Sheet Scaffold Research Articles

Open-Spaced Ridged Hydrogel Scaffolds Containing TiO2-Self-Assembled Monolayer of Phosphonates Promote Regeneration and Recovery Following Spinal Cord Injury.

The spinal cord has a poor ability to regenerate after an injury, which may be due to cell loss, cyst formation, inflammation, and scarring. A promising approach to treating a spinal cord injury (SCI) is the use of biomaterials. We have developed a novel hydrogel scaffold fabricated from oligo(poly(ethylene glycol) fumarate) (OPF) as a 0.08 mm thick sheet containing polymer ridges and a cell-attractive surface on the other side. When the cells are cultured on OPF via chemical patterning, the cells attach, align, and deposit ECM along the direction of the pattern. Animals implanted with the rolled scaffold sheets had greater hindlimb recovery compared to that of the multichannel scaffold control, which is likely due to the greater number of axons growing across it. The immune cell number (microglia or hemopoietic cells: 50-120 cells/mm2 in all conditions), scarring (5-10% in all conditions), and ECM deposits (Laminin or Fibronectin: approximately 10-20% in all conditions) were equal in all conditions. Overall, the results suggest that the scaffold sheets promote axon outgrowth that can be guided across the scaffold, thereby promoting hindlimb recovery. This study provides a hydrogel scaffold construct that can be used in vitro for cell characterization or in vivo for future neuroprosthetics, devices, or cell and ECM delivery.

The combination of a 3D-Printed porous Ti-6Al-4V alloy scaffold and stem cell sheet technology for the construction of biomimetic engineered bone at an ectopic site.

Cell sheet technology has been widely used in bone tissue engineering and regenerative medicine. However, controlling the shape and volume of large pieces of engineered bone tissue remains impossible without additional suitable scaffolds. Three-dimensional (3D) printed titanium (Ti) alloy scaffolds are mostly used as implant materials for repairing bone defects, but the unsatisfactory bioactivities of traditional Ti-based scaffolds severely limit their clinical applications. Herein, we hypothesize that the combination of bone marrow mesenchymal stem cell (BMSC) sheet technology and 3D porous Ti–6Al–4V (PT) alloy scaffolds could be used to fabricate biomimetic engineered bone. First, various concentrations of BMSCs were directly cocultured with PT scaffolds to obtain complexes of osteoblastic cell sheets and scaffolds. Then, as an experimental control, an osteoblastic BMSC sheet was prepared by continuous culturing under osteogenic conditions for 2 weeks without passaging and used to wrap the scaffolds. The BMSC sheet was composed of several layers of extracellular matrix (ECM) and a mass of BMSCs. The BMSCs exhibited excellent adherent, proliferative and osteogenic potential when cocultured with PT scaffolds, which may be attributed to the ability of the 3D microstructure of scaffolds to facilitate the biological behaviors of cells, as confirmed by the in vitro results. Moreover, the presence of BMSCs and ECM increased the angiogenic potential of PT scaffolds by the secretion of VEGF. Micro-CT and histological analysis confirmed the in vivo formation of biomimetic engineered bone when the complex of cocultured BMSCs and PT scaffolds and the scaffolds wrapped by prepared BMSC sheets were implanted subcutaneously into nude mice. Therefore, the combination of BMSC sheet technology and 3D-printed PT scaffolds could be used to construct customized biomimetic engineered bone, offering a novel and promising strategy for the precise repair of bone defects.

Effects of a graphene oxide-alginate sheet scaffold on rotator cuff tendon healing in a rat model

Natural polymer scaffolds used to promote rotator cuff healing have limitations in terms of their mechanical and biochemical properties. This animal study aimed to investigate the effects of combined graphene oxide (GO) and alginate scaffold and the toxicity of GO on rotator cuff healing in a rat model. First, the mechanical properties of a GO/alginate scaffold and a pure alginate scaffold were compared. The in vitro cytotoxicity of and proliferation of human tenocytes with the GO/alginate scaffold were evaluated by CCK-8 assay. For the in vivo experiment, 20 male rats were randomly divided into two groups (n = 10 each), and supraspinatus repair was performed: group 1 underwent supraspinatus repair alone, and group 2 underwent supraspinatus repair with the GO/alginate scaffold. Biomechanical and histological analyses were performed to evaluate the quality of tendon-to-bone healing 8weeks after rotator cuff repair. The GO/alginate scaffold exhibited an increased maximum load (p = .001) and tensile strength (p = .001). In the cytotoxicity test, the cell survival rate with the GO/alginate scaffold was 102.08%. The proliferation rate of human tenocytes was no significant difference between the GO/alginate and alginate groups for 1, 3, 5, and 7days. Biomechanically, group 2 exhibited a significantly greater ultimate failure load (p < .001), ultimate stress (p < .001), and stiffness (p < .001) than group 1. The histological analysis revealed that the tendon-to-bone interface in group 2 showed more collagen fibers bridging, tendon-to-bone integration, longitudinally oriented collagen fibers, and fibrocartilage formation than in group 1. A small amount of GO added to alginate improved the mechanical properties of the scaffold without evidence of cytotoxicity. At 8 weeks after rotator cuff repair, the GO/alginate scaffold improved tendon-to-bone healing without causing any signs of toxicity in a rat model.

Engineered models for placental toxicology: Emerging approaches based on tissue decellularization

Recent increases in prescriptions and illegal drug use as well as exposure to environmental contaminants during pregnancy have highlighted the critical importance of placental toxicology in understanding and identifying risks to both mother and fetus. Although advantageous for basic science, current in vitro models often fail to capture the complexity of placental response, likely due to their inability to recreate and monitor aspects of the microenvironment including physical properties, mechanical forces and stiffness, protein composition, cell-cell interactions, soluble and physicochemical factors, and other exogenous cues. Tissue engineering holds great promise in addressing these challenges and provides an avenue to better understand basic biology, effects of toxic compounds and potential therapeutics. The key to success lies in effectively recreating the microenvironment. One strategy to do this would be to recreate individual components and then combine them. However, this becomes challenging due to variables present according to conditions such as tissue location, age, health status and lifestyle. The extracellular matrix (ECM) is known to influence cellular fate by working as a storage of factors. Decellularized ECM (dECM) is a recent tool that allows usage of the original ECM in a refurbished form, providing a relatively reliable representation of the microenvironment. This review focuses on using dECM in modified forms such as whole organs, scaffold sheets, electrospun nanofibers, hydrogels, 3D printing, and combinations as building blocks to recreate aspects of the microenvironment to address general tissue engineering and toxicology challenges, thus illustrating their potential as tools for future placental toxicology studies.

A hydrogel sheet mask with tea tree essential oil entrapment and targeted dose delivery capability

Skin is a very aesthetic organ of our body which is significant for personal health. Skin problem specially face skin problem has the psychosocial effect. Skin problems like acne is one of the most common problems among adolescents. Commonly these are treated with benzoyl peroxide and erythromycin by physicians, but tea tree oil also has equal efficacy similar to them. But these oils are hard to administer because the essential oil composition are volatile and thus, does not stay over the skin for long duration as well as a larger dose of this oil is known to irritate the skin. Since hydrogel is well-known for its targeted drug delivery system, therefore, hydrogel patch or scaffold sheet can be used to deliver the tea tee oil to the skin in optimum quantity. But hydrogels are hydrophilic and cannot absorb essential oils. β-cyclodextrin has the capability to entrap volatile, unstable molecules inside its hydrophobic interior and can release on the target site. Whereas, chitosan oligosaccharide has the antimicrobial activity. Therefore, the present study deal with the formulation of hydrogel with β-cyclodextrin and chitosan oligosaccharide along with pectin and carboxymethyl cellulose (CMC) to enable this hydrogel to entrap essential oil and show enhanced antimicrobial activity. The synthesized hydrogel HCCHB1 has very good cytocompatibility according to MTT assay. The antimicrobial activity of tea tree oil entrapped HCCHB1 is greater than the tea tree oil alone. Hence, it can be helpful to deliver reduced dose of tea tree oil over the skin and minimize irritation without compromising the antimicrobial activity of the tea tree oil. The HCCHB1 has good porosity and high swelling property in water, pH 5.5 and essential oil with 1001%, 1033% and 94% respectively. Additionally, the successful interaction of polymers among each other has occurred successfully as represented by FTIR.

Ethanol treatment of nanoPGA/PCL composite scaffolds enhances human chondrocyte development in the cellular microenvironment of tissue-engineered auricle constructs.

A major obstacle for tissue engineering ear-shaped cartilage is poorly developed tissue comprising cell-scaffold constructs. To address this issue, bioresorbable scaffolds of poly-ε-caprolactone (PCL) and polyglycolic acid nanofibers (nanoPGA) were evaluated using an ethanol treatment step before auricular chondrocyte scaffold seeding, an approach considered to enhance scaffold hydrophilicity and cartilage regeneration. Auricular chondrocytes were isolated from canine ears and human surgical samples discarded during otoplasty, including microtia reconstruction. Canine chondrocytes were seeded onto PCL and nanoPGA sheets either with or without ethanol treatment to examine cellular adhesion in vitro. Human chondrocytes were seeded onto three-dimensional bioresorbable composite scaffolds (PCL with surface coverage of nanoPGA) either with or without ethanol treatment and then implanted into athymic mice for 10 and 20 weeks. On construct retrieval, scanning electron microscopy showed canine auricular chondrocytes seeded onto ethanol-treated scaffolds in vitro developed extended cell processes contacting scaffold surfaces, a result suggesting cell-scaffold adhesion and a favorable microenvironment compared to the same cells with limited processes over untreated scaffolds. Adhesion of canine chondrocytes was statistically significantly greater (p ≤ 0.05) for ethanol-treated compared to untreated scaffold sheets. After implantation for 10 weeks, constructs of human auricular chondrocytes seeded onto ethanol-treated scaffolds were covered with glossy cartilage while constructs consisting of the same cells seeded onto untreated scaffolds revealed sparse connective tissue and cartilage regeneration. Following 10 weeks of implantation, RT-qPCR analyses of chondrocytes grown on ethanol-treated scaffolds showed greater expression levels for several cartilage-related genes compared to cells developed on untreated scaffolds with statistically significantly increased SRY-box transcription factor 5 (SOX5) and decreased interleukin-1α (inflammation-related) expression levels (p ≤ 0.05). Ethanol treatment of scaffolds led to increased cartilage production for 20- compared to 10-week constructs. While hydrophilicity of scaffolds was not assessed directly in the present findings, a possible factor supporting the summary data is that hydrophilicity may be enhanced for ethanol-treated nanoPGA/PCL scaffolds, an effect leading to improvement of chondrocyte adhesion, the cellular microenvironment and cartilage regeneration in tissue-engineered auricle constructs.

A compositionally synergistic approach for the development of a multifunctional bilayer scaffold with antibacterial property for infected and chronic wounds

The increased complexity of the healing process and interplay of various pathological factors in cutaneous wounds, in particular, chronic wounds, entails a meticulous and spatiotemporally sufficient treatment to accomplish successful therapy. Here, we have developed the hybrid bilayer scaffold system using cryogelation and electrospinning techniques to closely resemble the native three-dimensional architecture of skin. The bilayer scaffold involves curcumin dextran nanoparticles (CDNp) and cerium oxide nanoparticles (CeO2) loaded gelatin cryogel layer and polyvinyl alcohol-poly (vinyl pyrrolidone)-iodine-potassium iodide (PPI-KI) electrospun fibrous layer towards the outer side. The developed scaffold exhibited high water uptake capability, degradation and hemocompatibility besides showing sustained and prolonged release of CDNp (65% ± 2.61 in 120 h) and iodine (66.47% ± 2.073 in 72 h). The in-vitro studies of bilayer scaffold, cryogels and electrospun sheet demonstrated enhanced cell viability and accelerated cell migration apart from displaying a strong antioxidant and antibacterial activity. The developed scaffolds elucidated a faster-wound closure rate in the in-vivo wound model in comparison to commercially available dressing and other groups. Further, in the infected wound model, the bilayer scaffold prevented wound infection and showed rapid and efficient healing of wounds. The developed dressing was observed to promote mature epithelialization with ample hair growth and increased deposition of mature collagen. Thus, the novel synergistic approach can remarkably provide new insights for the design of advanced and multifunctional wound dressing with potential application in accelerating the healing of cutaneous wounds.

Development of a novel meniscal sheet scaffold and its effectiveness for meniscal regeneration in a rabbit defect model.

This study evaluated the biomechanical strength of a novel two-layer meniscal sheet scaffold (MSS) consisting of polyglycolic acid and poly-Llactic acid/caprolactone and investigated meniscal healing using wrapping treatment for meniscal defect model in a rabbit. The ultimate failure load of the MSS was determined using a tensile testing machine, in vitro. A 2-mm cylindrical defects were created at the medial meniscus of rabbit knees (n = 40). Each knee was assigned to one of two groups. The defect group was not treated and the MSS group underwent wrapping treatment with MSS. Menisci were harvested at 2, 4, 8, and 12 weeks post-implantation. The regenerated meniscus and defect size were evaluated using macrophotographs. Ishida scores for regenerated tissue were determined using Safranin-O/Fast Green staining. Immunohistochemical analysis of Ki-67 for cell proliferation, anti-type I and II collagen antibodies for structure of the regenerated tissue was elucidated. Medial femoral cartilage was stained with Safranin-O/Fast Green and evaluated with Osteoarthritis Research Society International (OARSI) scores. The strength of MSS was maintained over 90% from initial time point to 4 weeks after hydrolysis and over 60% of the strength remained at 8 weeks. The surface area of the meniscus was larger and the defect size smaller in the MSS group than in the defect group at 8 and 12 weeks. Ishida scores revealed that the MSS group improved significantly compared to that of the defect group at all postsurgery time points evaluated. Ki-67 positive cell ratio was significantly higher in the MSS group. OARSI score of the defect group was significantly higher and the defect group showed progressive degeneration in the articular cartilage from 8 to 12 weeks. Overall, wrapping meniscus defects with MSS was useful for accelerating meniscal healing from an early stage and beneficial for tissue regeneration and promoting extracellular matrix maturation.

Engineering of axially vascularized bone tissue using natural coral scaffold and osteogenic bone marrow mesenchymal stem cell sheets

IntroductionBlood supply remains one of the obstacles to large bone tissue engineering. This study aimed to generate vascularized bone tissue by inducing axial vascularization into a construct combining natural coral scaffold and a bone marrow mesenchymal stem cells (BMSCs) sheet. Material and methodsIsolated BMSCs were cultured to form an osteogenic cell sheet using a continuous culture method. Natural coral scaffolds were prepared into customized shape with a cylinder of 20 mm length, 8 mm in outer diameter and 5 mm in inner diameter. Then, the freed superficial inferior epigastric vessel of rabbits was first wrapped with a cell sheet, and then inserted into the central passage of the scaffold, after being wrapped with another cell sheet, the complexes were implanted subcutaneously into a rabbit groin area. In contrast, the sheet-scaffold construct that implanted into groin subcutaneous area of the other side of the same rabbit with the distal end of the blood vessel was ligated, which was considered as control. New bone and vascularization formation were evaluated at 12 weeks postoperatively. ResultsThe volume of new bone formation and amount of capillary infiltration in the vascular circulation group were significantly greater than that in the vascular ligation group, which suggested that insertion of axial vessels could significantly promote angiogenesis and osteogenesis of the tissue-engineered bone. ConclusionsThese findings indicate that inserting an arteriovenous bundle into the constructs of mesenchymal stem cell sheet and coral has great potential for clinical applications to repair large bone defects.

Immunoassays on thiol-ene synthetic paper generate a superior fluorescence signal

The fluorescence-based detection of biological complexes on solid substrates is widely used in microarrays and lateral flow tests. Here, we investigate thiol-ene micropillar scaffold sheets (“synthetic paper”) as the solid substrate in such assays. Compared to state-of-the-art glass and nitrocellulose substrates, assays on synthetic paper provide a stronger fluorescence signal, similar or better reproducibility, lower limit of detection (LOD), and the possibility of working with lower immunoreagent concentrations. Using synthetic paper, we detected the antibiotic enrofloxacin in whole milk with a LOD of 1.64 nM, which is on par or better than the values obtained with other common tests, and much lower than the maximum level allowed by European Union regulations. The significance of these results lays in that they indicate that synthetically-derived microstructured substrate materials have the potential to improve the performance of diagnostic assays.

High Thermal Conductivity Enhancement of Polymer Composites with Vertically Aligned Silicon Carbide Sheet Scaffolds.

Owing to the growth of demand for highly integrated electronic devices, high heat dissipation of thermal management materials is essential. Epoxy composites have been prepared with vertically aligned (VA) three-dimensional (3D)-structured SiC sheet scaffolds. The required VA-SiC sheet scaffolds were prepared by a novel approach starting with a graphene oxide (GO) scaffold. The VA-GO scaffolds were reduced to VA-graphene scaffolds in an argon environment, and the latter were subsequently transformed into VA-SiC sheet scaffolds by a template-assisted chemical vapor deposition method. Epoxy resin was filled in the empty spaces of the 3D scaffold of SiC sheets to prepare the composite mass. The material so prepared shows anisotropic thermal property with ultrahigh through-plane conductivity of 14.32 W·m-1·K-1 at a SiC sheet content of 3.71 vol %. A thermal percolation is observed at 1.78 vol % SiC filler. The SiC sheet scaffold of covalently interconnected SiC nanoparticles plays a vital role in the formation of the thermal conductive network to significantly enhance the thermal conductivity of epoxy composites. The application of the VA-SiC/epoxy composite as an efficient thermal dissipating material has also been presented. The VA-SiC/epoxy composites have a strong potential for preparing heat-dissipating components in integrated microelectronics.

A simvastatin-releasing scaffold with periodontal ligament stem cell sheets for periodontal regeneration.

Simvastatin (SIM) has been documented to induce the osteogenic differentiation of periodontal ligament stem cells (PDLSCs). To establish an efficient release system for periodontal regeneration, a polycaprolactone (PCL) membrane scaffold containing SIM was electrospun and evaluated. The obtained PCL-SIM membrane scaffold showed sustained release up to 28 days, without deleterious effect on proliferation of PDLSCs on the scaffolds. PDLSCs were seeded onto scaffolds and their osteogenic differentiation was evaluated. After 21 days, expressions of collagen type I, alkaline phosphatase and bone sialoprotein genes were significantly upregulated and mineralized matrix formation was increased on the PCL-SIM scaffolds compared with the PCL scaffolds. In a heterotopic periodontal regeneration model, a cell sheet-scaffold construct was assembled by placement of multilayers of PDLSC sheets on PCL or PCL-SIM scaffolds, and these were then placed between dentin and ceramic bovine bone for subcutaneous implantation in athymic mice. After 8 weeks, the PCL-SIM membrane showed formation of significantly more ectopic cementum-like mineral on the dentin surface. These findings demonstrated that the PCL-SIM membrane scaffold promotes cementum-like tissue formation by sustained drug release, suggesting the feasibility of its therapeutic use with PDLSC sheets to improve periodontal regeneration.

Histological Analysis of the Wrapping Treatment for Meniscal Horizontal Tears in Rabbits.

To investigate meniscal regeneration and prevent cartilage degeneration using wrapping treatment for meniscal horizontal tears that have been difficult to repair in rabbits. Thirty knees from 15 Japanese white rabbits were divided into the horizontal (horizontal tears) or wrapping (horizontal tears with wrapping treatment) groups. Horizontal tears were created and wrapped with a sheet scaffold containing polyglycolic acid, polylactic acid, and polycaprolactone. The meniscus was stained with Safranin-O/Fast Green and evaluated with modified Pauli scores at 8, 12, and 16 weeks after implantation (n = 5). Cell morphology was determined with hematoxylin and eosin staining. Mature collagen was confirmed with Picrosirius Red staining. Furthermore, immunohistochemical analysis of inducible nitric oxide synthase (iNOS) for inflammation, Ki-67 for proliferation, and type II collagen for regeneration was performed. Medial femoral cartilage was stained with Safranin-O/Fast Green and evaluated with the Osteoarthritis Research Society International score at 8 and 16 weeks. The wrapping group had significantly better regeneration than the horizontal group, especially at 16 weeks (P < 0.05). Wrapping treatment induced fibrochondrocyte-like cells at 16 weeks. After wrapping treatment, iNOS was overexpressed at 8 weeks, Ki-67 at 8 and 12 weeks, and type II collagen at 16 weeks. Cartilage degeneration in the wrapping group did not progress significantly compared with that in the horizontal group at 16 weeks (P < 0.05). Wrapping treatment for meniscal horizontal tears induced meniscal regeneration as the sheet scaffold might induce intrinsic and extrinsic repair. Regaining the meniscal function by the wrapping treatment prevented cartilage degeneration.

Electrical stimulation of cell growth and neurogenesis using conductive and nonconductive microfibrous scaffolds.

The effect of exogenous electrical stimulation on cell viability, attachment, growth, and neurogenesis was examined using PC12 cells in microfibrous viscose-rayon scaffolds immersed in culture medium. The scaffolds were applied either in their nonconductive state or after coating the fibres with 200 nm of gold to give a scaffold sheet resistivity of (13 ± 1.3) Ω square-1. The cells were treated for 12 days using direct current electrical stimulation of 2 h per day. No cytotoxic effects were observed when up to 500 mV (8.3 mV mm-1) was applied to the scaffolds without gold, or when up to 100 mV (1.7 mV mm-1) was applied to the scaffolds with gold. Compared with unstimulated cells, whereas electrical stimulation significantly enhanced cell growth and attachment in the nonconductive scaffolds without gold, similar effects were not found for the conductive scaffolds with gold. Neural differentiation in the presence of nerve growth factor was improved by electrical stimulation in both scaffolds; however, neurite development and the expression of key differentiation markers were greater in the nonconductive scaffolds without gold than in the scaffolds with gold. Application of the same current to scaffolds with and without gold led to much higher levels of neurogenesis in the scaffolds without gold. This work demonstrates that substantial benefits in terms of cell growth and neural differentiation can be obtained using electric fields exerted across nonconductive microfibrous scaffolds, and that this approach to electrical stimulation can be more effective than when the stimulus is applied to cells on conductive scaffolds.

Construction of a Multilayered Mesenchymal Stem Cell Sheet with a 3D Dynamic Culture System.

Stem cell therapy shows a promising future in regenerating injured organ and tissues, and the cell sheet technique has been developed to improve the low cell retention and poor survival within the target zone. However, during the in vitro construction process, a solution for maintaining stem cell bioactivity and increasing the cell amount within the cell sheet is urgently needed. Here, this protocol presents a method for constructing a multilayered cell sheet with favorable stem cell bioactivity and optimal operability. Decellularized porcine pericardium (DPP) is prepared by phospholipase A2 (PLA2) decellularization method as the cell sheet scaffold, and rat bone marrow mesenchymal stem cells (BMSCs) are isolated and expanded as the seeded cells. The temporary multilayered cell sheet structure is constructed by using RAD16-I peptide hydrogel. Finally, the cell sheet is cultured with a dynamic perfusion system to stabilize the three-dimensional (3D) structure, and the cell sheet could be obtained following a 48-hour culture in vitro. This protocol provides an efficient and feasible method for constructing a multilayered stem cell sheet, and the cell sheet could be developed as a favorable stem cell therapy product in the future.

Collagen/Cellulose Nanofiber Blend Scaffolds Prepared at Various pH Conditions

Living tissues modules consist of highly organized cells and extracellular matrices (ECMs) in a hierarchical manner. Among these ECMs, type I collagen (COL), which is the most abundant protein, is widely accepted in the tissue engineering field due to its biocompatibility. However, improvement of mechanical properties of COL scaffolds still remains a challenge. We prepared the scaffold sheets with blends of COL and 2,2,6,6-tetramethylpiperidine-1-oxil(TEMPO)-oxidized cellulose nanofiber (TOCN). TOCNs with high mechanical properties reinforced COL scaffolds. Moreover, we prepared their blends under various pH conditions and investigated their mechanical properties and biocompatibilities. Sheets prepared at a higher pH possess a better mechanical performance. From Fourier transform infrared spectroscopy, X-ray diffraction, and scanning electron microscopy measurements, it is proved that the higher mechanical properties were attributed to COL triple inter helix structure, hydrogen interaction, and electrostatic interaction with TOCN. The biocompatibilities of COL/TOCN prepared at a higher pH were increased. We successfully demonstrated COL/TOCN blend materials with a high mechanical strength and high biocompatibility for scaffold materials.

Micrometer scale guidance of mesenchymal stem cells to form structurally oriented large-scale tissue engineered cartilage.

Tissue engineered cartilage constructs made with human mesenchymal stem cells (hMSCs), scaffolds and bioactive factors are a promising solution to treat cartilage defects. A major disadvantage of these constructs is their inferior mechanical properties compared to the native tissue, which is likely due to the lack of structural organization of the extracellular matrix of the engineered constructs. In this study, we developed three-dimensional (3-D) cartilage constructs from rectangular scaffold sheets containing hMSCs in micro-guidance channels and characterized their mechanical properties and metabolic requirements. The work led to a novel roll-up method to embed 2-D microscale structures in 3-D constructs. Further, micro-guidance channels incorporated within the 3-D cartilage constructs led to the production of aligned cell-produced matrix and enhanced mechanical function.

Fabrication and characterization of a decellularized bovine tendon sheet for tendon reconstruction.

Obtaining a performing decellularized tendon scaffold with proper dimensions and adequate availability is highly desirable. However, the combined study of complete decellularization and detailed characterization of native tendon extracellular matrix (ECM) from large animals is still lacking. In the present study, we developed a new decellularization protocol, including physical methods and enzymatic solutions for processing bovine Achilles tendons, and produced a decellularized bovine tendon sheet (DBTS) scaffold for tendon reconstruction. The decellularization effectiveness was demonstrated by DNA quantification and histological qualification. The removal of the alpha-gal epitopes was confirmed by ELISA analysis and immunohistochemical staining. After decellularization, there were no significant alterations of the native tendon extracellular matrix (ECM) properties, including the internal ultrastructure, biochemical compositions such as collagen, glycosaminoglycans (GAGs), basic fibroblast growth factor (bFGF) and transforming growth factor-β1 (TGF-β1), fibronectin and decorin, as well as substantial mechanical strength. Furthermore, the DBTS scaffold showed no cytotoxic and promoted the proliferation of NIH-3T3 fibroblasts in vitro. When implanted into rat subcutaneous tissue, the DBTS scaffold displayed excellent histocompatibility in vivo. Our results, while offering a new decellularization protocol for large tendons, can provide a promising biologic scaffold with a combination of mechanical strength and tendon ECM bioactive factors that may have many potential applications in tendon reconstruction. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2299-2311, 2017.

PLGA-PTMC-Cultured Bone Mesenchymal Stem Cell Scaffold Enhances Cartilage Regeneration in Tissue-Engineered Tracheal Transplantation.

The treatment of long-segment tracheal defect requires the transplantation of effective tracheal substitute, and the tissue-engineered trachea (TET) has been proposed as an ideal tracheal substitute. The major cause of the failure of segmental tracheal defect reconstruction by TET is airway collapse caused by the chondromalacia of TET cartilage. The key to maintain the TET structure is the regeneration of chondrocytes in cartilage, which can secrete plenty of cartilage matrices. To address the problem of the chondromalacia of TET cartilage, this study proposed an improved strategy. We designed a new cell sheet scaffold using the poly(lactic-co-glycolic acid) (PLGA) and poly(trimethylene carbonate) (PTMC) to make a porous membrane for seeding cells, and used the PLGA-PTMC cell-scaffold to pack the decellularized allogeneic trachea to construct a new type of TET. The TET was then implanted in the subcutaneous tissue for vascularization for 2 weeks. Orthotopic transplantation was then performed after implantation. The efficiency of the TET we designed was analyzed by histological examination and biomechanical analyses 4 weeks after surgery. Four weeks after surgery, both the number of chondrocytes and the amount of cartilage matrix were significantly higher than those contained in the traditional stem-cell-based TET. Besides, the coefficient of stiffness of TET was significantly larger than the traditional TET. This study provided a promising approach for the long-term functional reconstruction of long-segment tracheal defect, and the TET we designed had potential application prospects in the field of TET reconstruction.

Prefabrication of axially vascularized bone by combining β-tricalciumphosphate, arteriovenous loop, and cell sheet technique.

The repair of bone defects poses a great challenge for reconstructive surgeons. Although the development of tißsue engineering has exhibited promise in replacing damaged bone, the fabrication of large constructs with functional blood veßsels remains an obstacle. From the orthopedic surgeon's point of view, the generation of axially vascularized bone, which can anastomose with the recipient vessel, might be a solution to this medical problem. In this study, we aimed to prefabricate an axially vascularized bone by combining a ß-TCP scaffold, arteriovenous loop (AVL), and cell sheet in a bioreactor in vivo. Twelve rabbits were randomly allocated into two groups: the experimental group (presence of AVL) and the control group (absence of AVL). The constructs were explanted at 8 weeks postoperatively. The histomorphometric results showed 42.8±5.9% of the bone area in the AVL group and 26.6±3.5% in the control group. Similarly, vessel analysis revealed the average vessel density in the AVL group (12.5±3.3) was significantly more than that in the control group (6.1±1.5, p<0.05). Our research indicated that the combination of a ß-TCP scaffold, AVL and cell sheet might engineer vascularized bone. This prefabrication strategy might facilitate clinical translation of bone tissue engineering in reconstructing large bone defects.