Immobilization Of Gelatin Research Articles

Hydrophilic PCU scaffolds prepared by grafting PEGMA and immobilizing gelatin to enhance cell adhesion and proliferation.

Gelatin contains many functional motifs which can modulate cell specific adhesion, so we modified polycarbonate urethane (PCU) scaffold surface by immobilization of gelatin. PCU-g-gelatin scaffolds were prepared by direct immobilizing gelatins onto the surface of aminated PCU scaffolds. To increase the immobilization amount of gelatin, poly(ethylene glycol) methacrylate (PEGMA) was grafted onto PCU scaffolds by surface initiated atom transfer radical polymerization. Then, following amination and immobilization, PCU-g-PEGMA-g-gelatin scaffolds were obtained. Both modified scaffolds were characterized by chemical and biological methods. After immobilization of gelatin, the microfiber surface became rough, but the original morphology of scaffolds was maintained successfully. PCU-g-PEGMA-g-gelatin scaffolds were more hydrophilic than PCU-g-gelatin scaffolds. Because hydrophilic PEGMA and gelatin were grafted and immobilized onto the surface, the PCU-g-PEGMA-g-gelatin scaffolds showed low platelet adhesion, perfect anti-hemolytic activity and excellent cell growth and proliferation capacity. It could be envisioned that PCU-g-PEGMA-g-gelatin scaffolds might have potential applications in tissue engineering artificial scaffolds.

Improved immobilization of gelatin on a modified polyurethane urea

In this study, polyurethane urea was surface-modified to elevate cell recognition through immobilization of bioactive gelatin. The poly(urethane urea) was synthesized using poly(ε-caprolactone) diol in the absence of a chain extender. The synthesized polyurethane urea was then functionalized with gelatin (gelatin-grafted poly(urethane urea)) via aminolysis. Chemical changes at the polyurethane urea surface were monitored using titration, water contact angle. Fourier transform infrared, and zeta potential measurements. Significantly larger amounts of gelatin were grafted on the polyurethane urea surface compared to those previously reported for poly(ε-caprolactone) diol (three times more) and polyurethanes (two times more), while the mechanical properties were not compromised. Proliferation of human adipose–derived mesenchymal stem cells on the polyurethane urea and the gelatin-grafted polyurethane urea was evaluated through MTT assay. Although both samples enhanced human adipose–derived mesenchymal stem cells’ proliferation, gelatin-grafted polyurethane urea supported human adipose–derived mesenchymal stem cells’ proliferation at a remarkably higher rate.

Biofunctionalization of poly(l-lactide-co-glycolide) by post-plasma grafting of 2-aminoethyl methacrylate and gelatin immobilization

In the current study, the effect of post-plasma grafting of 2-aminoethyl methacrylate (AEMA) and the subsequent covalent immobilization of gelatin (GelB) on poly(l-lactide-co-glycolide) (PLGA) thin films were investigated. The applied modification resulted in surface chemistry changes of PLGA. More specifically, an increase of nitrogen from 0 at% to 14 at% with a concomitant decrease in carbon and oxygen concentration was observed. The samples were more hydrophilic after the treatment as reflected by a decrease of the water contact angle from 72° to 33° and more rough at the nanoscale as shown by atomic force microscopy (increase of Ra roughness from 0.7nm to 10nm). The growth of osteoblast-like MG-63 cells was enhanced on biofunctionalised PLGA-AEMA-GelB surfaces and the cells were more homogenously distributed than on non-modified PLGA. Our findings are especially important for tissue engineering applications, where substrates supporting homogenous cell cultures are particularly promising.

Radiation-induced biomimetic modification of dual-layered nano/microfibrous scaffolds for vascular tissue engineering

One of the interesting strategies for developing the artificial blood vessels is to generate multi-layered scaffolds for mimicking the structure of native blood vessels such as the intima, media, and adventitia. In this study, we prepared dual-layered poly(L-lactide-co-ɛ-caprolactone) (PLCL) scaffolds with micro- and nanofibers as a basic construct of the vessel using electrospinning methods, which was functionalized using a gelatin through acrylic acid (AAc) grafting by γ-ray irradiation. Based on the microfibrous platform (fiber diameter 5 μm), the thickness of the nanofibrous layer (fiber diameter 700 nm) was controlled from 1.1 ± 0.8 to 32.2 ± 1.7 μm, and the mechanical property of the scaffolds was almost maintained despite the increase in thickness of the nanofibrous layer. The successful AAc graft by γ-ray irradiation could allow the gelatin immobilization on the scaffolds. The proliferation of smooth muscle cells (SMC) on the scaffolds toward a microfibrous layer was approximately 1.3-times greater than in the other groups, and the infiltration was significantly increased, presenting a wide cell distribution in the cross-section. In addition, human umbilical vein endothelial cell (HUVEC) adhesion toward nanofibrous layer was well-managed over the entire surface, and the accelerated proliferation was observed on the gelatin-functionalized scaffolds presenting the well-organized gap-junctions. Therefore, our biomimetic dual-layered scaffolds may be the alternative tools for replacing the damaged blood vessels.

A Novel Route to Polycaprolactone Scaffold for Vascular Tissue Engineering

Polycaprolactone biocomposite scaffolds were prepared by a novel process of precoagulationevaporation and subsequent gelatin immobilization. Water was used as the additive for partial coagulation of the polycaprolactone-chloroform solution. The subsequent coating of the precoagulated solution on a glass surface followed by drying provided a tubular scaffold with regular porosity. The dried scaffolds were treated with carbon dioxide plasma in order to introduce functional groups for enhancing hydrophilicity of the scaffold. Subsequently, gelatin was immobilised on the scaffold by dip coating. Both chemical and morphological changes took place during plasma treatment and gelatin immobilization. The scaffolds were porous and turned out to be hydrophilic. The surfaces were characterized by scanning electron microscopy, atomic force microscopy, tensile measurements and contact angle measurements. These scaffolds were found to offer excellent surfaces for the growth of progenitor-derived endothelial cells.

Immobilization of Gelatin onto Acrylic Acid Grafted Polycaprolactone Monofilament

Immobilization of Gelatin onto Acrylic Acid Grafted Polycaprolactone Monofilament

Immobilization of Gelatin onto Poly(Glycidyl Methacrylate)-Grafted Polycaprolactone Substrates for Improved Cell–Material Interactions

To enhance the cytocompatibility of polycaprolactone (PCL), cell-adhesive gelatin is covalently immobilized onto the PCL film surface via two surface-modified approaches: a conventional chemical immobilization process and a surface-initiated atom transfer radical polymerization (ATRP) process. Kinetics studies reveal that the polymer chain growth from the PCL film using the ATRP process is formed in a controlled manner, and that the amount of immobilized gelatin increases with an increasing concentration of epoxide groups on the grafted P(GMA) brushes. In vitro cell adhesion and proliferation studies demonstrate that cell affinity and growth are significantly improved by the immobilization of gelatin on PCL film surfaces, and that this improvement is positively correlated to the amount of covalently immobilized gelatin. With the versatility of the ATRP process and tunable grafting efficacy of gelatin, this study offers a suitable methodology for the functionalization of biodegradable polyesters scaffolds to improve cell-material interactions.

Immobilization of gelatin onto natural nanofibers for tissue engineering scaffold applications without utilization of any crosslinking agent

Bacterial cellulose was oxidized by periodate oxidation to give rise to 2,3-dialdehyde bacterial cellulose (DABC) with 60.3 ± 0.5 % aldehyde content, which was further reacted with gelatin (Gel) for the immobilization of Gel to form DABC/Gel nanocomposites. The scanning electron microscopy and transmission electron microscopy revealed that DABC/Gel exhibited the refined 3D nano-network structures and the average thickness of Gel coatings in the composites was about 75 nm. FTIR and XPS were utilized to analyze the functional groups and chemical states of DABC/Gel nanocomposites. The results inferred that Gel was fixed on DABC nanofibers via the Schiff base reaction between –NH2 in Gel and –CHO in DABC backbone. NIH3T3 mice fibroblast cells were used for determining the cytocompatibility of the scaffolds. The morphology of the cells was observed through optical inverted microscopy. The results show that DABC/Gel can be used as scaffold material in tissue engineering.

Surface Modification of Smooth Poly(l-lactic acid) Films for Gelatin Immobilization

Poly(L-lactic acid) (PLLA) is widely used in drug delivery and medical implants. Surface modification of PLLA with functional groups to immobilize gelatin or other extracellular matrix proteins is commonly used to improve its cellular affinity. In this work, we use the oxygen plasma to treat PLLA film followed by modification with organosilanes with different functional groups, such as amine, epoxy, and aldehyde groups. Gelatin is then immobilized on the modified PLLA film, which is confirmed by water contact angle measurement, atomic force microscopy (AFM), and laser scanning confocal microscopy (LSCM). Among the used organosilanes, aminosilane is the best one for modification of PLLA used for immobilization of gelatin with the highest efficiency. Moreover, the cellular affinity of gelatin-immobilized PLLA is studied through the evaluation of cell proliferation and focal adhesion using the human umbilical vein endothelial cells (HUVECs). Our experimental results show that the gelatin immobilized on aminosilane- and aldehyde-silane-modified PLLA improves the cellular affinity of HUVECs, whereas that immobilized on epoxy-silane-modified PLLA does not show significant improvement on the cell proliferation.

Immobilization of gelatin on bacterial cellulose nanofibers surface via crosslinking technique

Bacterial cellulose is considered to be a potential material for tissue engineering. However, the absence of enough activity restricts its practical application as tissue engineering scaffold. This paper describes the synthesis of a novel bacterial cellulose/gelatin composite via crosslinking by procyanidin (PA). The morphology of the bacterial cellulose/gelatin composite was observed by field emission scanning electron microscopy (FE-SEM) and transmission electronic microscope (TEM). The composites were further characterized by fourier transformed infrared spectroscopy (FTIR) and X-ray diffraction (XRD). It was found that the 0.25wt.% Gel solution was the appropriate concentration for the BC/Gel composite. Furthermore, the proliferation, infiltration and adhesion of NIH3T3 cells on the BC/Gel-025 composite were evaluated. The results showed that the composite had better bioactivity than pure bacterial cellulose, and the composite supported cell growth.

Enhancement of cardiac myoblast responses onto electrospun PLCL fibrous matrices coated with polydopamine for gelatin immobilization

A variety of surface modification techniques have been proposed to improve the cell-biomaterial interactions. On the other hand, these processes may cleave long-chained polymers, and compromise their mechanical properties. In this study, dopamine was used as a bridge molecule to immobilize gelatin on the poly(L-lactide-co-ɛ-caprolactone) (PLCL) fibrous matrices, which may then be used as a cell delivery carrier. The PLCL fibrous matrices coated with polydopamine by dipping (D-PLCL) can subsequently immobilize gelatin (GD-PLCL). The D-PLCL matrices showed minimal changes in the mechanical properties with a tensile strain of 251.0±33.4% and 247.8±32.1% before and after the coating process, respectively. The cellular activities on the fibrous matrices increased in the order of PLCL<G-PLCL<D-PLCL<GD-PLCL; the H9c2 myoblasts on the GD-PLCL matrices showed approximately two-times higher adhesion and spreading than those on the PLCL matrices, and the proliferation was significantly greater on the GD-PLCL matrices than on the other matrices. Therefore, polydopamine can effectively immobilize the bioactive functional groups on the surface of electrospun fibrous matrices for the development of a tissue specific cell delivery carrier.

Design and Fabrication of Artificial Skin: Chitosan and Gelatin Immobilization on Silicone by Poly Acrylic Acid Graft Using a Plasma Surface Modification Method

Acrylic acid (AAc) was grafted onto the surfaces of polydimethylsiloxane (PDMS) films using a two-step oxygen plasma treatment. The first step of this method included oxygen plasma pretreatment of the PDMS films, immersion in AAc, and drying. The second step was carried out by plasma polymerization of the preadsorbed reactive AAc on the surfaces of the dried pretreated films. Then chitosan and gelatin were immobilized onto the poly(acrylic acid) grafted silicone through covalent bonding. The films were characterized by attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy and water contact angle measurements. Fibroblast cells (L929) were cultured onto the chitosan- and gelatin-immobilized poly(acrylic acid)-grafted silicone and poly(acrylic acid)-grafted silicone films. It was observed that the chitosan- and gelatin-immobilized surfaces showed significant cell growth in comparison with poly(acrylic acid)-grafted silicone samples. It seems that chitosan- and gelatin-immobilized surfaces may have an excellent potential to be used as a derm-like matrix.

Controlling the adhesion and differentiation of mesenchymal stem cells using hyaluronic acid-based, doubly crosslinked networks

We have created hyaluronic acid (HA)-based, cell-adhesive hydrogels that direct the initial attachment and the subsequent differentiation of human mesenchymal stem cells (MSCs) into pre-osteoblasts without osteogenic supplements. HA-based hydrogel particles (HGPs) with an average diameter of 5–6 μm containing an estimated 2.2 wt% gelatin (gHGPs) were synthesized by covalent immobilization of gelatin to HA HGPs prepared via an inverse emulsion polymerization technique. Separately, a photocrosslinkable HA macromer (HAGMA) was synthesized by chemical modification of HA with glycidyl methacrylate (GMA). Doubly crosslinked networks (DXNs) were engineered by embedding gHGPs in a secondary network established by HAGMA at a particle concentration of 2.5 wt%. The resultant composite gels, designated as HA-gHGP, have an average compressive modulus of 21 kPa, and are non-toxic to the cultured MSCs. MSCs readily attached to these gels, exhibiting an early stage of stress fiber assembly 3 h post seeding. By day 7, stellate-shaped cells with extended filopodia were found on HA-gHGP gels. Moreover, cells had migrated deep into the matrix, forming a three dimensional, branched and interconnected cell community. Conversely, MSCs on the control gels lacking gelatin moieties formed isolated spheroids with rounded cell morphology. After 28 days of culture on HA-gHGP, Type I collagen production and mineral deposition were detected in the absence of osteogenic supplements, suggesting induction of osteogenic differentiation. In contrast, cells on the control gels expressed markers for adipogenesis. Overall, the HA-gHGP composite matrix has great promise for directing the osteogenic differentiation of MSCs by providing an adaptable environment through the spatial presentation of cell-adhesive modules.

Gelatin Functionalization of Biomaterial Surfaces: Strategies for Immobilization and Visualization

In the present work, the immobilization of gelatin as biopolymer on two types of implantable biomaterials, polyimide and titanium, was compared. Both materials are known for their biocompatibility while lacking cell-interactive behavior. For both materials, a pre-functionalization step was required to enable gelatin immobilization. For the polyimide foils, a reactive succinimidyl ester was introduced first on the surface, followed by covalent grafting of gelatin. For the titanium material, methacrylate groups were first introduced on the Ti surface through a silanization reaction. The applied functionalities enabled the subsequent immobilization of methacrylamide modified gelatin. Both surface modified materials were characterized in depth using atomic force microscopy, static contact angle measurements, confocal fluorescence microscopy, attenuated total reflection infrared spectroscopy and X-ray photo-electron spectroscopy. The results indicated that the strategies elaborated for both material classes are suitable to apply stable gelatin coatings. Interestingly, depending on the material class studied, not all surface analysis techniques are applicable.

Surface modification of electrospun PLLA nanofibers by plasma treatment and cationized gelatin immobilization for cartilage tissue engineering

Electrospun poly(lactic acid) (PLLA) nanofibers (NF) were modified with cationized gelatin (CG) to improve their compatibility with chondrocytes and to show in vitro and in vivo the potential applications of CG-grafted PLLA nanofibrous membranes (CG-PLLA NFM) as a cartilage tissue engineering scaffold. PLLA NF were first treated with oxygen plasma to introduce –COOH groups on the surface, followed by covalent grafting of CG molecules onto the fiber surface, using water-soluble carbodiimide as the coupling agent. The effects of CG grafting and properties of NFM were characterized by scanning electron microscopy (SEM), transmission electron microscopy, thermogravimetric analysis, atomic force microscope, X-ray photoelectron spectra and Fourier transform infrared spectroscopy. In vitro studies indicated that CG-PLLA NFM could enhance viability, proliferation and differentiation of rabbit articular chondrocytes compared with pristine PLLA NFM. SEM observations of the cell–scaffold construct confirmed the tight attachment of chondrocytes to CG-PLLA NF and in-growth of cells into the interior of the membrane with proper maintenance of cell morphology. Improved cell differentiation in CG-PLLA NFM was confirmed by enhanced glycoaminoglycan and collagen secretion, histological analysis and reverse transcription-polymerase chain reaction studies, which showed that the cells were able to maintain the expression of characteristic markers (collagen II, aggregan and SOX 9) of chondrocytes. Subcutaneous implantation of the cell–scaffold constructs with autologous chondrocytes also confirmed the formation of ectopic cartilage tissues after 28 days by histological examination and immunostaining.

Surface Modification of Polystyrene Beads by Ultraviolet/Ozone Treatment and its Effect on Gelatin Coating

Problem statement: Polystyrene failed to provide any reactive functionality of surface hydrophilicity that is capable of binding proteins. It is known that polystyrene must be chemically modified to make its surface amenable to covalent crosslinking with protein. Approach: The aim of this study was to investigate the effects of UV/ozone treatment on gelatin coating. The surfaces of microsize polystyrene beads (150 µm) were modified by UV/ozone treatment system at different treatment time, ozone flow-rate and UV intensity was analyzed by Design expert software. The treated beads were characterized with ATR-FTIR analysis to determine the introduction of carbonyl (-C=O), carboxylic group (-COOH) and amide group (-CO-NH2) onto the polystyrene surface. Sample characterization was also carried out by SEM and densitometer. Gelatin immobilization was then preceded by incubating treated PS sample in gelatin solution and the total amount of gelatin coated on the modified surface was identified by Bradford assay. Results: The maximum amount of gelatin obtained was 63.75 µg mL-1 while the lowest amount obtained for untreated PS (9.947 µg mL-1). The introduction of carbonyl, hydroxyl and amide group on the polystyrene beads surface was confirmed by ATR-FTIR analysis and thus measures the importance of UV/ozone treatment. Conclusion: From the results, it has been found that time is the most significant factor to prepare samples for gelatin immobilization at reduced flow rate and at an increased Ultraviolet (UV) intensity in the ranges of study.

Covalently immobilized gelatin gradients within three-dimensional porous scaffolds

A stable gelatin gradient providing continuous increment of signaling for cell adhesion and proliferation was fabricated within 3D poly(L-lactic acid) (PLLA) scaffolds. The porous PLLA scaffold fabricated by NaCl particle leaching was vertically fixed on a glass vial. 1,6-Hexanediamine/propanol solution was continuously injected into the vial by a micropump to aminolyze the PLLA scaffold. As a result of reaction time difference, the introduced —NH2 groups increased continuously along with the longitude of the PLLA scaffold in the z-direction. After covalent immobilization of gelatin by glutaraldehyde coupling, the gelatin gradient scaffold was thus obtained. In vitro chondrocyte culture showed that the cells had higher viability and more extending morphology in the gelatin gradient scaffold than that in the uniform gelatin control.

Characterization and cytocompatibility of surface modified polyamide66.

The chemical modification of polyamide66 (PA66) membrane by graft polymerization with methacrylic acid (MAA) was initiated under ultraviolet light. Subsequently, covalent immobilization of bioactive surface was obtained by coupling gelatin to the MAA graft chains with the aid of a water-soluble carbodiimide (WSC). The existence of carboxyl groups grafted on PA66 surface was verified quantitatively by UV-vis spectroscopy. The chemical composition, surface topography, and wettability were investigated by Fourier transform infrared (FTIR) technique, X-ray photoelectron spectroscopy, atomic force microscopy, and water contact angle (WCA) measurement. Results showed that the WCA changed from the original value of 67.5 degrees to the minimum value of 30 degrees after grafting with PMAA. Original PA66 displayed a smooth surface morphology [root mean square (RMS) roughness was around 16 nm]. The modified PA66 surface exhibited an increase in roughness (RMS roughness around 21 nm). Simultaneously, the original and modified PA66 membranes were cultured with MG63 cells to investigate their cytocompatibility. The in vitro biological evaluation demonstrated that the immobilization of gelatin on PA66 membrane acted as a good template for the attachment and proliferation of cells. Also, the less toxic reagents and the moderate reaction conditions involved will be very helpful for the introduction of functional groups onto polymer surface.

Surface engineering of Ti–O films by photochemical immobilization of gelatin

Crystalline Ti–O films were prepared by unbalanced magnetron sputtering and the structure was confirmed by XRD. An organic layer of 3-aminopropylphosphonic acid (APP) was first introduced on the Ti–O films by self-assembling. The stability of the APP on Ti–O films was confirmed by XPS and FTIR analysis. Simultaneously, azido group was introduced in gelatin molecule to act as photoreactive point. The derivated gelatin was spin-coated onto the self-assembled layer and immobilized by UV irradiating. Chemical patterned surface was obtained by using a photomask when irradiating and confirmed by sirius red staining and surface profile analysis. Measured by surface profilometer, the thickness of the immobilized gelatin was about 5–20 nm. The adhering of human endothelial EVC304 cells on APP modified surface was enhanced in the cell culture test. Moreover, the adherence and growth of cells were prior on gelatin-immobilized region visually seen on the patterned surface. This result indicated gelatin-immobilized Ti–O surface can serve as a biocompatible biomaterial for endothelialization.

Gelatin Immobilization on Titanium Surface by Gamma Irradiation

Radiation induced graft polymerization of 2-hydroxyethyl acrylate (HEMA) on titanium surface and couple gelatin have been studied in the paper. The composition and properties of surface modified titanium were investigated by several surface sensitive techniques: X-ray photoelectron spectrum (XPS) and scanning electron microscopy (SEM). Results of the technological measurements show that surface chemistry is affected throughout the surface modification process, finally leading to a complete and homogeneous vinyl compound overlayer on top of the titanium samples. And the modified porous titanium exhibits an interesting mineralization in simulated body fluid (SBF) and an adsorptive property in bovine serum albumin (BSA).