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Robert L Mauck - One of the best experts on this subject based on the ideXlab platform.

  • pediatric laryngotracheal reconstruction with tissue Engineered Cartilage in a rabbit model
    Laryngoscope, 2016
    Co-Authors: Ian N Jacobs, Robert L Mauck, Robert A Redden, Rachel E Goldberg, Michael W Hast, Rebecca Salowe, Edward J Doolin
    Abstract:

    Objectives/Hypothesis To develop an effective rabbit model of in vitro- and in vivo-derived tissue-Engineered Cartilage for laryngotracheal reconstruction (LTR). Study Design 1) Determination of the optimal scaffold 1% hyaluronic acid (HA), 2% HA, and polyglycolic acid (PGA) and in vitro culture time course using a pilot study of 4 by 4-mm in vitro-derived constructs analyzed on a static culture versus zero-gravity bioreactor for 4, 8, and 12 weeks, with determination of compressive modulus and histology as outcome measures. 2) Three-stage survival rabbit experiment utilizing autologous auricular chondrocytes seeded in scaffolds, either 1% HA or PGA. The constructs were cultured for the determined in vitro time period and then cultured in vivo for 12 weeks. Fifteen LTRs were performed using HA Cartilage constructs, and one was performed with a PGA construct. All remaining specimens and the final reconstructed larynx underwent mechanical testing, histology, and glycosaminoglycan (GAG) content determination, and then were compared to cricoid control specimens (n = 13) and control LTR using autologous thyroid Cartilage (n = 18). Methods 1) One rabbit underwent an auricular punch biopsy, and its chondrocytes were isolated and expanded and then encapsulated in eight 4 by 4-mm discs of 1% HA, 2% HA, PGA either in rotary bioreactor or static culture for 4, 8, and 12 weeks, respectively, with determination of compressive modulus, GAG content, and histology. 2) Sixteen rabbits underwent ear punch biopsy; chondrocytes were isolated and expanded. The cells were seeded in 13 by 5 by 2.25-mm UV photopolymerized 1% HA (w/w) or calcium alginate encapsulated synthetic PGA (13 × 5 × 2 mm); the constructs were then incubated in vitro for 12 weeks (the optimal time period determined above in paragraph 1) on a shaker. One HA and one PGA construct from each animal was tested mechanically and histologically, and the remaining eight (4 HA and 4 PGA) were implanted in the neck. After 12 weeks in vivo, the most optimal-appearing HA construct was used as a graft for LTR in 15 rabbits and PGA in one rabbit. The seven remaining specimens underwent hematoxylin and eosin, Safranin O, GAG content determination, and flexural modulus testing. At 12 weeks postoperative, the animals were euthanized and underwent endoscopy. The larynges underwent mechanical and histological testing. All animals that died underwent postmortem examination, including gross and microhistological analysis of the reconstructed airway. Results Thirteen of the 15 rabbits that underwent LTR with HA in vitro- and in vivo-derived tissue-Engineered Cartilage constructs survived. The 1% HA specimens had the highest modulus and GAG after 12 weeks in vitro. The HA constructs became well integrated in the airway, supported respiration for the 12 weeks, and were histologically and mechanically similar to autologous Cartilage. Conclusions The engineering of in vitro- and in vivo-derived Cartilage with HA is a novel approach for laryngotracheal reconstruction. The data suggests that the in vitro- and in vivo-derived tissue-Engineered approaches may offer a promising alternative to current strategies used in pediatric airway reconstruction, as well as other head and neck applications. Level of Evidence NA. Laryngoscope, 2015

  • high mesenchymal stem cell seeding densities in hyaluronic acid hydrogels produce Engineered Cartilage with native tissue properties
    Acta Biomaterialia, 2012
    Co-Authors: Isaac E Erickson, Megan J Farrell, Jason A Burdick, Sydney R Kestle, Kilief H Zellars, Minwook Kim, Robert L Mauck
    Abstract:

    Abstract Engineered Cartilage based on adult mesenchymal stem cells (MSCs) is an alluring goal for the repair of articular defects. However, efforts to date have failed to generate constructs with sufficient mechanical properties to function in the demanding environment of the joint. Our findings with a novel photocrosslinked hyaluronic acid (HA) hydrogel suggest that stiff gels (high HA concentration, 5% w/v) foster chondrogenic differentiation and matrix production, but limit overall functional maturation due to the inability of the formed matrix to diffuse away from the point of production and form a contiguous network. In the current study, we hypothesized that increasing the MSC seeding density would decrease the required diffusional distance, and so expedite the development of functional properties. To test this hypothesis bovine MSCs were encapsulated at seeding densities of either 20,000,000 or 60,000,000 cells ml–1 in 1%, 3%, and 5% (w/v) HA hydrogels. Counter to our hypothesis the higher concentration HA gels (3% and 5%) did not develop more rapidly with increased MSC seeding density. However, the biomechanical properties of the low concentration (1%) HA constructs increased markedly (nearly 3-fold with a 3-fold increase in seeding density). To ensure that optimal nutrient access was delivered, we next cultured these constructs under dynamic culture conditions (with orbital shaking) for 9 weeks. Under these conditions 1% HA seeded at 60,000,000 MSCs ml–1 reached a compressive modulus in excess of 1 MPa (compared with 0.3–0.4 MPa for free swelling constructs). This is the highest level we have reported to date in this HA hydrogel system, and represents a significant advance towards functional stem cell-based tissue Engineered Cartilage.

  • coculture of human mesenchymal stem cells and articular chondrocytes reduces hypertrophy and enhances functional properties of Engineered Cartilage
    Tissue Engineering Part A, 2011
    Co-Authors: Liming Bian, Robert L Mauck, David Y Zhai, Jason A Burdick
    Abstract:

    Mesenchymal stem cells (MSCs) are being recognized as a viable cell source for Cartilage repair; however, it still remains a challenge to recapitulate the functional properties of native articular Cartilage using only MSCs. Additionally, MSCs may exhibit a hypertrophic phenotype under chondrogenic induction, resulting in calcification after ectopic transplantation. With this in mind, the objective of this study was to assess whether the addition of chondrocytes to MSC cultures influences the properties of tissue-Engineered Cartilage and MSC hypertrophy when cultured in hyaluronic acid hydrogels. Mixed cell populations (human MSCs and human chondrocytes at a ratio of 4:1) were encapsulated in the hydrogels and exhibited significantly higher Young's moduli, dynamic moduli, glycosaminoglycan levels, and collagen content than did constructs seeded with only MSCs or chondrocytes. Furthermore, the deposition of collagen X, a marker of MSC hypertrophy, was significantly lower in the coculture constructs than in the constructs seeded with MSCs alone. When MSCs and chondrocytes were cultured in distinct gels, but in the same wells, there was no improvement in biomechanical and biochemical properties of the Engineered tissue, implying that a close proximity is essential. This approach can be used to improve the properties and prevent calcification of Engineered Cartilage formed from MSC-seeded hydrogels with the addition of lower fractions of chondrocytes, leading to improved clinical outcomes.

  • mechanics and mechanobiology of mesenchymal stem cell based Engineered Cartilage
    Journal of Biomechanics, 2010
    Co-Authors: Alice H Huang, Megan J Farrell, Robert L Mauck
    Abstract:

    In this review, we outline seminal and recent work highlighting the potential of mesenchymal stem cells (MSCs) in producing Cartilage-like tissue equivalents. Specific focus is placed on the mechanical properties of Engineered MSC-based Cartilage and how these properties relate to that of Engineered Cartilage based on primary chondrocytes and to native tissue properties. We discuss current limitations and/or concerns that must be addressed for the clinical realization of MSC-based Cartilage therapeutics, and provide some insight into potential underpinnings for the observed deviations from chondrocyte-based Engineered constructs. We posit that these differences reveal specific deficits in terms of our description of chondrogenesis, and suggest that new benchmarks must be developed towards this end. Further, we describe the growing body of literature on the mechanobiology of MSC-based Cartilage, highlighting positive findings with regards to the furtherance of the chondrogenic phenotype. We likewise discuss the failure of early molecular changes to translate directly into Engineered constructs with improved mechanical properties. Finally, we highlight recent work from our group and others that may point to new strategies for enhancing the formation of Engineered Cartilage based on MSCs.

  • transient exposure to transforming growth factor beta 3 under serum free conditions enhances the biomechanical and biochemical maturation of tissue Engineered Cartilage
    Tissue Engineering Part A, 2008
    Co-Authors: Benjamin A Byers, Robert L Mauck, Ian E Chiang, Rocky S Tuan
    Abstract:

    A goal of Cartilage tissue engineering is the production of cell-laden constructs possessing sufficient mechanical and biochemical features to enable native tissue function. This study details a systematic characterization of a serum-free (SF) culture methodology employing transient growth factor supplementation to promote robust maturation of tissue-Engineered Cartilage. Bovine chondrocyte agarose hydrogel constructs were cultured under free-swelling conditions in serum-containing or SF medium supplemented continuously or transiently with varying doses of transforming growth factor beta 3 (TGF-beta3). Constructs were harvested weekly or bi-weekly and assessed for mechanical and biochemical properties. Transient exposure (2 weeks) to low concentrations (2.5-5 ng/mL) of TGF-beta3 in chemically defined medium facilitated robust and highly reproducible construct maturation. Constructs receiving transient TGF-beta3 exposure achieved native tissue levels of compressive modulus (0.8 MPa) and proteoglycan content (6-7% of wet weight) after less than 2 months of in vitro culture. This maturation response was far superior to that observed after continuous growth factor supplementation or transient TGF-beta3 treatment in the presence of serum. These findings represent a significant advance in developing an ex vivo culture methodology to promote production of clinically relevant and mechanically competent tissue-Engineered Cartilage constructs for implantation to repair damaged articular surfaces.

Lawrence J Bonassar - One of the best experts on this subject based on the ideXlab platform.

  • heterogeneous matrix deposition in human tissue Engineered Cartilage changes the local shear modulus and resistance to local construct buckling
    Journal of Biomechanics, 2020
    Co-Authors: Jill M Middendorf, Itai Cohen, Caroline Dugopolski, Stephen Kennedy, Eric Blahut, Lawrence J Bonassar
    Abstract:

    Abstract Human tissue Engineered Cartilage is a promising solution for focal Cartilage defects, but these constructs do not have the same local mechanical properties as native tissue. Most clinically relevant Engineered Cartilage constructs seed human chondrocytes onto a collagen scaffold, which buckles at low loads and strains. This buckling creates local regions of high strain that could cause cell death and damage the Engineered tissue. Since human tissue Engineered Cartilage is commonly grown in-vivo prior to implantation, new matrix deposition could improve the local implant mechanics and prevent local tissue buckling. However, the relationship between local biochemical composition and the local mechanics or local buckling probability has never been quantified. Therefore, this study correlated the local biochemical composition of human tissue Engineered Cartilage constructs using Fourier transform infrared spectroscopy (FTIR) with the local shear modulus and local buckling probability. The local shear modulus and local buckling probability were obtained using a confocal elastography technique. The local shear modulus increased with increases in local aggrecan content in the interior region (inside the scaffold). A minimum amount of aggrecan was required to prevent local construct buckling at physiologic strains. Since the original scaffold was primarily composed of collagen, increases in collagen content due to new matrix deposition was minimal and had little effect on the mechanical properties. Thus, we concluded that aggrecan deposition inside the scaffold pores is the most effective way to improve the mechanical function and prevent local tissue damage in human tissue Engineered Cartilage constructs.

  • multiscale mechanics of tissue Engineered Cartilage grown from human chondrocytes and human induced pluripotent stem cells
    Journal of Orthopaedic Research, 2020
    Co-Authors: Jill M Middendorf, Itai Cohen, Caroline Dugopolski, Stephen Kennedy, Nicole Diamantides, Sonya Shortkroff, Lawrence J Bonassar
    Abstract:

    Tissue-Engineered Cartilage has shown promising results in the repair of focal Cartilage defects. However, current clinical techniques rely on an extra surgical procedure to biopsy healthy Cartilage to obtain human chondrocytes. Alternatively, induced pluripotent stem cells (iPSCs) have the ability to differentiate into chondrocytes and produce cartilaginous matrix without the need to biopsy healthy Cartilage. However, the mechanical properties of tissue-Engineered Cartilage with iPSCs are unknown and might be critical to long-term tissue function and health. This study used confined compression, Cartilage on glass tribology, and shear testing on a confocal microscope to assess the macroscale and microscale mechanical properties of two constructs seeded with either chondrocyte-derived iPSCs (Ch-iPSCs) or native human chondrocytes. Macroscale properties of Ch-iPSC constructs provided similar or better mechanical properties than chondrocyte constructs. Under compression, Ch-iPSC constructs had an aggregate modulus that was two times larger than chondrocyte constructs and was closer to native tissue. No differences in the shear modulus and friction coefficients were observed between Ch-iPSC and chondrocyte constructs. On the microscale, Ch-iPSC and chondrocyte constructs had different depth-dependent mechanical properties, neither of which matches native tissue. These observed depth-dependent differences may be important to the function of the implant. Overall, this comparison of multiple mechanical properties of Ch-iPSC and chondrocyte constructs shows that using Ch-iPSCs can produce equivalent or better global mechanical properties to chondrocytes. Therefore, iPSC-seeded Cartilage constructs could be a promising solution to repair focal Cartilage defects. The chondrocyte constructs used in this study have been implanted into humans for clinical trials. Therefore, Ch-iPSC constructs could also be used clinically in place of the current chondrocyte construct.

  • adhesion and integration of tissue Engineered Cartilage to porous polyethylene for composite ear reconstruction
    Journal of Biomedical Materials Research Part B, 2015
    Co-Authors: N A Osullivan, Lawrence J Bonassar, Michael J Yaremchuk, Shinji Kobayashi, Mitun P Ranka, Katherine L Zaleski, Mark A Randolph
    Abstract:

    The objective of this study was to assess the ability of tissue Engineered Cartilage to adhere to and integrate with porous polyethylene (PPE) in vivo and to evaluate the biomechanical integrity of the bond formed at the interface. Porcine auricular, articular, and costal chondrocytes were suspended in fibrin gel polymer and placed between discs of PPE to form tri-layer constructs. Controls consisted of fibroblasts suspended in gel or gel alone between the discs. Constructs were implanted into nude mice for 6, 12, and 18 weeks. Upon harvest, specimens were evaluated for neoCartilage formation and integration into the PPE, using histological, dimensional (mass, thickness, diameter), and biomechanical (adhesion strength, interfacial stiffness, failure energy and failure strain) analyses. Neotissue was formed in all experimental constructs, consisting mostly of neoCartilage integrating with discs of PPE. Control samples contained only fibrous tissue. Biomechanical analyses demonstrated that adhesion strength, interfacial stiffness, and failure energy were all significantly higher in the chondrocyte-seeded samples than in fibroblast-seeded controls, with the exception of costal constructs at 12 weeks, which were not significantly greater than controls. In general, failure strains did not vary between groups. In conclusion, porous polyethylene supported the growth of neoCartilage that formed mechanically functional bonds with the PPE.

  • Insights into interstitial flow, shear stress, and mass transport effects on ECM heterogeneity in bioreactor-cultivated Engineered Cartilage hydrogels
    Biomechanics and modeling in mechanobiology, 2011
    Co-Authors: Tony Chen, Lawrence J Bonassar, Mark R. Buckley, Itai Cohen, Hani A Awad
    Abstract:

    Interstitial flow in articular Cartilage is secondary to compressive and shear deformations during joint motion and has been linked with the well-characterized heterogeneity in structure and composition of its extracellular matrix. In this study, we investigated the effects of introducing gradients of interstitial flow on the evolution of compositional heterogeneity in Engineered Cartilage. Using a parallel-plate bioreactor, we observed that Poiseuille flow stimulation of chondrocyte-seeded agarose hydrogels led to an increase in glycosaminoglycan and type II collagen deposition in the surface region of the hydrogel exposed to flow. Experimental measurements of the interstitial flow fields based on the fluorescence recovery after photobleaching technique suggested that the observed heterogeneity in composition is associated with gradients in interstitial flow in a boundary layer at the hydrogel surface. Interestingly, the interstitial flow velocity profiles were nonlinearly influenced by flow rate, which upon closer examination led us to the original observation that the apparent hydrogel permeability decreased exponentially with increased interfacial shear stress. We also observed that interstitial flow enhances convective mass transport irrespective of molecular size within the boundary layer near the hydrogel surface and that the convective contribution to transport diminishes with depth in association with interstitial flow gradients. The implications of the nonlinearly inverse relationship between the interfacial shear stress and the interstitial flux and permeability and its consequences for convective transport are important for tissue engineering, since porous scaffolds comprise networks of Poiseuille channels (pores) through which interstitial flow must navigate under mechanical stimulation or direct perfusion.

  • role for interleukin 1α in the inhibition of chondrogenesis in autologous implants using polyglycolic acid polylactic acid scaffolds
    Tissue Engineering, 2005
    Co-Authors: Nicole Rotter, Martin P Vacanti, Roland D Eavey, Charles A Vacanti, Lawrence J Bonassar
    Abstract:

    Significant challenges remain in generating tissue-Engineered Cartilage in immunocompetent animals. Scaffold materials such as polyglycolic acid lead to significant inflammatory reactions, inhibiti...

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

  • heterogeneous Engineered Cartilage growth results from gradients of media supplemented active tgf β and is ameliorated by the alternative supplementation of latent tgf β
    Biomaterials, 2016
    Co-Authors: Michael B Albro, Gordana Vunjaknovakovic, Clark T Hung, Alexander D Cigan, Robert J Nims, Krista M Durney, Jae K Shim, Gerard A Ateshian
    Abstract:

    Transforming growth factor beta (TGF-β) has become one of the most widely utilized mediators of Engineered Cartilage growth. It is typically exogenously supplemented in the culture medium in its active form, with the expectation that it will readily transport into tissue constructs through passive diffusion and influence cellular biosynthesis uniformly. The results of this investigation advance three novel concepts regarding the role of TGF-β in Cartilage tissue engineering that have important implications for tissue development. First, through the experimental and computational analysis of TGF-β concentration distributions, we demonstrate that, contrary to conventional expectations, media-supplemented exogenous active TGF-β exhibits a pronounced concentration gradient in tissue constructs, resulting from a combination of high-affinity binding interactions and a high cellular internalization rate. These gradients are sustained throughout the entire culture duration, leading to highly heterogeneous tissue growth; biochemical and histological measurements support that while biochemical content is enhanced up to 4-fold at the construct periphery, enhancements are entirely absent beyond 1 mm from the construct surface. Second, construct-encapsulated chondrocytes continuously secrete large amounts of endogenous TGF-β in its latent form, a portion of which undergoes cell-mediated activation and enhances biosynthesis uniformly throughout the tissue. Finally, motivated by these prior insights, we demonstrate that the alternative supplementation of additional exogenous latent TGF-β enhances biosynthesis uniformly throughout tissue constructs, leading to enhanced but homogeneous tissue growth. This novel demonstration suggests that latent TGF-β supplementation may be utilized as an important tool for the translational engineering of large Cartilage constructs that will be required to repair the large osteoarthritic defects observed clinically.

  • differences in Engineered Cartilage from human chondrocytes and mesenchymal stem cells in pellet and construct culture
    Volume 1A: Abdominal Aortic Aneurysms; Active and Reactive Soft Matter; Atherosclerosis; BioFluid Mechanics; Education; Biotransport Phenomena; Bone J, 2013
    Co-Authors: Grace D Oconnell, G Palmer, Clark T Hung
    Abstract:

    Articular Cartilage serves as the load-bearing material of joints. One approach to functional tissue engineering is to recapitulate the biochemical and mechanical function of healthy native Cartilage in vitro, prior to implantation. We have been successful in cultivating Engineered Cartilage with compressive mechanical properties and glycosaminoglycan (GAG) content near native values by encapsulating chondrocytes or stem cells in a clinically relevant hydrogel [1, 2]. Clinical application of functional Engineered Cartilage will likely use of chondrocytes (AC) from osteoarthritic tissue or mesenchymal stem cells (MSCs), which have been shown to have chondrogenic potential. That is, it is may be more feasible to differentiate healthy MSCs towards a chondrogenic lineage than to ‘reprogram’ ACs acquired from an osteoarthritic joint.Copyright © 2013 by ASME

  • insulin ascorbate and glucose have a much greater influence than transferrin and selenous acid on the in vitro growth of Engineered Cartilage in chondrogenic media
    Tissue Engineering Part A, 2013
    Co-Authors: Alexander D Cigan, Gordana Vunjaknovakovic, Clark T Hung, Robert J Nims, Michael B Albro, John D Esau, Marissa P Dreyer, Gerard A Ateshian
    Abstract:

    The primary goal of this study was to characterize the response of chondrocyte-seeded agarose constructs to varying concentrations of several key nutrients in a chondrogenic medium, within the overall context of optimizing the key nutrients and the placement of nutrient channels for successful growth of Cartilage tissue constructs large enough to be clinically relevant in the treatment of osteoarthritis (OA). To this end, chondrocyte-agarose constructs (o4×2.34 mm, 30×10(6) cells/mL) were subjected to varying supplementation levels of insulin (0× to 30× relative to standard supplementation), transferrin (0× to 30×), selenous acid (0× to 10×), ascorbate (0× to 30×), and glucose (0× to 3×). The quality of resulting Engineered tissue constructs was evaluated by their compressive modulus (E(-Y)), tensile modulus (E(+Y)), hydraulic permeability (k), and content of sulfated glycosaminoglycans (sGAG) and collagen (COL); DNA content was also quantified. Three control groups from two separate castings of constructs (1× concentrations of all medium constituents) were used. After 42 days of culture, values in each of these controls were, respectively, E(-Y)=518±78, 401±113, 236±67 kPa; E(+Y)=1420±430, 1140±490, 1240±280 kPa; k=2.3±0.8×10(-3), 5.4±7.0×10(-3), 3.3±1.3×10(-3) mm(4)/N·s; sGAG=7.8±0.3, 6.3±0.4, 4.1±0.5%/ww; COL=1.3±0.2, 1.1±0.3, 1.4±0.4%/ww; and DNA=11.5±2.2, 12.1±0.6, 5.2±2.8 μg/disk. The presence of insulin and ascorbate was essential, but their concentrations may drop as low as 0.3× without detrimental effects on any of the measured properties; excessive supplementation of ascorbate (up to 30×) was detrimental to E(-Y), and 30× insulin was detrimental to both E(+Y) and E(-Y). The presence of glucose was similarly essential, and matrix elaboration was significantly dependent on its concentration (p<10(-6)), with loss of functional properties, composition, and cellularity observed at ≤0.3×; excessive glucose supplementation (up to 3×) showed no detrimental effects. In contrast, transferrin and selenous acid had no influence on matrix elaboration. These findings suggest that adequate distributions of insulin, ascorbate, and glucose, but not necessarily of transferrin and selenous acid, must be ensured within large Engineered Cartilage constructs to produce a viable substitute for joint tissue lost due to OA.

  • insulin and ascorbate have a much greater influence than transferrin and selenous acid on the growth of Engineered Cartilage in chondrogenic media
    ASME 2012 Summer Bioengineering Conference Parts A and B, 2012
    Co-Authors: Alexander D Cigan, Clark T Hung, Robert J Nims, Michael B Albro, Sarah L Breves, Gerard A Ateshian
    Abstract:

    Tissue engineering of Cartilage, which is a much sought-after approach for treatment of osteoarthritis and Cartilage defects, requires appreciable culture time. Chemically defined chondrogenic media (CM) are commonly employed as they offer a promising alternative to serum-based media, and often include insulin, transferrin, and selenous acid (ITS) as well as ascorbate [1]. Concentrations of ITS constituents have been optimized based upon their ability to stimulate proliferation of a variety of cell types [2]. However, little is reflected in the literature as to the influences of various ITS constituent concentrations upon Cartilage matrix deposition by chondrocytes. In Engineered Cartilage constructs that seek to match compositional and mechanical properties of native Cartilage, knowledge of such influences would be highly desirable, especially when optimizing nutrient, hormone and vitamin supply for large constructs wherein rates of transport and consumption become more critical. Furthermore, this information would prove useful in modeling growth and remodeling of Engineered tissues. Therefore, this study seeks to elucidate mechanical and biochemical properties as a direct result of modulating ITS and ascorbate concentrations within chondrocyte-agarose constructs.Copyright © 2012 by ASME

  • response of Engineered Cartilage to mechanical insult depends on construct maturity
    Osteoarthritis and Cartilage, 2010
    Co-Authors: Elizabeth Y Dong, Gerard A Ateshian, Clark T Hung
    Abstract:

    Summary Injury to articular Cartilage leads to degenerative changes resulting in a loss of mechanical and biochemical properties. In Engineered Cartilage, the injury response of developing constructs is unclear. Objective To characterize the cellular response of tissue-Engineered constructs cultured in chemically-defined medium after mechanical insult, either by compression-induced cracking, or by cutting, as a function of construct maturity. Methods Primary immature bovine articular chondrocytes (4–6 weeks) were encapsulated in agarose hydrogel (2%, 30 millioncells/mL) and cultured in chemically-defined medium supplemented with Transforming growth factor (TGF)-β3 (10ng/mL, first 2 weeks). At early (5 days) and late (35 days) times in culture, subsets of constructs were exposed to mechanical overload to produce a crack in the tissue or were exposed to a sharp wound with a perpendicular cut. Constructs were returned to culture and allowed to recover in static conditions. Mechanical and biochemical properties were evaluated at 2-week intervals to day 70, and cellular viability was assessed at 2-week intervals to day 85. Results Constructs injured early in culture recovered their mechanical stiffness back to control values, regardless of the mode of injury. Later in culture, when constructs exhibited properties similar to those of native Cartilage, compression-induced cracking catastrophically damaged the bulk matrix of the tissue and resulted in permanent mechanical failure with persistent cell death. No such detrimental outcomes were observed with cutting. Biochemical content was similar across all groups irrespective of mode or time of injury. Conclusions Unlike native Cartilage, Engineered Cartilage constructs exhibit a reparative capacity when the bulk integrity of the developing tissue is preserved after injury.

Ran Xiao - One of the best experts on this subject based on the ideXlab platform.

  • bone marrow mesenchymal stem cell based Engineered Cartilage ameliorates polyglycolic acid polylactic acid scaffold induced inflammation through m2 polarization of macrophages in a pig model
    Stem Cells Translational Medicine, 2016
    Co-Authors: Jinping Ding, Ning Kang, Qian Wang, Bo Chen, Tao Lv, Xin Fu, Ran Xiao
    Abstract:

    UNLABELLED: : The regeneration of tissue-Engineered Cartilage in an immunocompetent environment usually fails due to severe inflammation induced by the scaffold and their degradation products. In the present study, we compared the tissue remodeling and the inflammatory responses of Engineered Cartilage constructed with bone marrow mesenchymal stem cells (BMSCs), chondrocytes, or both and scaffold group in pigs. The Cartilage-forming capacity of the constructs in vitro and in vivo was evaluated by histological, biochemical, and biomechanical analyses, and the inflammatory response was investigated by quantitative analysis of foreign body giant cells and macrophages. Our data revealed that BMSC-based Engineered Cartilage suppressed in vivo inflammation through the alteration of macrophage phenotype, resulting in better tissue survival compared with those regenerated with chondrocytes alone or in combination with BMSCs. To further confirm the macrophage phenotype, an in vitro coculture system established by Engineered Cartilage and macrophages was studied using immunofluorescence, enzyme-linked immunosorbent assay, and gene expression analysis. The results demonstrated that BMSC-based Engineered Cartilage promoted M2 polarization of macrophages with anti-inflammatory phenotypes including the upregulation of CD206, increased IL-10 synthesis, decreased IL-1β secretion, and alterations in gene expression indicative of M1 to M2 transition. It was suggested that BMSC-seeded constructs have the potential to ameliorate scaffold-induced inflammation and improve cartilaginous tissue regeneration through M2 polarization of macrophages. SIGNIFICANCE: Finding a strategy that can prevent scaffold-induced inflammation is of utmost importance for the regeneration of tissue-Engineered Cartilage in an immunocompetent environment. This study demonstrated that bone marrow mesenchymal stem cell (BMSC)-based Engineered Cartilage could suppress inflammation by increasing M2 polarization of macrophages, resulting in better tissue survival in a pig model. Additionally, the effect of BMSC-based Cartilage on the phenotype conversion of macrophages was further studied through an in vitro coculture system. This study could provide further support for the regeneration of Cartilage engineering in immunocompetent animal models and provide new insight into the interaction of tissue-Engineered Cartilage and macrophages.

  • effects of co culturing bmscs and auricular chondrocytes on the elastic modulus and hypertrophy of tissue Engineered Cartilage
    Biomaterials, 2012
    Co-Authors: Ning Kang, Yilin Cao, Xia Liu, Yue Guan, Jian Wang, Fuxing Gong, Xun Yang, Li Yan, Qian Wang, Ran Xiao
    Abstract:

    Co-culture of BMSCs and chondrocytes is considered as a promising strategy to generate tissue Engineered Cartilage as chondrocytes induce the chondrogenesis of BMSCs and inhibit the hypertrophy of Engineered Cartilage. Because the tissue specific stem/progenitor cells have been isolated from mature tissues including auricular Cartilage, we hypothesized that adding stem cells to auricular chondrocytes in co-culture would also enhance the quality of Engineered Cartilage. In the present study, using the histological assay, biomechanical evaluation, and quantitative analysis of gene expression, we compared different strategies of auricular chondrocytes, BMSCs induction, and co-culture at different ratios on PGA/PLA scaffolds to construct tissue Engineered elastic Cartilage in vitro and in vivo. The up-regulation of RUNX2 and down-regulation of SOX9 were found in BMSCs chondrogenic induction group, which might imply a regulatory mechanism for the hypertrophy and potential osteogenic differentiation. Engineered Cartilage in co-culture 5:5 group showed the densest elastic fibers and the highest Young's modulus, which were consistent with the expression profile of Cartilage matrix-related genes including DCN and LOXL2 genes. Moreover, the better proliferative and chondrogenic potentials of Engineered Cartilage in co-culture 5:5 group were demonstrated by the stronger expression of Ki67 and Dlk1.

Charles A Vacanti - One of the best experts on this subject based on the ideXlab platform.

  • role for interleukin 1α in the inhibition of chondrogenesis in autologous implants using polyglycolic acid polylactic acid scaffolds
    Tissue Engineering, 2005
    Co-Authors: Nicole Rotter, Martin P Vacanti, Roland D Eavey, Charles A Vacanti, Lawrence J Bonassar
    Abstract:

    Significant challenges remain in generating tissue-Engineered Cartilage in immunocompetent animals. Scaffold materials such as polyglycolic acid lead to significant inflammatory reactions, inhibiti...

  • tissue Engineered Cartilage as a graft source for laryngotracheal reconstruction a pig model
    Archives of Otolaryngology-head & Neck Surgery, 2004
    Co-Authors: Syed H Kamil, Martin P Vacanti, Roland D Eavey, Charles A Vacanti, Christopher J Hartnick
    Abstract:

    Objective To evaluate the feasibility of using tissue-Engineered Cartilage for laryngotracheal reconstruction in the pig model. Design Auricular Cartilage was harvested from 3 young swine. The Cartilage was digested, processed, and suspended and a cell culture was obtained. The cells were then suspended in 3 mL of a 30% solution of a biodegradable polymer (Pluronic F-127) (polyethylene oxide/polypropylene oxide copolymer) at a cellular concentration of 50 × 106cells/mL. This suspension was then implanted subcutaneously into each pig's dorsum. Eight weeks after implantation, the Cartilage was harvested with the surrounding perichondrial capsule. An anterior Cartilage graft laryngotracheal reconstruction was performed. Bronchoscopy was performed at 3 postoperative weeks to demonstrate airway patency. The animals were killed at 3 months, and specimens were obtained for histological analysis. Setting An animal research facility. Subjects Three young Yorkshire swine. Results All 3 pigs survived to the 3-month postoperative interval with no evidence of stridor or airway distress. Interval bronchoscopy revealed a normal patent airway with a mucosalized graft. Histopathologic analysis revealed incorporation of the tissue-Engineered Cartilage graft in the cricoid area, which correlated with results of bronchoscopic evaluation. Conclusion Tissue-Engineered auricular Cartilage served as a viable graft in the pig model and might be an alternative Cartilage source for laryngotracheal reconstruction.

  • transplantation of chondrocytes utilizing a polymer cell construct to produce tissue Engineered Cartilage in the shape of a human ear
    Plastic and Reconstructive Surgery, 1997
    Co-Authors: Joseph P Vacanti, Keith T Paige, Joseph Upton, Charles A Vacanti
    Abstract:

    This study evaluates the feasibility of growing tissue-Engineered Cartilage in the shape of a human ear using chondrocytes seeded onto a synthetic biodegradable polymer fashioned in the shape of a 3-year-old child's auricle. A polymer template was formed in the shape of a human auricle using a nonwo

  • transplantation of chondrocytes utilizing a polymer cell construct to produce tissue Engineered Cartilage in the shape of a human ear
    Plastic and Reconstructive Surgery, 1997
    Co-Authors: Yilin Cao, Keith T Paige, Joseph Upton, Joseph P Vacanti, Charles A Vacanti
    Abstract:

    This study evaluates the feasibility of growing tissue-Engineered Cartilage in the shape of a human ear using chondrocytes seeded onto a synthetic biodegradable polymer fashioned in the shape of a 3-year-old child's auricle. A polymer template was formed in the shape of a human auricle using a nonwoven mesh of polyglycolic acid molded after being immersed in a 1% solution of polylactic acid. Each polyglycolic acid-polylactic acid template was seeded with chondrocytes isolated from bovine articular Cartilage and then implanted into subcutaneous pockets on the dorsa of 10 athymic mice. The three-dimensional structure was well maintained after removal of an external stent that had been applied for 4 weeks. Specimens harvested 12 weeks after implantation and subjected to gross morphologic and histologic analysis demonstrated new Cartilage formation. The overall geometry of the experimental specimens closely resembled the complex structure of the child's auricle. These findings demonstrate that polyglycolic acid-polylactic acid constructs can be fabricated in a very intricate configuration and seeded with chondrocytes to generate new Cartilage that would be useful in plastic and reconstructive surgery.

  • experimental tracheal replacement using tissue Engineered Cartilage
    Journal of Pediatric Surgery, 1994
    Co-Authors: Charles A Vacanti, Keith T Paige, Joseph Upton, Woo Seob Kim, Junichi Sakata, Joseph P Vacanti
    Abstract:

    The authors tested the feasibility of using tissue-Engineered Cartilage, grown in the shape of cylinders, for replacing large circumferential defects of the cervical trachea in rats. Chondrocytes obtained from the shoulder of newborn calves were seeded onto a synthetic nonwoven mesh, 100 microns thick, of polyglycolic acid fibers 15 microns in diameter, cut into pieces of 2.5 x 4 cm. Twenty cell-polymer constructs were wrapped around silastic tubes and implanted into 10 nude mice for 4 weeks. Specimens were then excised and evaluated grossly and histologically for the presence of new Cartilage, and biomechanically for their ability to resist collapse upon application of negative pressure. Six cylinders of tissue-Engineered Cartilage were then sutured into large circumferential defects created in the cervical tracheas of nude rats to replace the excised trachea. Implantation of cell-polymer constructs resulted in the formation of cylinders of hyaline Cartilage. When placed within the lumen of a segment of bowel denuded of its mucosal lining, the hollow cylinders resisted collapse in all instances upon administration of negative 200 mm Hg pressure. The Cartilage was grossly and histologically identical to that from which the cells had been initially isolated. Four of the six animals receiving these Cartilage cylinders as tracheal replacements survived the procedure and were able to breathe in an unassisted fashion. Three of these animals never recovered fully from the anesthetic and the operation, and expired at 24, 48, and 72 hours. The fourth animal fully recovered from the procedure, and breathed spontaneously for 1 week, with no apparent limitations. Increasing respiratory distress then developed, and the animal died.(ABSTRACT TRUNCATED AT 250 WORDS)