Cevat Erisken

Protein-releasing polymeric scaffolds induce fibrochondrocytic differentiation of endogenous cells for knee meniscus regeneration in sheep

Sheep knee meniscus was regenerated by spatiotemporal delivery of two recombinant human proteins in 3D-printed bioresorbable scaffold. Bioprinting Cartilage Scaffolds Weak in the knees? You may need more cartilage. Because donor cartilage as well as synthetic products are scarce, Lee et al. devised a way to regenerate the knee meniscus, using the native tissue from sheep knees as inspiration. The authors three-dimensionally (3D) printed poly-ε-caprolactone (PCL) scaffolds based on the anatomically correct dimensions of the native meniscus. These scaffolds were loaded at specific locations with polymeric microspheres containing one of two growth factors, to be released sequentially. Connective tissue growth factor (CTGF) was released first, followed by transforming growth factor–β3 (TGFβ3). Control over both spatial and temporal protein release allowed for zone-specific matrix development: type I collagen in the outer zone and type II collagen in the inner zone, similar to the native meniscus. It is believed that such growth factor release induced endogenous stem cells to differentiate into fibrochondrocytes—the cells that make up the meniscus. In vivo in a large animal (sheep) model, the implanted scaffolds led to tissue regeneration with desired mechanical properties. If translated to humans, this acellular biomaterial would fulfill the major unmet need of repairing deteriorated cartilage in not only the knees but also tendon-bone junctions, the intervertebral discs of the spine, and the temporomandibular joint. Regeneration of complex tissues, such as kidney, liver, and cartilage, continues to be a scientific and translational challenge. Survival of ex vivo cultured, transplanted cells in tissue grafts is among one of the key barriers. Meniscus is a complex tissue consisting of collagen fibers and proteoglycans with gradient phenotypes of fibrocartilage and functions to provide congruence of the knee joint, without which the patient is likely to develop arthritis. Endogenous stem/progenitor cells regenerated the knee meniscus upon spatially released human connective tissue growth factor (CTGF) and transforming growth factor–β3 (TGFβ3) from a three-dimensional (3D)–printed biomaterial, enabling functional knee recovery. Sequentially applied CTGF and TGFβ3 were necessary and sufficient to propel mesenchymal stem/progenitor cells, as a heterogeneous population or as single-cell progenies, into fibrochondrocytes that concurrently synthesized procollagens I and IIα. When released from microchannels of 3D-printed, human meniscus scaffolds, CTGF and TGFβ3 induced endogenous stem/progenitor cells to differentiate and synthesize zone-specific type I and II collagens. We then replaced sheep meniscus with anatomically correct, 3D-printed scaffolds that incorporated spatially delivered CTGF and TGFβ3. Endogenous cells regenerated the meniscus with zone-specific matrix phenotypes: primarily type I collagen in the outer zone, and type II collagen in the inner zone, reminiscent of the native meniscus. Spatiotemporally delivered CTGF and TGFβ3 also restored inhomogeneous mechanical properties in the regenerated sheep meniscus. Survival and directed differentiation of endogenous cells in a tissue defect may have implications in the regeneration of complex (heterogeneous) tissues and organs.

Biomimetic scaffold design for functional and integrative tendon repair

Rotator cuff tears represent the most common shoulder injuries in the United States. The debilitating effect of this degenerative condition coupled with the high incidence of failure associated with existing graft choices underscores the clinical need for alternative grafting solutions. The 2 critical design criteria for the ideal tendon graft would require the graft to not only exhibit physiologically relevant mechanical properties but also be able to facilitate functional graft integration by promoting the regeneration of the native tendon-to-bone interface. Centered on these design goals, this review will highlight current approaches to functional and integrative tendon repair. In particular, the application of biomimetic design principles through the use of nanofiber- and nanocomposite-based scaffolds for tendon tissue engineering will be discussed. This review will begin with nanofiber-based approaches to functional tendon repair, followed by a section highlighting the exciting research on tendon-to-bone interface regeneration, with an emphasis on implementation of strategic biomimicry in nanofiber scaffold design and the concomitant formation of graded multi-tissue systems for integrative soft-tissue repair. This review will conclude with a summary and discussion of future directions.

Scaffold Fiber Diameter Regulates Human Tendon Fibroblast Growth and Differentiation

The diameter of collagen fibrils in connective tissues, such as tendons and ligaments is known to decrease upon injury or with age, leading to inferior biomechanical properties and poor healing capacity. This study tests the hypotheses that scaffold fiber diameter modulates the response of human tendon fibroblasts, and that diameter-dependent cell responses are analogous to those seen in healthy versus healing tissues. Particularly, the effect of the fiber diameter (320 nm, 680 nm, and 1.80 μm) on scaffold properties and the response of human tendon fibroblasts were determined over 4 weeks of culture. It was observed that scaffold mechanical properties, cell proliferation, matrix production, and differentiation were regulated by changes in the fiber diameter. More specifically, a higher cell number, total collagen, and proteoglycan production were found on the nanofiber scaffolds, while microfibers promoted the expression of phenotypic markers of tendon fibroblasts, such as collagen I, III, V, and tenomodulin. It is possible that the nanofiber scaffolds of this study resemble the matrix in a state of injury, stimulating the cells for matrix deposition as part of the repair process, while microfibers represent the healthy matrix with micron-sized collagen bundles, thereby inducing cells to maintain the fibroblastic phenotype. The results of this study demonstrate that controlling the scaffold fiber diameter is critical in the design of scaffolds for functional and guided connective tissue repair, and provide new insights into the role of matrix parameters in guiding soft tissue healing.

Viscoelastic and Biomechanical Properties of Osteochondral Tissue Constructs Generated From Graded Polycaprolactone and Beta-Tricalcium Phosphate Composites

The complex micro-/nanostructure of native cartilage-to-bone insertion exhibits gradations in extracellular matrix components, leading to variations in the viscoelastic and biomechanical properties along its thickness to allow for smooth transition of loads under physiological movements. Engineering a realistic tissue for osteochondral interface would, therefore, depend on the ability to develop scaffolds with properly graded physical and chemical properties to facilitate the mimicry of the complex elegance of native tissue. In this study, polycaprolactone nanofiber scaffolds with spatially controlled concentrations of beta-tricalcium phosphate nanoparticles were fabricated using twin-screw extrusion-electrospinning process and seeded with MC3T3-E1 cells to form osteochondral tissue constructs. The objective of the study was to evaluate the linear viscoelastic and compressive properties of the native bovine osteochondral tissue and the tissue constructs formed in terms of their small-amplitude oscillatory shear, unconfined compression, and stress relaxation behavior. The native tissue, engineered tissue constructs, and unseeded scaffolds exhibited linear viscoelastic behavior for strain amplitudes less than 0.1%. Both native tissue and engineered tissue constructs demonstrated qualitatively similar gel-like behavior as determined using linear viscoelastic material functions. The normal stresses in compression determined at 10% strain for the unseeded scaffold, the tissue constructs cultured for four weeks, and the native tissue were 0.87+/-0.08 kPa, 3.59+/-0.34 kPa, and 210.80+/-8.93 kPa, respectively. Viscoelastic and biomechanical properties of the engineered tissue constructs were observed to increase with culture time reflecting the development of a tissuelike structure. These experimental findings suggest that viscoelastic material functions of the tissue constructs can provide valuable inputs for the stages of in vitro tissue development.

Mechanical and burning properties of highly loaded composite propellants

An improvement in the performance of solid rocket motors was achieved by increasing the oxidizer content of HTPB-based solid propellants. To minimize the adverse changes in the mechanical and rheological properties due to the increased amount of hard solid particles in the soft polymeric binder matrix, the optimum combi- nation of the particle sizes and volume fractions of the bimodal ammonium perchlorate and the aluminum powder in the solid load was obtained from the results of testing a series of propellant samples prepared by using ammonium perchlorate in four different average particle sizes, 9.22, 31.4, 171, and 323 mm. The maximum packing density of solids in the binder matrix was determined by changing the sizes and the volume fractions of fine and coarse ammonium perchlorate at constant solid loading. The aver- age size (10.4 mm) and concentration of aluminum powder used as metallic fuel were maintained constant for ballistic requirements. Optimum sizes and fine-to-coarse ratio of ammonium perchlorate particles were determined to be at mean diameters of 31.4 and 323 mm and fine-to-coarse ratio of 35/65. Solid content of the propellant was then increased from 75 to 85.6% by volume by using the predetermined optimum sizes and fine to coarse ratio of ammonium perchlorate. Mechanical properties of the propellant samples were measured by using an Instron tester with a crosshead speed of 50 mm/ min at 257C. The effect of oxidizer content and fine-to-coarse ratio of oxidizer on the burning rate of the propellant was also investigated by using a strand burner at various pressures. From experiments in which the size and the fine-to-coarse ratio of ammo- nium perchlorate were changed at constant solid loading, a minimum value of initial modulus was obtained for each fine-to-coarse ratio, indicating that the solids packing fraction is maximum at this ratio. The tensile strength and the burning rate increase, while the elongation at maximum stress decreases with increasing fine-to-coarse ratio of ammonium perchlorate. Experiments in which the total solid loading was increased at constant fine-to-coarse ratio of ammonium perchlorate show that the modulus, the tensile strength and the burning rate increase, while the elongation at maximum stress decreases with increasing solid loading. Propellants having solid loading of up to 82% exhibit acceptable mechanical properties and improved burning properties suitable for rocket applications. q 1998 John Wiley & Sons, Inc. J Appl Polym Sci 67: 1457-1464, 1998

Synthesis and characterization of polycaprolactone for anterior cruciate ligament regeneration

Anterior cruciate ligament (ACL) is the most frequently torn ligament in the knee, and complete healing is unlikely due to lack of vascularization. Current approaches for the treatment of ACL injuries include surgical interventions and grafting, however recent reports show that surgeries have 94% recurrency, and that repaired tissues are biomechanically inferior to the native tissue. These necessitate the need for new strategies for scar-free repair/regeneration of ACL injuries. Polycaprolactone (PCL) is a biodegradable and biocompatible synthetic polymer, which has been widely used in the connective tissue repair/regeneration attempts. Here, we report on the synthesis of PCL via ring opening polymerization using ε-caprolactone as the monomer, and ammonium heptamolybdate as a catalyst. The synthesized PCL was characterized using Fourier Transform Infrared Spectroscopy (FTIR) and Nuclear Magnetic Resonance (NMR) spectroscopy. It was then processed using electrospinning to form nanofiber-based scaffolds. These scaffolds were characterized in terms of surface as well as mechanical properties, and compared to the properties of commercially available PCL, and of native ACL tissue harvested from sheep. In addition, scaffolds fabricated with synthesized PCL were evaluated regarding their cell attachment capacity using human bone marrow mesenchymal stem cells (hBMSCs). Our findings demonstrated that the synthesized PCL is similar to its commercially available counterpart in terms of surface morphology and mechanical properties. In addition, fibrous scaffolds generated with electrospinning showed weaker mechanical properties visa vis native ACL tissue in terms of ultimate stress, and elastic modulus. Also, the synthesized PCL can accommodate cell attachment when tested with hBMSCs. Putting together, these observations reveal that the PCL synthesized in this study could be a good candidate as a biomaterial for ligament repair or regeneration.

Processing of polycaprolactone and hydroxyapatite to fabricate graded electrospun composites for tendon-bone interface regeneration

Abstract The process of electrospinning is utilized with different approaches including conventional electrospinning, extrusion electrospinning, and electroblowing to form nanofibrous meshes and composites. Here, we report on the quality and properties of spatially graded polycaprolactone (PCL) and nano-hydroxyapatite (nHA) composite meshes fabricated with multiple-spinneret electrospinning. The composite meshes were characterized in terms of the amount of spatially allocated nHA concentration across the mesh, fiber diameter, porosity, pore size, and hydrophilicity of meshes. Results show that linearly and continuously varying nHA concentration distribution, i.e. graded structure, can be accomplished across the mesh thickness using multiple-spinneret electrospinning, which is in accordance with the change of mineral concentration observed in native tendon-bone interface. Furthermore, incorporation of nanoparticles into nanofibers led to increased fiber diameter as depicted by a shift in fiber diameter distribution, a significant increase in mean fiber diameter from 361±9 nm to 459±21 nm, and an increase in contact angle from 120.01±2.77° to 115.24±1.17°. These findings suggest that the composite meshes formed in this study could serve as model systems to be used as scaffolds in tendon-bone tissue engineering application in particular, and for other tissue-tissue interfaces in a broader context.

Characterization of Human Dental Pulp Tissue Under Oscillatory Shear and Compression

Availability of material as well as biological properties of native tissues is critical for biomaterial design and synthesis for regenerative engineering. Until recently, selection of biomaterials and biomolecule carriers for dental pulp regeneration has been done randomly or based on experience mainly due to the absence of benchmark data for dental pulp tissue. This study, for the first time, characterizes the linear viscoelastic material functions and compressive properties of human dental pulp tissue harvested from wisdom teeth, under oscillatory shear and compression. The results revealed a gel-like behavior of the pulp tissue over the frequency range of 0.1-100 rps. Uniaxial compression tests generated peak normal stress and compressive modulus values of 39.1 ± 20.4 kPa and 5.5 ± 2.8 kPa, respectively. Taken collectively, the linear viscoelastic and uniaxial compressive properties of the human dental pulp tissue reported here should enable the better tailoring of biomaterials or biomolecule carriers to be employed in dental pulp regeneration.

Effects of QMix and ethylenediaminetetraacetic acid on decalcification and erosion of root canal dentin

The aim of this study was to evaluate the effect of initial NaOCl on the decalcification and erosion ability of EDTA and QMix. Sixty‐maxillary‐incisors were bisected longitudinally and the tooth‐halves were used. The experiment was conducted in two‐sets. In set‐I, 80‐tooth halves were treated in the presence or absence of initial NaOCl and EDTA. In set‐II, 40‐tooth halves were immersed in NaOCl and QMix. After each treatment, calcium‐ion release was determined with flame photometry. The erosion was imaged using SEM. Initial NaOCl led to concentration‐ and time‐dependent increase in calcium removal effect of 17% EDTA (p < .05). The rate of calcium removal and root canal wall erosion was considerably more severe with the use of 5% NaOCl for 3 min (p < .05). QMix as a final solution showed less decalcification and erosion than 17% EDTA when used 5% NaOCl as an initial irrigant (p < .05). Optimizing the concentration and application time of NaOCl can decrease the decalcification effect of chelating agents.

Cartilage-Bone Interface Features, Scaffold and Cell Options for Regeneration

Tissues with different material and biological properties are connected to one another through interfaces, which can be generally categorized as soft-to-soft tissue interfaces (muscle-tendon, etc.), softto-hard tissue interfaces (cartilage-bone, tendon-bone, etc.) and hardto-hard tissue interfaces (dentin-enamel, etc.). Since these interfaces merge biological materials, i.e., tissues, having distinct composition, structure and function, they possess complexities associated with their hierarchical structures, and when injured their healing/regeneration pathways follow more intricate phenomena compared to single tissues making up the interfaces. Findings reveal that injuries related to tissues connected in series occur mostly at the interfaces due to the mismatch between material properties of individual tissues. Therefore, interface tissue engineering has recently attracted significant attention from academia to be able to understand the mechanism of cell-materials interactions relevant to interfaces. This paper reviews the structure, composition and function of cartilage-bone interface in conjunction with the scaffold and cell options for its regeneration.