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C A Van Blitterswijk - One of the best experts on this subject based on the ideXlab platform.

  • 3d fiber deposited scaffolds for tissue engineering influence of pores geometry and architecture on dynamic mechanical properties
    Biomaterials, 2006
    Co-Authors: Lorenzo Moroni, J R De Wijn, C A Van Blitterswijk
    Abstract:

    One of the main issues in tissue engineering is the fabrication of scaffolds that closely mimic the biomechanical properties of the tissues to be regenerated. Conventional fabrication techniques are not sufficiently suitable to control scaffold structure to modulate mechanical properties. Within novel scaffold fabrication processes 3D fiber deposition (3DF) showed great potential for tissue engineering applications because of the precision in making reproducible 3D scaffolds, characterized by 100% interconnected pores with different shapes and sizes. Evidently, these features also affect mechanical properties. Therefore, in this study we considered the influence of different structures on dynamic mechanical properties of 3DF scaffolds. Pores were varied in size and shape, by changing fibre diameter, spacing and orientation, and layer thickness. With increasing porosity, dynamic mechanical analysis (DMA) revealed a decrease in elastic properties such as dynamic stiffness and Equilibrium Modulus, and an increase of the viscous parameters like damping factor and creep unrecovered strain. Furthermore, the Poisson's ratio was measured, and the shear Modulus computed from it. Scaffolds showed an adaptable degree of compressibility between sponges and incompressible materials. As comparison, bovine cartilage was tested and its properties fell in the fabricated scaffolds range. This investigation showed that viscoelastic properties of 3DF scaffolds could be modulated to accomplish mechanical requirements for tailored tissue engineered applications.

  • 3d fiber deposited scaffolds for tissue engineering influence of pores geometry and architecture on dynamic mechanical properties
    Biomaterials, 2006
    Co-Authors: Lorenzo Moroni, J R De Wijn, C A Van Blitterswijk
    Abstract:

    One of the main issues in tissue engineering is the fabrication of scaffolds that closely mimic the biomechanical properties of the tissues to be regenerated. Conventional fabrication techniques are not sufficiently suitable to control scaffold structure to modulate mechanical properties. Within novel scaffold fabrication processes 3D fiber deposition (3DF) showed great potential for tissue engineering applications because of the precision in making reproducible 3D scaffolds, characterized by 100% interconnected pores with different shapes and sizes. Evidently, these features also affect mechanical properties. Therefore, in this study we considered the influence of different structures on dynamic mechanical properties of 3DF scaffolds. Pores were varied in size and shape, by changing fibre diameter, spacing and orientation, and layer thickness. With increasing porosity, dynamic mechanical analysis (DMA) revealed a decrease in elastic properties such as dynamic stiffness and Equilibrium Modulus, and an increase of the viscous parameters like damping factor and creep unrecovered strain. Furthermore, the Poisson's ratio was measured, and the shear Modulus computed from it. Scaffolds showed an adaptable degree of compressibility between sponges and incompressible materials. As comparison, bovine cartilage was tested and its properties fell in the fabricated scaffolds range. This investigation showed that viscoelastic properties of 3DF scaffolds could be modulated to accomplish mechanical requirements for tailored tissue engineered applications.

  • design of porous scaffolds for cartilage tissue engineering using a three dimensional fiber deposition technique
    Biomaterials, 2004
    Co-Authors: Tim F Woodfield, Jos Malda, J R De Wij, Fabienne Peters, J Riesle, C A Van Blitterswijk
    Abstract:

    In this study, we present and characterize a fiber deposition technique for producing three-dimensional poly(ethylene glycol)-terephthalate—poly(butylene terephthalate) (PEGT/PBT) block co-polymer scaffolds with a 100% interconnecting pore network for engineering of articular cartilage. The technique allowed us to “design-in” desired scaffold characteristics layer by layer by accurately controlling the deposition of molten co-polymer fibers from a pressure-driven syringe onto a computer controlled x–y–z table. By varying PEGT/PBT composition, porosity and pore geometry, 3D-deposited scaffolds were produced with a range of mechanical properties. The Equilibrium Modulus and dynamic stiffness ranged between 0.05–2.5 and 0.16–4.33 MPa, respectively, and were similar to native articular cartilage explants (0.27 and 4.10 MPa, respectively). 3D-deposited scaffolds seeded with bovine articular chondrocytes supported a homogeneous cell distribution and subsequent cartilage-like tissue formation following in vitro culture as well as subcutaneous implantation in nude mice. This was demonstrated by the presence of articular cartilage extra cellular matrix constituents (glycosaminoglycan and type II collagen) throughout the interconnected pore volume. Similar results were achieved with respect to the attachment of expanded human articular chondrocytes, resulting in a homogenous distribution of viable cells after 5 days dynamic seeding. The processing methods and model scaffolds developed in this study provide a useful method to further investigate the effects of scaffold composition and pore architecture on articular cartilage tissue formation.

Clark T Hung - One of the best experts on this subject based on the ideXlab platform.

  • silk microfiber reinforced silk hydrogel composites for functional cartilage tissue repair
    Acta Biomaterialia, 2015
    Co-Authors: Supansa Yodmuang, Clark T Hung, Pen-hsiu Grace Chao, Stephanie L Mcnamara, Adam B Nover, Biman B Mandal, Monica Agarwal, Terriann N Kelly, David L Kaplan, Gordana Vunjaknovakovic
    Abstract:

    Abstract Cartilage tissue lacks an intrinsic capacity for self-regeneration due to slow matrix turnover, a limited supply of mature chondrocytes and insufficient vasculature. Although cartilage tissue engineering has achieved some success using agarose as a scaffolding material, major challenges of agarose-based cartilage repair, including non-degradability, poor tissue–scaffold integration and limited processing capability, have prompted the search for an alternative biomaterial. In this study, silk fiber–hydrogel composites (SF–silk hydrogels) made from silk microfibers and silk hydrogels were investigated for their potential use as a support material for engineered cartilage. We demonstrated the use of 100% silk-based fiber–hydrogel composite scaffolds for the development of cartilage constructs with properties comparable to those made with agarose. Cartilage constructs with an Equilibrium Modulus in the native tissue range were fabricated by mimicking the collagen fiber and proteoglycan composite architecture of native cartilage using biocompatible, biodegradable silk fibroin from Bombyx mori . Excellent chondrocyte response was observed on SF–silk hydrogels, and fiber reinforcement resulted in the development of more mechanically robust constructs after 42 days in culture compared to silk hydrogels alone. Thus, we demonstrate the versatility of silk fibroin as a composite scaffolding material for use in cartilage tissue repair to create functional cartilage constructs that overcome the limitations of agarose biomaterials, and provide a much-needed alternative to the agarose standard.

  • tissue engineered articular cartilage exhibits tension compression nonlinearity reminiscent of the native cartilage
    Journal of Biomechanics, 2013
    Co-Authors: Terriann N Kelly, Gerard A. Ateshian, Eric G Lima, Brendan L Roach, Zachary Weidner, Charles R Mackenziesmith, Grace D Oconnell, Aaron M Stoker, James L Cook, Clark T Hung
    Abstract:

    The tensile Modulus of articular cartilage is much larger than its compressive Modulus. This tension-compression nonlinearity enhances interstitial fluid pressurization and decreases the frictional coefficient. The current set of studies examines the tensile and compressive properties of cylindrical chondrocyte-seeded agarose constructs over different developmental stages through a novel method that combines osmotic loading, video microscopy, and uniaxial unconfined compression testing. This method was previously used to examine tension-compression nonlinearity in native cartilage. Engineered cartilage, cultured under free-swelling (FS) or dynamically loaded (DL) conditions, was tested in unconfined compression in hypertonic and hypotonic salt solutions. The apparent Equilibrium Modulus decreased with increasing salt concentration, indicating that increasing the bath solution osmolarity shielded the fixed charges within the tissue, shifting the measured moduli along the tension-compression curve and revealing the intrinsic properties of the tissue. With this method, we were able to measure the tensile (401±83 kPa for FS and 678±473 kPa for DL) and compressive (161±33 kPa for FS and 348±203 kPa for DL) moduli of the same engineered cartilage specimens. These moduli are comparable to values obtained from traditional methods, validating this technique for measuring the tensile and compressive properties of hydrogel-based constructs. This study shows that engineered cartilage exhibits tension-compression nonlinearity reminiscent of the native tissue, and that dynamic deformational loading can yield significantly higher tensile properties.

  • electrostatic and non electrostatic contributions of proteoglycans to the compressive Equilibrium Modulus of bovine articular cartilage
    Journal of Biomechanics, 2010
    Co-Authors: Clare Canal Guterl, Clark T Hung, Gerard A. Ateshian
    Abstract:

    Abstract This study presents direct experimental evidence for assessing the electrostatic and non-electrostatic contributions of proteoglycans to the compressive Equilibrium Modulus of bovine articular cartilage. Immature and mature bovine cartilage samples were tested in unconfined compression and their depth-dependent Equilibrium compressive Modulus was determined using strain measurements with digital image correlation analysis. The electrostatic contribution was assessed by testing samples in isotonic and hypertonic saline; the combined contribution was assessed by testing untreated and proteoglycan-depleted samples. Though it is well recognized that proteoglycans contribute significantly to the compressive stiffness of cartilage, results demonstrate that the combined electrostatic and non-electrostatic contributions may add up to more than 98% of the Modulus, a magnitude not previously appreciated. Of this contribution, about two thirds arises from electrostatic effects. The compressive Modulus of the proteoglycan-depleted cartilage matrix may be as low as 3 kPa, representing less than 2% of the normal tissue Modulus; experimental evidence also confirms that the collagen matrix in digested cartilage may buckle under compressive strains, resulting in crimping patterns. Thus, it is reasonable to model the collagen as a fibrillar matrix that can sustain only tension. This study also demonstrates that residual stresses in cartilage do not arise exclusively from proteoglycans, since cartilage remains curled relative to its in situ geometry even after proteoglycan depletion. These increased insights on the structure–function relationships of cartilage can lead to improved constitutive models and a better understanding of the response of cartilage to physiological loading conditions.

  • genipin enhances the mechanical properties of tissue engineered cartilage and protects against inflammatory degradation when used as a medium supplement
    Journal of Biomedical Materials Research Part A, 2009
    Co-Authors: Eric G Lima, Gerard A. Ateshian, Kacey G. Marra, Alicia J. Defail, Andrea R Tan, Timon Tai, Clark T Hung
    Abstract:

    Osteoarthritis (OA) is a common and painful disorder characterized by the degeneration of the articular surfaces of diarthrodial joints. One of the most promising approaches to treat OA is the design of engineered tissue that reproduces the functional properties of healthy cartilage. This tissue engineering approach typically entails the use of living cells embedded in a three-dimensional scaffold that is cultured over time in a growth medium supplemented with various physical or biological stimulants to encourage development. Recently our laboratory has been able to cultivate engineered cartilage with an Equilibrium Modulus (EY) and an glycosaminoglycan (GAG) content that match the native tissue using the temporal application of growth factors over a six-week culture period1. While these findings are encouraging, the dynamic Modulus (G*) and the collagen content for these constructs amount to less than a quarter of those of native articular cartilage. Of the two mechanical measurements, G*, which is correlated with the collagen content, is considered to be the more physiologically relevant as it measures the behavior of the tissue during the application of cyclic loads and better captures fluid pressurization within the biphasic tissue. No group has currently been able to reproduce native values of collagen regardless of the tissue engineering strategy employed. Obtaining native values of G* and collagen therefore remains an important but elusive challenge. Cross-linking agents such as glutaraldehyde, formaldehyde, or epoxy are typically used to chemically-treat (or fix) devitalized xenografts and allografts to reduce the immune rejection or the enzymatic degradation that is typical of transplanted bioprostheses2,3. Typically, however, cross-linking agents are lethal to cells. The use of genipin, a naturally occurring crosslinker with toxicity levels ten thousand fold less than glutaraldehyde4 as a biocompatible and stable cross-linker has been established by other groups5,6 and provides a promising new avenue to explore for tissue-engineering. Genipin cross-linking works by forming intra- and intermolecular cross-links of the amino residues on tropocollagen or proteoglycan molecules. A modified cyclic form of genipin can reside stably within the extracellular network, adding bridges across adjacent fibers. As such, previous groups have explored the use of genipin as a onetime treatment to fix tissue prior to implantation7, for the assembly of tissue-engineering scaffolds prior to cell seeding8,9, or in order to modulate the release of growth factors from degradable scaffolds or beads5. Genipin has also been studied for its anti-inflammatory effects, either administered directly to different cell lines10,11, or administered orally to animals12,13. Moreover, genipin cross-linking has been shown to affect the mechanical properties of biological tissues, increasing the tensile strength of bovine pericardium14, porcine tendon15, and type I collagen gels16. Our laboratory uses agarose hydrogel as a scaffold system for cartilage tissue engineering. This gel has been used extensively in chondrocyte biology studies and has shown some promise for tissue engineering applications. Agarose is a neutrally charged polysaccharide and as such is unaffected by genipin. However the chondrocytes embedded in the gel elaborate an extracellular matrix over time in culture that exhibits amine groups, which are subject to genipin cross-linking. In this set of studies we propose a novel use for genipin: not as a scaffold cross-linker, but as a medium supplement to promote cross-linking of de novo cell products as they are produced. We hypothesize that the application of genipin will stabilize the extracellular matrix components and increase the mechanical properties of developing cartilaginous tissue in our agarose hydrogels. We hypothesize two mechanisms through which the physical enhancement of tissue properties is fostered: (1) by reorganization and enhanced retention of cell synthesized extracellular matrix components, and (2) through reduction of the loss of extracellular matrix components by increasing their resilience to catabolic degradation.

  • the beneficial effect of delayed compressive loading on tissue engineered cartilage constructs cultured with tgf β3
    Osteoarthritis and Cartilage, 2007
    Co-Authors: Eric G Lima, Robert L. Mauck, Gerard A. Ateshian, Liming Bian, Benjamin A Byers, Rocky S Tuan, Clark T Hung
    Abstract:

    Summary Objective To determine whether the functional properties of tissue-engineered constructs cultured in a chemically-defined medium supplemented briefly with TGF-β3 can be enhanced with the application of dynamic deformational loading. Methods Primary immature bovine cells (2–3 months old) were encapsulated in agarose hydrogel (2%, 30×10 6 cells/ml) and cultured in chemically-defined medium supplemented for the first 2 weeks with transforming growth factor beta 3 (TGF-β3) (10μg/ml). Physiologic deformational loading (1Hz, 3h/day, 10% unconfined deformation initially and tapering to 2% peak-to-peak deformation by day 42) was applied either concurrent with or after the period of TGF-β3 supplementation. Mechanical and biochemical properties were evaluated up to day 56. Results Dynamic deformational loading applied concurrently with TGF-β3 supplementation yielded significantly lower (−90%) overall mechanical properties when compared to free-swelling controls. In contrast, the same loading protocol applied after the discontinuation of the growth factor resulted in significantly increased (+10%) overall mechanical properties relative to free-swelling controls. Equilibrium Modulus values reach 1306±79kPa and glycosaminoglycan levels reach 8.7±1.6% w.w. during this 8-week period and are similar to host cartilage properties (994±280kPa, 6.3±0.9% w.w.). Conclusions An optimal strategy for the functional tissue engineering of articular cartilage, particularly to accelerate construct development, may incorporate sequential application of different growth factors and applied deformational loading.

Gordana Vunjaknovakovic - One of the best experts on this subject based on the ideXlab platform.

  • silk microfiber reinforced silk hydrogel composites for functional cartilage tissue repair
    Acta Biomaterialia, 2015
    Co-Authors: Supansa Yodmuang, Clark T Hung, Pen-hsiu Grace Chao, Stephanie L Mcnamara, Adam B Nover, Biman B Mandal, Monica Agarwal, Terriann N Kelly, David L Kaplan, Gordana Vunjaknovakovic
    Abstract:

    Abstract Cartilage tissue lacks an intrinsic capacity for self-regeneration due to slow matrix turnover, a limited supply of mature chondrocytes and insufficient vasculature. Although cartilage tissue engineering has achieved some success using agarose as a scaffolding material, major challenges of agarose-based cartilage repair, including non-degradability, poor tissue–scaffold integration and limited processing capability, have prompted the search for an alternative biomaterial. In this study, silk fiber–hydrogel composites (SF–silk hydrogels) made from silk microfibers and silk hydrogels were investigated for their potential use as a support material for engineered cartilage. We demonstrated the use of 100% silk-based fiber–hydrogel composite scaffolds for the development of cartilage constructs with properties comparable to those made with agarose. Cartilage constructs with an Equilibrium Modulus in the native tissue range were fabricated by mimicking the collagen fiber and proteoglycan composite architecture of native cartilage using biocompatible, biodegradable silk fibroin from Bombyx mori . Excellent chondrocyte response was observed on SF–silk hydrogels, and fiber reinforcement resulted in the development of more mechanically robust constructs after 42 days in culture compared to silk hydrogels alone. Thus, we demonstrate the versatility of silk fibroin as a composite scaffolding material for use in cartilage tissue repair to create functional cartilage constructs that overcome the limitations of agarose biomaterials, and provide a much-needed alternative to the agarose standard.

  • engineering of functional cartilage tissue using stem cells from synovial lining a preliminary study
    Clinical Orthopaedics and Related Research, 2008
    Co-Authors: Ming Pei, V L Kish, Gordana Vunjaknovakovic
    Abstract:

    Stem cells derived from synovial lining—synovial lining-derived stem cells or SDSCs—are a promising cell source for cartilage tissue engineering. We hypothesized that negatively selected SDSCs would form cartilage constructs and conventionally passaged SDSCs would be contaminated with macrophages, inhibiting SDSC-based chondrogenesis. We mixed SDSCs with fibrin gel and seeded the cells into polyglycolic acid scaffolds. After 3 days of incubation with a proliferative growth factor cocktail (containing transforming growth factor β1 [TGF-β1], insulin-like growth factor I [IGF-I], and basic fibroblast growth factor [FGF-2]), the cell-fibrin-polyglycolic acid constructs were transferred into rotating bioreactor systems and cultured with a chondrogenic growth factor cocktail (TGF-β1/IGF-I) for up to 4 weeks. Tissue constructs based on negatively selected SDSCs had cartilaginous characteristics; were rich in glycosaminoglycans and collagen II; exhibited high expression of mRNA and protein for collagen II, aggrecan, and Sox 9; exhibited a negligible level of mRNA and protein for collagens I and X; and had an Equilibrium Modulus in the range of values measured for native human cartilage. Conventional passage yielded SDSCs with contaminating macrophages, which adversely affected the quality of tissue-engineered cartilage. We thus propose functional cartilage constructs could be engineered in vitro through the use of negatively isolated SDSCs.

  • bioreactor studies of native and tissue engineered cartilage
    Biorheology, 2002
    Co-Authors: Gordana Vunjaknovakovic, Ivan Martin, Bojana Obradovic, Lisa E Freed
    Abstract:

    Functional tissue engineering of cartilage involves the use of bioreactors designed to provide a controlled in vitro environment that embodies some of the biochemical and physical signals known to regulate chondrogenesis. Hydrodynamic conditions can affect in vitro tissue formation in at least two ways: by direct effects of hydrodynamic forces on cell morphology and function, and by indirect flow-induced changes in mass transfer of nutrients and metabolites. In the present work, we discuss the effects of three different in vitro environments: static flasks (tissues fixed in place, static medium), mixed flasks (tissues fixed in place, unidirectional turbulent flow) and rotating bioreactors (tissues dynamically suspended in laminar flow) on engineered cartilage constructs and native cartilage explants. As compared to static and mixed flasks, dynamic laminar flow in rotating bioreactors resulted in the most rapid tissue growth and the highest final fractions of glycosaminoglycans and total collagen in both tissues. Mechanical properties (Equilibrium Modulus, dynamic stiffness, hydraulic permeability) of engineered constructs and explanted cartilage correlated with the wet weight fractions of glycosaminoglycans and collagen. Current research needs in the area of cartilage tissue engineering include the utilization of additional physiologically relevant regulatory signals, and the development of predictive mathematical models that enable optimization of the conditions and duration of tissue culture.

  • bioreactor cultivation conditions modulate the composition and mechanical properties of tissue engineered cartilage
    Journal of Orthopaedic Research, 1999
    Co-Authors: Gordana Vunjaknovakovic, Alan J Grodzinsky, Ivan Martin, Bojana Obradovic, Steven Treppo, Robert Langer, Lisa E Freed
    Abstract:

    Summary: Cartilaginous constructs have been grown in vifm with use of isolated cells, biodegradable polymer scaffolds, and bioreactors. In the present work, the relationships between the composition and mechanical properties of engineered cartilage constructs were studied by culturing bovine calf articular chondrocytes on fibrous polyglycolic acid scaffolds (5 mm in diameter, 2-mm thick; and 97% porous) in three different environments: static flasks, mixed flasks. and rotating vessels. After 6 weeks of cultivation, the composition, morphology, and mechanical function of the constructs in radially confined static and dynamic compression all depended on the conditions of in vitro cultivation. Static culture yielded small and fragile constructs, while turbulent flow in mixed flasks yielded constructs with fibrous outer capsules: both environments resulted in constructs with poor mechanical properties. The constructs that were cultured freely suspended in a dynamic laminar flow field in rotating vessels were the largest, contained continuous cartilage-like extracellular matrices with the highest fractions of glycosarninoglycan and collagen, and had the best mechanical properties. The Equilibrium Modulus, hydraulic permeability, dynamic stiffness, and streaming potential correlated with the wet-weight fractions of glycosaminoglycan, collagen, and water. These findings suggest that the hydrodynamic conditions in tissue-culture bioreactors can modulate the composition. morphology, mechanical properties, and electromechanical function of engineered cartilage.

Zhen Tong - One of the best experts on this subject based on the ideXlab platform.

  • preferential adsorption of poly ethylene glycol on hectorite clay and effects on poly n isopropylacrylamide hectorite nanocomposite hydrogels
    Langmuir, 2010
    Co-Authors: Tao Wang, Xinxing Liu, Lijun Xiong, Chaoyang Wang, Zhen Tong
    Abstract:

    Poly(N-isopropylacrylamide)/Laponite nanocomposite hydrogel (NC gel) was synthesized via in situ polymerization in the Laponite suspension containing PEG. The adsorption of PEG on Laponite platelets was characterized by zeta-potential, which decreased with the PEG adsorption. The tensile strength decreased and elongation at break increased with increasing PEG concentration. The effective network chain density of PNIPAm/Laponite NC gels determined from the Equilibrium Modulus G(e) decreased upon adsorption of PEG on the Laponite. All of these results revealed the preferential adsorption of PEG on the Laponite platelets occupying the active sites for the PNIPAm chain anchoring, which hindered their cross-linking effect in the NC gels. However, the temperature sensitive swelling behavior still remained in the PNIPAm/Laponite NC gels containing PEG with higher swelling volume below the LCST due to the lower cross-linker density. By adjusting the amount of added PEG, we can easily control the properties of the PNIPAm/Laponite NC gels.

  • ultrahigh deformability and transparence of hectorite clay nanocomposite hydrogels with nimble ph response
    Macromolecules, 2009
    Co-Authors: Lijun Xiong, Xinxing Liu, Meina Zhu, Zhen Tong
    Abstract:

    Nimble pH response nanocomposite hydrogels (NC gel) with ultrahigh tensibility and high transparency were synthesized via in situ copolymerization of acrylamide and sodium acrylate (SA) in the aqueous suspension of hectorite clay Laponite RDS with a minute amount of N,N′-methylenebisacrylamide (BIS). The stability of the Laponite suspension containing ionic monomers and the tensile properties of the NC gels were investigated. The addition of ionic monomer SA was found to reduce the ζ potential and stability of the suspension. The tensile strength and elongation at break of these ionic NC gels obviously decreased when SA was greater than 10 mol % in the monomers. Interestingly, the addition of a minute amount of BIS (≤0.05 mol %) enhanced the homogeneity of the ionic NC gels and thus improved their transparency (transmittance >90%), tensile strength (>100 kPa), and elongation (>2000%). The relaxation Modulus of the ionic NC gels was fit with G(t) = Ge[1 + (t/λ0)−n], where Ge was the Equilibrium Modulus and...

  • concentration dependence of critical exponents for gelation in gellan gum aqueous solutions upon cooling
    European Polymer Journal, 2008
    Co-Authors: Lin Dai, Xinxing Liu, Yiliao Liu, Zhen Tong
    Abstract:

    Abstract The sol–gel transition in aqueous gellan gum solutions induced upon cooling was investigated by rheology measurements. The gelation temperature was determined from the crossover point of storage and loss moduli, i.e., G ′ =  G ′′ ( T c ) and from the Winter’s criterion ( T gel ), respectively, which increased with gellan concentration. T gel was higher than T c and the difference became larger as the gellan concentration got higher. The relaxation critical exponent n was estimated with the Winter’s method and the self-similarity was observed from the critical gel. The scaling for the zero-shear viscosity η 0 before the gel point and the Equilibrium Modulus G e after the gel point was established against the relative distance e from the gel point over the gellan concentration C g of 1.0–2.5 wt%, giving the critical exponents k and z . The critical exponent n calculated from k and z agrees well with n from the Winter’s criterion. However, no universal n was found for the gelation in aqueous gellan gum solutions, indicating that this gelation should be classified into the cross-linking category for the physical gelation. The critical exponent n decreased with increasing C g for the gellan gum solution. The fractal dimension d f calculated from n with the screened hydrodynamic interaction and the excluded volume effect suggested a denser structure in the critical gel with higher C g .

Gerard A. Ateshian - One of the best experts on this subject based on the ideXlab platform.

  • tissue engineered articular cartilage exhibits tension compression nonlinearity reminiscent of the native cartilage
    Journal of Biomechanics, 2013
    Co-Authors: Terriann N Kelly, Gerard A. Ateshian, Eric G Lima, Brendan L Roach, Zachary Weidner, Charles R Mackenziesmith, Grace D Oconnell, Aaron M Stoker, James L Cook, Clark T Hung
    Abstract:

    The tensile Modulus of articular cartilage is much larger than its compressive Modulus. This tension-compression nonlinearity enhances interstitial fluid pressurization and decreases the frictional coefficient. The current set of studies examines the tensile and compressive properties of cylindrical chondrocyte-seeded agarose constructs over different developmental stages through a novel method that combines osmotic loading, video microscopy, and uniaxial unconfined compression testing. This method was previously used to examine tension-compression nonlinearity in native cartilage. Engineered cartilage, cultured under free-swelling (FS) or dynamically loaded (DL) conditions, was tested in unconfined compression in hypertonic and hypotonic salt solutions. The apparent Equilibrium Modulus decreased with increasing salt concentration, indicating that increasing the bath solution osmolarity shielded the fixed charges within the tissue, shifting the measured moduli along the tension-compression curve and revealing the intrinsic properties of the tissue. With this method, we were able to measure the tensile (401±83 kPa for FS and 678±473 kPa for DL) and compressive (161±33 kPa for FS and 348±203 kPa for DL) moduli of the same engineered cartilage specimens. These moduli are comparable to values obtained from traditional methods, validating this technique for measuring the tensile and compressive properties of hydrogel-based constructs. This study shows that engineered cartilage exhibits tension-compression nonlinearity reminiscent of the native tissue, and that dynamic deformational loading can yield significantly higher tensile properties.

  • electrostatic and non electrostatic contributions of proteoglycans to the compressive Equilibrium Modulus of bovine articular cartilage
    Journal of Biomechanics, 2010
    Co-Authors: Clare Canal Guterl, Clark T Hung, Gerard A. Ateshian
    Abstract:

    Abstract This study presents direct experimental evidence for assessing the electrostatic and non-electrostatic contributions of proteoglycans to the compressive Equilibrium Modulus of bovine articular cartilage. Immature and mature bovine cartilage samples were tested in unconfined compression and their depth-dependent Equilibrium compressive Modulus was determined using strain measurements with digital image correlation analysis. The electrostatic contribution was assessed by testing samples in isotonic and hypertonic saline; the combined contribution was assessed by testing untreated and proteoglycan-depleted samples. Though it is well recognized that proteoglycans contribute significantly to the compressive stiffness of cartilage, results demonstrate that the combined electrostatic and non-electrostatic contributions may add up to more than 98% of the Modulus, a magnitude not previously appreciated. Of this contribution, about two thirds arises from electrostatic effects. The compressive Modulus of the proteoglycan-depleted cartilage matrix may be as low as 3 kPa, representing less than 2% of the normal tissue Modulus; experimental evidence also confirms that the collagen matrix in digested cartilage may buckle under compressive strains, resulting in crimping patterns. Thus, it is reasonable to model the collagen as a fibrillar matrix that can sustain only tension. This study also demonstrates that residual stresses in cartilage do not arise exclusively from proteoglycans, since cartilage remains curled relative to its in situ geometry even after proteoglycan depletion. These increased insights on the structure–function relationships of cartilage can lead to improved constitutive models and a better understanding of the response of cartilage to physiological loading conditions.

  • genipin enhances the mechanical properties of tissue engineered cartilage and protects against inflammatory degradation when used as a medium supplement
    Journal of Biomedical Materials Research Part A, 2009
    Co-Authors: Eric G Lima, Gerard A. Ateshian, Kacey G. Marra, Alicia J. Defail, Andrea R Tan, Timon Tai, Clark T Hung
    Abstract:

    Osteoarthritis (OA) is a common and painful disorder characterized by the degeneration of the articular surfaces of diarthrodial joints. One of the most promising approaches to treat OA is the design of engineered tissue that reproduces the functional properties of healthy cartilage. This tissue engineering approach typically entails the use of living cells embedded in a three-dimensional scaffold that is cultured over time in a growth medium supplemented with various physical or biological stimulants to encourage development. Recently our laboratory has been able to cultivate engineered cartilage with an Equilibrium Modulus (EY) and an glycosaminoglycan (GAG) content that match the native tissue using the temporal application of growth factors over a six-week culture period1. While these findings are encouraging, the dynamic Modulus (G*) and the collagen content for these constructs amount to less than a quarter of those of native articular cartilage. Of the two mechanical measurements, G*, which is correlated with the collagen content, is considered to be the more physiologically relevant as it measures the behavior of the tissue during the application of cyclic loads and better captures fluid pressurization within the biphasic tissue. No group has currently been able to reproduce native values of collagen regardless of the tissue engineering strategy employed. Obtaining native values of G* and collagen therefore remains an important but elusive challenge. Cross-linking agents such as glutaraldehyde, formaldehyde, or epoxy are typically used to chemically-treat (or fix) devitalized xenografts and allografts to reduce the immune rejection or the enzymatic degradation that is typical of transplanted bioprostheses2,3. Typically, however, cross-linking agents are lethal to cells. The use of genipin, a naturally occurring crosslinker with toxicity levels ten thousand fold less than glutaraldehyde4 as a biocompatible and stable cross-linker has been established by other groups5,6 and provides a promising new avenue to explore for tissue-engineering. Genipin cross-linking works by forming intra- and intermolecular cross-links of the amino residues on tropocollagen or proteoglycan molecules. A modified cyclic form of genipin can reside stably within the extracellular network, adding bridges across adjacent fibers. As such, previous groups have explored the use of genipin as a onetime treatment to fix tissue prior to implantation7, for the assembly of tissue-engineering scaffolds prior to cell seeding8,9, or in order to modulate the release of growth factors from degradable scaffolds or beads5. Genipin has also been studied for its anti-inflammatory effects, either administered directly to different cell lines10,11, or administered orally to animals12,13. Moreover, genipin cross-linking has been shown to affect the mechanical properties of biological tissues, increasing the tensile strength of bovine pericardium14, porcine tendon15, and type I collagen gels16. Our laboratory uses agarose hydrogel as a scaffold system for cartilage tissue engineering. This gel has been used extensively in chondrocyte biology studies and has shown some promise for tissue engineering applications. Agarose is a neutrally charged polysaccharide and as such is unaffected by genipin. However the chondrocytes embedded in the gel elaborate an extracellular matrix over time in culture that exhibits amine groups, which are subject to genipin cross-linking. In this set of studies we propose a novel use for genipin: not as a scaffold cross-linker, but as a medium supplement to promote cross-linking of de novo cell products as they are produced. We hypothesize that the application of genipin will stabilize the extracellular matrix components and increase the mechanical properties of developing cartilaginous tissue in our agarose hydrogels. We hypothesize two mechanisms through which the physical enhancement of tissue properties is fostered: (1) by reorganization and enhanced retention of cell synthesized extracellular matrix components, and (2) through reduction of the loss of extracellular matrix components by increasing their resilience to catabolic degradation.

  • the beneficial effect of delayed compressive loading on tissue engineered cartilage constructs cultured with tgf β3
    Osteoarthritis and Cartilage, 2007
    Co-Authors: Eric G Lima, Robert L. Mauck, Gerard A. Ateshian, Liming Bian, Benjamin A Byers, Rocky S Tuan, Clark T Hung
    Abstract:

    Summary Objective To determine whether the functional properties of tissue-engineered constructs cultured in a chemically-defined medium supplemented briefly with TGF-β3 can be enhanced with the application of dynamic deformational loading. Methods Primary immature bovine cells (2–3 months old) were encapsulated in agarose hydrogel (2%, 30×10 6 cells/ml) and cultured in chemically-defined medium supplemented for the first 2 weeks with transforming growth factor beta 3 (TGF-β3) (10μg/ml). Physiologic deformational loading (1Hz, 3h/day, 10% unconfined deformation initially and tapering to 2% peak-to-peak deformation by day 42) was applied either concurrent with or after the period of TGF-β3 supplementation. Mechanical and biochemical properties were evaluated up to day 56. Results Dynamic deformational loading applied concurrently with TGF-β3 supplementation yielded significantly lower (−90%) overall mechanical properties when compared to free-swelling controls. In contrast, the same loading protocol applied after the discontinuation of the growth factor resulted in significantly increased (+10%) overall mechanical properties relative to free-swelling controls. Equilibrium Modulus values reach 1306±79kPa and glycosaminoglycan levels reach 8.7±1.6% w.w. during this 8-week period and are similar to host cartilage properties (994±280kPa, 6.3±0.9% w.w.). Conclusions An optimal strategy for the functional tissue engineering of articular cartilage, particularly to accelerate construct development, may incorporate sequential application of different growth factors and applied deformational loading.

  • functional tissue engineering of articular cartilage through dynamic loading of chondrocyte seeded agarose gels
    Journal of Biomechanical Engineering-transactions of The Asme, 2000
    Co-Authors: Robert L. Mauck, Clark T Hung, Christopher C B Wang, Pen-hsiu Grace Chao, Michael A Soltz, Dennis D Wong, Wilmot B Valhmu, Gerard A. Ateshian
    Abstract:

    Due to its avascular nature, articular cartilage exhibits a very limited capacity to regenerate and to repair. Although much of the tissue-engineered cartilage in existence has been successful in mimicking the morphological and biochemical appearance of hyaline cartilage, it is generally mechanically inferior to the natural tissue. In this study, we tested the hypothesis that the application of dynamic deformational loading at physiological strain levels enhances chondrocyte matrix elaboration in cell-seeded agarose scaffolds to produce a more functional engineered tissue construct than in free swelling controls. A custom-designed bioreactor was used to load cell-seeded agarose disks dynamically in unconfined compression with a peak-to-peak compressive strain amplitude of 10 percent, at a frequency of 1 Hz. 3 × (1 hour on, 1 hour off)/day, 5 days/week for 4 weeks. Results demonstrated that dynamically loaded disks yielded a sixfold increase in the Equilibrium aggregate Modulus over free swelling controls after 28 days of loading (100±16 k Pa versus 15±8 kPa, p<0.0001). This represented a 21-fold increase over the Equilibrium Modulus of day 0 (4.8±2.3 k Pa). Sulfated glycosaminoglycan content and hydroxyproline content was also found to be greater in dynamically loaded disks compared to free swelling controls at day 21 (p<0.0001 and p=0.002, respectively).