Tri-layered Scaffold Research Articles

Co- and Triaxial Electrospinning for Stem Cell-based Bone Regeneration.

Bone tissue is composed of organic minerals and cells. It has the capacity to heal for certain minor damages, but when the bone defects surpass the critical threshold, they need fixing. Bone regeneration through natural and synthetic biodegradable materials requires various steps, such as manufacturing methods and materials selection. A successful biodegradable bone graft should have a high surface area/ volume ratio, strength, and a biocompatible, porous structure capable of promoting cell adhesion, proliferation, and differentiation. Considering these requirements, the electrospinning technique is promising for creating functional nano-sized scaffolds. The multi-axial methods, such as coaxial and triaxial electrospinning, are the most popular techniques to produce double or tri-layered scaffolds, respectively. Recently, stem cell culture on scaffolds and the application of osteogenic differentiation protocols on these scaffolds have opened new possibilities in the field of biomaterials research. This review discusses an overview of the progress in coaxial and triaxial technology through biodegradable composite bone materials. The review also carefully elaborates the osteogenic differentiation using stem cells and their performance with nano-sized scaffolds.

Fabrication of a Triple-Layer Bionic Vascular Scaffold via Hybrid Electrospinning.

Tissue engineering aims to develop bionic scaffolds as alternatives to autologous vascular grafts due to their limited availability. This study introduces a novel wet-electrospinning fabrication technique to create small-diameter, uniformly aligned tubular scaffolds. By combining this innovative method with conventional electrospinning, a bionic tri-layer scaffold that mimics the zonal structure of vascular tissues is produced. The inner and outer layers consist of PCL/Gelatin and PCL/PLGA fibers, respectively, while the middle layer is crafted using PCL through Wet Vertical Magnetic Rod Electrospinning (WVMRE). The scaffold's morphology is analyzed using Scanning Electron Microscopy (SEM) to confirm its bionic structure. The mechanical properties, degradation profile, wettability, and biocompatibility of the scaffold are also characterized. To enhance hemocompatibility, the scaffold is crosslinked with heparin. The results demonstrate sufficient mechanical properties, good wettability of the inner layer, proper degradability of the inner and middle layers, and overall good biocompatibility. In conclusion, this study successfully develops a small-diameter tri-layer tubular scaffold that meets the required specifications.

Biomimicking trilayer scaffolds with controlled estradiol release for uterine tissue regeneration.

Scaffold-based tissue engineering provides an efficient approach for repairing uterine tissue defects and restoring fertility. In the current study, a novel trilayer tissue engineering scaffold with high similarity to the uterine tissue in structure was designed and fabricated via 4D printing, electrospinning and 3D bioprinting for uterine regeneration. Highly stretchable poly(l-lactide-co-trimethylene carbonate) (PLLA-co-TMC, "PTMC" in short)/thermoplastic polyurethane (TPU) polymer blend scaffolds were firstly made via 4D printing. To improve the biocompatibility, porous poly(lactic acid-co-glycolic acid) (PLGA)/gelatin methacryloyl (GelMA) fibers incorporated with polydopamine (PDA) particles were produced on PTMC/TPU scaffolds via electrospinning. Importantly, estradiol (E2) was encapsulated in PDA particles. The bilayer scaffolds thus produced could provide controlled and sustained release of E2. Subsequently, bone marrow derived mesenchymal stem cells (BMSCs) were mixed with gelatin methacryloyl (GelMA)-based inks and the formulated bioinks were used to fabricate a cell-laden hydrogel layer on the bilayer scaffolds via 3D bioprinting, forming ultimately biomimicking trilayer scaffolds for uterine tissue regeneration. The trilayer tissue engineering scaffolds thus formed exhibited a shape morphing ability by transforming from the planar shape to tubular structures when immersed in the culture medium at 37°C. The trilayer tissue engineering scaffolds under development would provide new insights for uterine tissue regeneration.

A 3D printed biomimetic scaffold for cartilage regeneration with lubrication, load-bearing, and adhesive fixation properties

Reconstructing the tri-layer structure of articular cartilage (AC) and replicating its functions pose significant challenges. In response, we propose a novel approach utilizing 3D printing technology to replicate and reconstruct the complex structure of AC. This is achieved by introducing three distinct functional hydrogels, each providing lubrication, load-bearing, and adhesive fixation functions. We have systematically characterized the structure, chemical composition, and function of each layer. The results demonstrate the effectiveness of the tri-layer scaffold, which exhibits low friction, mechanical properties similar to natural AC, excellent adhesion capacity, and good biocompatibility. This study paves the way for innovation in the traditional methods of preparing biomimetic AC and has broad applications in osteochondral (OC) defect repair.

Development of a highly porous bioscaffold by the combination of bubble entrapping and freezing-thawing techniques to fabricate hyaluronic acid/gelatin tri-layer wound dressing

The electrospun nanofibers and porous scaffolds hold great promise in regenerative medicine. A novel nanofiber-hydrogel‑silicone tri-layer wound dressing has been designed and fabricated to address the limitations of each platform. The bottom nanofiber layer with a 110 mm diameter meets the wound surface and regulates cell attachment and migration. The middle hydrogel layer was fabricated through the optimization of chemical crosslink formation and freezing-thawing cycles (physical crosslink). The fabricated hydrogel with interconnected porous structure has optimized properties (gel fraction (89.45 %) and porosity (80 %)) for wound dressing application. The silicone layer on the outer surface was designed to fix the wound dressing on the skin and prevent the penetration of pathogens. The scanning electron microscope micrograph showed structural integrity in the tri-layer scaffold. In vivo data showed that the tri-layer scaffold accelerates wound healing in the mice model and angiogenesis in the chorioallantoic membrane model. Therefore, the designed scaffold inspired by the skin's structure can be used as a wound dressing to treat wounds.

Direction-oriented fiber guiding with a tunable tri-layer-3D scaffold for periodontal regeneration.

Multilayered scaffolds mimicking mechanical and biological host tissue architectures are the current prerequisites for successful tissue regeneration. We propose our tunable tri-layered scaffold, designed to represent the native periodontium for potential regenerative applications. The fused deposition modeling platform is used to fabricate the novel movable three-layered polylactic acid scaffold mimicking in vivo cementum, periodontal ligament, and alveolar bone layers. The scaffold is further provided with multiple angulated fibers, offering directional guidance and facilitating the anchorage dependence on cell adhesion. Additionally, surface modifications of the scaffold were made by incorporating coatings like collagen and different concentrations of gelatin methacryloyl to enrich the cell adhesion and proliferation. The surface characterization of our designed scaffolds was performed using tribological studies, atomic force microscopy, contact angle measurement, scanning electron microscopy, and micro-computed tomography. Furthermore, the material characterization of this scaffold was investigated by attenuated total reflectance-Fourier transformed infrared spectroscopy. The scaffold's mechanical characterization, such as strength and compression modulus, was demonstrated by compression testing. The L929 mouse fibroblast cells and MG63 human osteosarcoma cells have been cultured on the scaffold. The scaffold's superior biocompatibility was evaluated using fluorescence dye with fluorescence microscopy, scanning electron microscopy, in vitro wound healing assay, MTT assay, and flow cytometry. The mineralization capability of the scaffolds was also studied. In conclusion, our study demonstrated the construction of a multilayered movable scaffold, which is highly biocompatible and most suitable for various downstream applications such as periodontium and in situ tissue regeneration of complex, multilayered tissues.

Fabrication of a tri-layer scaffold with dual release of heparin and PRP for tissue engineering of small‐diameter blood vessels

A tri-layer scaffold was fabricated according to functionally graded materials (FGM) theory from poly(glycerol sebcate)/poly(L-lactic acid) blend by the increasing polylactic acid ratio from the inner to the outer layer. Heparin and platelet-rich plasma were loaded in separate layers to achieve proper remodeling and patency. The scaffold showed a linear degradation trend. The elastic modulus was 13.8 ± 1.11 Mpa that was in the favorable range for constructing artificial vessels. The integrity of the layers was preserved due to its FGM structure. The scaffold showed full blood compatibility by minimal platelet adhesion and hemolysis due to relatively slow release of Heparin.

Fabrication, Characterization, and In Vitro Cytotoxicity Assessment of Tri-Layered Multifunctional Scaffold for Effective Chronic Wound Healing.

Chronic wounds have been a global health risk that demands intensive exploration. A tri-layered biomaterial scaffold has been developed for skin wounds. The top layer of the scaffold is superhydrophobic, and the bottom layer is hydrophilic, both of which were electrospun using recycled expanded polystyrene (EPS) and monofilament fishing line (MFL), respectively. The intermediate layer of the scaffold comprised hydrogel by cross-linking chitosan (CS) with polyethylene glycol. The surface morphology, surface chemistry, thermal degradation, and wettability characteristics of each layer of the scaffold were examined. Also, the antibacterial activity and in vitro cytotoxicity study on the combined tri-layered scaffold were assessed against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). Data revealed exceptional water repellency of the heat-treated electrospun top superhydrophobic layer (TSL) with a high-water contact angle (WCA) of 172.44°. A TSL with 15 wt% of micro-/nano-inclusions had the best thermal stability above 400 °C. The bottom hydrophilic layer (BHL) displayed a WCA of 9.91°. Therapeutically, the synergistic effect of the combined tri-layered scaffold significantly inhibited bacteria growth by 70.5% for E. coli and 68.6% for S. aureus. Furthermore, cell viability is enhanced when PEG is included as part of the intermediate CS hydrogel layer (ICHL) composition.

Sandwich-like electro-conductive polyurethane-based gelatin/soybean oil nanofibrous scaffolds with a targeted release of simvastatin for cardiac tissue engineering

Cardiac tissue engineering (CTE) is a promising way for the restoration of injured cardiac tissue in the healthcare system. The development of biodegradable scaffolds with appropriate chemical, electrical, mechanical, and biological properties is an unmet need for the success of CTE. Electrospinning is a versatile technique that has shown potential applications in CTE. Herein, four different types of multifunctional scaffolds, including synthetic-based poly (glycerol sebacate)-polyurethane (PGU), PGU-Soy scaffold, and a series of trilayer scaffolds containing two outer layers of PGU-Soy and a middle (inner) layer of gelatin (G) as a natural and biodegradable macromolecule without simvastatin (S) and with simvastatin (GS), an anti-inflammatory agent, were fabricated in the sandwich-like structure using electrospinning technique. This approach offers a combination of the advantages of both synthetic and natural polymers to enhance the bioactivity and the cell-to-cell and cell-to-matrix intercommunication. An in vitro drug release analysis was performed after the incorporation of soybean oil (Soy) and G. Soy is used as a semiconducting material was introduced to improve the electrical conductivity of nanofibrous scaffolds. The physicochemical properties, contact angle, and biodegradability of the electrospun scaffolds were also assessed. Moreover, the blood compatibility of nanofibrous scaffolds was studied through activated partial thromboplastin time (APTT), prothrombin time (PT), and hemolytic assay. The results showed that all scaffolds exhibited defect-free morphologies with mean fiber diameters in the range of 361 ± 109 to 417 ± 167 nm. A delay in blood clotting was observed, demonstrating the anticoagulant nature of nanofibrous scaffolds. Furthermore, rat cardiomyoblast cell lines (H9C2) were cultured on scaffolds for 7 days, and the morphology and cell arrangement were monitored. Data indicated an appropriate cytocompatibility. Of note, in the PGU-Soy/GS nanofibrous scaffold, a high survival rate was indicated compared to other groups. Our findings exhibited that the simvastatin-loaded polymeric system had positive effects on cardiomyoblasts attachment and growth and could be utilized as a drug release carrier in the field of CTE.Graphical

Evaluation of a co-culture of rapidly isolated chondrocytes and stem cells seeded on tri-layered collagen-based scaffolds in a caprine osteochondral defect model

Cartilage has poor regenerative capacity and thus damage to the joint surfaces presents a major clinical challenge. Recent research has focussed on the development of tissue-engineered and cell-based approaches for the treatment of cartilage and osteochondral injuries, with current clinically available cell-based approaches including autologous chondrocyte implantation and matrix-assisted autologous chondrocyte implantation. However, these approaches have significant disadvantages due to the requirement for a two-stage surgical procedure and an in vitro chondrocyte expansion phase which increases logistical challenges, hospital times and costs. In this study, we hypothesized that seeding biomimetic tri-layered scaffolds, with proven regenerative potential, with chondrocyte/infrapatellar fat pad stromal cell co-cultures would improve their regenerative capacity compared to scaffolds implanted cell-free. Rapid cell isolation techniques, without the requirement for long term in vitro culture, were utilised to achieve co-cultures of chondrocytes and stromal cells and thus overcome the limitations of existing cell-based techniques. Cell-free and cell-seeded scaffolds were implanted in osteochondral defects, created within the femoral condyle and trochlear ridge, in a translational large animal goat model. While analysis showed trends towards delayed subchondral bone healing in the cell-seeded scaffold group, by the 12 month timepoint the cell-free and cell-seeded groups yield cartilage and bone tissue with comparable quality and quantity. The results of the study reinforce the potential of the biomimetic tri-layered scaffold to repair joint defects but failed to demonstrate a clear benefit from the addition of the CC/FPMSC co-culture to this scaffold. Taking into consideration the additional cost and complexity associated with the cell-seeded scaffold approach, this study demonstrates that the treatment of osteochondral defects using cell-free tri-layered scaffolds may represent a more prudent clinical approach.

Development of Tri-Layered Biomimetic Atelocollagen Scaffolds with Interfaces for Osteochondral Tissue Engineering.

The development of biomimetic scaffolds containing cartilage, calcified cartilage, and bone regeneration for precise osteochondral repair remains a challenge. Herein, a novel tri-layered scaffold-with a top layer containing type II atelocollagen and chondroitin sulphate for cartilage regeneration, an intermediate layer with type II atelocollagen and hydroxyapatite for calcified cartilage formation, and a bottom layer with type I atelocollagen and hydroxyapatite for bone growth-that can be built using liquid-phase cosynthesis, is described. The tri-layered scaffolds are mechanically demonstrably superior and have a lower risk of delamination than monolayer scaffolds. This is due to higher cohesion arising from the interfaces between each layer. In vitro results show that although monolayer scaffolds can stimulate bone marrow stem cells to differentiate and form cartilage, calcified cartilage, and bone separately (detected using quantitative polymerase chain reaction analysis and staining with safranin-O and Alizarin Red S), the tri-layered scaffolds support the regeneration of cartilage, calcified cartilage, and bone simultaneously after 2 and 4 months of implantation (detected using gross and micro-computed tomography images, histological staining, and Avizo, a software used to detect microlevel defects in metals). This work presents data on a promising approach in devising strategies for the precise repair of osteochondral defects.

The Incorporation of Etanercept into a Porous Tri-Layer Scaffold for Restoring and Repairing Cartilage Tissue.

Cartilage diseases currently affect a high percentage of the world’s population. Almost all of these diseases, such as osteoarthritis (OA), cause inflammation of this soft tissue. However, this could be controlled with biomaterials that act as an anti-inflammatory delivery system, capable of dosing these drugs over time in a specific area. The objective of this study was to incorporate etanercept (ETA) into porous three-layer scaffolds to decrease the inflammatory process in this soft tissue. ETA is a blocker of pro-inflammatory cytokines, such as tumour necrosis factor alpha (TNF-α) and interleukin 6 (IL-6). For this reason, the scaffold was built based on natural polymers, including chitosan and type I collagen. The scaffold was grafted next to subchondral bone using hydroxyapatite as filler. One of the biomaterials obtained was also crosslinked to compare its mechanical properties with the non-treated one. Both samples’ physicochemical properties were studied with SEM, micro-CT and photoacoustic imaging, and their rheological properties were also compared. The cell viability and proliferation of the human chondrocyte C28/I2 cell line were studied in vitro. An in vitro and in vivo controlled release study was evaluated in both specimens. The ETA anti-inflammatory effect was also studied by in vitro TNF-α and IL-6 production. The crosslinked and non-treated scaffolds had rheological properties suitable for this application. They were non-cytotoxic and favoured the in vitro growth of chondrocytes. The in vitro and in vivo ETA release showed desirable results for a drug delivery system. The TNF-α and IL-6 production assay showed that this drug was effective as an anti-inflammatory agent. In an in vivo OA mice model, safranin-O and fast green staining was carried out. The OA cartilage tissue improved when the scaffold with ETA was grafted in the damaged area. These results demonstrate that this type of biomaterial has high potential for clinical applications in tissue engineering and as a controlled drug delivery system in OA articular cartilage.

Layer-specific stem cell differentiation in tri-layered tissue engineering biomaterials: Towards development of a single-stage cell-based approach for osteochondral defect repair.

Successful repair of osteochondral defects is challenging, due in part to their complex gradient nature. Tissue engineering approaches have shown promise with the development of layered scaffolds that aim to promote cartilage and bone regeneration within the defect. The clinical potential of implanting these scaffolds cell-free has been demonstrated, whereby cells from the host bone marrow MSCs infiltrate the scaffolds and promote cartilage and bone regeneration within the required regions of the defect. However, seeding the cartilage layer of the scaffold with a chondrogenic cell population prior to implantation may enhance cartilage tissue regeneration, thus enabling the treatment of larger defects. Here the development of a cell seeding approach capable of enhancing articular cartilage repair without the requirement for in vitro expansion of the cell population is explored. The intrinsic ability of a tri-layered scaffold previously developed in our group to direct stem cell differentiation in each layer of the scaffold was first demonstrated. Following this, the optimal chondrogenic cell seeding approach capable of enhancing the regenerative capacity of the tri-layered scaffold was demonstrated with the highest levels of chondrogenesis achieved with a co-culture of rapidly isolated infrapatellar fat pad MSCs (FPMSCs) and chondrocytes (CCs). The addition of FPMSCs to a relatively small number of CCs led to a 7.8-fold increase in the sGAG production over chondrocytes in mono-culture. This cell seeding approach has the potential to be delivered within a single-stage approach, without the requirement for costly in vitro expansion of harvested cells, to achieve rapid repair of osteochondral defects.

Scaffold With Natural Calcified Cartilage Zone for Osteochondral Defect Repair in Minipigs

Background: Long-term outcomes of current clinical interventions for osteochondral defect are less than satisfactory. One possible reason is an ignorance of the interface structure between cartilage and subchondral bone, the calcified cartilage zone (CCZ). However, the importance of natural CCZ in osteochondral defects has not been directly described. Purpose: To explore the feasibility of fabricating trilayer scaffold containing natural CCZ for osteochondral defects and the role of CCZ in the repair process. Study Design: Controlled laboratory study. Methods: The scaffold was prepared by cross-linking lyophilized type II collagen sponge and acellular normal pig subchondral bone with or without natural CCZ. Autologous bone marrow stem cells (BMSCs) of minipig were mixed with type II collagen gel and injected into the cartilage layer of the scaffold before operation. Thirty minipigs were randomly divided into CCZ (n = 10), non-CCZ (n = 10), and blank control (n = 10) groups. An 8 mm–diameter full-thickness osteochondral defect was created on the trochlear surface, and scaffold containing BMSCs was transplanted into the defect according to grouping requirements. At 12 and 24 weeks postoperatively, specimens were assessed by macroscopic observation, magnetic resonance imaging examination, and histological observations (hematoxylin and eosin, Safranin O–fast green, type II collagen immunohistochemical, and Sirius red staining). Semiquantitative cartilage repair scoring was conducted using the MOCART (Magnetic Resonance Observation of Cartilage Repair Tissue) system and the O’Driscoll repaired cartilage value system. Results: The defects in the blank control and non-CCZ groups were filled with fibrous tissue, while the cartilage layer of the CCZ group was mainly repaired by hyaline cartilage at 24 weeks postoperatively. The superior repair outcome of the CCZ group was confirmed by MOCART and O’Driscoll score. Conclusion: The trilayer scaffold containing natural CCZ obtained the best repair effect compared with the non-CCZ scaffold and the blank control, indicating the importance of the CCZ in osteochondral tissue engineering. Clinical Relevance: This study demonstrates the necessity to reconstruct CCZ in clinical osteochondral defect repair and provides a possible strategy for osteochondral tissue engineering.

The Effects of Crosslinking on the Rheology and Cellular Behavior of Polymer-Based 3D-Multilayered Scaffolds for Restoring Articular Cartilage

Polymer-based tri-layered (bone, intermediate and top layers) scaffolds used for the restoration of articular cartilage were prepared and characterized in this study to emulate the concentration gradient of cartilage. The scaffolds were physically or chemically crosslinked. In order to obtain adequate scaffolds for the intended application, the impact of the type of calcium phosphate used in the bone layer, the polymer used in the intermediate layer and the interlayer crosslinking process were analyzed. The correlation among SEM micrographs, physical-chemical characterization, swelling behavior, rheological measurements and cell studies were examined. Storage moduli at 1 Hz were 0.3–1.7 kPa for physically crosslinked scaffolds, and 4–5 kPa (EDC/NHS system) and 15–20 kPa (glutaraldehyde) for chemically crosslinked scaffolds. Intrinsic viscoelasticity and poroelasticity were considered in discussing the physical mechanism dominating in different time/frequency scales. Cell evaluation showed that all samples are available as alternatives to repair and/or substitute cartilage in articular osteoarthritis.

Utilizing an integrated tri-layered scaffold with Titanium-Mesh-Cage base to repair cartilage defects of knee in goat model

The application of multilayered monolithic scaffolds to repair osteochondral defects has become a hot research topic due to the advantage of simulating native osteochondral physiological structure. However, it was difficult to promote proliferation and migration of different kinds of cells in the corresponding layers under the same culture condition. Besides, the methods utilized in previous researches to construct tissue-engineered osteochondral construct were time-consuming, laborious, complicated, costly and difficult to ensure the quality and integration of multilayered construction. Inspired by the application of titanium mesh cage (TMC) for bone graft in spinal fusion surgery, we attempt to 3D print an integrated tri-layered scaffold with Titanium-Mesh-Cage base which could be filled with autogenous cancellous bone for bone graft to the osteochondral defect. Finally, the results of repairing cartilage defects of knee with this scaffold in goat model were evaluated. The results of in-vivo experiments suggested that utilizing the integrated tri-layered scaffold with Titanium-Mesh-Cage base to repair osteochondral defect was a convenient, cost-effective and time-saving procedures, which could achieve a satisfactory repair result as good as that of Mosaicplasty.

Regional and sustained dual-release of growth factors from biomimetic tri-layered scaffolds for the repair of large-scale osteochondral defects

Mesenchymal stem cells (MSCs) are considered to be important cell sources for tissue regeneration. Growth factors (GFs) are key mediators of MSCs chondrogenesis and osteogenesis in the enhancement of osteochondral repair. However, uncontrolled delivery and release of these bioactive factors may reduce their bioavailability and lead to off-target side effects. In this study, tri-layered scaffolds, based on the chondrocyte extracellular matrix, nano-hydroxyapatite and silk fibroin were prepared by imitating the structural layers of the osteochondral unit. Particularly, tri-layered scaffolds were functioned for the local and sustained release of transforming growth factor-β3 to the chondral layer and bone morphogenetic protein-2 to the bony layer. In conjunction with engrafted human umbilical cord mesenchymal stem cells (hUCMSCs), osteochondral regeneration was enhanced. In vitro experiments indicated that scaffolds supported hUCMSCs proliferation, vitality and adhesion, and facilitated hUCMSCs lineage differentiation toward chondrocytes or osteoblasts, dependent on stimulation by sustained GFs release, at chondral layer or bony layer, respectively. Furthermore, evaluation of osteochondral repair in vivo after 8- and 16-weeks post-implantation indicated that the tri-layered scaffolds with sustained release function and hUCMSC delivery significantly accelerated the repair of large-size osteochondral defects, compared with other scaffolds. In summary, our studies showed that tri-layered scaffolds with the biomimetic structure of osteochondral unit, with sustained GFs release and seeded with hUCMSCs were more conducive to osteochondral regeneration, providing new materials and strategies for tissue engineering for the repair large-scale osteochondral defects.

Design, construction, and biological testing of an implantable porous trilayer scaffold for repairing osteoarthritic cartilage

Various tissue engineering systems for cartilage repair have been designed and tested over the past two decades, leading to the development of many promising cartilage grafts. However, no one has yet succeeded in devising an optimal system to restore damaged articular cartilage. Here, the design, assembly, and biological testing of a porous, chitosan/collagen-based scaffold as an implant to repair damaged articular cartilage is reported. Its gradient composition and trilayer structure mimic variations in natural cartilage tissue. One of its layers includes hydroxyapatite, a bioactive component that facilitates the integration of growing tissue on local bone in the target area after scaffold implantation. The scaffold was evaluated for surface morphology; rheological performance (storage, loss, complex, and time-relaxation moduli at 1 kHz); physiological stability; in vitro activity and cytotoxicity (on a human chondrocyte C28 cell line); and in vivo performance (tissue growth and biodegradability), in a murine model of osteoarthritis. The scaffold was shown to be mechanically resistant and noncytotoxic, favored tissue growth in vivo, and remained stable for 35 days postimplantation in mice. These encouraging results highlight the potential of this porous chitosan/collagen scaffold for clinical applications in cartilage tissue engineering.

A Biomimetic Basement Membrane Substitute Based on Tri-Layered Nanofibrous Scaffold for Skin Reconstruction.

Developing basement membranes (BMs) substitute remains major problem for constructing functional tissue engineered skin because of its complex structure and multifunction of regulating cellular behavior. Herein, a stable electrospinning method was employed to generate a biomimetic model of natural BMs based on novel scaffold electrospun from Poly(ɛ-caprolactone) (PCL) and cellulose acetate (CA) incorporated with chitosan (CS). The morphology, structure, surface hydrophilicity, roughness and mechanical tensile strength of prepared monolayer and tri-layered scaffold were comprehensively compared. Besides, co-culture system via seeding keratinocytes (Kcs) and fibroblasts (Fbs) on opposite side of tri-layered scaffold revealed more effective segregation of both cell types within the central nanofibrous barrier together with enhanced cell attachment and proliferation than that on the monolayer scaffold. Moreover, the deposition of type VII collagen and laminin-5 was examined in comparison with normal skin BMs. Furthermore, the histological studies revealed characteristics of reconstructing BM zone at the junction of dermis-epidermis after in vivo implantation for 2 weeks, and wound healing while the seeded cells interacted with the endogenous cells. Additionally, the expression of active integrin β1 and phosphorylated focal adhesion kinase (FAK) was promoted with treatment of tri-layered scaffold. This study stressed that this tri-layer scaffold might provoke biomimetic responses of Kcs and Fbs and thus be applied for future development of BMs containing tissues.

Bioengineered smart trilayer skin tissue substitute for efficient deep wound healing.

Skin substitutes for deep wound healing require meticulous designing and fabrication to ensure proper structural and functional regeneration of the tissue. Range of physical and mechanical properties conducive for regeneration of different layers of skin is a prerequisite of an ideal scaffold. However, single or bilayer substitutes, lacking this feature, fail to heal full thickness wound. Complete scar free regeneration of skin is still a big challenge. This study reports fabrication of a trilayer scaffold, from biodegradable polymers that can provide the right ambience for simultaneous regeneration of all the three layers of skin. The scaffold was developed through optimization of different fabrication techniques, namely, casting, electrospinning and lyophilisation, for obtaining a tailored trilayer structure. It has mechanical strength similar to skin layers, can maintain a porosity-gradient and provides microenvironments suitable for simultaneous regeneration of epidermis, dermis and hypodermis. A co-culture model, of keratinocytes and dermal fibroblasts, confirms the efficiency of the scaffold in supporting proliferation and differentiation of different types of cells, into organized tissue. The scaffold showed improved and expedited wound healing in-vivo. Taken together, these compelling evidences successfully established the engineered trilayer scaffold as a promising template for skin tissue regeneration in case of deep wound.