Materials For Tissue Engineering Applications Research Articles

Synergistic Effects of Psidium Guajava and Copper Nanoparticles Reinforced Hybrid Hydrogel for Tissue Engineering

Hydrogels are biopolymers proficient in engrossing much water in their 3D network structure. However, single-polymer hydrogels frequently experience poor physio-mechanical properties, confining their border applications. The present work concentrated on developing chemically crosslinked hydrogels using the terpolymerization of gelatin (GEL), guar gum (GGM), and polyvinyl alcohol (PVA). Ethanolic extract of Psidium guajava leaf (EPG) and copper nanoparticles (CuNPs) were added to enhance the biomechanical properties of the developed hydrogels. Hydrogels' viscoelastic, mechanical, swelling, and cytotoxicity properties were assessed. All the hydrogels exhibited a porous-like structure with a swelling index of 230 to 280%. A compressive strength of 5 MPa with splendid chondrocyte viability was noticed in the hydrogels comprised of EPG and CuNPs. The multiple interactions among the polymer chains impart better frequency and shear strain-dependent behavior. The time-dependent frictional behavior of hydrogel under the lubrication of artificial synovial fluid reveals the decreased coefficient of friction over time. The performance of the hybrid hydrogel enhanced with EPG and CuNPs was superior, making it a promising material for tissue engineering applications.

Biocompatibility and antimicrobial efficacy of silver-doped borosilicate bioactive glass for tissue engineering application

Borate bioactive glasses are an auspicious material for tissue engineering applications due to their enhanced dissolution rate, bioactivity, and capacity to integrate therapeutic ions. In this study, borate-based S49B4 bioactive glass doped with silver at mass fraction of 0.5, 1, and 3 wt% were studied for bioactivity, degradation, antibacterial, and cytocompatibility. Thermogravimetric analysis revealed that the bioactive glasses were thermally stable between 600 and 700 °C. Fourier transform infrared spectroscopy and X-ray diffraction confirmed the successful synthesis of an amorphous phase of the doped borosilicate bioactive glasses and the incorporation of silver ion crystals within the structure, as well as associated contributions from borate and silicate network formers in the borosilicate bioactive glass. Morphological evaluation revealed that the borosilicate bioactive glasses exhibit a uniform and spherical shape across all formulations, with the mean particle size varying from 65 to 76 nm. An in-vitro acellular bioactivity in simulated body fluid medium showed that increasing the silver content increased the degradation rate and pH. Besides, scanning electron microscopy and Energy dispersive X-ray spectroscopy analysis revealed an upsurge in apatite production on the BBGs' surfaces as well as incremental Calcium-Phosphate ratio values of 1.50, 1.65 and 1.70 as the silver content increases. The antibacterial effect was tested against Escherichia coli and Staphylococcus aureus, while cytocompatibility was tested against human gingival epithelial cells. Silver integration at 1 wt percent yielded the most promising outcomes, which were particularly bactericidal at 79.8 % for Escherichia coli and 93.41 % for Staphylococcus aureus. Similarly, its Lactate Dehydrogenase percentage is significantly similar to the negative control employed in the study, indicating its biocompatibility. In contrast, 3 wt% silver exhibited the maximum bactericidal activity while also exhibiting mild cytotoxicity. In summary, our research indicates that elevated silver concentration enhances the bioactivity and antimicrobial characteristics of borosilicate bioactive glasses; nevertheless, a higher silver weight percent in this study also increases the possibility of cytotoxicity. It is therefore essential to carefully regulate the amount of silver doping at lower concentrations in order to maximize antibacterial action and minimize toxicity to human cells. The results presented here contribute to our understanding of the prospective use of silver doped borosilicate bioactive glasses as a possible material for tissue engineering applications.

Fabrication, characterization and numerical validation of a novel thin-wall hydrogel vessel model for cardiovascular research based on a patient-specific stenotic carotid artery bifurcation

In vitro vascular models, primarily made of silicone, have been utilized for decades for studying hemodynamics and supporting the development of implants for catheter-based treatments of diseases such as stenoses and aneurysms. Hydrogels have emerged as prominent materials in tissue-engineering applications, offering distinct advantages over silicone models for fabricating vascular models owing to their viscoelasticity, low friction, and tunable mechanical properties. Our study evaluated the feasibility of fabricating thin-wall, anatomical vessel models made of polyvinyl alcohol hydrogel (PVA-H) based on a patient-specific carotid artery bifurcation using a combination of 3D printing and molding technologies. The model’s geometry, elastic modulus, volumetric compliance, and diameter distensibility were characterized experimentally and numerically simulated. Moreover, a comparison with silicone models with the same anatomy was performed. A PVA-H vessel model was integrated into a mock circulatory loop for a preliminary ultrasound-based assessment of fluid dynamics. The vascular model's geometry was successfully replicated, and the elastic moduli amounted to 0.31 ± 0.007 MPa and 0.29 ± 0.007 MPa for PVA-H and silicone, respectively. Both materials exhibited nearly identical volumetric compliance (0.346 and 0.342% mmHg−1), which was higher compared to numerical simulation (0.248 and 0.290% mmHg−1). The diameter distensibility ranged from 0.09 to 0.20% mmHg−1 in the experiments and between 0.10 and 0.18% mmHg−1 in the numerical model at different positions along the vessel model, highlighting the influence of vessel geometry on local deformation. In conclusion, our study presents a method and provides insights into the manufacturing and mechanical characterization of hydrogel-based thin-wall vessel models, potentially allowing for a combination of fluid dynamics and tissue engineering studies in future cardio- and neurovascular research.

Self-Reinforced PTLG Copolymer with Shish Kebab Structures and a Bionic Surface as Bioimplant Materials for Tissue Engineering Applications.

Green and biodegradable materials with great mechanical properties and biocompatibility will offer new opportunities for next-generation high-performance biological materials. Herein, the novel oriented shish kebab crystals of a novel poly(trimethylene carbonate-lactide-glycolide) (PTLG) vascular stent are first reported to be successfully fabricated through a feasible solid-state drawing process to simultaneously enhance the mechanical performance and biocompatibility. The crystal structure of this self-reinforced vascular stent was transformed from spherulites to a shish kebab crystal, which indicates the mechanical interlocking effect and prevents the lamellae from slipping with a significant improvement of mechanical strength to 333 MPa. Meanwhile, it is different from typical biomedical polymers with smooth surface structures, and the as-obtained PTLG vascular stent exhibits a bionic surface morphology with a parallel micro groove and ridge structure. These ridges and grooves were attributed to the reorganization of cytoskeleton fiber bundles following the direction of blood flow shear stress. The structure and parameters of these morphologies were highly similar to the inner surface of blood vessels of the human, which facilitates cell adhesion growth to improve its proliferation, differentiation, and activity on the surface of PTLG.

Advanced Bioink Materials for Tissue Engineering Applications

Advanced Bioink Materials for Tissue Engineering Applications

Chiral nanomaterials in tissue engineering.

Addressing significant medical challenges arising from tissue damage and organ failure, the field of tissue engineering has evolved to provide revolutionary approaches for regenerating functional tissues and organs. This involves employing various techniques, including the development and application of novel nanomaterials. Among them, chiral nanomaterials comprising non-superimposable nanostructures with their mirror images have recently emerged as innovative biomaterial candidates to guide tissue regeneration due to their unique characteristics. Chiral nanomaterials including chiral fibre supramolecular hydrogels, polymer-based chiral materials, self-assembling peptides, chiral-patterned surfaces, and the recently developed intrinsically chiroptical nanoparticles have demonstrated remarkable ability to regulate biological processes through routes such as enantioselective catalysis and enhanced antibacterial activity. Despite several recent reviews on chiral nanomaterials, limited attention has been given to the specific potential of these materials in facilitating tissue regeneration processes. Thus, this timely review aims to fill this gap by exploring the fundamental characteristics of chiral nanomaterials, including their chiroptical activities and analytical techniques. Also, the recent advancements in incorporating these materials in tissue engineering applications are highlighted. The review concludes by critically discussing the outlook of utilizing chiral nanomaterials in guiding future strategies for tissue engineering design.

Multicomponent Peptide-Based Hydrogels Containing Chemical Functional Groups as Innovative Platforms for Biotechnological Applications.

Multicomponent hydrogels (HGs) based on ultrashort aromatic peptides have been exploited as biocompatible matrices for tissue engineering applications, the delivery of therapeutic and diagnostic agents, and the development of biosensors. Due to its capability to gel under physiological conditions of pH and ionic strength, the low molecular-weight Fmoc-FF (Nα-fluorenylmethoxycarbonyl-diphenylalanine) homodimer is one of the most studied hydrogelators. The introduction into the Fmoc-FF hydrogel of additional molecules like protein, organic compounds, or other peptide sequences often allows the generation of novel hydrogels with improved mechanical and functional properties. In this perspective, here we studied a library of novel multicomponent Fmoc-FF based hydrogels doped with different amounts of the tripeptide Fmoc-FFX (in which X= Cys, Ser, or Thr). The insertion of these tripeptides allows to obtain hydrogels functionalized with thiol or alcohol groups that can be used for their chemical post-derivatization with bioactive molecules of interest like diagnostic or biosensing agents. These novel multicomponent hydrogels share a similar peptide organization in their supramolecular matrix. The hydrogels' biocompatibility, and their propensity to support adhesion, proliferation, and even cell differentiation, assessed in vitro on fibroblast cell lines, allows us to conclude that the hybrid hydrogels are not toxic and can potentially act as a scaffold and support for cell culture growth.

Electrospinning and nanofibre applications: fundamentals and recent status

A field that is constantly growing is the bio fabrication of biomimetic materials for tissue engineering applications. Particularly intriguing are the mechanical and structural features that nano fibrous scales can emulate (e.g., collagen fibres). This review provides a broad overview of the production of nanofibers with a focus on the creation and use of electrospun nano fibrous scaffolds. Electrospinning allows for the creation of mats with precise fibre arrangements and structural integrity using a variety of biodegradable biopolymers. This review also lists some production process benefits and drawbacks. Also, the characteristics of the nanofibers that can be created using each process are illustrated together with the electrospinning techniques for producing nanofibers.

Optimization of Gelatin Methacryloyl Hydrogel Properties through an Artificial Neural Network Model.

Gelatin methacryloyl (GelMA) hydrogels are promising materials for tissue engineering applications due to their biocompatibility and tunable properties. However, the time-consuming process of preparing GelMA hydrogels with desirable properties for specific biomedical applications limits their clinical use. Visible-light-induced cross-linking is a well-known method for the preparation of GelMA hydrogels; however, a comprehensive investigation on the influence of critical parameters such as Eosin Y (EY), triethanolamine (TEA), and N-vinyl-2-pyrrolidone (NVP) concentrations on the stiffness and gelation time has yet to be performed. In this study, we systematically investigated the effect of these critical parameters on the stiffness and gelation time of GelMA hydrogels. We developed an artificial neural network (ANN) model with three input variables, EY, TEA, and NVP concentrations, and two output variables, Young's modulus and gelation time, derived from our experimental design. Through the alteration of individual chemical concentrations, [EY] between 0.005 and 0.5 mM and [TEA] and [NVP] between 10 and 1000 mM, we studied the impact of these alterations on the real-time values of stiffness and gelation time. Furthermore, we demonstrated the validity of the ANN model in predicting the properties of GelMA hydrogels. We also studied cell survival to establish nontoxic concentration ranges for each component, enabling safer use of GelMA hydrogels in relevant biomedical applications. Our results showed that the ANN model can accurately predict the properties of GelMA hydrogels, allowing for the synthesis of hydrogels with desirable stiffness for various biomedical applications. In conclusion, our study provides a comprehensive library that characterizes the stiffness and gelation time and demonstrates the potential of the ANN model to predict these properties of GelMA hydrogels depending on the critical parameters. The ANN models developed here can facilitate the optimization of GelMA hydrogels with the most efficient mechanical properties that resemble a native extracellular matrix and better address the need in the in vivo microenvironment. The approach of this study is to bring research about the synthesis of GelMA hydrogels to a new level where the synthesis of these hydrogels can be standardized with minimum cost and effort.

Biopolymeric conjugation with synthetic fibers and applications

Abstract Presently, several different kinds of polymer composite materials of varying properties have been developed and these composite materials play a vital role in construction and automotive industries. Polymer composites are normally preferred owing to some of their unique properties such as light weight, low cost, good surface finishes, more durability, and non-corrosiveness. But it is a challenge to environmental sustainability, therefore researchers are emphasizing on development of new modified biodegradable polymer composite materials. The biopolymer matrix reinforced by synthetic fibers is a viable alternative, which exhibits adequate mechanical properties and biodegradability. Although various advanced and improved composite materials are developed by using synthetic fibers, natural fibers, and nanoparticles, the use of synthetic fibers as reinforcing material is cost effective and shows improved performance. Among the various kinds of synthetic fibers, normally glass fibers (GF) in the form of short fiber are the most widely used reinforcing material, which is cost effective, provides good impact resistance, stiffness, strength, thermal stability, and chemical resistance. For requirement of high stiffness of the composite material, carbon fibers (CF) are more suitable than GF. Some other synthetic fibers such as aramid (AF), polypropylene fibers (PP-F), polyacrylonitrile fiber (PAN-F), basalt (BF), and polyethylene terephthalate fiber (PET-F) are some cases used as reinforcing material for synthesis of composites. The composite reinforced with synthetic fibers are used as a highly suitable material for manufacturing of various components in cars, space vehicles and railways. Recently some new hybrid composite materials are developed by using both natural and synthetic fibers as reinforcing material, which exhibits dynamic thermal, mechanical properties and potentially suitable from automobile to construction industry. Recently, numerous new biomaterial composite has been developed by using biopolymer as matrix with reinforcement of various kinds of synthetic fibers, which are used as good implant material for tissue engineering applications.

Ethylene glycol-based thermoresponsive block copolymer brushes with cell-affinity peptides for thermally controlled interaction with target cells

Tissue engineering has recently attracted attention as a potential remedy for intractable diseases. To be effective, such treatments require cell separation methods that do not modify cellular surfaces. In this study, we developed cell separation materials using ethylene glycol-based thermoresponsive block copolymer brushes and cell-affinity peptides. A poly(2-hydroxyethyl methacrylate-co-propargyl acrylate) (P(HEMA-co-PgA)) brush was grafted onto a glass substrate through atom transfer radical polymerization (ATRP). Subsequently, poly(2-(2-methoxyethoxy)ethyl methacrylate) (PMEO2MA), P(MEO2MA-co-HEMA-co-poly(ethylene glycol) methacrylate [PEGMA]), or P(MEO2MA-co-PEGMA) was grafted onto the P(HEMA-co-PgA) brush-coated substrates through a second ATRP. A Gly-Gly-Gly-Arg-Glu-Asp-Val (GGGREDV) peptide was conjugated to the copolymer brush via a click reaction. The prepared copolymer brushes exhibited thermoresponsive properties. The block copolymer brushes having the P(MEO2MA-co-HEMA-co-PEGMA) and P(MEO2MA-co-PEGMA) segment exhibited effective human umbilical vein endothelial cells (HUVECs) adhesion and normal human dermal fibroblasts (NHDFs) repulsion at 37 °C. By reducing temperature to 20 °C, adherent HUVECs were successfully recovered from the copolymer brushes. Using these copolymer brushes, HUVECs were separated from contaminant NHDFs and smooth muscle cells with these simple changes in temperature. The development of thermoresponsive ethylene glycol-based copolymer brushes with affinity peptides could be a useful cell separation material for tissue engineering applications.

Biomedical Applications of the Biopolymer Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV): Drug Encapsulation and Scaffold Fabrication.

Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) is a biodegradable and biocompatible biopolymer that has gained popularity in the field of biomedicine. This review provides an overview of recent advances and potential applications of PHBV, with special emphasis on drug encapsulation and scaffold construction. PHBV has shown to be a versatile platform for drug delivery, offering controlled release, enhanced therapeutic efficacy, and reduced side effects. The encapsulation of various drugs, such as anticancer agents, antibiotics, and anti-inflammatory drugs, in PHBV nanoparticles or microspheres has been extensively investigated, demonstrating enhanced drug stability, prolonged release kinetics, and increased bioavailability. Additionally, PHBV has been used as a scaffold material for tissue engineering applications, such as bone, cartilage, and skin regeneration. The incorporation of PHBV into scaffolds has been shown to improve mechanical properties, biocompatibility, and cellular interactions, making them suitable for tissue engineering constructs. This review highlights the potential of PHBV in drug encapsulation and scaffold fabrication, showing its promising role in advancing biomedical applications.

Dual-Crosslinking of Gelatin-Based Hydrogels: Promising Compositions for a 3D Printed Organotypic Bone Model.

Gelatin-based hydrogels have emerged as a popular scaffold material for tissue engineering applications. The introduction of variable crosslinking methods has shown promise for fabricating stable cell-laden scaffolds. In this work, we examine promising composite biopolymer-based inks for extrusion-based 3D bioprinting, using a dual crosslinking approach. A combination of carefully selected printable hydrogel ink compositions and the use of photoinduced covalent and ionic crosslinking mechanisms allows for the fabrication of scaffolds of high accuracy and low cytotoxicity, resulting in unimpeded cell proliferation, extracellular matrix deposition, and mineralization. Three selected bioink compositions were characterized and the respective cell-laden scaffolds were bioprinted. Temporal stability, morphology, swelling, and mechanical properties of the scaffolds were thoroughly studied and the biocompatibility of the constructs was assessed using rat mesenchymal stem cells while focusing on osteogenesis. Experimental results showed that the composition of 1% alginate, 4% gelatin, and 5% (w/v) gelatine methacrylate, was found to be optimal among the examined, with shape fidelity of 88%, large cell spreading area and cell viability at around 100% after 14 days. The large pore diameters that exceed 100 µm, and highly interconnected scaffold morphology, make these hydrogels extremely potent in bone tissue engineering and bone organoid fabrication.

A native extracellular matrix material for tissue engineering applications: Characterization of pericardial fluid.

Tissue engineering applications are widely used to repair and regenerate damaged tissues and organs. A scaffold, which is an important component in tissue engineering, provides a 3D environment for cells. In this study, the usability of PF components for the production of an ideal scaffold was investigated. For this aim, pericardial fluid (PF) was harvested from the bovine heart, then its structure and components were characterized. The results of Raman spectroscopy analysis, histological staining, and scanning electron microscopy (SEM) shows that the pericardial fluid contains collagen type I and IV, elastin, fibrin, and glycosaminoglycan (GAG), which are native extracellular matrix (ECM) components. The results demonstrated that (i) PF contains native ECM proteins and GAG such as collagen types I, III, and IV, elastin, and fibrin. (ii) The PF is highly similar to the native ECM structure. (iii) PF can significantly contribute to many tissue engineering studies as a native ECM material to increase the biocompatibility of biomaterials and to several in vitro/in vivo cell culture studies. (iv) PF containing multiple ECM molecules, can be used alone or together with hyaluronic acid, poly(ethylene glycol) (PEG), alginate, chitosan, matrigel, and gelatin methacryloyl (GelMA) materials in bioprinting systems for eliminating the disadvantages of these materials.

3D printed bio-ceramic loaded PEGDA/vitreous carbon composite: Fabrication, characterization, and life cycle assessment

Stereolithography (SLA) is gaining vast popularity in the development of three-dimensional parts with customized materials for tissue engineering applications. Consequently, developing customized materials like bio-composites (bio-polymers and bio-ceramics) is the basic pillar that meets the application requirement. Photo-crosslinkable poly(ethylene glycol) diacrylate (PEGDA) has outstanding biocompatibility and biophysical properties for tissue engineering. But, because of its poor mechanical properties, its scope is limited to load-bearing applications. This research aims to enhance the mechanical and tribological properties of PEGDA by reinforcing Vitreous Carbon (VC) bioceramic. Therefore, 1 to 5 wt % of VC have been added in PEGDA and developed novel PEGDA/VC composite resins for SLA. The rheological and sedimentation test have been performed to check the suitability for SLA printing. Afterward, printed materials have been characterized by fourier transform infrared spectrometer, X-ray diffraction, thermogravimetric analysis, optical profilometer, and scanning electron microscope. Moreover, tensile compression, flexural, and tribological properties have been evaluated. It was found that the addition of VC in PEGDA enhanced its mechanical, thermal, and tribological properties. Besides, a life cycle assessment of materials and energy resources in SLA process has been performed to investigate the environmental impact.

Naturally Derived Maleic Chitosan/Thiolated Sodium Hyaluronate Hydrogels as Potential 3D Printing Tissue Engineering Materials

Abstract3D printing is an attractive method to accurately construct artificial organs or alternative materials with complicated structures and functional performance. Naturally derived hydrogels have emerged as promising materials for the preparation of biomimetic 3D organization or scaffolds by 3D printing due to their good biocompatibility, high water content, and fascinating 3D network. However, the poor printing properties and weak structural stability of naturally derived hydrogels limit their applications. In this study, photopolymerizable hydrogels are designed based on maleic chitosan (MCS) and thiolated sodium hyaluronate (SHHA). The Michael addition between MCS and SHHA improves the viscosity of the mixed solution. Moreover, it benefits the 3D printing process, followed by photopolymerization (acrylate‐thiol step‐chain polymerization and acrylate–acrylate chain polymerization) to form a stable covalent network rapidly. The rheological property, swelling behaviors, microstructure, and in vitro degradation are tuned by adjusting the molar ratio of the thiol group and acrylate group. In addition, MCS/SHHA hydrogel scaffolds with good accuracy and enhanced structural stability are prepared using extrusion‐based 3D printing and photopolymerization technology. The hydrogels display excellent cytocompatibility and can support adherence of L929 cells, which can be used as prospective materials for tissue engineering applications.

An injectable, self-healing and degradable hydrogel scaffold as a functional biocompatible material for tissue engineering applications

Injectable hydrogels derived from natural extracellular matrices exhibit excellent adhesion to endothelial cells in vitro and are ideal for many biomedical applications. However, their applicability in vivo is limited by the risk of infection or immunogenicity, and the current injectables also suffer from degradation, viscosity, and drug release. In this study, a multifunctional hydrogel scaffold (COB hydrogels) was constructed by incorporating bioactive glass nanoparticles with a Schiff base crosslinking-based hydrogel composed of carboxymethyl chitosan and oxidized cellulose. The incorporation of nanoparticles not only shortened the gelation time of the COB hydrogels, but also enhanced the performance of the hydrogel in terms of function, such as drug loading capacity. The prepared hydrogels also have self-healing ability, injectability, drug loading and sustained release, antibacterial properties and biocompatibility. In addition, given their no cytotoxicity and mild inflammation in vivo, the hydrogel scaffolds will be important for tissue engineering and drug delivery applications.

Polyhydroxybutyrate (PHB) in nanoparticulate form improves physical and biological performance of scaffolds

Polyhydroxyalkanoates (PHAs) are natural polyesters produced by microorganisms as a source of intracellular energy reserves. Due to their desirable material characteristics, these polymers have been thoroughly investigated for tissue engineering and drug delivery applications. A tissue engineering scaffold serves as a substitute of the native extracellular matrix (ECM) and plays a crucial role in tissue regeneration by providing temporary support for cells during natural ECM formation. In this study, porous, biodegradable scaffolds were prepared using native polyhydroxybutyrate (PHB) and PHB in nanoparticulate form using salt leaching method, to investigate the differences in the physicochemical properties such as crystallinity, hydrophobicity, surface morphology, roughness, and surface area and biological properties of the prepared scaffolds. As per the BET analysis, PHB nanoparticles-based (PHBN) scaffolds presented a significant difference in the surface area as compare to PHB scaffolds. PHBN scaffolds showed decreased crystallinity and improved mechanical strength as compared to PHB scaffolds. Thermogravimetry analysis shows delayed degradation of PHBN scaffolds. An examination of Vero cell lines' cell viability and adhesion over time revealed enhanced performance of PHBN scaffolds. Our research suggests that scaffold made of PHB nanoparticles could serve as a superior material for tissue engineering applications than its native form.

Porous Thermoplastic Molded Regenerated Silk Crosslinked by the Addition of Citric Acid.

Thermoplastic molded regenerated silk fibroin was proposed as a structural material in tissue engineering applications, mainly for application in bone. The protocol allows us to obtain a compact non-porous material with a compression modulus in the order of a Giga Pascal in dry conditions (and in the order of tens of MPa in wet conditions). This material is produced by compressing a lyophilized silk fibroin powder or sponge into a mold temperature higher than the glass transition temperature. The main purpose of the produced resin was the osteofixation and other structural applications in which the lack of porosity was not an issue. In this work, we introduced the use of citric acid in the thermoplastic molding protocol of silk fibroin to obtain porosity inside the structural material. The citric acid powder during the compression acted as a template for the pore formation. The mean pore diameter achieved by the addition of the higher amount of citric acid was around 5 μm. In addition, citric acid could effectively crosslink the silk fibroin chain, improving its mechanical strength. This effect was proved both by evaluating the compression modulus (the highest value recorded was 77 MPa in wet conditions) and by studying the spectra obtained by Fourier transform infrared spectroscopy. This protocol may be applied in the near future to the production of structural bone scaffolds.

Gelatin methacryloyl (GelMA) loaded with concentrated hypoxic pretreated adipose-derived mesenchymal stem cells(ADSCs) conditioned medium promotes wound healing and vascular regeneration in aged skin

BackgroundAging skin is characterized by a disturbed structure and lack of blood supply, which makes it difficult to heal once injured. ADSCs secrete large amounts of cytokines, which promote wound healing and vascular regeneration through paracrine secretion, and the number of cytokines can be elevated by hypoxic pretreating. However, the components of ADSCs are difficult to retain in wounds. Gelatin methacrylate (GelMA) is a photopolymerizable hydrogel synthesized from gelatin and has recently emerged as a potentially attractive material for tissue engineering applications. GelMA loaded with concentrated hypoxic pretreated ADSCs conditioned medium could provide a new method of treating wounds in aged skin.MethodsPrimary ADSCs were isolated from human adipose tissue and characterized by flow cytometry and differentiation test. ADSCs in passages 4-6 were pretreated in the hypoxic and normoxic environments to collect conditioned medium, the conditioned medium was then concentrated to prepare concentrated ADSCs conditioned medium(cADSC-CM)(the one collected from ADSCs under hypoxia was called hypo-CM ,and the one from normoxia was called nor-CM). The concentration of cytokines was detected. After treated with cADSC-CM, the abilities of proliferation, migration, and tube formation of human umbilical vascular endothelial cells (HUVECs) were assayed, and Akt/mTOR and MAPK signal pathway was detected using western blotting. GelMA+hypo-CM hydrogel was prepared, and a comprehensive evaluation of morphology, protein release efficiency, degradation rate, mechanical properties, and rheology properties were performed. Full-thickness skin wounds were created on the backs of 20-month-old mice. After surgery, GelMA, GelMA+F12, GelMA+hypo-CM, and GelMA+nor-CM were applied to the wound surface respectively. H&E, Masson, and immunohistochemistry staining were performed, and a laser Doppler perfusion imager was used to evaluate the blood perfusion. The student’s t-test was used for analysis between two groups and a one-way analysis of variance (ANOVA) was used for analysis among multi groups.ResultsOur results revealed that 1) wounds in aged skin healed more slowly than that in young skin and exhibited poorer perfusion; 2) hypoxic pretreated ADSCs secreted more cytokines including VEGF by activating HIF1α; 3) hypo-CM promoted proliferation and migration of HUVECs through VEGF/Akt/mTOR and MAPK signal pathway; 4) GelMA-hypoCM accelerated wound healing and angiogenesis in aged skin in vivo.ConclusionGelMA loaded with concentrated hypoxic pretreated adipose-derived mesenchymal stem cells conditioned medium could accelerate wound healing in aged skin by promoting angiogenesis.