Evaluation of Cross-Linking Methods for Electrospun Gelatin on Cell Growth and Viability (original) (raw)
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In vitro evaluation of crosslinked electrospun fish gelatin scaffolds
Materials Science and Engineering: C, 2013
Gelatin from cold water fish skin was electrospun, crosslinked and investigated as a substrate for the adhesion and proliferation of cells. Gelatin was first dissolved in either water or concentrated acetic acid and both solutions were successfully electrospun. Cross-linking was achieved via three different routes: glutaraldehyde vapor, genipin and dehydrothermal treatment. Solution's properties (surface tension, electrical conductivity and viscosity) and scaffold's properties (chemical bonds, weight loss and fiber diameters) were measured. Cellular viability was analyzed culturing 3T3 fibroblasts plated on the scaffolds and grown up to 7 days. The cells were fixed and observed with SEM or stained for DNA and F-actin and observed with confocal microscopy. In all scaffolds, the cells attached and spread with varying degrees. The evaluation of cell viability showed proliferation of cells until confluence in scaffolds crosslinked by glutaraldehyde and genipin; however the rate of growth in genipin crosslinked scaffolds was slow, recovering only by day five. The results using the dehydrothermal treatment were the less satisfactory. Our results show that glutaraldehyde treated fish gelatin is the most suitable substrate, of the three studied, for fibroblast adhesion and proliferation.
… Research Part A, 2006
Engineering functional three-dimensional (3-D) tissue constructs for the replacement and/or repair of damaged native tissues using cells and scaffolds is one of the ultimate goals of tissue engineering. In this study, non-woven fibrous scaffolds were electrospun from the synthetic biodegradable polymer poly(lactic-co-glycolic acid) (PLGA) and natural proteins, gelatin (denatured collagen) and elastin. In the absence of cross-linking agent, the average PGE fiber diameter increased from 347 ± 103 nm to 999 ± 123 nm upon wetting as measured by scanning electron microscopy. Rat bone marrow stromal cells (rBMSC) were used paradigmatically to study the 3-D cell culture properties of PGE scaffolds. Consistent with the observed properties of the individual fibers, PGE scaffolds initially swelled in aqueous culture medium, however rBMSC seeded PGE scaffolds contracted to < 50% of original size. Time course histological analysis demonstrated uniform seeding of rBMSC into PGE scaffolds and complete cell penetration into the fibrous architecture over 4 weeks of in vitro culture.
Can we achieve biomimetic electrospun scaffolds with gelatin alone?
Frontiers in Bioengineering and Biotechnology, 2023
Introduction: Gelatin is a natural polymer commonly used in biomedical applications in combination with other materials due to its high biocompatibility, biodegradability, and similarity to collagen, principal protein of the extracellular matrix (ECM). The aim of this study was to evaluate the suitability of gelatin as the sole material to manufacture tissue engineering scaffolds by electrospinning. Methods: Gelatin was electrospun in nine different concentrations onto a rotating collector and the resulting scaffold's mechanical properties, morphology and topography were assessed using mechanical testing, scanning electron microscopy and white light interferometry, respectively. After characterizing the scaffolds, the effects of the concentration of the solvents and crosslinking agent were statistically evaluated with multivariate analysis of variance and linear regressions. Results: Fiber diameter and inter-fiber separation increased significantly when the concentration of the solvents, acetic acid (HAc) and dimethyl sulfoxide (DMSO), increased. The roughness of the scaffolds decreased as the concentration of dimethyl sulfoxide increased. The mechanical properties were significantly affected by the DMSO concentration. Immersed crosslinked scaffolds did not degrade until day 28. The manufactured gelatin-based electrospun scaffolds presented comparable mechanical properties to many human tissues such as trabecular bone, gingiva, nasal periosteum, oesophagus and liver tissue. Discussion: This study revealed for the first time that biomimetic electrospun scaffolds with gelatin alone can be produced for a significant number of human tissues by appropriately setting up the levels of factors and their interactions. These findings also extend statistical relationships to a form that would be an excellent starting point for future research that could optimize factors and interactions using both traditional statistics and machine learning techniques to further develop specific human tissue.
The Review on Electrospun Gelatin Fiber Scaffold
Journal of Research Updates in Polymer Science, 2012
The fabrication of the Guided Tissue Regeneration (GTR) membrane materials have become the key technique of the tissue engineering scaffold study. The cells adhere well on the fibers whose dimension is below their own so that the porous three dimension scaffold material can mimic the strueture of the natural extracellular matrix better and have the potential to be an ideal GTR membrane material. Gelatin, a kind of protein obtained from hydrolyzed and denatured animal skin, is a condensation polymer of a variety of amino acids and so it is a kind of bio-polymer with good water-solubility. Gelatin fiber mats with submicro and nanometer scale can simulate extracellular matrix structure of the human tissues and organs and can be used widely in the tissue engineering field because of their excellent bio-affinity. Electrospinning is a very attractive method for preparing polymer or composite nanofibers and so electrospinning technique was developed to prepare nanofibrous gelatin matrix. The electrospun of gelatin to fabricate the scaffold material has obtained more attention recently because of its biocompatibility, high surface area-to-volume ratio, degradability and less immunogenic property. The structure and performance of the electrospinning gelatin fiber mats which were manufactured by different solvents, electrospinning process, cross-linking process were reviewed. The properties and application of the two-component and multicomponent gelatin fiber mats were analyzed.
Materials Letters, 2019
Phytic-acid (PA), in addition to antioxidant and anti-cancer effects, can be considered as a natural crosslinker to improve mechanical properties of gelatin(Ge)/poly(e-caprolactone) (PCL) scaffolds with no toxicity. PA, which is an abundant and naturally available crosslinker, can be an appropriate alternative for costly natural crosslinkers such as genipin for gelatin-based scaffolds. In this study, the mixture of polymers (Ge/PCL: 60/40), containing different ratios of PA, were electrospun. The ability of PA in crosslinking gelatin has been investigated by FTIR, XRD, and DSC where the successful crosslinking process was proved. SEM, porosity, and toxicity tests revealed that fabricated scaffolds could be appropriate options for skin tissue engineering. To evaluate mechanical stabilities, Young's modulus, ultimate strength, and breaking strain of scaffolds have been measured to show a significant elevation of mechanical parameters upon crosslinking. According to results, the optimum amount of PA (7.5%) could increase Young's modulus and breaking strain significantly with non-toxicity effects. Consequently, PA can be applied as an in-situ crosslinker for gelatin-based nanocomposites for manufacturing skin tissue scaffolds.
Journal of Applied Polymer Science, 2015
Gelatin fibers were prepared by electrospinning of gelatin/acetic acid/water ternary mixtures with the aim of studying the feasibility of fabricating gelatin nanofiber mats at room temperature using an alternative benign solvent by significantly reducing the acetic acid concentration. The results showed that gelatin nanofibers can be optimally electrospun with low acetic acid concentration (25% v/v) combined with gelatin concentrations higher than 300 mg/ml. Both gelatin solutions and electrospun gelatin mats (prepared with different acetic acid aqueous solutions) were analyzed by FTIR and DSC techniques in order to determine the chemical and structure changes of the polymer. The electrospun gelatin mats fabricated from solutions with low acetic acid content showed some advantages as the maintenance of the decomposition temperature of the pure gelatin (~230ºC) and the reduction of the acid content on electrospun mats, which allowed to reach a cell viability upper than 90% (analyzed by cell viability test using human dermal fibroblast and embryonic kidney cells). This study has also analyzed the influence of gelatin and acetic acid concentration both on the solution viscosity and the electrospun fiber diameter, obtaining a clear relationship between these parameters.
Crosslinking of the electrospun gelatin nanofibers
Polymer, 2006
Gelatin (Gt) nanofibers have been prepared by using an electrospinning process in a previous study. In order to improve their water-resistant ability and thermomechnical performance for potential biomedical applications, in this article, the electrospun gelatin nanofibers were crosslinked with saturated glutaraldehyde (GTA) vapor at room temperature. An exposure of this nanofibrous material in the GTA vapor for 3 days generated a crosslinking extent sufficient to preserve the fibrous morphology tested by soaking in 37 8C warm water. On the other hand, the crosslinking also led to improved thermostability and substantial enhancement in mechanical properties. The denaturation temperature corresponding to the helix to coil transition of the air-dried samples increased by about 11 8C and the tensile strength and modulus were nearly 10 times higher than those of the as-electrospun gelatin fibers. Furthermore, cytotoxicity was evaluated based on a cell proliferation study by culturing human dermal fibroblasts (HDFs) on the crosslinked gelatin fibrous scaffolds for 1, 3, 5 and 7 days. It was found cell expansion took place and almost linearly increased during the course of whole period of the cell culture. The initial inhibition of cell expansion on the crosslinked gelatin fibrous substrate suggested some cytotoxic effect of the residual GTA on the cells.
Journal of Biomedical Materials Research Part A, 2010
Nanofibrous scaffolds were prepared from gelatin solutions and were further cross-linked with glutaraldehyde (GA). The fiber diameter was varied from 100 to 1000 nm by controlling the applied voltage (4-15 kV) and the concentration of the gelatin solution (4-15%). The tensile moduli and the tensile strength of the noncross-linked scaffolds varied from 20 to 120 MPa and 0.5 to 3.5 MPa, respectively. Cross-linking with GA led to an increase in both the tensile modulus and strength and correlated with cross-linker concentration. Gelatin-based matrices were characterized by Fourier transform infrared spectroscopy and differential scanning calorimetry. High cellular viabilities and rounded morphology of chondrocytes was observed at the end of 7 days in culture with added matrix deposition and flattening of cells at 15 days. Matrix stiffness was noted to impact cell densities and the expression of chondrocytic markers, especially aggrecan. The ratios of collagen-II (C-II) to collagen-I (C-I) of 0.62 and 1.33 were noted on gelatin nanofibrous scaffolds cross-linked with 0.1% GA at the end of 7 and 15 days in culture, respectively. C-II/C-I ratios of 1.30 and 2.58 were noted on scaffolds cross-linked with 1.0% GA at the end of 7 and 15 days in culture, respectively.
Acta Biomaterialia, 2011
The development of suitable biomimetic three-dimensional scaffolds is a fundamental requirement of tissue engineering. This paper presents the first successful attempt to obtain electrospun gelatin nanofibers cross-linked with a low toxicity agent, genipin, and able to retain the original nanofiber morphology after water exposure. The optimized procedure involves an electrospinning solution containing 30 wt.% gelatin in 60/40 acetic acid/water (v/v) and a small amount of genipin, followed by further cross-linking of the as-electrospun mats in 5% genipin solution for 7 days, rinsing in phosphate-buffered saline and then air drying at 37°C. The results of scanning electron microscopy investigations indicated that the cross-linked nanofibers were defect free and very regular and they also maintained the original morphology after exposure to water. Genipin addition to the electrospinning solution dramatically reduced the extensibility of the as-electrospun mats, which displayed further remarkable improvements in elastic modulus and stress at break after successive cross-linking up to values of about 990 and 21 MPa, respectively. The results of the preliminary in vitro tests carried out using vascular wall mesenchymal stem cells indicated good cell viability and adhesion to the gelatin scaffolds.