The effect of BMP‐2 and VEGF loading of gelatin‐pectin‐BCP scaffolds to enhance osteoblast proliferation

A composite scaffold of gelatine (Gel)‐pectin (Pec)‐biphasic calcium phosphate (BCP) was successfully fabricated. Growth factors such as bone morphogenetic protein‐2 (BMP‐2) and vascular endothelial growth factor (VEGF) were loaded into the Gel‐Pec‐BCP hydrogel scaffolds by freeze‐drying. The surface morphology was investigated by scanning electron microscopy, and BCP dispersion in the hydrogel scaffolds was measured by energy dispersive and X‐ray diffraction spectroscopy. The results obtained from Fourier transform infrared spectroscopy and quantitative measurements showed successfully loading of BMP‐2 and VEGF into the Gel‐Pec‐BCP hydrogel scaffolds. In addition MC3T3‐E1 preosteoblasts were cultivated on the three types of scaffolds to investigate the effects of BMP‐2 and VEGF on cell viability and proliferation. The Gel‐Pec‐BCP scaffolds loaded with VEGF and BMP‐2 demonstrated more cell spreading and proliferation compared to those of the Gel‐Pec‐BCP scaffolds. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 41241.     

Silk fibroin‐chondroitin sulfate‐alginate porous scaffolds: Structural properties and in vitro studies

The development of porous biodegradable scaffolds is of great interest in tissue engineering. In this regard, exploration of novel biocompatible materials is needed. Silk fibroin‐chondroitin sulfate‐sodium alginate (SF‐CHS‐SA) porous hybrid scaffolds were successfully prepared via lyophilization method and crosslinked by 1‐ethyl‐3‐(3‐dimethylaminopropyl)carbodiimide‐ethanol treatment. According to the scanning electron microscopy studies, mean pore diameters of the scaffolds were in the range of 60–187 μm. The porosity percentage of the scaffold with SF‐CHS‐SA ratio of 70 : 15 : 15 (w/w/w %) was 92.4 ± 3%. Attenuated total reflectance Fourier transform infrared spectroscopy, X‐ray diffraction, and differential scanning calorimetry results confirmed the transition from amorphous random coil to crystalline β‐sheet in treated SF‐CHS‐SA scaffold. Compressive modulus was significantly improved in hybrid scaffold with SF‐CHS‐SA ratio of 70 : 15 : 15 (3.35 ± 0.15 MPa). Cytotoxicity assay showed that the scaffolds have no toxic effects on chondrocytes. Attachment of chondrocytes was much more improved within the SF‐CHS‐SA hybrid scaffold. Real‐time polymerase chain reaction analyses showed a significant increase in gene expression of collagen type II, aggrecan, and SOX9 and decrease in gene expression of collagen type I for SF‐CHS‐SA compared with SF scaffold. This novel hybrid scaffold can be a good candidate to be utilized as an efficient scaffold for cartilage tissue engineering. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014 , 131, 41048.     

Chitosan/gelatin porous bone scaffolds made by crosslinking treatment and freeze‐drying technology: Effects of crosslinking durations on the porous structure,compressive strength,and in vitro cytotoxicity

In this study, freezing was used to separate a solute (polymer) and solvent (deionized water). The polymer in the ice crystals was then crosslinked with solvents, and this diminished the linear pores to form a porous structure. Gelatin and chitosan were blended and frozen, after which crosslinking agents were added, and the whole was frozen again and then freeze‐dried to form chitosan/gelatin porous bone scaffolds. Stereomicroscopy, scanning electron microscopy, compressive strength testing, porosity testing, in vitro biocompatibility, and cytotoxicity were used to evaluate the properties of the bone scaffolds. The test results show that both crosslinking agents, glutaraldehyde (GA) and 1‐ethyl‐3‐(3‐dimethylaminopropyl) carbodiimide, were able to form a porous structure. In addition, the compressive strength increased as a result of the increased crosslinking time. However, the porosity and cell viability were not correlated with the crosslinking times. The optimal porous and interconnected pore structure occurred when the bone scaffolds were crosslinked with GA for 20 min. It was proven that crosslinking the frozen polymers successfully resulted in a division of the linear pores, and this resulted in interconnected multiple pores and a compressively strong structure. The 48‐h cytotoxicity did not affect the cell viability. This study successfully produced chitosan/gelatin porous materials for biomaterials application. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 41851.     

Alcohol-triggered silk fibroin hydrogels having random coil and β-turn structures enhanced for cytocompatible cell response

Alcohol additive is one of the stimulants that induces the fast gelation of silk fibroin solution. Based on our previous report, different alcohol types influence the gelation kinetic and the properties of resulting silk fibroin hydrogels. Here, the effects of alcohol concentrations on the silk fibroin gelation and cell response were reported. All fibroin hydrogels prepared with various alcohol additives showed cell biocompatibility, especially the fibroin hydrogel prepared with 10 wt % n-butanol. Results on the mechanical properties of hydrogels, n-butanol additive enhanced a higher compressive modulus up to ~ 22 times in comparison to non-alcoholic fibroin hydrogel. Fourier transform infrared analysis and peak deconvolution showed a possible formation of more β-turn linkage and random coil structure of fibroin segments in alcoholic fibroin hydrogel. So, the micro-segmental structure of fibroin hydrogel caused the higher compressive modulus, prolonged deformation of the hydrogels, and efficient cell growth on the fibroin hydrogel. © 2019 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2020 , 137, 48731.     

Assessment of cell proliferation in knitting scaffolds with respect to pore‐size heterogeneity,surface wettability,and surface roughness

In this study, various types of poly(ε‐caprolactone) (PCL) knitting scaffolds were fabricated and analyzed to assess the cell‐culturing characteristics of knitting scaffolds with respect to pore‐size heterogeneity, surface wettability, and surface roughness. First, control knitting scaffolds were fabricated using 150‐µm‐diameter PCL monofilaments. Using chloroform and NaOH, PCL knitting scaffolds with varying roughness, pore‐size heterogeneity, and surface wettability were fabricated. Cell‐culture assessments were performed on these six types of PCL knitting scaffolds. Saos‐2 cells were used for cell assessments and cultured for 14 days on each scaffold. Consequently, heterogeneous pore‐size distribution and high surface wettability were found to enhance cell proliferation in knitting scaffolds. In addition, for highly hydrophobic knitting scaffolds exhibiting water contact angles greater than 110 degrees, smaller surface roughness was found to enhance cell proliferation. According to this study, in the case of knitting scaffold, NaOH‐treated knitting scaffold, without any control for the pore‐size homogenization, could be a candidate as the optimal knitting scaffold. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42566.     

Study on porous silk fibroin materials. II. Preparation and characteristics of spongy porous silk fibroin materials

Porous silk fibroin materials, with average pore size 10 ∼ 300 μm, pore density 1 ∼ 2000/mm², and porosity 35 ∼ 70%, were prepared by freeze drying aqueous solution of silk fibroin obtained by dissolving silk fibroin in ternary solvent CaCl₂ · CH₃CH₂OH · H₂O. Pore size distribution of such materials mostly accorded with logarithmic normal distribution. It is possible to control the aforementioned structural parameters and the physical properties of moisture permeability, compressibility, strength, elongation, etc., by adjusting freezing temperature and concentration of silk fibroin solution. Above glass transition zone (−34 ∼ −20°C) of silk fibroin, the freezing temperature has more significant effect on the structure and properties of porous silk fibroin materials. © 2001 John Wiley & Sons, Inc. J Appl Polym Sci 79: 2192–2199, 2001     

Physical and mechanical properties of the fully interconnected chitosan ice‐templated scaffolds

Porous chitosan scaffolds were prepared with a freeze‐casting technique with different concentrations, 1.5 and 3 wt %, and also different cooling rates, 1 and 4°C/min. The pore morphology, porosity, pore size, mechanical properties, and water absorption characteristics of the scaffolds were studied. Scanning electron microscopy images showed that the freeze‐cast scaffolds were fully interconnected because of the existence of pores on the chitosan walls in addition to many unidirectionally elongated pores. Increases in the chitosan concentration and freezing rate led to elevations in the thickness of the chitosan walls and reductions in the pores size, respectively. These two results led to the enhancement of the compressive strength from 34 to 110 kPa for the scaffolds that had 96–98% porosity. Also, augmentation of the chitosan concentration and decreases in the freezing rate led to the reduction of the number of pores on the chitosan walls. Furthermore, the volume of water absorption increased with a reduction in the chitosan concentration and cooling rate from 690 to 1020%. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 41476.     

Differences in regenerated silk fibroin prepared with different solvent systems: From structures to conformational changes

Dissolution is an important part of silk fibroin (SF) reprocessing, and it is the only way to process it into films, gels, porous scaffold materials, and electrostatic spinning silk fibers. There are a variety of dissolution systems used to dissolve SF. However, few studies have focused on the differences between these different solvent systems. The dissolution of SF with different solvent systems was investigated in this study. Regenerated silk fibroin (RSF) solutions and films were characterized by dynamic light scattering, Fourier transform infrared spectroscopy, differential scanning calorimetry, X‐ray diffraction, and scanning electron microscopy. The results show that the RSF film structures changed with the solvent system, especially LiBr–H₂O. The characterization proved that the random coil did not change into a β‐sheet structure during film formation, and this indicated that its crystal structure and thermal stability was different from others. Interestingly, the differences in the morphologies of all of the RSF films prepared with different solvents were outstanding. Because the mechanism and force of the ion in the solvent systems were different, the SF molecule was hydrolyzed differently in individual solvent systems and produced different hydrolyzed SF molecular chains. These chains had different self‐assembly processes and would lead to RSF products with different microstructures and properties. This suggests that a suitable solvent should be chosen for different uses. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 41959.     

Platelet lysate and adipose mesenchymal stromal cells on silk fibroin nonwoven mats for wound healing

In this work platelet lysate (PL) and adipose‐derived mesenchymal stromal cells (ASCs) seeded on nonwoven fibroin mats were in vitro and in vivo evaluated for tissue regenerative applications. Nonwoven mats obtained by a large scale water entanglement technique were characterized for their physico‐chemical properties. Results indicated a high purity of fibroin fibers, their stability after sterilization process and appropriate technological properties suitable for tissue engineering. Moreover, the scaffolds in vitro supported adhesion and migration of ASCs and the presence of PL improved the cell proliferation. The products were then applied on epithelial/dermal wounds carried out on the dorsal surface of rabbit: the skin reparative process was solved in 9 days, with a completely restitutio ad integrum of the epithelium in animals treated with PL alone; ASCs did not further improve the wound healing. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2016 , 133, 42942.     

Study on the properties of nano‐TiO2 particles modified silk fibroin porous films

Using the freeze‐drying method, Nano‐TiO₂/silk fibroin porous films were synthesized with different ratios of TiO₂ to silk fibroin solution. Through scanning electron microscopy (SEM), X‐ray diffraction (XRD), thermogravimetric analysis (TGA), tensile strain, and water‐solubility tests, the structures and properties of these porous films were characterized. The SEM results indicated that the pores of the nano‐TiO₂/silk fibroin porous films were uniformly distributed by the freeze‐drying method. The XRD analysis indicated that the formation of nano‐TiO₂ particles might induce a conformational transition of silk fibroin from the typical Silk I to the typical Silk II structure partly with an increase in the crystallinity of the porous films. Compared with the pure silk fibroin porous films, the mechanical properties of nano‐TiO₂/silk fibroin porous films were improved, and its heat transition temperature was also enhanced; however, the water‐solubility of this material was diminished. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2010     

Chitosan‐finished Antheraea mylitta silk fibroin nonwoven composite films for wound dressing

In this study, we aimed to produce nonwoven wound‐dressing films made of Antheraea mylitta (tasar) silk fibroin by a solution‐casting method. These nonwoven films were finished with chitosan solutions of different concentrations ranging from 0.75 to 2% w/v with a pad–dry method to fabricate nonwoven composite films. Chitosan‐finished tasar fibroin nonwoven composite films (CMTFFs) showed higher mechanical and dynamic mechanical properties as compared to nonwoven tasar fibroin. The physical, structural, and thermal properties of the films were investigated. The hemocompatibility, cytocompatibility, and biodegradation tests showed that the CMTFF was a promising material for use as a wound dressing. © 2016 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2016 , 133, 44341.     

Biocomposite membranes of sodium alginate and silk fibroin fibers for biomedical applications

Biocomposite membranes from biodegradable and biocompatible natural polymers were prepared from sodium alginate solution reinforced with silk fibroin fibers in several fiber content by casting and solvent evaporation. The properties of these biocomposites were investigated by scanning electron microscopy, swelling test, water vapor transmission, mechanical and thermal analyses, and cytotoxicity test. A biocomposite with uniform fiber dispersion and good fiber–matrix interaction was obtained through the incorporation of fibroin fibers in the alginate membrane, even though the fibers were used without any surface treatment to enhance the interfacial adhesion. The incorporation of fibroin fibers improved the tensile strength and also provided a new property to the alginate, that is, the resistance to tear. Moreover, the use of silk fibroin fibers in polymeric composites can result in a material with adequate characteristics for application in the biomaterial field, especially as wound dressings, because of its nontoxic effect to cells, flexibility, and resistance to tear. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 130: 3451–3457, 2013     

Preparation of acrylamide‐based poly‐HIPEs with enhanced mechanical strength using PVDBM‐b‐PEG‐emulsified CO2‐in‐water emulsions

Poly(vinyl acetate‐alt‐dibutyl maleate)‐block‐poly(ethylene glycol) (PVDBM‐b‐PEG) copolymers were synthesized via reversible addition–fragmentation chain transfer radical polymerization and used as emulsifiers to form stable CO₂‐in‐water high internal phase emulsions (C/W HIPEs). Then, highly interconnected cellular polyacrylamide (PAM) and poly(acrylamide‐co‐N‐hydroxymethyl acrylamide) [P(AM‐co‐HMAM)] poly‐HIPEs with enhanced mechanical strength were prepared based on the stable C/W HIPEs. The porous structures of the PAM poly‐HIPEs, as well as morphology and compressive modulus, could be influenced by the surfactant concentration and the length of the CO₂‐philic tails of the surfactants. PAM poly‐HIPEs with the smallest average pore diameter (11.12 ± 0.62 μm) and the highest compressive modulus (22.65 ± 0.10 MPa) could be obtained by using the short CO₂‐philic chains of the PVDBM‐b‐PEG surfactant at a high concentration (1.0 wt %). Moreover, with the copolymerization of N‐hydroxymethyl acrylamide (HMAM) comonomers with acrylamide, the compressive modulus of the obtained P(AM‐co‐HMAM) poly‐HIPEs was three times higher than that of PAM poly‐HIPEs. Both PAM and P(AM‐co‐HMAM) poly‐HIPEs were employed as scaffolds to guide H9c2 cardiac muscle cellular growth. Fluorescence images showed that a smaller average pore size and a narrower pore‐size distribution were helpful for cell growth and proliferation on these materials. © 2018 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2018 , 135, 46346.     

Immobilization of lipase with alginate hydrogel beads and the lipase‐catalyzed kinetic resolution of α‐phenyl ethanol

The immobilization of enzymes is one of the key issues in both the field of enzymatic research and industrialization. In this article, we report a facile method for immobilizing Candida antarctica lipase B in an alginate carrier. In the presence of calcium cations, an enzyme–alginate suspension was crosslinked to form beads with a porous structure at room temperature, and the enzymes were well dispersed in the beads. The chiral resolution of α‐phenyl ethanol in the organic phase was tested by the enzyme–alginate beads. The effects of the reaction parameters, such as the enzyme concentration, temperature, and molar ratio of the substrate to the solvent, on the resolution behavior are discussed. Reuse cycle experiments for the chiral resolution of α‐phenyl ethanol demonstrated that the activity of the enzyme–alginate beads was maintained without marked deactivation up to five repeated cycles. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014 , 131, 40178.     

The inclusion of fetal bovine serum in gelatin/PCL electrospun scaffolds reduces short‐term osmotic stress in HEK 293 cells caused by scaffold components

Components of gelatin/polycaprolactone (PCL) electrospun scaffolds are released to surrounding media and cause osmotic changes that adversely affect cell viability and proliferation. In this study, the physiological properties of gelatin/PCL scaffolds were investigated by qRT‐PCR and by performing cellular studies on HEK 293 cells. Components released from gelatin/PCL scaffolds were found to induce osmotic stress response in these cells. However, osmotic stress was inhibited by adding fetal bovine serum (FBS) to scaffolds. In addition, focal adhesion related genes were found to be up‐regulated in HEK 293 cells on gelatin/PCL/20% FBS scaffolds, and this induced the down‐regulations of cell‐death related genes. Furthermore, the inclusion of 20% FBS improved the viabilities of HEK 293 cells on gelatin/PCL scaffolds. This study indicates that adding FBS to gelatin/PCL scaffolds improves scaffold bio‐affinity. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2013     

Facile fabrication of near-infrared light-responsive shape memory nanocomposite scaffolds with hierarchical porous structures

In this work, the near-infrared (NIR) light-responsive shape memory scaffolds with hierarchical porous structures are designed and facilely formed by freeze drying of 3D printed viscous gel-like pickering emulsions, which are stabilized by hydrophobically modified graphene oxide (g-GO) and silica nanoparticles, and contain thermo-responsive poly(d , l -lactic acid-co-trimethylene carbonate) (PLMC) in the oil phase. The prepared scaffolds display an interconnected filament structure with hierarchical pores and high porosity. The incorporation of g-GO nanoparticles into PLMC matrix prompts that the scaffold shape memory can be triggered by NIR light with fast shape recovery. Moreover, the in vitro mineralization experiment shows that the scaffolds have biological activity, and the drug release study demonstrates that the scaffolds can be used as drug carriers with efficient drug release capacity. Furthermore, cell culture assays based on mouse bone mesenchymal stem cells exhibit that the scaffolds own good cytocompatibility. Therefore, the facile preparation and remote activation of the shape memory nanocomposite scaffolds with hierarchical porous structure and multifunctionality represents a highly attractive candidate as minimally invasive implantation scaffolds for bone tissue engineering applications.     

Effective immobilization of lipase onto a porous gelatin‐co‐Poly(vinyl alcohol) copolymer and evaluation of its hydrolytic properties

Crosslinked copolymers of gelatin and poly(vinyl alcohol) (PVA) with excellent water absorption and water retention abilities were successfully synthesized using ⁶⁰Co γ radiation. Ammonium persulfate (APS), as a water‐soluble initiator and sodium bicarbonate (NaHCO₃) as a foaming agent were used. The best synthesis conditions were evaluated with regard to the maximum percentage of swelling as a function of the APS concentration, NaHCO₃ concentration, amount of water, and reaction time. The maximum swelling percentage (1694.59%) of the copolymer gelatin‐co‐PVA, was obtained at the optimum parameters [APS] = 2.92 × 10^?1 mol/L, [NaHCO₃] = 7.94 × 10^?2 mol/L, and 1.5 mL of water with 31.104 kGy of the γ radiation dose. The copolymer was characterized by Fourier transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM) methods. The SEM analysis showed a highly nanoporous and cellular structure of the copolymer. The copolymer was used as a support for lipase immobilization. The optimization of the reaction conditions, including the pH and temperature for immobilization, on the basis of the hydrolysis of p‐nitrophenyl palmitate, was carried out. An excellent efficiency for protein loading (70%) at pH 8.5 by the copolymer was observed. The results observed during the evaluation of the hydrolytic properties showed excellent activity of the bound lipase. The porous gelatin‐co‐PVA bound lipase was found to be stable at 75°C and pH 8.5; it displayed 2.326 ± 0.005 U/g of lipase activity. The stability and activity of the copolymer‐bound lipase were also studied as a function of the time at 75°C, and the biocatalyst was found to be stable and active up to 1 h, beyond which the activity decreased. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014 , 131, 39622.     

Immobilization of β‐galactosidase from Escherichia coli onto modified natural silk fibers

A polymeric support based on the natural silk fibers was prepared and characterized for covalent immobilization of β‐galactosidase from Escherichia coli. The silk fibers were grafted using polyacrylonitrile in presence of benzophenone as a photo‐initiator. The grafted fibers were then activated by treatment with hydrazine hydrate followed by glyoxal cross‐linker. FTIR spectra, scanning electron microscope (SEM) in addition to the staining test derived from the Coomassie protein assay were utilized for investigation of the modification and immobilization steps. Also, the activity of both free and immobilized β‐galactosidase was evaluated as a function of the various important parameters like grafting percentage, pH, and temperature. In addition, the kinetic parameters K_m and v_max for both free and immobilized enzyme were anticipated using Michaelis–Menten equation. The results in this study indicated that the prepared modified woven silk fibers could be used effectively as a polymeric support for immobilization of β‐galactosidase. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 130: 2923–2931, 2013     

Fabrication of co‐continuous poly(ε‐caprolactone)/polyglycolide blend scaffolds for tissue engineering

The apparent inability of a single biomaterial to meet all the requirements for tissue engineering scaffolds has led to continual research in novel engineered biomaterials. One method to provide new materials and fine‐tune their properties is via mixing materials. In this study, a biodegradable powder blend of poly(ε‐caprolactone) (PCL), polyglycolide (PGA), and poly(ethylene oxide) (PEO) was prepared and three‐dimensional interconnected porous PCL/PGA scaffolds were fabricated by combining cryomilling and compression molding/polymer leaching techniques. The resultant porous scaffolds exhibited co‐continuous morphologies with ～50% porosity. Mean pore sizes of 24 and 56 μm were achieved by varying milling time. The scaffolds displayed high mechanical properties and water uptake, in addition to a remarkably fast degradation rate. The results demonstrate the potential of this fabrication approach to obtain PCL/PGA blend scaffolds with interconnected porosity. In general, these results provide significant insight into an approach that will lead to the development of new composites and blends in scaffold manufacturing. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42471.     

Investigations on rheological properties and gelation of tasar regenerated silk fibroin solution

Tasar silk is a variety of non‐mulberry silk indigenous to the Indian subcontinent. We present the measured frequency‐dependent viscoelastic moduli of Tasar regenerated silk fibroin (RSF) solution using optical tweezers at two concentrations (0.16% and 0.25% w/v) and extend these measurements to the low frequency regime using a video microscopy technique. We extend the investigation on the rheological behavior of Tasar RSF for four more RSF concentrations, viz., 0.50%, 1.00%, 2.50% and 5.00% using video microscopy. In all the RSF samples, both storage and loss moduli are found to increase with frequency. At lower frequencies the loss modulus is more than the storage modulus and exhibit similar behavior until a crossover frequency beyond which the storage modulus exceeds the loss modulus at all frequencies. The relaxation time which is inversely related to the crossover frequency is found to rise sharply at 5% w/v, indicating the onset of gelation in the sample. These results are examined in relation to the viscoelastic parameters of mulberry silk, wherein the larger crossover frequencies at the same higher concentrations indicate relaxation times that are an order of magnitude smaller than those measured for Tasar RSF. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014 , 131, 40104.