 15 min was observed from the hydrogels with CNCs, compared with 92% strain recovery of the neat cross-linked PVA hydrogels. The incorporation of CNCs did not affect the swelling ratio and thermal stability of the hydrogels. These results suggest the cross-linked CNC-PVA hydrogels have potential for use in biomedical and tissue engineering applications.     

Effect of aluminum oxide doping on the structural,electrical, and optical properties of zinc oxide (AOZO) nanofibers synthesized by electrospinning

Zinc oxide nanofibers doped with aluminum oxide were prepared by sol–gel processing and electrospinning techniques using polyvinylpyrrolidone (PVP), zinc acetate and aluminum acetate as precursors. The resulting nanofibers were characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), UV–Vis spectroscopy, and current–voltage (I–V) properties. The nanofibers had diameters in the range of 60–150 nm. The incorporation of aluminum oxide resulted in a decrease in the crystallite sizes of the zinc oxide nanofibers. Aluminum oxide doped zinc oxide (AOZO) nanofibers exhibited lower bandgap energies compared to undoped zinc oxide nanofibers. However, as the aluminum content (Al/(Al + Zn) × 100%) was increased from 1.70 at.% to 3.20 at.% in the electrospinning solution, the bandgap energy increased resulting in lower conductivity. The electrical conductivity of the AOZO samples was found to depend on the amount of aluminum dopant in the matrix as reflected in the changes in oxidation state elucidated from XPS data. Electrospinning was found to be a productive, simple, and easy method for tuning the bandgap energy and conductivity of zinc oxide semiconducting nanofibers.     

Biomimetic acellular detoxified glutaraldehyde cross-linked bovine pericardium for tissue engineering

Glutaraldehyde (GLUT) processing, cellular antigens, calcium ions in circulation, and phospholipids present in the native tissue are predominantly responsible for calcification, degeneration, and lack of natural microenvironment for host progenitor cell migration in tissue implants. The study presents an improved methodology for adhesion and proliferation of endothelial progenitor cells (EPCs) without significant changes in biomechanical and biodegradation properties of the processed acellular bovine pericardium. The anti-calcification potential of the processed tissue was enhanced by detoxification of GLUT-cross-linked bovine pericardium by decellularization, pretreating it with ethanol or removing the free aldehydes by citric acid treatment and lyophilization. The treated tissues were assessed for biomechanical properties, GLUT ligand quantification, adhesion, proliferation of EPCs, and biodegradability. The results indicate that there was no significant change in biomechanical properties and biodegradability when enzymatic hydrolysis (p > 0.05) is employed in detoxified acellular GLUT cross-linked tissue (DBP–G–CA–ET), compared with the native detoxified GLUT cross-linked bovine pericardium (NBP–G–CA–ET). DBP–G–CA–ET exhibited a significant (p > 0.05) increase in the viability of EPCs and cell adhesion as compared to acellular GLUT cross-linked bovine pericardium (p < 0.05). Lyophilized acellular detoxified GLUT cross-linked bovine pericardium, employed in our study as an alternative to conventional GLUT cross-linked bovine pericardium, might provide longer durability and better biocompatibility, and reduce calcification. The developed bovine pericardium patches could be used in cardiac reconstruction and repair, arteriotomy, soft tissue repair, and general surgical procedures with tissue regeneration dimensions.     

Crosslinked gelatin nanofibres: Preparation,characterisation and in vitro studies using glial-like cells

Gelatin (GL) nanofibrous matrices mimicking the complex biological structure of the natural extracellular matrix (ECM) were prepared from aqueous solutions by electrospinning technique. GL nanofibres with a diameter size of around 300 nm were obtained optimising the process and solution parameters. To increase the GL stability in aqueous environment γ-glycidoxypropyltrimethoxysilane (GPTMS) was used as GL crosslinker. GPTMS crosslinking did not modify the nanofibrous matrix morphology: fibre diameter and membrane pores size were 327 ± 45 nm and 1.64 ± 0.37 μm, respectively. The produced GPTMS crosslinked GL nanofibres (GL/GPTMS_NF) were found to support the in vitro adhesion, proliferation and survival of neonatal olfactory bulb ensheating cells (NOBECs).     

Comparison of titanium soaked in 5 M NaOH or 5 M KOH solutions

Commercially pure titanium plates/coupons and pure titanium powders were soaked for 24 h in 5 M NaOH and 5 M KOH solutions, under identical conditions, over the temperature range of 37° to 90 °C. Wettability of the surfaces of alkali-treated cpTi coupons was studied by using contact angle goniometry. cpTi coupons soaked in 5 M NaOH or 5 M KOH solutions were found to have hydrophilic surfaces. Hydrous alkali titanate nanofibers and nanotubes were identified with SEM/EDXS and grazing incidence XRD. Surface areas of Ti powders increased > 50–220 times, depending on the treatment, when soaked in the above solutions. A solution was developed to coat amorphous calcium phosphate, instead of hydroxyapatite, on Ti coupon surfaces. In vitro cell culture tests were performed with osteoblast-like cells on the alkali-treated samples.