Fabrication of hybrid scaffolds obtained from combinations of PCL with gelatin or collagen via electrospinning for skeletal muscle tissue engineering.

The creation of skeletal muscle tissue in vitro is a major topic of interest today in the field of biomedical research, due to the lack of treatments for muscle loss due to traumatic accidents or disease. For this reason, the intrinsic properties of nanofibrillar structures to promote cell adhesion, proliferation, and cell alignment presents an attractive tool for regenerative medicine to recreate organized tissues such as muscle. Electrospinning is one of the processing techniques often used for the fabrication of these nanofibrous structures and the combination of synthetic and natural polymers is often required to achieve optimal mechanical and physiochemical properties. Here, polycaprolactone (PCL) is selected as a synthetic polymer used for the fabrication of scaffolds, and the effect of protein addition on the final scaffolds' properties is studied. Collagen and gelatin were the proteins selected and two different concentrations were analyzed (2 and 4 wt/vol%). Different PCL/protein systems were prepared, and a structural, mechanical and functional characterization was performed. The influence of fiber alignment on the properties of the final scaffolds was assessed through morphological, mechanical and biological evaluations. A bioreactor was used to promote cell proliferation and differentiation within the scaffolds. The results revealed that protein addition produced a decrease in the fiber size of the membranes, an increase in their hydrophilicity, and a softening of their mechanical properties. The biological study showed the ability of the selected systems to harbor cells, allow their growth and, potentially, develop musculoskeletal tissues.

Tunable physical and mechanical properties of gelatin hydrogel after transglutaminase crosslinking on two gelatin types.

The crosslinking and related gel properties of 3 wt% gelatin (type-A and type-B) catalyzed by microbial transglutaminase (MTG, dose of 0-20 U/g gelatin) have been investigated. A MTG-depended increase in the molecular weight and mean diameter of both gelatins was observed, where type-A presented a higher crosslinking efficiency than type-B due to more acyl donors of the former. As MTG concentration increased, the surface hydrophobicity and thermal stability of type-A gelatin increased. Textural profile analysis (TPA) of type-A gelatin hydrogel showed a decrease in hardness and slight increase in springiness, while type-B gelatin gel was not affected generally. Rheological measurements confirmed the melting point of type-A gelatin hydrogel continually increased until the disappearance of gel thermo-reversibility at higher MTG levels (≥12 U/g gelatin), while type-B gelatin hydrogel always showed a sol-gel transition, suggesting that the gel performance was depended on the dominance of whether physical crosslinking or chemical crosslinking. Scanning electron microscope (SEM) results showed that the network structure of the type-A gelatin became more irregular as MTG increasing which indicated that introducing additional covalent cross-links within or between gelatin chains had a profound influence on gel's network structure, closely associated with the gel properties mentioned above. In summary, the superiority of type-A in MTG-crosslinking efficiency than type-B, can be used to modulate the physical and mechanical properties of gelatin hydrogel, governing by the combing of weak physical crosslinking and strong covalent crosslinking, which will be suitable for numerous industrial applications.

Microstructure and characteristic properties of dogfish skin gelatin gels prepared by freeze/spray-drying methods.

The effects of two pretreatments (microwaves or oven-drying) on the dogfish (Squalus acanthias) skin as well as two drying processes (freeze-drying or spray-drying) on the extracted gelatins were studied. Thus six types of gelatins were obtained, three of which were freeze-dried (FG) and the others were spray-dried (SG), from the untreated skin (US), microwaves-pretreated skin (MS) and oven-pretreated skin (OS). The highest yield (8.67%) was obtained for the OSFG, while the lowest one (3.06%) was measured for the OSSG. Interestingly, all gelatins exhibited relatively high protein (84.02-89.53%), and low lipid (0.50-1.71%) and ash (3.05-7.17%) contents. In addition, gelatins were analyzed by the Fourier transform infrared and the spectra displayed important differences in some specific peaks, particularly in the amide I, amide II and amide III. The gelatins extracted from the untreated skin, regardless the drying method, presented the highest foaming capacity. The textural profile analysis showed that USSG was the hardest (213.6 g) and the chewier (23.8 N × mm) gelatin. Moreover, analysis of thermal properties showed that USSG also has the highest glass-transition temperature. The interesting properties of gelatin extracted from dogfish skin encourage their future use as a functional ingredient in industrial food formulations.

Synergistic effects of acetylated distarch adipate and sesbania gum on gelatinization and retrogradation of wheat starch.

The synergistic effects of the combination of acetylated distarch adipate and sesbania gum (C-ADA-SG) with the ratio of 5:0, 4:1, 2.5:2.5, 1:4, 0:5, accounting for 5% (w/w) of the starch on gelatinization and retrogradation properties of wheat starch (WS) were studied. Scanning Electron Microscopy (SEM) showed that the microstructure of gelatinized WASs (WS added with C-ADA-SG) tended to be smoother. Based on the results of Rapid Visco-Analyzer (RVA) and Differential Scanning Calorimetry (DSC), the pasting characteristics of WS were affected and the gelatinization process was retarded by C-ADA-SG. After seven-day storage at 4 °C, compared to WS, the gel firmness, syneresis, retrogradation enthalpy, and the relative crystallinity of WASs clearly decreased by 17.47-44.36 g, 11.16-17.93%, 0.22-0.80 J·g-1, and 4.06-8.61%, respectively. However, the band ratio 1639:1157 cm-1 by FTIR and loss tangent (tan δ) value were increased with C-ADA-SG addition. Meanwhile, X-Ray Diffraction (XRD) reflected that native WS showed A-type crystal structure, which transferred to B + V type after retrogradation. Furthermore, Low-Field Nuclear Magnetic Resonance (LF-NMR) declared that C-ADA-SG increased the water mobility and limited the diffusion and seepage of water during storage. Generally, ADA and SG produced a synergistic effect on retarding the gelatinization and retrogradation of WS.

Preparation of Electrospun Gelatin Mat with Incorporated Zinc Oxide/Graphene Oxide and Its Antibacterial Activity.

Current wound dressings have poor antimicrobial activities and are difficult to degrade. Therefore, biodegradable and antibacterial dressings are urgently needed. In this article, we used the hydrothermal method and side-by-side electrospinning technology to prepare a gelatin mat with incorporated zinc oxide/graphene oxide (ZnO/GO) nanocomposites. The resultant fibers were characterized by field emission environment scanning electron microscopy (FESEM), energy-dispersive X-ray spectroscopy (EDX), X-ray diffractometry (XRD) and Fourier transform infrared spectroscopy (FTIR). Results indicated that the gelatin fibers had good morphology, and ZnO/GO nanocomposites were uniformly dispersed on the fibers. The loss of Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) viability were observed to more than 90% with the incorporation of ZnO/GO. The degradation process showed that the composite fibers completely degraded within 7 days and had good controllable degradation characteristics. This study demonstrated the potential applicability of ZnO/GO-gelatin mats with excellent antibacterial properties as wound dressing material.

Preparation and Characterization of ß-glucosidase Films for Stabilization and Handling in Dry Configurations.

BACKGROUND: Although the stability of proteins is of significance to maintain protein function for therapeutical applications, this remains a challenge. Herein, a general method of preserving protein stability and function was developed using gelatin films. METHODS: Enzymes immobilized onto films composed of gelatin and Ethylene Glycol (EG) were developed to study their ability to stabilize proteins. As a model functional protein, ß-glucosidase was selected. The tensile properties, microstructure, and crystallization behavior of the gelatin films were assessed. RESULTS: Our results indicated that film configurations can preserve the activity of ß-glucosidase under rigorous conditions (75% relative humidity and 37°C for 47 days). In both control films and films containing 1.8 % ß-glucosidase, tensile strength increased with increased EG content, whilst the elongation at break increased initially, then decreased over time. The presence of ß-glucosidase had a negligible influence on tensile strength and elongation at break. Scanning electron-microscopy (SEM) revealed that with increasing EG content or decreasing enzyme concentrations, a denser microstructure was observed. CONCLUSION: In conclusion, the dry film is a promising candidate to maintain protein stabilization and handling. The configuration is convenient and cheap, and thus applicable to protein storage and transportation processes in the future.

Induced Aggregation of Epoxy Polysiloxane Grafted Gelatin by Organic Solvent and Green Application.

In this paper, we studied the aggregation of amphiphilic polymer epoxy-terminated polydimethylsiloxane (PDMS-E) grafted gelatin (PGG) in water induced by methanol, ethanol, 2-propanol, acetone, tetrahydrofuran (THF), and 1,4-dioxane. The aggregation pattern of the polymer was monitored by infrared spectroscopy, X-ray diffraction, transmission electron microscopy, and scanning electron microscopy. It was revealed that the aggregate morphology showed clear dependence on the solvent polarity. The PGG aggregates had regular spherical morphology in polar solvents, including water, methanol, ethanol, 2-propanol, and acetone. The coating performance was evaluated by X-ray photoelectron spectroscopy and friction experiment, and PGG and acetone coating exhibited excellent coating performance on the surface of pigskin. Gel was formed in acetone and tetrahydrofuran (THF) with the slow evaporation of solvent, and this property can possibly be applied to industrial sewage treatment. White precipitate and soft film were formed in non-polar 1,4-dioxane.

Influence of gelatinization process and HMDSO plasma treatment on the chemical changes and water vapor permeability of corn starch films.

In this study, surface, chemical, physicochemical and barrier properties of films treated with hexamethyldisiloxane (HMDSO) cold plasma were investigated. Normal and high amylose starches were gelatinized at different level to obtain films with different amount of free amylopectin. The obtained films were subjected to HMDSO plasma treatment. XPS analysis indicated chemical changes including substitution and crosslinking of the starch molecule, as reflected by the CSi bond increasing and the C-OH bonds reduction on treated films. These changes modified the thermal transitions (Tm and ΔH). The highest amount of CSi bonds was more noticeable in the TF50 film, suggesting a better interaction between active species of plasma and the free amylopectin released into the continuous phase of the film. Moreover, active species of plasma increased the crystallinity in all films. These results suggested that a higher helical packaging, crosslinking and hydrophobic blocking groups (CSi) of starch molecules resulted in films with improved barrier performance against water molecules.

Amination degree of gelatin is critical for establishing structure-property-function relationships of biodegradable thermogels as intracameral drug delivery systems.

Glaucoma is a lifelong disorder that necessitates continuous medical therapy to manage its symptoms and preserve the vision of patients; accordingly, it is highly beneficial to develop a long-acting injectable depot system that can exhibit better drug delivery capability. This study aims to investigate the effect of the amination degree of gelatin on the carbodiimide-mediated grafting of thermo-responsive poly(N-isopropylacrylamide) segments onto biodegradable protein backbone molecules. Moreover, the potential applications of these carrier materials for intracameral pilocarpine administration in glaucomatous subjects will be considered. The gelatins with different amination degrees that are prepared by controlling the feed amount of adipic acid dihydrazide are further used for the synthesis of graft copolymers. The results of chemical characterization and electron microscopy studies showed that both grafting reaction effectiveness and gelling carrier ultrastructure vary in response to biomaterial amination. Compared to unmodified biopolymer thermogel without gel formation, graft copolymers that are composed of aminated gelatin networks showed a more remarkable temperature-triggered pilocarpine capture under physiological conditions. This could create more stable depot-forming carrier systems with improved in vivo pharmacological efficacy. Although the increase in amination degree enhances the biodegradation resistance of graft copolymers for achieving extended drug release profiles and provides significant therapeutic benefits, carriers with excess positive charges may potentiate the cytotoxic actions of oxidative stress signals and may cause damage in cellular barrier integrity. Consequently, unfavorable ocular tissue responses and poor treatment outcomes are observed in glaucomatous rabbits. For the first time, our findings suggest that the amination degree of gelatin performs a crucial function in guiding the development of structure-property-function relationships of biodegradable thermogels as intracameral drug delivery systems.

Gelatin-Based Antimicrobial Films Incorporating Pomegranate (Punica granatum L.) Seed Juice by-Product.

Pomegranate (Punica granatum L.) seed juice by-product (PSP) was added as reinforcing and antimicrobial agent to fish gelatin (FG) films as a promising eco-friendly active material for food packaging applications. A complete linkage analysis of polysaccharides in PSP showed xylan and cellulose as main components. This residue showed also high total phenolic content and antioxidant activity. Three formulations were processed by adding PSP to FG (0, 10, 30 wt. %) by the casting technique, showing films with 10 wt. % of PSP the best performance. The addition of PSP decreased elongation at break and increased stiffness in the FG films, particularly for 30 wt. % loading. A good compatibility between FG and PSP was observed by SEM. No significant (p < 0.05) differences were obtained for barrier properties to oxygen and water vapour permeability compared to the control with the incorporation of PSP, whereas water resistance considerably increased and transparency values decreased (p < 0.05). High thermal stability of films and inhibition against S. aureus were observed. The addition of PSP at 10 wt. % into FG was shown as a potential strategy to maintain the integrity of the material and protect food against lipid oxidation, reducing huge amounts of pomegranate and fish wastes.

Novel hard capsule prepared by tilapia (Oreochromis niloticus) scale gelatin and konjac glucomannan: Characterization, and in vitro dissolution.

A novel material for making capsules was prepared with tilapia scale gelatin and konjac glucomannan (KGM). Rheological behaviors of gelatin with KGM at different levels (0.1%, 0.15%, and 0.2%, w/w) were investigated. The highest values (P < 0.05) of gel strength (518.33 ± 6.17 g) and melting temperature (39.7 ± 0.11 °C) could be observed at the gelatin solution with 0.15% (w/w) KGM. The tensile strength (TS) and elongation at break (EAB) of the films increased with the increasing of KGM. The water contact angle was enhanced with the increasing of KGM, indicating the decrease of the film hydrophilicity. XRD and FTIR showed the interactions between gelatin and KGM. Gelatin solution with 0.15% KGM is suitable for preparing capsules. Drug dissolution test in vitro showed the shell rupture time is in range of 3-5 min, and 80% of the drugs were released within 10 min. Therefore, the composite materials made of tilapia scale gelatin and KGM can be utilized for hard capsules.

Rheological behavior, emulsifying properties and structural characterization of phosphorylated fish gelatin.

Rheological, microstructural and emulsifying properties of fish gelatin phosphorylated using sodium trimetaphosphate (STMP) were studied. Phosphorylation was carried out at 50 °C for 0, 0.5, 1 or 2 h. Rheological behaviors indicated that phosphorylation decreased gelation rate constant (kgel) and apparent viscosity of gelatin solutions. Phosphorylation time was inversely proportional to tan δ; gelling and melting points of fish gelatin gels; however gel properties could be improved by short time of phosphorylation. Scanning electron microscopy and atomic force microscopy revealed that longer time of phosphorylation resulted in looser gel network with more aggregation. Longer phosphorylation time could stabilize fish gelatin emulsions, and endowed emulsions with smaller particle size and lower coefficient viscosity, but higher ζ-potential values. These results suggested that phosphorylation could be applied to obtain fish gelatin with varying functional properties suitable for numerous industrial applications.

Physico-mechanical and structural properties of eggshell membrane gelatin- chitosan blend edible films.

This study investigated the physico-mechanical and structural properties of composite edible films based on eggshell membrane gelatin (G) and chitosan (Ch) (75G:25Ch, 50G:50Ch, 25G:75Ch). The results demonstrated that the addition of Ch increased elongation at break significantly (p<0.05), but resulted in no significant change in tensile strength (TS) using 75G:25Ch, 50G:50Ch mixtures in comparison with gelatin-based film. The water solubility and water vapor permeability of the 50G:50Ch film decreased significantly compared to plain films (100G:0Ch and 0G:100Ch) and other composite films (p<0.05). Fourier transform infrared spectroscopy evaluation of structural properties showed that both polymers are totally miscible. Scanning electron microscopy was used to study the morphology of the composite films; it revealed a homogenous and compact structure in 75G:25Ch and 50G:50 Ch. Also, the chemical interactions introduced by the addition of chitosan to eggshell membrane gelatin as new resources could improve the films' functional properties.

Physico-mechanical properties of gelatin films modified with Lysine, Arginine and Histidine.

The aim of this work was to evaluate the plasticizing effect of Lysine (Lys), Arginine (Arg) and Histidine (His) on gelatin films. Gelatin films incorporated with Lys, Arg and His at different levels (0, 0.5 and 1.0%) were prepared. The films without amino acids were prepared as controls. The addition of Lys increased the elongation at break (EAB) but decreased the tensile strength (TS) compared to the control, especially at 1.0% incorporated level. Arg had cross-linking effect on gelatin films through hydrogen bonds demonstrated by FTIR Analysis, which increased TS of the films. His films showed worse EAB, transmission and transparence compared to the control. Furthermore, the addition of Lys and His reduced L* value and a* value with the coincidentally increased b* and ΔE* values compared with the control. Thermal stability (DSC) and UV light barrier of gelatin films were improved by the addition of three kinds of amino acids. Thus, Lys and Arg had potential to improve the properties of gelatin films as natural plasticizer and cross-linker, respectively.

Osteoinductivity of nanostructured hydroxyapatite-functionalized gelatin modulated by human and endogenous mesenchymal stromal cells.

The demand of new strategies for the induction of bone regeneration is continuously increasing. Biomimetic porous gelatin-nanocrystalline hydroxyapatite scaffolds with tailored properties were previously developed, showing a positive response in terms of cell adhesion, proliferation, and differentiation. In the present paper, we focused on their osteoinductive properties. The effect of scaffolds on osteogenic differentiation of human mesenchymal stromal cells (hMSCs) was investigated in vitro. hMSCs were seeded on GEL (type A gelatin) and GEL containing 10 wt% hydroxyapatite (GEL-HA) and cultured in osteogenic medium. Results showed that GEL and GEL-HA10 sustained hMSC differentiation, with an increased ALP activity and a higher expression of bone specific genes. The osteoinductive ability of these scaffolds was then studied in vivo in a heterotopic bone formation model in nude mice. The influence of hMSCs within the implants was examined as well. Both GEL and GEL-HA10 scaffolds mineralized when implanted without hMSCs. On the contrary, the presence of hMSC abolished or reduced mineralization of GEL and GEL-HA10 scaffolds. However, we could observe a species-specific response to the presence of HA, which stimulated osteogenic differentiation of human cells only. In conclusion, the scaffolds showed promising osteoinductive properties and may be suitable for use in confined critical defects. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 914-923, 2018.

Preparation and characterization of alginate and gelatin microcapsules containing Lactobacillus rhamnosus.

This paper describes the preparation and characterization of alginate beads coated with gelatin and containing Lactobacillus rhamnosus. Capsules were obtained by extrusion method using CaCl2 as cross linker. An experimental design was performed using alginate and gelatin concentrations as the variables investigated, while the response variable was the concentration of viable cells. Beads were characterized in terms of size, morphology, scanning electron microscopy (SEM), moisture content, Fourier Transform Infrared Spectrometry (FTIR), thermal behavior and cell viability during storage. The results showed that the highest concentration of viable cells (4.2 x 109 CFU/g) was obtained for 1 % w/v of alginate and 0.1 % w/v of gelatin. Capsules were predominantly spherical with a rough surface, a narrow size distribution ranging from 1.53 to 1.90 mm and a moisture content of 97.70 ± 0.03 %. Furthermore, FTIR and thermogravimetric analysis indicated an interaction between alginate-gelatin. Cell concentration of alginate/gelatin microcapsules was 105 CFU/g after 4 months of storage at 8 oC.

Preparation and characterization of alginate and gelatin microcapsules containing Lactobacillus rhamnosus

ABSTRACT This paper describes the preparation and characterization of alginate beads coated with gelatin and containing Lactobacillus rhamnosus. Capsules were obtained by extrusion method using CaCl2 as cross linker. An experimental design was performed using alginate and gelatin concentrations as the variables investigated, while the response variable was the concentration of viable cells. Beads were characterized in terms of size, morphology, scanning electron microscopy (SEM), moisture content, Fourier Transform Infrared Spectrometry (FTIR), thermal behavior and cell viability during storage. The results showed that the highest concentration of viable cells (4.2 x 109 CFU/g) was obtained for 1 % w/v of alginate and 0.1 % w/v of gelatin. Capsules were predominantly spherical with a rough surface, a narrow size distribution ranging from 1.53 to 1.90 mm and a moisture content of 97.70 ± 0.03 %. Furthermore, FTIR and thermogravimetric analysis indicated an interaction between alginate-gelatin. Cell concentration of alginate/gelatin microcapsules was 105 CFU/g after 4 months of storage at 8 oC.

A novel biomimetic composite substitute of PLLA/gelatin nanofiber membrane for dura repairing.

OBJECTIVE: Biomimetic design will significantly improve growth and regeneration of dural cells and tissue for better repairing effects and fewer complications in repairing the native dura. This study designed a novel composite, biomimetic substitute based on the characteristics of native dura extracellular matrix. METHODS AND RESULTS: This substitute is expected to rapidly induce cell adhesion, migration, and fast regeneration of neotissue. The material characteristics (contact angle, surface charge, and zeta potential were evaluated), in vitro biological characteristics (cell stretch, connections between cells, cell proliferation) and in vivo tissue regeneration capability of this substitute were evaluated, compared to those of collagen dura substitute, the mostly used dura substitute. The results showed that the surface properties of this composite substitute were more biomimetic to native extracellular matrix than collagen substitute did, together with better cytocompatibility, tissue ingrowth, and neoangiogenesis. This composite substitute further demonstrated in clinical case study its ideal repair effect with no CSF leakage or other adverse reactions. CONCLUSION: In conclusion, the new biomimetic composite substitute provides alternative substitute for dura repairing.

Effect of drying method on mechanical, thermal and water absorption properties of enzymatically crosslinked gelatin hydrogels.

Enzymatically crossliked gelatin hydrogel was submitted to two different drying methods: air drying and freeze drying. The resulting polymeric tridimensional arrangement (compact or porous, respectively) led to different thermal and swelling properties. Significant differences (p < 0.05) on thermal and mechanical characteristics as well as swelling in non-enzymatic gastric and intestinal simulated fluids (37 ºC) were detected. Water absorption data in such media was modelled according to Higuchi, Korsmeyer-Peppas, and Peppas-Sahlin equations. Freeze dried hydrogel showed Fickian diffusion behavior while air dried hydrogels presented poor adjustment to Higuchi model suggesting the importance of the relaxation mechanism at the beginning of swelling process. It was possible to conclude that the same gelatin hydrogel may be suitable to different applications depending on the drying process used.

Effect of drying method on mechanical, thermal and water absorption properties of enzymatically crosslinked gelatin hydrogels

ABSTRACT Enzymatically crossliked gelatin hydrogel was submitted to two different drying methods: air drying and freeze drying. The resulting polymeric tridimensional arrangement (compact or porous, respectively) led to different thermal and swelling properties. Significant differences (p < 0.05) on thermal and mechanical characteristics as well as swelling in non-enzymatic gastric and intestinal simulated fluids (37 ºC) were detected. Water absorption data in such media was modelled according to Higuchi, Korsmeyer-Peppas, and Peppas-Sahlin equations. Freeze dried hydrogel showed Fickian diffusion behavior while air dried hydrogels presented poor adjustment to Higuchi model suggesting the importance of the relaxation mechanism at the beginning of swelling process. It was possible to conclude that the same gelatin hydrogel may be suitable to different applications depending on the drying process used.