Surfactant-assisted photo-crosslinked silk fibroin sponges: A versatile platform for the design of bone scaffolds.

Critical size bone defects cannot heal without aid and current clinical approaches exhibit some limitations, underling the need for novel solutions. Silk fibroin, derived from silkworms, is widely utilized in tissue engineering and regenerative medicine due to its remarkable properties, making it a promising candidate for bone tissue regeneration in vitro and in vivo. However, the clinical translation of silk-based materials requires refinements in 3D architecture, stability, and biomechanical properties. In earlier research, improved mechanical resistance and stability of chemically crosslinked methacrylate silk fibroin (Sil-Ma) sponges over physically crosslinked counterparts were highlighted. Furthermore, the influence of photo-initiator and surfactant concentrations on silk properties was investigated. However, the characterization of sponges with Sil-Ma solution concentrations above 10 % (w/V) was hindered by production optimization challenges, with only cell viability assessed. This study focuses on the evaluation of methacrylate sponges' suitability as temporal bone tissue regeneration scaffolds. Sil-Ma sponge fabrication at a fixed concentration of 20 % (w/V) was optimized and the impact of photo-initiator (LAP) concentrations and surfactant (Tween 80) presence/absence was studied. Their effects on pore formation, silk secondary structure, mechanical properties, and osteogenic differentiation of hBM-MSCs were investigated. We demonstrated that, by tuning silk sponges' composition, the optimal combination boosted osteogenic gene expression, offering a strategy to tailor biomechanical properties for effective bone regeneration. Utilizing Design of Experiment (DoE), correlations between sponge composition, porosity, and mechanical properties are established, guiding tailored material outcomes. Additionally, correlation matrices elucidate the microstructure's influence on gene expressions, providing insights for personalized approaches in bone tissue regeneration.

Nanocomposite Methacrylated Silk Fibroin-Based Scaffolds for Bone Tissue Engineering.

The treatment of bone defects is a clinical challenge. Bone tissue engineering is gaining interest as an alternative to current treatments, with the development of 3D porous structures (scaffolds) helpful in promoting bone regeneration by ensuring temporary functional support. In this work, methacrylated silk fibroin (SilMA) sponges were investigated as scaffolds for bone tissue engineering by exploiting the combination of physical (induced by NaCl salt during particulate leaching) and chemical crosslinking (induced by UV-light exposure) techniques. A biomimetic approach was adopted to better simulate the extracellular matrix of the bone by introducing either natural (mussel shell-derived) or synthetic-origin hydroxyapatite nanoparticles into the SilMA sponges. The obtained materials were characterized in terms of pore size, water absorption capability and mechanical properties to understand both the effect of the inclusion of the two different types of nanoparticles and the effect of the photocrosslinking. Moreover, the SilMA sponges were tested for their bioactivity and suitability for bone tissue engineering purposes by using osteosarcoma cells, studying their metabolism by an AlamarBlue assay and their morphology by scanning electron microscopy. Results indicate that photocrosslinking helps in obtaining more regular structures with bimodal pore size distributions and in enhancing the stability of the constructs in water. Moreover, the addition of naturally derived hydroxyapatite was observed to be more effective at activating osteosarcoma cell metabolism than synthetic hydroxyapatite, showing a statistically significant difference in the AlamarBlue measurement on day 7 after seeding. The methacrylated silk fibroin/hydroxyapatite nanocomposite sponges developed in this work were found to be promising tools for targeting bone regeneration with a sustainable approach.

Silk fibroin-based inks for in situ 3D printing using a double crosslinking process.

The shortage of tissues and organs for transplantation is an urgent clinical concern. In situ 3D printing is an advanced 3D printing technique aimed at printing the new tissue or organ directly in the patient. The ink for this process is central to the outcomes, and must meet specific requirements such as rapid gelation, shape integrity, stability over time, and adhesion to surrounding healthy tissues. Among natural materials, silk fibroin exhibits fascinating properties that have made it widely studied in tissue engineering and regenerative medicine. However, further improvements in silk fibroin inks are needed to match the requirements for in situ 3D printing. In the present study, silk fibroin-based inks were developed for in situ applications by exploiting covalent crosslinking process consisting of a pre-photo-crosslinking prior to printing and in situ enzymatic crosslinking. Two different silk fibroin molecular weights were characterized and the synergistic effect of the covalent bonds with shear forces enhanced the shift in silk secondary structure toward ß-sheets, thus, rapid stabilization. These hydrogels exhibited good mechanical properties, stability over time, and resistance to enzymatic degradation over 14 days, with no significant changes over time in their secondary structure and swelling behavior. Additionally, adhesion to tissues in vitro was demonstrated.

Polymeric Hydrogels for Intervertebral Disc Replacement/Integration: Playing with the Chemical Composition for Tuning Shear Behavior and Hydrophilicity.

Damages to the intervertebral disc (IVD) due to improper loading or degeneration result in back pain, which is a common disease affecting an increasing number of patients. Different strategies for IVD remediation have been developed, from surgical treatment to disc replacement, by using both metallic and non-metallic materials. Hydrogels are very attractive materials due to their ability to simulate the properties of many soft tissues; moreover, their chemical composition can be varied in order to assure performances similar to the natural disc. In particular, for the replacement of the IVD outer ring, namely, the anulus fibrosus, the shear properties are of paramount importance. In this work, we produced hydrogels through the photo-induced crosslinking of different mixtures composed of two hydrophilic monofunctional and difunctional polymers, namely, poly(ethyleneglycol) methyl ether methacrylate (PEGMEMA) and poly(ethyleneglycol) dimethacrylate (PEGDMA), together with a hydrophobic molecule, i.e., tert-butyl acrylate (tBA). By changing the ratio among the precursors, we demonstrated the tunability of both the shear properties and hydrophilicity. The structural properties of hydrogels were studied by solid-state nuclear magnetic resonance (NMR). These experiments provided insights on both the structure and molecular dynamics of polymeric networks and, together with information obtained by differential scanning calorimetry (DSC), allowed for correlating the physical properties of the hydrogels with their chemical composition.

Ice Recrystallization Inhibition Activity of Silk Proteins.

The cryopreservation of cells, tissue, and organs is essential in both fundamental research and practical applications, such as modern regenerative medicine and technological applications. However, the formation of ice crystals during ice recrystallization can have harmful or even fatal effects on biological systems. To address this challenge, we explore the ice recrystallization inhibition (IRI) activity of two natural silk proteins of Bombyx mori, fibroin and sericin. We found that silk fibroin (SF) had higher ice recrystallization inhibition activity than silk sericin (SS). Moreover, SF aqueous solutions perform better in inhibiting ice recrystallization than SF phosphate-buffered saline solutions. Sum-frequency generation spectroscopy shows that stronger electrostatic interactions are responsible for the higher IRI ability of SF. This work is significant for broadening the applications of silk proteins in biomedical fields.

Silk wastes and autoclaved degumming as an alternative for a sustainable silk process.

Silk degumming is considered the first point in the preparation of silk-based materials since this process could modify the silk fiber and the properties of its related products. This study evaluated the differences in morphology, secondary structure, amino acid content, thermal stability, and mechanical properties of two types of raw materials, defective cocoons (DC) and silk fibrous waste (SW), degummed by chemical (C) and autoclaving (A) methods. Subsequently, silk fibroin films were prepared by dissolving each type of degummed fibers, and thermal and structural films properties were determined. The findings demonstrated that autoclaving is an efficient alternative to remove silk sericin, as the resulting fibers presented improved structural, thermal, and mechanical properties compared to those obtained by the chemical method. For films preparation, autoclave resulted in a good option, but dissolution parameters need to be adjusted for defective cocoons. Furthermore, similarities between the physicochemical properties of fibers and films from both fibrous wastes suggest that SW is a promising raw material for producing fibrous resources and regenerated silk fibroin materials. Overall, these findings suggest new recycling methods for fibrous waste and by-products generated in the silk textile production process.

Functionalised-biomatrix for wound healing and cutaneous regeneration: future impactful medical products in clinical translation and precision medicine.

Skin tissue engineering possesses great promise in providing successful wound injury and tissue loss treatments that current methods cannot treat or achieve a satisfactory clinical outcome. A major field direction is exploring bioscaffolds with multifunctional properties to enhance biological performance and expedite complex skin tissue regeneration. Multifunctional bioscaffolds are three-dimensional (3D) constructs manufactured from natural and synthetic biomaterials using cutting-edge tissue fabrication techniques incorporated with cells, growth factors, secretomes, antibacterial compounds, and bioactive molecules. It offers a physical, chemical, and biological environment with a biomimetic framework to direct cells toward higher-order tissue regeneration during wound healing. Multifunctional bioscaffolds are a promising possibility for skin regeneration because of the variety of structures they provide and the capacity to customise the chemistry of their surfaces, which allows for the regulated distribution of bioactive chemicals or cells. Meanwhile, the current gap is through advanced fabrication techniques such as computational designing, electrospinning, and 3D bioprinting to fabricate multifunctional scaffolds with long-term safety. This review stipulates the wound healing processes used by commercially available engineered skin replacements (ESS), highlighting the demand for a multifunctional, and next-generation ESS replacement as the goals and significance study in tissue engineering and regenerative medicine (TERM). This work also scrutinise the use of multifunctional bioscaffolds in wound healing applications, demonstrating successful biological performance in the in vitro and in vivo animal models. Further, we also provided a comprehensive review in requiring new viewpoints and technological innovations for the clinical application of multifunctional bioscaffolds for wound healing that have been found in the literature in the last 5 years.

Plasma-Polymerised Antibacterial Coating of Ovine Tendon Collagen Type I (OTC) Crosslinked with Genipin (GNP) and Dehydrothermal-Crosslinked (DHT) as a Cutaneous Substitute for Wound Healing.

Tissue engineering products have grown in popularity as a therapeutic approach for chronic wounds and burns. However, some drawbacks include additional steps and a lack of antibacterial capacities, both of which need to be addressed to treat wounds effectively. This study aimed to develop an acellular, ready-to-use ovine tendon collagen type I (OTC-I) bioscaffold with an antibacterial coating for the immediate treatment of skin wounds and to prevent infection post-implantation. Two types of crosslinkers, 0.1% genipin (GNP) and dehydrothermal treatment (DHT), were explored to optimise the material strength and biodegradability compared with a non-crosslinked (OTC) control. Carvone plasma polymerisation (ppCar) was conducted to deposit an antibacterial protective coating. Various parameters were performed to investigate the physicochemical properties, mechanical properties, microstructures, biodegradability, thermal stability, surface wettability, antibacterial activity and biocompatibility of the scaffolds on human skin cells between the different crosslinkers, with and without plasma polymerisation. GNP is a better crosslinker than DHT because it demonstrated better physicochemical properties (27.33 ± 5.69% vs. 43 ± 7.64% shrinkage), mechanical properties (0.15 ± 0.15 MPa vs. 0.07 ± 0.08 MPa), swelling (2453 ± 419.2% vs. 1535 ± 392.9%), biodegradation (0.06 ± 0.06 mg/h vs. 0.15 ± 0.16 mg/h), microstructure and biocompatibility. Similarly, its ppCar counterpart, GNPppCar, presents promising results as a biomaterial with enhanced antibacterial properties. Plasma-polymerised carvone on a crosslinked collagen scaffold could also support human skin cell proliferation and viability while preventing infection. Thus, GNPppCar has potential for the rapid treatment of healing wounds.

Modeling a Dynamic Printability Window on Polysaccharide Blend Inks for Extrusion Bioprinting.

Extrusion-based bioprinting is one of the most widespread technologies due to its affordability, wide range of processable materials, and ease of use. However, the formulation of new inks for this technique is based on time-consuming trial-and-error processes to establish the optimal ink composition and printing parameters. Here, a dynamic printability window was modeled for the assessment of the printability of polysaccharide blend inks of alginate and hyaluronic acid with the intent to build a versatile predictive tool to speed up the testing procedures. The model considers both the rheological properties of the blends (viscosity, shear thinning behavior, and viscoelasticity) and their printability (in terms of extrudability and the ability of forming a well-defined filament and detailed geometries). By imposing some conditions on the model equations, it was possible to define empirical bands in which the printability is ensured. The predictive capability of the built model was successfully verified on an untested blend of alginate and hyaluronic acid chosen to simultaneously optimize the printability index and minimize the size of the deposited filament.

Silk-Based Matrices and c-Kit-Positive Cardiac Progenitor Cells for a Cellularized Silk Fibroin Scaffold: Study of an in vivo Model.

The production of a cellularized silk fibroin scaffold is very difficult because it is actually impossible to differentiate cells into a well-organized cardiac tissue. Without vascularization, not only do cell masses fail to grow, but they may also exhibit an area of necrosis, indicating a lack of oxygen and nutrients. In the present study, we used the so-called tyrosine protein kinase kit (c-Kit)-positive cardiac progenitor cells (CPCs) to generate cardiac cellularized silk fibroin scaffolds, multipotent cells isolated from the adult heart to date that can show some degree of differentiation toward the cardiac phenotype. To test their ability to differentiate into the cardiac phenotype in vivo as well, CPC and collagen organoid-like masses were implanted into nude mice and their behavior observed. Since the 3-dimensional structure of cardiac tissue can be preserved by scaffolds, we prepared in parallel different silk fibroin scaffolds with 3 different geometries and tested their behavior in 3 different models of immunosuppressed animals. Unfortunately, CPC cellularized silk fibroin scaffolds cannot be used in vivo. CPCs implanted alone or in collagen type I gel were destroyed by CD3+ lymphocyte aggregates, whereas the porous and partially oriented scaffolds elicited a consistent foreign body response characterized by giant cells. Only the electrospun meshes were resistant to the foreign body reaction. In conclusion, c-Kit-positive CPCs, although expressing a good level of cardiac differentiation markers in vitro with or without fibroin meshes, are not suitable for an in vivo model of cardiac organoids because they are degraded by a T-cell-mediated immune response. Even scaffolds which may preserve the survival of these cells in vivo also induced a host response. However, among the tested scaffolds, the electrospun meshes (F-scaffold) induced a lower response compared to all the other tested structures.

Engineered-Skin of Single Dermal Layer Containing Printed Hybrid Gelatin-Polyvinyl Alcohol Bioink via 3D-Bioprinting: In Vitro Assessment under Submerged vs. Air-Lifting Models.

Three-dimensional (3D) in vitro skin models are frequently employed in cosmetic and pharmaceutical research to minimize the demand for animal testing. Hence, three-dimensional (3D) bioprinting was introduced to fabricate layer-by-layer bioink made up of cells and improve the ability to develop a rapid manufacturing process, while maintaining bio-mechanical scaffolds and microstructural properties. Briefly, gelatin-polyvinyl alcohol (GPVA) was mixed with 1.5 × 106 and 3.0 × 106 human dermal fibroblast (HDF) cell density, together with 0.1% genipin (GNP), as a crosslinking agent, using 3D-bioprinting. Then, it was cultured under submerged and air-lifting conditions. The gross appearance of the hydrogel's surface and cross-section were captured and evaluated. The biocompatibility testing of HDFs and cell-bioink interaction towards the GPVA was analyzed by using live/dead assay, cell migration activity, cell proliferation assay, cell morphology (SEM) and protein expression via immunocytochemistry. The crosslinked hydrogels significantly demonstrated optimum average pore size (100-199 µm). The GPVA crosslinked with GNP (GPVA_GNP) hydrogels with 3.0 × 106 HDFs was proven to be outstanding, compared to the other hydrogels, in biocompatibility testing to promote cellular interaction. Moreover, GPVA-GNP hydrogels, encapsulated with 3.0 × 106 HDFs under submerged cultivation, had a better outcome than air-lifting with an excellent surface cell viability rate of 96 ± 0.02%, demonstrated by 91.3 ± 4.1% positively expressed Ki67 marker at day 14 that represented active proliferative cells, an average of 503.3 ± 15.2 µm for migration distance, and maintained the HDFs' phenotypic profiles with the presence of collagen type I expression. It also presented with an absence of alpha-smooth muscle actin positive staining. In conclusion, 3.0 × 106 of hybrid GPVA hydrogel crosslinked with GNP, produced by submerged cultivation, was proven to have the excellent biocompatibility properties required to be a potential bioinks for the rapid manufacturing of 3D in vitro of a single dermal layer for future use in cosmetic, pharmaceutic and toxicologic applications.

Quercetin-Embedded Gelastin Injectable Hydrogel as Provisional Biotemplate for Future Cutaneous Application: Optimization and In Vitro Evaluation.

Chronic wounds have become an epidemic in millions of patients and result in amputations. In order to overcome this, immediate treatment is a realistic strategy to minimize the risk of complications and aid in the healing rate of the cutaneous wound. Functionalized engineered biomaterials are proven to be a potential approach to embarking on skin wound management. Thus, this study aimed to evaluate the efficacy of a quercetin-embedded gelatin−elastin (Gelastin) injectable hydrogel to act as a provisional biotemplate with excellent physicochemical properties, to be utilized for future cutaneous application. Briefly, the hydrogel was homogenously pre-mixed with genipin (GNP), followed by the incorporation of quercetin (QC). The physicochemical properties comprised the contact angle, swelling ratio, crosslinking degree, enzymatic biodegradation, and water vapor transmission rate (WVTR), as well as chemical characterization. Energy-dispersive X-ray (EDX), XRD, and Fourier transform infra-red (FTIR) analyses were conducted. Briefly, the findings demonstrated that the crosslinked hybrid biomatrix demonstrated better resilience at >100%, a contact angle of >20°, a swelling ratio average of 500 ± 10%, a degradation rate of <0.05 mg/hour, and a successful crosslinking degree (<70%free amine group), compared to the non-crosslinked hybrid biomatrix. In addition, the WVTR was >1500 g/m2 h, an optimal moisture content designed to attain regular cell function and proliferation. The outcomes convey that Gelastin-QC hydrogels deliver the optimum features to be used as a provisional biotemplate for skin tissue engineering purposes.

The Discovery and Development of Natural-Based Biomaterials with Demonstrated Wound Healing Properties: A Reliable Approach in Clinical Trials.

Current research across the globe still focuses strongly on naturally derived biomaterials in various fields, particularly wound care. There is a need for more effective therapies that will address the physiological deficiencies underlying chronic wound treatment. The use of moist bioactive scaffolds has significantly increased healing rates compared to local and traditional treatments. However, failure to heal or prolonging the wound healing process results in increased financial and social stress imposed on health institutions, caregivers, patients, and their families. The urgent need to identify practical, safe, and cost-effective wound healing scaffolding from natural-based biomaterials that can be introduced into clinical practice is unequivocal. Naturally derived products have long been used in wound healing; however, clinical trial evaluations of these therapies are still in their infancy. Additionally, further well-designed clinical trials are necessary to confirm the efficacy and safety of natural-based biomaterials in treating wounds. Thus, the focus of this review is to describe the current insight, the latest discoveries in selected natural-based wound healing implant products, the possible action mechanisms, and an approach to clinical studies. We explore several tested products undergoing clinical trials as a novel approach to counteract the debilitating effects of impaired wound healing.

A Comprehensive Review on Collagen Type I Development of Biomaterials for Tissue Engineering: From Biosynthesis to Bioscaffold.

Collagen is the most abundant structural protein found in humans and mammals, particularly in the extracellular matrix (ECM). Its primary function is to hold the body together. The collagen superfamily of proteins includes over 20 types that have been identified. Yet, collagen type I is the major component in many tissues and can be extracted as a natural biomaterial for various medical and biological purposes. Collagen has multiple advantageous characteristics, including varied sources, biocompatibility, sustainability, low immunogenicity, porosity, and biodegradability. As such, collagen-type-I-based bioscaffolds have been widely used in tissue engineering. Biomaterials based on collagen type I can also be modified to improve their functions, such as by crosslinking to strengthen the mechanical property or adding biochemical factors to enhance their biological activity. This review discusses the complexities of collagen type I structure, biosynthesis, sources for collagen derivatives, methods of isolation and purification, physicochemical characteristics, and the current development of collagen-type-I-based scaffolds in tissue engineering applications. The advancement of additional novel tissue engineered bioproducts with refined techniques and continuous biomaterial augmentation is facilitated by understanding the conventional design and application of biomaterials based on collagen type I.

Silk fibroin films with embedded magnetic nanoparticles: evaluation of the magneto-mechanical stimulation effect on osteogenic differentiation of stem cells.

We report about a biomaterial in the form of film ∼10 µm thick, consisting of a silk fibroin matrix with embedded iron oxide superparamagnetic nanoparticles, for prospective applications as bioactive coating in regenerative medicine. Films with different load of magnetic nanoparticles are produced (nanoparticles/silk fibroin nominal ratio = 5, 0.5 and 0 wt%) and the structural, mechanical and magnetic properties are studied. The nanoparticles form aggregates in the silk fibroin matrix and the film stiffness, as tested by nanoindentation, is spatially inhomogeneous, but the protein structure is not altered. In vitro biological tests are carried out on human bone marrow-derived mesenchymal stem cells cultured on the films up to 21 days, with and without an applied static uniform magnetic field. The sample with the highest nanoparticles/silk fibroin ratio shows the best performance in terms of cell proliferation and adhesion. Moreover, it promotes a faster and better osteogenic differentiation, particularly under magnetic field, as indicated by the gene expression level of typical osteogenic markers. These findings are explained in light of the results of the physical characterization, combined with numerical calculations. It is established that the applied magnetic field triggers a virtuous magneto-mechanical mechanism in which dipolar magnetic forces between the nanoparticle aggregates give rise to a spatial distribution of mechanical stresses in the silk fibroin matrix. The film with the largest nanoparticle load, under cell culture conditions (i.e. in aqueous environment), undergoes matrix deformations large enough to be sensed by the seeded cells as mechanical stimuli favoring the osteogenic differentiation.

Powder 3D Printing of Bone Scaffolds with Uniform and Gradient Pore Sizes Using Cuttlebone-Derived Calcium Phosphate and Glass-Ceramic.

The pore geometry of bone scaffolds has a major impact on their cellular response; for this reason, 3D printing is an attractive technology for bone tissue engineering, as it allows for the full control and design of the porosity. Calcium phosphate materials synthesized from natural sources have recently attracted a certain interest because of their similarity to natural bone, and they were found to show better bioactivity than synthetic compounds. Nevertheless, these materials are very challenging to be processed by 3D printing due to technological issues related to their nanometric size. In this work, bone scaffolds with different pore geometries, with a uniform size or with a size gradient, were fabricated by binder jetting 3D printing using a biphasic calcium phosphate (BCP) nanopowder derived from cuttlebones. To do so, the nanopowder was mixed with a glass-ceramic powder with a larger particle size (45-100 µm) in 1:10 weight proportions. Pure AP40mod scaffolds were also printed. The sintered scaffolds were shown to be composed mainly by hydroxyapatite (HA) and wollastonite, with the amount of HA being larger when the nanopowder was added because BCP transforms into HA during sintering at 1150 °C. The addition of bio-derived powder increases the porosity from 60% to 70%, with this indicating that the nanoparticles slow down the glass-ceramic densification. Human mesenchymal stem cells were seeded on the scaffolds to test the bioactivity in vitro. The cells' number and metabolic activity were analyzed after 3, 5 and 10 days of culturing. The cellular behavior was found to be very similar for samples with different pore geometries and compositions. However, while the cell number was constantly increasing, the metabolic activity on the scaffolds with gradient pores and cuttlebone-derived powder decreased over time, which might be a sign of cell differentiation. Generally, all scaffolds promoted fast cell adhesion and proliferation, which were found to penetrate and colonize the 3D porous structure.

Use of Bombyx mori silk fibroin in tissue engineering: From cocoons to medical devices, challenges, and future perspectives.

Silk fibroin has become a prominent material in tissue engineering (TE) over the last 20 years with almost 10,000 published works spanning in all the TE applications, from skeleton to neuronal regeneration. Fibroin is an extremely versatile biopolymer that, due to its ease of processing, has enabled the development of an entire plethora of materials whose properties and architectures can be tailored to suit target applications. Although the research and development of fibroin TE materials and devices is mature, apart from sutures, only a few medical products made of fibroin are used in the clinical routines. <40 clinical trials of Bombyx mori silk-related products have been reported by the FDA and few of them resulted in a commercialized device. In this review, after explaining the structure and properties of silk fibroin, we provide an overview of both fibroin constructs existing in the literature and fibroin devices used in clinic. Through the comparison of these two categories, we identified the burning issues faced by fibroin products during their translation to the market. Two main aspects will be considered. The first is the standardization of production processes, which leads both to the standardization of the characteristics of the issued device and the correct assessment of its failure. The second is the FDA regulations, which allow new devices to be marketed through the 510(k) clearance by demonstrating their equivalence to a commercialized medical product. The history of some fibroin medical devices will be taken as a case study. Finally, we will outline a roadmap outlining what actions we believe are needed to bring fibroin products to the market.

Enthesis Healing Is Dependent on Scaffold Interphase Morphology-Results from a Rodent Patellar Model.

The use of multiphasic scaffolds to treat injured tendon-to-bone entheses has shown promising results in vitro. Here, we used two versions of a biphasic silk fibroin scaffold to treat an enthesis defect created in a rat patellar model in vivo. One version presented a mixed transition between the bony and the tendon end of the construct (S-MT) while this transition was abrupt in the second version (S-AT). At 12 weeks after surgery, the S-MT scaffold promoted better healing of the injured enthesis, with minimal undesired ossification of the insertion area. The expression of tenogenic and chondrogenic markers was sustained for longer in the S-MT-treated group and the tangent modulus of the S-MT-treated samples was similar to the native tissue at 12 weeks while that of the S-AT-treated enthesis was lower. Our study highlights the important role of the transition zone of multiphasic scaffolds in the treatment of complex interphase tissues such as the tendon-to-bone enthesis.

Development of alginate-based hydrogels for blood vessel engineering.

Vascular diseases are among the primary causes of death worldwide. In serious conditions, replacement of the damaged vessel is required. Autologous grafts are preferred, but their limited availability and difficulty of the harvesting procedures favour synthetic alternatives' use. However, as synthetic grafts may present significant drawbacks, tissue engineering-based solutions are proposed. Herein, tubular hydrogels of alginate combined with collagen type I and/or silk fibroin were prepared by ionotropic gelation using gelatin hydrogel sacrificial moulds loaded with calcium ions (Ca2+). The time of exposure of alginate solutions to Ca2+-loaded gelatin was used to control the wall thickness of the hydrogels (0.47 ± 0.10 mm-1.41 ± 0.21 mm). A second crosslinking step with barium chloride prevented their degradation for a 14 day period and improved mechanical properties by two-fold. Protein leaching tests showed that collagen type I, unlike silk fibroin, was strongly incorporated in the hydrogels. The presence of silk fibroin in the alginate matrix, containing or not collagen, did not significantly improve hydrogels' properties. Conversely, hydrogels enriched only with collagen were able to better support EA.hy926 and MRC-5 cells' growth and characteristic phenotype. These results suggest that a two-step crosslinking procedure combined with the use of collagen type I allow for producing freestanding vascular substitutes with tuneable properties in terms of size, shape and wall thickness.