Sample records for bovine cartilage explants

  1. Effects of electromagnetic fields on proteoglycan metabolism of bovine articular cartilage explants

    NARCIS (Netherlands)

    De Mattei, M; Pasello, M; Pellati, A; Stabellini, G; Massari, L; Gemmati, D; Caruso, A


    Electromagnetic field (EMF) exposure has been proposed for the treatment of osteoarthritis. In this study, we investigated the effects of EMF (75 Hz, 2,3 mT) on proteoglycan (PG) metabolism of bovine articular cartilage explants cultured in vitro, both under basal conditions and in the presence of

  2. The effect of protease inhibitors on the induction of osteoarthritis-related biomarkers in bovine full-depth cartilage explants

    DEFF Research Database (Denmark)

    He, Yi; Zheng, Qinlong; Jiang, Mengmeng


    Objective The specific degradation of type II collagen and aggrecan by matrix metalloproteinase (MMP)-9, -13 and ADAMTS-4 and -5 (aggrecanase-1 and -2) in the cartilage matrix is a critical step in pathology of osteoarthritis (OA). The aims of this study were: i) To investigate the relative...... contribution of ADAMTS-4 and ADAMTS-5 to cartilage degradation upon catabolic stimulation; ii) To investigate the effect of regulating the activities of key enzymes by mean of broad-spectrum inhibitors. Methods Bovine full-depth cartilage explants stimulated with tumor necrosis factor alpha (TNF...... protease for the generation of 374ARGS aggrecan fragment in the TNF-α/OSM stimulated bovine cartilage explants. This study addresses the need to determine the roles of ADAMTS-4 and ADAMTS-5 in human articular degradation in OA and hence identify the attractive target for slowing down human cartilage...

  3. Suppression of MMP activity in bovine cartilage explants cultures has little if any effect on the release of aggrecanase-derived aggrecan fragments

    DEFF Research Database (Denmark)

    Wang, Bijue; Chen, Pingping; Jensen, Anne-Christine Bay


    -epitope specific immunoassays; (1) sandwich (342)FFGVG-G2 ELISA, (2) competition NITEGE(373)ELISA (3) sandwich G1-NITEGE(373 )ELISA (4) competition (374)ARGSV ELISA, and (5) sandwich (374)ARGSV-G2 ELISA all detecting aggrecan fragments, and (6) sandwich CTX-II ELISA, detecting C-telopeptides of type II collagen......- and aggrecanase-derived fragments of aggrecan and type II collagen into the supernatant of bovine cartilage explants cultures using neo-epitope specific immunoassays, and to associate the release of these fragments with the activity of proteolytic enzymes using inhibitors. FINDINGS: Bovine cartilage explants were....... We found that (1) aggrecanase-derived aggrecan fragments are released in the early (day 2-7) and mid phase (day 9-14) into the supernatant from bovine explants cultures stimulated with catabolic cytokines, (2) the release of NITEGE(373 )neo-epitopes are delayed compared to the corresponding (374...

  4. Suppression of MMP activity in bovine cartilage explants cultures has little if any effect on the release of aggrecanase-derived aggrecan fragments

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    Sondergaard Bodil-Cecilie


    Full Text Available Abstract Background Progressive loss of articular cartilage is a central hallmark in many joint disease, however, the relative importance of individual proteolytic pathways leading to cartilage erosion is at present unknown. We therefore investigated the time-dependant release ex vivo of MMP- and aggrecanase-derived fragments of aggrecan and type II collagen into the supernatant of bovine cartilage explants cultures using neo-epitope specific immunoassays, and to associate the release of these fragments with the activity of proteolytic enzymes using inhibitors. Findings Bovine cartilage explants were cultured in the presence or absence of the catabolic cytokines oncostatin M (OSM and tumor necrosis factor alpha (TNFα. In parallel, explants were co-cultured with protease inhibitors such as GM6001, TIMP1, TIMP2 and TIMP3. Fragments released into the supernatant were determined using a range of neo-epitope specific immunoassays; (1 sandwich 342FFGVG-G2 ELISA, (2 competition NITEGE373ELISA (3 sandwich G1-NITEGE373 ELISA (4 competition 374ARGSV ELISA, and (5 sandwich 374ARGSV-G2 ELISA all detecting aggrecan fragments, and (6 sandwich CTX-II ELISA, detecting C-telopeptides of type II collagen. We found that (1 aggrecanase-derived aggrecan fragments are released in the early (day 2-7 and mid phase (day 9-14 into the supernatant from bovine explants cultures stimulated with catabolic cytokines, (2 the release of NITEGE373 neo-epitopes are delayed compared to the corresponding 374ARGSV fragments, (3 the MMP inhibitor GM6001 did not reduce the release of aggrecanase-derived fragment, but induced a further delay in the release of these fragments, and finally (4 the MMP-derived aggrecan and type II collagen fragments were released in the late phase (day 16-21 only. Conclusion Our data support the model, that aggrecanases and MMPs act independently in the processing of the aggrecan molecules, and furthermore that suppression of MMP-activity had little if

  5. Intermittent cyclic loading of cartilage explants modulates fibronectin metabolism. (United States)

    Steinmeyer, J; Ackermann, B; Raiss, R X


    The aim of this study was to evaluate systematically the effect of tissue load, its amplitude, time of intermittence and duration of loading on the biosynthesis and release of fibronectin by intermittently loaded mature bovine articular cartilage explants. Cyclic compressive pressure was introduced using a sinusoidal waveform of 0.5 Hz-frequency with a peak stress of 0.1, 0.5 or 1.0 MPa for a period of 10 s followed by an unloaded period lasting 10, 100 or 1000 s. Fibronectin and total proteins were radiolabeled with 10 microCi/ml [3H]-phenylalanine during the final 18 h of the 1, 3 or 6 day experiments. The content of endogenous fibronectin was determined using enzyme-linked immunosorbant assay (ELISA), whereas the viability of explants was measured using sections of cartilage explants stained with fluorescein diacetate and propidium iodide. The deformation of loaded explants was determined using a load-displacement transducer system. The mechanical factor time of intermittence significantly altered the synthesis and release of fibronectin by cartilage explants, whereas the tested range of load magnitudes, as well as the duration of loading, seemed to be of subordinate importance. Loading affected the viability of the superficial zone in the cartilage, whereas the chondrocytes of the intermediate and deep zone remained viable. The compression of loaded explants was dependent on the magnitude of stress, as well as on the duration of unloading between each loading cycle. Synthesis of fibronectin, the retention of newly synthesized fibronectin within the extracellular matrix, and the portion of newly synthesized proteins that were fibronectin was significantly increased in cartilage explants which were cyclically compressed with 0.5 MPa for 10 s followed by a period of unloading lasting 100 s. Previous studies reporting that cartilage explants of human and animal osteoarthritic joints synthesize and retain elevated amounts of fibronectin imply that in our experiments

  6. The metabolic dynamics of cartilage explants over a long-term culture period

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    E.K Moo


    Full Text Available INTRODUCTION: Although previous studies have been performed on cartilage explant cultures, the generalized dynamics of cartilage metabolism after extraction from the host are still poorly understood due to differences in the experimental setups across studies, which in turn prevent building a complete picture. METHODS: In this study, we investigated the response of cartilage to the trauma sustained during extraction and determined the time needed for the cartilage to stabilize. Explants were extracted aseptically from bovine metacarpal-phalangeal joints and cultured for up to 17 days. RESULTS: The cell viability, cell number, proteoglycan content, and collagen content of the harvested explants were analyzed at 0, 2, 10, and 17 days after explantation. A high percentage of the cartilage explants were found to be viable. The cell density initially increased significantly but stabilized after two days. The proteoglycan content decreased gradually over time, but it did not decrease to a significant level due to leakage through the distorted peripheral collagen network and into the bathing medium. The collagen content remained stable for most of the culture period until it dropped abruptly on day 17. CONCLUSION: Overall, the tested cartilage explants were sustainable over long-term culture. They were most stable from day 2 to day 10. The degradation of the collagen on day 17 did not reach diseased levels, but it indicated the potential of the cultures to develop into degenerated cartilage. These findings have implications for the application of cartilage explants in pathophysiological fields.

  7. Experimentally induced cartilage degeneration treated by pulsed electromagnetic field stimulation; an in vitro study on bovine cartilage. (United States)

    Veronesi, Francesca; Fini, Milena; Giavaresi, Gianluca; Ongaro, Alessia; De Mattei, Monica; Pellati, Agnese; Setti, Stefania; Tschon, Matilde


    Osteoarthritis (OA) is the final result of progressive alterations to articular cartilage structure, composition and cellularity, followed by an increase in the concentration of pro-inflammatory cytokines in joint synovial fluid. Even though the effect of pulsed electromagnetic field (PEMF) stimulation in counteracting OA progression and inflammation is of increasing interest, because of its anabolic and anti-inflammatory properties, the present study aimed to improve the knowledge on cartilage extracellular matrix (ECM) and chondrocyte changes related to the exposure of PEMF, from a histological and histomorphometric point of view. An in vitro OA model was realized, culturing bovine cartilage explants with a high dose of interleukin 1β (IL1β, 50 ng/ml) at different experimental times (24 h, and 7 and 21 days). The effects of PEMFs (75 Hz, 1.5 mT) were evaluated in cartilage explants treated with IL1β or not (control), in terms of cartilage structure, cellularity and proteoglycans, glycosaminoglycans, collagen II and transforming growth factor β1 synthesis by using histology, histomorphometry and immunohistochemistry. Making a comparison with control cartilage, IL1β-treated explants showed a decrease in cartilage matrix, structure and cellularity parameters. PEMFs were able to counteract the progression of OA acting on both cartilage cellularity and ECM in cartilage previously treated with IL1β. Normal distribution (Kolmogroc-Smirnov test) and homoscedasticity (Levene test) of data were verified, then, the non-parametric Kruskal Wallis test followed by Mann-Whiteny U test for pairwise comparisons were performed. The p-value was adjusted according to the Dunn-Sidak correction. These results, obtained by culturing and treating cartilage explants from two different joints, confirmed that PEMF stimulation can be used as adjuvant therapy to preserve cartilage from detrimental effects of high inflammatory cytokine levels during OA.

  8. Effects of sodium hyaluronate and methylprednisolone acetate on proteoglycan synthesis in equine articular cartilage explants. (United States)

    Doyle, Aimie J; Stewart, Allison A; Constable, Peter D; Eurell, Jo Ann C; Freeman, David E; Griffon, Dominique J


    To determine effects of sodium hyaluronate (HA) on corticosteroid-induced cartilage matrix catabolism in equine articular cartilage explants. 30 articular cartilage explants from fetlock joints of 5 adult horses without joint disease. Articular cartilage explants were treated with control medium or medium containing methylprednisolone acetate (MPA; 0.05, 0.5, or 5.0 mg/mL), HA (0.1, 1.0, or 1.5 mg/mL), or both. Proteoglycan (PG) synthesis was measured by incorporation of sulfur 35-labeled sodium sulphate into PGs, and PG degradation was measured by release of radiolabeled PGs into the medium. Total glycosaminoglycan (GAG) content in media and explants and total explant DNA were determined. Methylprednisolone acetate caused a decrease in PG synthesis, whereas HA had no effect. Only the combination of MPA at a concentration of 0.05 mg/mL and HA at a concentration of 1.0 mg/mL increased PG synthesis, compared with control explants. Methylprednisolone acetate increased degradation of newly synthesized PGs into the medium, compared with control explants, and HA alone had no effect. Hyaluronate had no effect on MPA-induced PG degradation and release into media. Neither MPA alone nor HA alone had an effect on total cartilage GAG content. Methylprednisolone acetate caused an increase in release of GAG into the medium at 48 and 72 hours after treatment. In combination, HA had no protective effect on MPA-induced GAG release into the medium. Total cartilage DNA content was not affected by treatments. Our results indicate that HA addition has little effect on corticosteroid-induced cartilage matrix PG catabolism in articular cartilage explants.

  9. Bovine explant model of degeneration of the intervertebral disc

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    Sivan Sarit


    Full Text Available Abstract Background Many new treatments for degeneration of the intervertebral disc are being developed which can be delivered through a needle. These require testing in model systems before being used in human patients. Unfortunately, because of differences in anatomy, there are no ideal animal models of disc degeneration. Bovine explant model systems have many advantages but it is not possible to inject any significant volume into an intact disc. Therefore we have attempted to mimic disc degeneration in an explant bovine model via enzymatic digestion. Methods Bovine coccygeal discs were incubated with different concentrations of the proteolytic enzymes, trypsin and papain, and maintained in culture for up to 3 weeks. A radio-opaque solution was injected to visualise cavities generated. Degenerative features were monitored histologically and biochemically (water and glycosaminoglycan content, via dimethylmethylene blue. Results and Conclusion The central region of both papain and trypsin treated discs was macro- and microscopically fragmented, with severe loss of metachromasia. The integrity of the surrounding tissue was mostly in tact with cells in the outer annulus appearing viable. Biochemical analysis demonstrated greatly reduced glycosaminoglycan content in these compared to untreated discs. We have shown that bovine coccygeal discs, treated with proteolytic enzymes can provide a useful in vitro model system for developing and testing potential new treatments of disc degeneration, such as injectable implants or biological therapies.

  10. Low-intensity pulsed ultrasound (LIPUS) and pulsed electromagnetic field (PEMF) treatments affect degeneration of cultured articular cartilage explants

    NARCIS (Netherlands)

    Tan, Lijun; Tan, Lijun; Ren, Yijin; van Kooten, Theo G.; Grijpma, Dirk W.; Kuijer, Roel


    Purpose: Articular cartilage has some capacity for self-repair. Clinically used low-intensity pulsed ultrasound (LIPUS) and pulsed electromagnetic field (PEMF) treatments were compared in their potency to prevent degeneration using an explant model of porcine cartilage. Methods: Explants of porcine

  11. Low-intensity pulsed ultrasound (LIPUS) and pulsed electromagnetic field (PEMF) treatments affect degeneration of cultured articular cartilage explants

    NARCIS (Netherlands)

    Tan, Lijun; Ren, Yijin; van Kooten, Theo G.; Grijpma, Dirk W.; Kuijer, Roelof

    PURPOSE: Articular cartilage has some capacity for self-repair. Clinically used low-intensity pulsed ultrasound (LIPUS) and pulsed electromagnetic field (PEMF) treatments were compared in their potency to prevent degeneration using an explant model of porcine cartilage. METHODS: Explants of porcine

  12. Cartilage (Bovine and Shark) (PDQ®)—Patient Version (United States)

    Expert-reviewed information summary about the use of bovine and shark cartilage as a treatment for people with cancer. Note: The information in this summary is no longer being updated and is provided for reference purposes only.

  13. Cartilage (Bovine and Shark) (PDQ®)—Health Professional Version (United States)

    Expert-reviewed information summary about the use of bovine and shark cartilage as a treatment for people with cancer. Note: The information in this summary is no longer being updated and is provided for reference purposes only.

  14. Proteomic Profile of Brucella abortus-Infected Bovine Chorioallantoic Membrane Explants (United States)

    Mol, Juliana P. S.; Pires, Simone F.; Chapeaurouge, Alexander D.; Perales, Jonas; Santos, Renato L.; Andrade, Hélida M.; Lage, Andrey P.


    Brucella abortus is the etiological agent of bovine brucellosis, a zoonotic disease that causes significant economic losses worldwide. The differential proteomic profile of bovine chorioallantoic membrane (CAM) explants at early stages of infection with B. abortus (0.5, 2, 4, and 8 h) was determined. Analysis of CAM explants at 0.5 and 4 h showed the highest differences between uninfected and infected CAM explants, and therefore were used for the Differential Gel Electrophoresis (DIGE). A total of 103 spots were present in only one experimental group and were selected for identification by mass spectrometry (MALDI/ToF-ToF). Proteins only identified in extracts of CAM explants infected with B. abortus were related to recognition of PAMPs by TLR, production of reactive oxygen species, intracellular trafficking, and inflammation. PMID:27104343

  15. Facilitation of granulocyte migration into bovine pulmonary artery intimal explants by intact viable endothelium.


    Niedermeyer, M E; Meyrick, B.; Parl, F.F.; Brigham, K L


    To characterize the role of normal endothelium in granulocyte chemotaxis, the authors measured granulocyte adherence to and migration into bovine pulmonary artery intimal explants. Explants were placed, endothelium uppermost, in chemotaxis chambers with zymosan-activated plasma in the lower well and 5 X 10(6)/ml 51Cr-labeled granulocytes in the upper well. After 15, 30, 60, 120, 180, or 240 minutes incubation at 37 C, granulocyte adherence was measured by removal of adherent granulocytes from...

  16. Effects of dosage titration of methylprednisolone acetate and triamcinolone acetonide on interleukin-1-conditioned equine articular cartilage explants in vitro. (United States)

    Dechant, J E; Baxter, G M; Frisbie, D D; Trotter, G W; McIlwraith, C W


    Osteoarthritis is a frequent sequela of joint disease, especially with severe injuries or if attempts at therapy are unsuccessful. Negative and positive effects of corticosteroid treatment of articular cartilage have been demonstrated by in vitro and in vivo studies. To assess the metabolic effects of varying dosages of methylprednisolone acetate (MPA) and triamcinolone acetonide (TA) on interleukin-1alpha (IL-1) conditioned equine cartilage explants. Our hypothesis was that lower dosages of corticosteroids would be less detrimental to cartilage metabolism than higher dosages. TA would be less detrimental to cartilage metabolism than MPA. Treatment groups included articular cartilage explants with no IL-1 (control), IL-1 alone, and IL-1 plus 10, 5, 1 and 0.5 mg/ml MPA or 1.2, 0.6, 0.12 and 0.06 mg/ml TA. Explants were labelled with 35SO4 prior to the beginning and end of the experiment to assess glycosaminoglycan (GAG) degradation and synthesis, respectively. Total GAG content in media and explants and total cartilage DNA were also analysed. MPA and TA reduced GAG synthesis compared to control and IL-1 alone. The highest dosage of MPA (10 mg/ml) reduced GAG synthesis less than lower dosages of MPA and all dosages of TA. Compared to IL-1 alone, all dosages of TA and lower dosages of MPA increased GAG degradation. MPA at 10 mg/ml reduced GAG degradation. Both MPA and TA increased media GAG content compared to control and IL-1 explants. Total cartilage GAGs were unchanged with MPA, but reduced with TA, compared with IL-1 alone. Total cartilage DNA was decreased with MPA and increased with TA compared to IL-1 and control explants. MPA and TA did not counteract the negative effects of IL-1 and did not maintain cartilage metabolism at control levels. Lower dosages of MPA and TA were not less detrimental to cartilage metabolism than higher dosages. TA did not appear to be less harmful than MPA on cartilage metabolism. The results of this study differ from the findings of

  17. Histologic Evaluation of Explanted Tissue-Engineered Bovine Pericardium (CardioCel). (United States)

    Prabhu, Sudesh; Armes, Jane E; Bell, Douglas; Justo, Robert; Venugopal, Prem; Karl, Tom; Alphonso, Nelson


    CardioCel is a bovine pericardium that is subjected to a novel anticalcification tissue-engineering process. We present the histopathologic findings of human explants of CardioCel that were used in operations for congenital heart disease in children. Six explants were identified from 140 patients undergoing CardioCel implants from October 2012 to March 2015. CardioCel explants were evaluated histologically using hematoxylin and eosin, Masson trichrome, and immunohistochemical staining. A variable inflammatory response was seen in the surrounding native tissue, but not within the CardioCel graft in any of the explants. A neointimal layer of varying thickness developed on the visceral surface of 5 CardioCel explants with endothelialization of the longest duration explant. A granulation tissue layer developed on the parietal surface of the graft (consistently thicker than the neointima). Maintained collagen fiber architecture (laminated) and variable fibroblastic invasion (which increased with the age of the implant) were identified in all 6 cases. Scattered capillary vessels were noted in the majority of the explants with new collagen fibers in one, suggesting early remodeling. Calcium was seen in 1 explant at the interface of the graft and inflammatory response on its parietal surface. Evidence of graft remodeling was noted in the majority of the explants without inflammatory cells or calcification within the explanted graft material. A noticeable feature was the differential thickness of the host reaction to the parietal compared with the visceral surface of the graft. We will continue to evaluate CardioCel as a cardiovascular substitute for extracardiac and intracardiac reconstructions. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. A dose titration of triamcinolone acetonide on insulin-like growth factor-1 and interleukin-1-conditioned equine cartilage explants. (United States)

    Sandler, E A; Frisbie, D D; McIlwraith, C W


    Previous in vitro pilot studies have defined a potentially beneficial effect of insulin-like growth factor-1 (IGF-1) and triamcinolone acetonide (TA) on interleukin-1 (IL-1)-conditioned equine cartilage. Furthermore, an optimal dose for IGF-1 treatment alone has been documented previously using the same test system as in the current project. To perform a dose titration of TA on IL-1-conditioned equine articular cartilage explants in the presence of an optimised IGF-1 dose, in order to optimise a triamcinolone concentration for use in combination with IGF-1 for future investigations. Cartilage explants were harvested from the distal femur of a normal horse. The effect of a clinically relevant TA dose range was evaluated in the presence of IL-1 and IGF-1 through measurement of proteoglycan (PG) matrix metabolism (synthesis and degradation). TA and IGF-1 in combination inhibited the IL-1-induced release of PG matrix components (glycosaminoglycan or GAG) from the articular cartilage, as well as producing a significant increase in GAG synthesis. This experiment provided proof of principle that a combination treatment appears to be able to combat the IL-1-induced matrix depletion, while enhancing anabolic metabolism within the articular cartilage. The use of IGF-1 in conjunction with TA in vivo has the potential to provide beneficial anabolic effects not seen with TA alone.

  19. Effects of compression on the loss of newly synthesized proteoglycans and proteins from cartilage explants

    Energy Technology Data Exchange (ETDEWEB)

    Sah, R.L.; Doong, J.Y.; Grodzinsky, A.J.; Plaas, A.H.; Sandy, J.D. (Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology Harvard-M.I.T., Cambridge (United States))


    The effects of mechanical compression of calf cartilage explants on the catabolism and loss into the medium of proteoglycans and proteins radiolabeled with (35S)sulfate and (3H)proline were examined. A single 2- or 12-h compression of 3-mm diameter cartilage disks from a thickness of 1.25 to 0.50 mm, or slow cyclic compression (2 h on/2 h off) from 1.25 mm to 1.00, 0.75, or 0.50 mm for 24 h led to transient alterations and/or sustained increases in loss of radiolabeled macromolecules. The effects of imposing or removing loads were consistent with several compression-induced physical mediators including fluid flow, diffusion, and matrix disruption. Cyclic compression induced convective fluid flow and enhanced the loss of 35S- and 3H-labeled macromolecules from tissue into medium. In contrast, prolonged static compression induced matrix consolidation and appeared to hinder the diffusional transport and loss of 35S- and 3H-labeled macromolecules. Since high amplitude cyclic compression led to a sustained increase in the rate of loss of 3H- and 35S-labeled macromolecules that was accompanied by an increase in the rate of loss of (3H)hydroxyproline residues and an increase in tissue hydration, such compression may have caused disruption of the collagen meshwork. The 35S-labeled proteoglycans lost during such cyclic compression were of smaller average size than those from controls, but contained a similarly low proportion (approximately 15%) that could form aggregates with excess hyaluronate and link protein. The size distribution and aggregability of the remaining tissue proteoglycans and 35S-labeled proteoglycans were not markedly affected. The loss of tissue proteoglycan paralleled the loss of 35S-labeled macromolecules.

  20. In vitro and in vivo modulation of cartilage degradation by a standardized Centella asiatica fraction.

    NARCIS (Netherlands)

    Hartog, A.; Smit, H.F.; Kraan, P.M. van der; Hoijer, M.A.; Garssen, J.


    Osteoarthritis (OA) is a degenerative joint disease in which focal cartilage destruction is one of the primary features. The present study aims to evaluate the effect of a Centella asiatica fraction on in vitro and in vivo cartilage degradation. Bovine cartilage explants and bovine chondrocytes

  1. Effect of retinoic acid on proteoglycan turnover in bovine articular cartilage cultures

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    Campbell, M.A.; Handley, C.J.


    This paper describes proteoglycan catabolism by adult bovine articular cartilage treated with retinoic acid as a means of stimulating the loss of this macromolecule from the extracellular matrix of cartilage. Addition of retinoic acid (10(-12)-10(-6) M) to adult bovine articular cartilage which had been labeled with (/sup 35/S)sulfate for 6 h after 5 days in culture, resulted in a dose-dependent increase in the rate of loss of /sup 35/S-labeled proteoglycans from the matrix of the tissue. Concomitant with this loss was a decrease in the proteoglycan content of the tissue. Incubation of cultures treated with 1 microM retinoic acid, at 4 degrees C, or with 0.5 mM cycloheximide, resulted in a significant decrease in the rate of retinoic acid-induced loss of proteoglycans and demonstrated cellular involvement in this process. Analysis of the /sup 35/S-labeled proteoglycans remaining in the matrix showed that the percentage of radioactivity associated with the small proteoglycan species extracted from the matrix of articular cartilage explants labeled with (/sup 35/S)sulfate after 5 days in culture was 15% and this increased to 22% in tissue maintained in medium alone. In tissue treated with 1 microM retinoic acid for 6 days, the percentage of radioactivity associated with the small proteoglycan was 58%. Approximately 93% of the /sup 35/S-labeled proteoglycans released into the medium of control and retinoic acid-treated cultures was recovered in high density fractions after CsCl gradient centrifugation and eluted on Sepharose CL-2B as a broad peak with a Kav of 0.30-0.37. Less than 17% of these proteoglycans was capable of aggregating with hyaluronate. These results indicate that in both control and retinoic acid-treated cultures the larger proteoglycan species is lost to the medium at a greater rate than the small proteoglycan species. The effect of retinoic acid on proteoglycan turnover was shown to be reversible.

  2. Preliminary investigation of intrinsic UV fluorescence spectroscopic changes associated with proteolytic digestion of bovine articular cartilage (United States)

    Lewis, William; Padilla-Martinez, Juan-Pablo; Ortega-Martinez, Antonio; Franco, Walfre


    Degradation and destruction of articular cartilage is the etiology of osteoarthritis (OA), an entity second only to cardiovascular disease as a cause of disability in the United States. Joint mechanics and cartilage biochemistry are believed to play a role in OA; an optical tool to detect structural and chemical changes in articular cartilage might offer benefit for its early detection and treatment. The objective of the present study was to identify the spectral changes in intrinsic ultraviolet (UV) fluorescence of cartilage that occur after proteolytic digestion of cartilage. Bovine articular cartilage samples were incubated in varying concentrations of collagenase ranging from 10ug/mL up to 5mg/mL for 18 hours at 37°C, a model of OA. Pre- and post-incubation measurements were taken of the UV excitation-emission spectrum of each cartilage sample. Mechanical tests were performed to determine the pre- and post-digestion force/displacement ratio associated with indentation of each sample. Spectral changes in intrinsic cartilage fluorescence and stiffness of the cartilage were associated with proteolytic digestion. In particular, changes in the relative intensity of fluorescence peaks associated with pentosidine crosslinks (330 nm excitation, 390 nm emission) and tryptophan (290 nm excitation, 340 nm emission) were found to correlate with different degrees of cartilage digestion and cartilage stiffness. In principle, it may be possible to use UV fluorescence spectral data for early detection of damage to articular cartilage, and as a surrogate measure for cartilage stiffness.

  3. Controlled release of C-type natriuretic peptide by microencapsulation dampens proinflammatory effects induced by IL-1β in cartilage explants. (United States)

    Peake, Nick J; Pavlov, Anton M; D'Souza, Alveena; Pingguan-Murphy, Belinda; Sukhorukov, Gleb B; Hobbs, Adrian J; Chowdhury, Tina T


    C-type natriuretic peptide (CNP) exhibits potent anti-inflammatory effects in chondrocytes that have the potential to repair cartilage damage observed in osteoarthritis (OA). However, treatments for OA have been challenging due to poor targeting and delivery of therapeutics. The present study fabricated polyelectrolyte microcapsules loaded with CNP and examined whether the layer-by-layer (LbL) approach could have protective effects in cartilage explants treated with the pro-inflammatory cytokine, interleukin-1β (IL-1β). SEM showed uniform, 2 to 3 μm spherical microcapsules with morphological characteristic similar to templates loaded with or without CNP. The protein was localized around the external surface of the microcapsules with encapsulation efficiencies >82.9%. CNP release profiles were broadly similar following 9 days of culture. The presence of CNP microcapsules did not significantly affect cell viability (80%) with DNA values that remained stable throughout the culture conditions. Confocal imaging showed clustering of microcapsules in chondrocytes to natriuretic peptide receptor (Npr) 2 and 3. Treatment of cartilage explants with CNP microcapsules led to concentration-dependent inhibition of NO release in response to IL-1β and restoration of matrix synthesis. In summary, we demonstrate controlled delivery of CNP to dampen pro-inflammatory effects induced by IL-1β in cartilage explants. The LbL approach has the potential to promote cartilage repair in vivo.

  4. Chondrogenic Priming at Reduced Cell Density Enhances Cartilage Adhesion of Equine Allogeneic MSCs - a Loading Sensitive Phenomenon in an Organ Culture Study with 180 Explants. (United States)

    Spaas, Jan H; Broeckx, Sarah Y; Chiers, Koen; Ferguson, Stephen J; Casarosa, Marco; Van Bruaene, Nathalie; Forsyth, Ramses; Duchateau, Luc; Franco-Obregón, Alfredo; Wuertz-Kozak, Karin


    Clinical results of regenerative treatments for osteoarthritis are becoming increasingly significant. However, several questions remain UNANSWERED concerning mesenchymal stem cell (MSC) adhesion and incorporation into cartilage. To this end, peripheral blood (PB) MSCs were chondrogenically induced and/or stimulated with pulsed electromagnetic fields (PEMFs) for a brief period of time just sufficient to prime differentiation. In an organ culture study, PKH26 labelled MSCs were added at two different cell densities (0.5 x106 vs 1.0 x106). In total, 180 explants of six horses (30 per horse) were divided into five groups: no lesion (i), lesion alone (ii), lesion with naïve MSCs (iii), lesion with chondrogenically-induced MSCs (iv) and lesion with chondrogenically-induced and PEMF-stimulated MSCs (v). Half of the explants were mechanically loaded and compared with the unloaded equivalents. Within each circumstance, six explants were histologically evaluated at different time points (day 1, 5 and 14). COMP expression was selectively increased by chondrogenic induction (p = 0.0488). PEMF stimulation (1mT for 10 minutes) further augmented COL II expression over induced values (p = 0.0405). On the other hand, MSC markers remained constant over time after induction, indicating a largely predifferentiated state. In the unloaded group, MSCs adhered to the surface in 92.6% of the explants and penetrated into 40.7% of the lesions. On the other hand, physiological loading significantly reduced surface adherence (1.9%) and lesion filling (3.7%) in all the different conditions (p < 0.0001). Remarkably, homogenous cell distribution was characteristic for chondrogenic induced MSCs (+/- PEMFs), whereas clump formation occurred in 39% of uninduced MSC treated cartilage explants. Finally, unloaded explants seeded with a moderately low density of MSCs exhibited greater lesion filling (p = 0.0022) and surface adherence (p = 0.0161) than explants seeded with higher densities of MSCs. In

  5. Characterization of bovine cartilage by fiber Bragg grating-based stress relaxation measurements (United States)

    Baier, V.; Marchi, G.; Foehr, P.; Burgkart, R.; Roths, J.


    A fiber-based device for testing mechanical properties of cartilage is presented within this study. The measurement principle is based on stepwise indentation into the tissue and observing of corresponding relaxation of the stress. The indenter tip is constituted of a cleaved optical fiber that includes a fiber Bragg grating which is used as the force sensor. Stress relaxation measurements at 25 different positions on a healthy bovine cartilage sample were performed to assess the behavior of healthy cartilage. For each indentation step a good agreement was found with a viscoelastic model that included two time constants. The model parameters showed low variability and a clear dependence with indentation depth. The parameters can be used as reference values for discriminating healthy and degenerated cartilage.

  6. Chondrogenic Priming at Reduced Cell Density Enhances Cartilage Adhesion of Equine Allogeneic MSCs - a Loading Sensitive Phenomenon in an Organ Culture Study with 180 Explants

    Directory of Open Access Journals (Sweden)

    Jan H. Spaas


    Full Text Available Background: Clinical results of regenerative treatments for osteoarthritis are becoming increasingly significant. However, several questions remain unanswered concerning mesenchymal stem cell (MSC adhesion and incorporation into cartilage. Methods: To this end, peripheral blood (PB MSCs were chondrogenically induced and/or stimulated with pulsed electromagnetic fields (PEMFs for a brief period of time just sufficient to prime differentiation. In an organ culture study, PKH26 labelled MSCs were added at two different cell densities (0.5 x106 vs 1.0 x106. In total, 180 explants of six horses (30 per horse were divided into five groups: no lesion (i, lesion alone (ii, lesion with naïve MSCs (iii, lesion with chondrogenically-induced MSCs (iv and lesion with chondrogenically-induced and PEMF-stimulated MSCs (v. Half of the explants were mechanically loaded and compared with the unloaded equivalents. Within each circumstance, six explants were histologically evaluated at different time points (day 1, 5 and 14. Results: COMP expression was selectively increased by chondrogenic induction (p = 0.0488. PEMF stimulation (1mT for 10 minutes further augmented COL II expression over induced values (p = 0.0405. On the other hand, MSC markers remained constant over time after induction, indicating a largely predifferentiated state. In the unloaded group, MSCs adhered to the surface in 92.6% of the explants and penetrated into 40.7% of the lesions. On the other hand, physiological loading significantly reduced surface adherence (1.9% and lesion filling (3.7% in all the different conditions (p Conclusion: The present study demonstrates that primed chondrogenic induction of MSCs at a lower cell density without loading results in significantly enhanced and homogenous MSC adhesion and incorporation into equine cartilage.

  7. Noninvasive assessment of articular cartilage surface damage using reflected polarized light microscopy (United States)

    Huynh, Ruby N.; Nehmetallah, George; Raub, Christopher B.


    Articular surface damage occurs to cartilage during normal aging, osteoarthritis, and in trauma. A noninvasive assessment of cartilage microstructural alterations is useful for studies involving cartilage explants. This study evaluates polarized reflectance microscopy as a tool to assess surface damage to cartilage explants caused by mechanical scraping and enzymatic degradation. Adult bovine articular cartilage explants were scraped, incubated in collagenase, or underwent scrape and collagenase treatments. In an additional experiment, cartilage explants were subject to scrapes at graduated levels of severity. Polarized reflectance parameters were compared with India ink surface staining, features of histological sections, changes in explant wet weight and thickness, and chondrocyte viability. The polarized reflectance signal was sensitive to surface scrape damage and revealed individual scrape features consistent with India ink marks. Following surface treatments, the reflectance contrast parameter was elevated and correlated with image area fraction of India ink. After extensive scraping, polarized reflectance contrast and chondrocyte viability were lower than that from untreated explants. As part of this work, a mathematical model was developed and confirmed the trend in the reflectance signal due to changes in surface scattering and subsurface birefringence. These results demonstrate the effectiveness of polarized reflectance microscopy to sensitively assess surface microstructural alterations in articular cartilage explants.

  8. Altered osmotic swelling behavior of proteoglycan-depleted bovine articular cartilage using high frequency ultrasound

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Q; Zheng, Y P; Leung, G; Mak, A F T [Department of Health Technology and Informatics, Hong Kong Polytechnic University, Kowloon, Hong Kong (China); Lam, W L; Guo, X [Department of Rehabilitation Sciences, Hong Kong Polytechnic University, Hong Kong (China); Lu, H B; Qin, L [Musculoskeletal Research Laboratory, Department of Orthopaedics and Traumatology, Chinese University of Hong Kong, Hong Kong (China)], E-mail:


    Swelling behavior is an electrochemical mechanical property of articular cartilage. It plays an important role in weight bearing and joint lubrication. In this study, the altered transient and inhomogeneous swelling behavior of the degenerated articular cartilage was observed and quantified in situ using ultrasound. Three groups of bovine patellar articular cartilage samples (n = 10 x 3) were obtained and digested by trypsin for 10, 20 and 30 min respectively to mimic different levels of degeneration. The osmotic-free shrinkage and swelling behavior induced by changing the concentration of the bathing saline solution from 0.15 M to 2 M and then back to 0.15 M were characterized using high-frequency ultrasound (central frequency = 35 MHz) before and after digestion. It was found that the degenerated cartilage specimens showed a weaker shrinkage-swelling behavior compared with the normal cartilage samples. However, no significant differences in the peak shrinkage or swelling strains were observed between different groups. The absolute values of the peak shrinkage strain significantly (p < 0.05) decreased by 45.4%, 42.1% and 50.6% respectively after the trypsin digestion for 10, 20 and 30 min, but such significance was not demonstrated for the peak swelling strains. Due to the potential alterations in the collagen-PG matrix during trypsin digestion, the correlation between the swelling strain and the shrinkage strain of the degenerated samples changed slightly in comparison with the normal samples. The proposed ultrasound method has been successfully used to measure the transient and inhomogeneous swelling behavior of the degenerated articular cartilage and has the potential for the characterization of osteoarthritis.

  9. Analysis of mass spectrometry data from the secretome of an explant model of articular cartilage exposed to pro-inflammatory and anti-inflammatory stimuli using machine learning. (United States)

    Swan, Anna L; Hillier, Kirsty L; Smith, Julia R; Allaway, David; Liddell, Susan; Bacardit, Jaume; Mobasheri, Ali


    Osteoarthritis (OA) is an inflammatory disease of synovial joints involving the loss and degeneration of articular cartilage. The gold standard for evaluating cartilage loss in OA is the measurement of joint space width on standard radiographs. However, in most cases the diagnosis is made well after the onset of the disease, when the symptoms are well established. Identification of early biomarkers of OA can facilitate earlier diagnosis, improve disease monitoring and predict responses to therapeutic interventions. This study describes the bioinformatic analysis of data generated from high throughput proteomics for identification of potential biomarkers of OA. The mass spectrometry data was generated using a canine explant model of articular cartilage treated with the pro-inflammatory cytokine interleukin 1 β (IL-1β). The bioinformatics analysis involved the application of machine learning and network analysis to the proteomic mass spectrometry data. A rule based machine learning technique, BioHEL, was used to create a model that classified the samples into their relevant treatment groups by identifying those proteins that separated samples into their respective groups. The proteins identified were considered to be potential biomarkers. Protein networks were also generated; from these networks, proteins pivotal to the classification were identified. BioHEL correctly classified eighteen out of twenty-three samples, giving a classification accuracy of 78.3% for the dataset. The dataset included the four classes of control, IL-1β, carprofen, and IL-1β and carprofen together. This exceeded the other machine learners that were used for a comparison, on the same dataset, with the exception of another rule-based method, JRip, which performed equally well. The proteins that were most frequently used in rules generated by BioHEL were found to include a number of relevant proteins including matrix metalloproteinase 3, interleukin 8 and matrix gla protein. Using this

  10. Small-angle neutron scattering studies from solutions of bovine nasal cartilage proteoglycan (United States)

    Patel, A.; Stivala, S. S.; Damle, S. P.; Gregory, J. D.; Bunick, G. J.; Uberbacher, E. C.


    Small-angle neutron scattering, SANS, of the proteoglycan subunit of bovine nasal cartilage in 0.15 N LiC1 at 25°C yielded the radius of gyration, R g, radius of gyration of the cross-section, R q, persistence length, a *, and the molecular weight, M. The following values were obtained: M = 3.9 × 10 6, R g = 745 Å, R q = 34.6 Å and a * = 35.2 Å. These values compare favorably with those that were obtained from small angle X-ray scattering, SAXS, of a similar extract. The scattering curve of the proteoglycan subunit in D 2O showed a characteristic broad peak in the specified angular range similar to that observed from SAXS, thus confirming the polyelectrolyte nature of the proteoglycan.

  11. Cartilage. (United States)

    Caplan, Arnold I.


    Cartilage is a fundamental biological material that helps to shape the body and then helps to support it. Its fundamental properties of strength and resilience are explained in terms of the tissue's molecular structure. (JN)

  12. Tensorial Electrokinetics in Articular Cartilage (United States)

    Reynaud, Boris; Quinn, Thomas M.


    Electrokinetic phenomena contribute to biomechanical functions of articular cartilage and underlie promising methods for early detection of osteoarthritic lesions. Although some transport properties, such as hydraulic permeability, are known to become anisotropic with compression, the direction-dependence of cartilage electrokinetic properties remains unknown. Electroosmosis experiments were therefore performed on adult bovine articular cartilage samples, whereby fluid flows were driven by electric currents in directions parallel and perpendicular to the articular surface of statically compressed explants. Magnitudes of electrokinetic coefficients decreased slightly with compression (from ∼−7.5 μL/As in the range of 0–20% compression to −6.0 μL/As in the 35–50% range) consistent with predictions of microstructure-based models of cartilage material properties. However, no significant dependence on direction of the electrokinetic coupling coefficient was detected, even for conditions where the hydraulic permeability tensor is known to be anisotropic. This contrast may also be interpreted using microstructure-based models, and provides insights into structure-function relationships in cartilage extracellular matrix and physical mediators of cell responses to tissue compression. Findings support the use of relatively simple isotropic modeling approaches for electrokinetic phenomena in cartilage and related materials, and indicate that measurement of electrokinetic properties may provide particularly robust means for clinical evaluation of cartilage matrix integrity. PMID:16798804

  13. Infrapatellar fat pad aggravates degeneration of acute traumatized cartilage: a possible role for interleukin-6. (United States)

    He, J; Jiang, Y; Alexander, P G; Ulici, V; Zhu, Y; Wu, S; Tuan, R S


    The infrapatellar fat pad (IPFP), which is located underneath the patella, close to cartilage surfaces, functions in distributing mechanical load and has been shown to produce cytokines. This study aims to assess the involvement of the IPFP in the progression of post-traumatic osteoarthritis (OA) through investigating the crosstalk between the IPFP and injured cartilage in vitro. A single blunt impact (36 MPa) on healthy bovine articular cartilage explants was used to generate traumatized cartilage. Conditioned media from IPFP and traumatized cartilage (FP-CM and TC-CM) were prepared separately. After culturing in FP-CM, the posttraumatic cartilage explants were analyzed for expression of cartilage degeneration associated genes and secretion of the interleukin (IL)-6, into the culture medium. The effect of traumatized cartilage on IPFP was studied by treating IPFP-derived adipocytes and IPFP adipose-derived stromal cells (ADSC) with TC-CM followed by analysis of cytokine expression. FP-CM aggravated glycosaminoglycan (GAG) release in traumatized cartilage, but did not significantly affect healthy cartilage. FP-CM raised gene expression of cyclooxygenase-2, inducible nitric oxide synthase, and IL-6 in traumatized cartilage explants, and lowered expression of tissue inhibitor of metalloproteinases-1, 2, 3, compared to non-conditioned medium. Of particular significance is that medium IL-6 levels increased substantially in both FP-CM and FP-CM treated traumatized cartilage cultures. Extrinsic IL-6 treatment of traumatized cartilage simulated part of the effects of FP-CM. TC-CM elevated levels of IL-6 expression in IPFP derived adipocytes and ADSCs. IPFP aggravates post-traumatized cartilage degeneration, and IL-6 is a candidate tissue degeneration mediator. Copyright © 2016 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

  14. The development and characterization of a competitive ELISA for measuring active ADAMTS-4 in a bovine cartilage ex vivo model

    DEFF Research Database (Denmark)

    He, Yi; Zheng, Qinlong; Simonsen, Ole


    a monoclonal antibody against this neoepitope of active ADAMTS-4. Furthermore, we established and characterized a competitive ELISA for measuring active ADAMTS-4 form applying the specific antibody. We used this assay to profile the presence of active ADAMTS-4 and its aggrecan degradation product (NITEGE(373......)) in a bovine cartilage ex vivo model. We found that after stimulation with catabolic factors, the cartilage initially released high levels of aggrecanase-derived aggrecan fragments into supernatant but subsequently decreased to background levels. The level of active ADAMTS-4 released into the supernatant...... of osteoarthritis stained strongly for active ADAMTS-4 where surface fibrillation and clustered chondrocytes were observed. This assay could be an effective tool for studying ADAMTS-4 activity and for screening drugs regulating ADAMTS-4 activation. Moreover, it could be a potential biomarker for degenerative joint...

  15. Early transcriptional responses of bovine chorioallantoic membrane explants to wild type, ΔvirB2 or ΔbtpB Brucella abortus infection.

    Directory of Open Access Journals (Sweden)

    Juliana P S Mol

    Full Text Available The pathogenesis of the Brucella-induced inflammatory response in the bovine placenta is not completely understood. In this study we evaluated the role of the B. abortus Type IV secretion system and the anti-inflammatory factor BtpB in early interactions with bovine placental tissues. Transcription profiles of chorioallantoic membrane (CAM explants inoculated with wild type (strain 2308, ΔvirB2 or ΔbtpB Brucella abortus were compared by microarray analysis at 4 hours post infection. Transcripts with significant variation (>2 fold change; P<0.05 were functionally classified, and transcripts related to defense and inflammation were assessed by quantitative real time RT-PCR. Infection with wild type B. abortus resulted in slightly more genes with decreased than increased transcription levels. Conversely, infection of trophoblastic cells with the ΔvirB2 or the ΔbtpB mutant strains, that lack a functional T4SS or that has impaired inhibition of TLR signaling, respectively, induced more upregulated than downregulated genes. Wild type Brucella abortus impaired transcription of host genes related to immune response when compared to ΔvirB and ΔbtpB mutants. Our findings suggest that proinflammatory genes are negatively modulated in bovine trophoblastic cells at early stages of infection. The virB operon and btpB are directly or indirectly related to modulation of these host genes. These results shed light on the early interactions between B. abortus and placental tissue that ultimately culminate in inflammatory pathology and abortion.

  16. Intra-Articular Injections of Polyphenols Protect Articular Cartilage from Inflammation-Induced Degradation: Suggesting a Potential Role in Cartilage Therapeutics (United States)

    Natarajan, Venkatachalam; Madhan, Balaraman; Tiku, Moti L.


    Arthritic diseases, such as osteoarthritis and rheumatoid arthritis, inflict an enormous health care burden on society. Osteoarthritis, a degenerative joint disease with high prevalence among older people, and rheumatoid arthritis, an autoimmune inflammatory disease, both lead to irreversible structural and functional damage to articular cartilage. The aim of this study was to investigate the effect of polyphenols such as catechin, quercetin, epigallocatechin gallate, and tannic acid, on crosslinking type II collagen and the roles of these agents in managing in vivo articular cartilage degradation. The thermal, enzymatic, and physical stability of bovine articular cartilage explants following polyphenolic treatment were assessed for efficiency. Epigallocatechin gallate and tannic acid-treated explants showed >12 °C increase over native cartilage in thermal stability, thereby confirming cartilage crosslinking. Polyphenol-treated cartilage also showed a significant reduction in the percentage of collagen degradation and the release of glycosaminoglycans against collagenase digestion, indicating the increase physical integrity and resistance of polyphenol crosslinked cartilage to enzymatic digestion. To examine the in vivo cartilage protective effects, polyphenols were injected intra-articularly before (prophylactic) and after (therapeutic) the induction of collagen-induced arthritis in rats. The hind paw volume and histomorphological scoring was done for cartilage damage. The intra-articular injection of epigallocatechin gallate and tannic acid did not significantly influence the time of onset or the intensity of joint inflammation. However, histomorphological scoring of the articular cartilage showed a significant reduction in cartilage degradation in prophylactic- and therapeutic-groups, indicating that intra-articular injections of polyphenols bind to articular cartilage and making it resistant to degradation despite ongoing inflammation. These studies establish

  17. Anisotropy of collagen fibre alignment in bovine cartilage: comparison of polarised light microscopy and spatially resolved diffusion-tensor measurements. (United States)

    de Visser, S K; Bowden, J C; Wentrup-Byrne, E; Rintoul, L; Bostrom, T; Pope, J M; Momot, K I


    To compare collagen fibre alignment angles obtained from polarised light microscopy (PLM) and diffusion-tensor imaging (DTI) in bovine articular cartilage. Five samples of bovine articular cartilage from five different animals were studied using magnetic resonance imaging and PLM techniques. T(2)-weighted, diffusion-tensor (DT), and PLM images were acquired for each sample and average depth profiles of the PLM and DTI angles, as well as the banding patterns observed in T(2)-weighted magnetic resonance (MR) images, were compared. Statistical properties of the distributions of the DTI and PLM angles were examined. The samples exhibited a range of alignment morphologies. In the samples with the "conventional" three-zone alignment pattern, a correlation between the PLM and DTI alignment zones and the banding in T(2)-weighted MR images was observed. The shapes of the depth profiles of the PLM and DTI alignment angles were qualitatively similar for each sample. Three samples showed good quantitative correlation between the DT and PLM alignment angles. The correlation between the diffusion and PLM alignment angles was best in the regions of low degree of disorder of fibre alignment. This study provides the first quantitative comparison of DTI of cartilage with the more established PLM techniques. The correlation between alignment angles derived from PLM and DTI data was evident across a wide range of alignment morphologies. The results support the use of DTI for the quantitative measurement of collagen fibre alignment. The microscopic-scale (~10 microm) dispersion of fibre alignment angles appears to be an important factor for understanding the extent of quantitative correlation between PLM and DTI results.

  18. Hyaline Cartilage Tissue Is Formed through the Co-culture of Passaged Human Chondrocytes and Primary Bovine Chondrocytes (United States)

    Taylor, Drew W.; Ahmed, Nazish; Hayes, Anthony J.; Ferguson, Peter; Gross, Allan E.; Caterson, Bruce


    To circumvent the problem of a sufficient number of cells for cartilage engineering, the authors previously developed a two-stage culture system to redifferentiate monolayer culture-expanded dedifferentiated human articular chondrocytes by co-culture with primary bovine chondrocytes (bP0). The aim of this study was to analyze the composition of the cartilage tissue formed in stage 1 and compare it with bP0 grown alone to determine the optimal length of the co-culture stage of the system. Biochemical data show that extracellular matrix accumulation was evident after 2 weeks of co-culture, which was 1 week behind the bP0 control culture. By 3 to 4 weeks, the amounts of accumulated proteoglycans and collagens were comparable. Expression of chondrogenic genes, Sox 9, aggrecan, and collagen type II, was also at similar levels by week 3 of culture. Immunohistochemical staining of both co-culture and control tissues showed accumulation of type II collagen, aggrecan, biglycan, decorin, and chondroitin sulfate in appropriate zonal distributions. These data indicate that co-cultured cells form cartilaginous tissue that starts to resemble that formed by bP0 after 3 weeks, suggesting that the optimal time to terminate the co-culture stage, isolate the now redifferentiated cells, and start stage 2 is just after 3 weeks. PMID:22610463

  19. In vitro and in vivo modulation of cartilage degradation by a standardized Centella asiatica fraction. (United States)

    Hartog, Anita; Smit, H Friso; van der Kraan, Peter M; Hoijer, Maarten A; Garssen, Johan


    Osteoarthritis (OA) is a degenerative joint disease in which focal cartilage destruction is one of the primary features. The present study aims to evaluate the effect of a Centella asiatica fraction on in vitro and in vivo cartilage degradation. Bovine cartilage explants and bovine chondrocytes cultured in alginate were stimulated with IL-1 beta in the presence or absence of different concentrations (2, 5 and 10 microg/ml) of a standardized Centella asiatica triterpenes (CAT) fraction. The CAT fraction inhibited the IL-1 beta-induced proteoglycan (PG) release and nitric oxide (NO) production by cartilage explants in a dose-dependent manner. The IL-1 beta-induced reduction in PG synthesis and proliferation of chondrocytes cultured in alginate were counteracted by the CAT fraction at a concentration of 10 microg/ml. In a zymosan-induced acute arthritis model, the CAT fraction inhibited PG depletion without modulating joint swelling and inflammatory cell infiltration. In conclusion, the present study demonstrated for the first time that the tested Centella asiatica fraction was able to inhibit the zymosan-induced cartilage degradation in vivo without affecting the zymosan-induced inflammatory cell infiltration and joint swelling. The in vitro data indicate that the cartilage protective activity might at least partially be induced by the inhibition of NO production. The overall results indicate a possible disease modifying osteoarthritic activity of the Centella asiatica fraction.

  20. Quantitative Mass Spectrometry To Study Inflammatory Cartilage Degradation and Resulting Interactions with the Complement System. (United States)

    Melin Fürst, Camilla; Åhrman, Emma; Bratteby, Klas; Waldemarson, Sofia; Malmström, Johan; Blom, Anna M


    Joint diseases are often characterized by inflammatory processes that result in pathological changes in joint tissues, including cartilage degradation and release of components into the synovial fluid. The complement system plays a central role in promoting the inflammation. Because several cartilage proteins are known to interact with complement, causing either activation or inhibition of the system, we aimed to investigate these interactions comprehensively. Bovine cartilage explants were cultured with IL-1α to induce cartilage degradation, followed by incubation with human serum. Label-free selected reaction monitoring mass spectrometry was used to specifically quantify complement proteins interacting with the cartilage explant. In parallel, the time-dependent degradation of cartilage was detected using mass spectrometry analysis (liquid chromatography-tandem mass spectrometry). Complement proteins resulting from activation of the classical, alternative, and terminal pathways were detected on IL-1α-stimulated cartilage at time points when clear alterations in extracellular matrix composition had occurred. Increased levels of the complement activation product C4d, as detected by ELISA in serum after incubation with IL-1α-stimulated cartilage, confirmed the selected reaction monitoring results indicating complement activation. Further, typical activated (cleaved) C3 fragments were detected by Western blotting in extracts of IL-1α-stimulated cartilage. No complement activation was triggered by cartilage cultured in the absence of IL-1α. Components released from IL-1α-stimulated cartilage during culture had an inhibitory effect on complement activation. These were released after a longer incubation period with IL-1α and may represent a feedback reaction to cartilage-triggered complement activation observed after a shorter incubation period. Copyright © 2016 by The American Association of Immunologists, Inc.

  1. Chondroprotective effect of zinc oxide nanoparticles in conjunction with hypoxia on bovine cartilage-matrix synthesis. (United States)

    Mirza, Eraj Humayun; Pan-Pan, Chong; Wan Ibrahim, Wan Mohd Azhar Bin; Djordjevic, Ivan; Pingguan-Murphy, Belinda


    Articular cartilage is a tissue specifically adapted to a specific niche with a low oxygen tension (hypoxia), and the presence of such conditions is a key factor in regulating growth and survival of chondrocytes. Zinc deficiency has been linked to cartilage-related disease, and presence of Zinc is known to provide antibacterial benefits, which makes its inclusion attractive in an in vitro system to reduce infection. Inclusion of 1% zinc oxide nanoparticles (ZnONP) in poly octanediol citrate (POC) polymer cultured in hypoxia has not been well determined. In this study we investigated the effects of ZnONP on chondrocyte proliferation and matrix synthesis cultured under normoxia (21% O2 ) and hypoxia (5% O2 ). We report an upregulation of chondrocyte proliferation and sulfated glycosaminoglycan (S-GAG) in hypoxic culture. Results demonstrate a synergistic effect of oxygen concentration and 1% ZnONP in up-regulation of anabolic gene expression (Type II collagen and aggrecan), and a down regulation of catabolic (MMP-13) gene expression. Furthermore, production of transcription factor hypoxia-inducible factor 1A (HIF-1A) in response to hypoxic condition to regulate chondrocyte survival under hypoxia is not affected by the presence of 1% ZnONP. Presence of 1% ZnONP appears to act to preserve homeostasis of cartilage in its hypoxic environment. © 2015 Wiley Periodicals, Inc.

  2. Concentration determination of collagen and proteoglycan in bovine nasal cartilage by Fourier transform infrared imaging and PLS (United States)

    Zhang, Xuexi; Xiao, Zhi-Yan; Yin, Jianhua; Xia, Yang


    Fourier transform infrared imaging (FTIRI) combined with chemometrics can be used to detect the structure of bio-macromolecule, measure the concentrations of some components, and so on. In this study, FTIRI with Partial Least-Squares (PLS) regression was applied to study the concentration of two main components in bovine nasal cartilage (BNC), collagen and proteoglycan. An infrared spectrum library was built by mixing the collagen and chondroitin 6-sulfate (main of proteoglycan) at different ratios. Some pretreatments are needed for building PLS model. FTIR images were collected from BNC sections at 6.25μm and 25μm pixel size. The spectra extracted from BNC-FTIR images were imported into the PLS regression program to predict the concentrations of collagen and proteoglycan. These PLS-determined concentrations are agreed with the result in our previous work and biochemical analytical results. The prediction shows that the concentrations of collagen and proteoglycan in BNC are comparative on the whole. However, the concentration of proteoglycan is a litter higher than that of collagen, to some extent.

  3. Co-culture with infrapatellar fat pad differentially stimulates proteoglycan synthesis and accumulation in cartilage and meniscus tissues. (United States)

    Nishimuta, James F; Bendernagel, Monica F; Levenston, Marc E


    Although osteoarthritis is widely viewed as a disease of the whole joint, relatively few studies have focused on interactions among joint tissues in joint homeostasis and degeneration. In particular, few studies have examined the effects of the infrapatellar fat pad (IFP) on cartilaginous tissues. The aim of this study was to test the hypothesis that co-culture with healthy IFP would induce degradation of cartilage and meniscus tissues. Bovine articular cartilage, meniscus, and IFP were cultured isolated or as cartilage-fat or meniscus-fat co-cultures for up to 14 days. Conditioned media were assayed for sulfated glycosaminoglycan (sGAG) content, nitrite content, and matrix metalloproteinase (MMP) activity, and explants were assayed for sGAG and DNA contents. Co-cultures exhibited increased cumulative sGAG release and sGAG release rates for both cartilage and meniscus, and the cartilage (but not meniscus) exhibited a substantial synergistic effect of co-culture (sGAG release in co-culture was significantly greater than the summed release from isolated cartilage and fat). Fat co-culture did not significantly alter the sGAG content of either cartilage or meniscus explants, indicating that IFP co-culture stimulated net sGAG production by cartilage. Nitrite release was increased relative to isolated tissue controls in co-cultured meniscus, but not the cartilage, with no synergistic effect of co-culture. Interestingly, MMP-2 production was decreased by co-culture for both cartilage and meniscus. This study demonstrates that healthy IFP may modulate joint homeostasis by stimulating sGAG production in cartilage. Counter to our hypothesis, healthy IFP did not promote degradation of either cartilage or meniscus tissues.

  4. Full-Length Recombinant Human Proteoglycan 4 Interacts with Hyaluronan to Provide Cartilage Boundary Lubrication. (United States)

    Abubacker, Saleem; Dorosz, Samuel G; Ponjevic, Dragana; Jay, Gregory D; Matyas, John R; Schmidt, Tannin A


    Proteoglycan 4 (PRG4) is a mucin-like glycoprotein present in synovial fluid and at the surface of articular cartilage. The objectives of this study were to (1) assess the articular cartilage surface adsorption and in vitro cartilage boundary lubricating ability of full-length recombinant human PRG4 (rhPRG4), and (2) cartilage boundary lubricating ability of purified rhPRG4, both alone and in combination with hyaluronan (HA). rhPRG4 adsorption onto articular cartilage explants was assessed by immunohistochemistry and dot blot. An in vitro cartilage-cartilage friction test was used to assess rhPRG4's cartilage boundary lubricating ability compared to bovine PRG4, and that of purified rhPRG4 both alone and in combination with HA. rhPRG4 was able to adsorb to the articular surface, as well as the cut surface, of cartilage explants. The kinetic coefficient of friction of rhPRG4 was similar to that of PRG4 (p = 0.16) and lower than phosphate-buffered saline (p < 0.05), while that of purified rhPRG4 + HA was significantly lower than rhPRG4 alone (p < 0.05). This study demonstrates that rhPRG4 can adsorb to an intact articular cartilage surface and functions as an effective boundary lubricant, both alone and with HA, and provides the foundation for in vivo evaluation of this clinically relevant full-length rhPRG4 for treatment of osteoarthritis.

  5. Direct Quantification of Solute Diffusivity in Agarose and Articular Cartilage Using Correlation Spectroscopy. (United States)

    Shoga, Janty S; Graham, Brian T; Wang, Liyun; Price, Christopher


    Articular cartilage is an avascular tissue; diffusive transport is critical for its homeostasis. While numerous techniques have been used to quantify diffusivity within porous, hydrated tissues and tissue engineered constructs, these techniques have suffered from issues regarding invasiveness and spatial resolution. In the present study, we implemented and compared two separate correlation spectroscopy techniques, fluorescence correlation spectroscopy (FCS) and raster image correlation spectroscopy (RICS), for the direct, and minimally-invasive quantification of fluorescent solute diffusion in agarose and articular cartilage. Specifically, we quantified the diffusional properties of fluorescein and Alexa Fluor 488-conjugated dextrans (3k and 10k) in aqueous solutions, agarose gels of varying concentration (i.e. 1, 3, 5%), and in different zones of juvenile bovine articular cartilage explants (i.e. superficial, middle, and deep). In agarose, properties of solute diffusion obtained via FCS and RICS were inversely related to molecule size, gel concentration, and applied strain. In cartilage, the diffusional properties of solutes were similarly dependent upon solute size, cartilage zone, and compressive strain; findings that agree with work utilizing other quantification techniques. In conclusion, this study established the utility of FCS and RICS as simple and minimally invasive techniques for quantifying microscale solute diffusivity within agarose constructs and articular cartilage explants.

  6. Effect of partial H2O-D2O replacement on the anisotropy of transverse proton spin relaxation in bovine articular cartilage.

    Directory of Open Access Journals (Sweden)

    Sirisha Tadimalla

    Full Text Available Anisotropy of transverse proton spin relaxation in collagen-rich tissues like cartilage and tendon is a well-known phenomenon that manifests itself as the "magic-angle" effect in magnetic resonance images of these tissues. It is usually attributed to the non-zero averaging of intra-molecular dipolar interactions in water molecules bound to oriented collagen fibers. One way to manipulate the contributions of these interactions to spin relaxation is by partially replacing the water in the cartilage sample with deuterium oxide. It is known that dipolar interactions in deuterated solutions are weaker, resulting in a decrease in proton relaxation rates. In this work, we investigate the effects of deuteration on the longitudinal and the isotropic and anisotropic contributions to transverse relaxation of water protons in bovine articular cartilage. We demonstrate that the anisotropy of transverse proton spin relaxation in articular cartilage is independent of the degree of deuteration, bringing into question some of the assumptions currently held over the origins of relaxation anisotropy in oriented tissues.

  7. Mycobacterial antigens stimulate rheumatoid mononuclear cells to cartilage proteoglycan depletion

    NARCIS (Netherlands)

    Wilbrink, B.; Bijlsma, J. W.; Huber-Bruning, O.; van Roy, J. L.; den Otter, W.; van Eden, W.


    In a coculture with porcine articular cartilage explants unstimulated blood mononuclear cells (BMC) from patients with rheumatoid arthritis (RA), but not from healthy controls, induced proteoglycan depletion of dead cartilage. Specific stimulation of the RA BMC with Mycobacterium tuberculosis (MT),

  8. Induction of increased cAMP levels in articular chondrocytes blocks matrix metalloproteinase-mediated cartilage degradation, but not aggrecanase-mediated cartilage degradation

    DEFF Research Database (Denmark)

    Karsdal, Morten Asser; Sumer, Eren Ufuk; Wulf, Helle


    -dependently inhibited by forskolin and IBMX. The highest concentration of IBMX lowered cytokine-induced release of sGAG by 72%. CONCLUSION: Levels of cAMP in chondrocytes play a key role in controlling catabolic activity. Increased cAMP levels in chondrocytes inhibited MMP expression and activity and consequently......OBJECTIVE: Calcitonin has been suggested to have chondroprotective effects. One signaling pathway of calcitonin is via the second messenger cAMP. We undertook this study to investigate whether increased cAMP levels in chondrocytes would be chondroprotective. METHODS: Cartilage degradation...... was induced in bovine articular cartilage explants by 10 ng/ml oncostatin M (OSM) and 20 ng/ml tumor necrosis factor (TNF). In these cultures, cAMP levels were augmented by treatment with either forskolin (4, 16, or 64 microM) or 3-isobutyl-1-methyl xanthine (IBMX; 4, 16, or 64 microM). Cartilage degradation...

  9. The effect of collagen degradation on chondrocyte volume and morphology in bovine articular cartilage following a hypotonic challenge. (United States)

    Turunen, S M; Lammi, M J; Saarakkala, S; Han, S-K; Herzog, W; Tanska, P; Korhonen, R K


    Collagen degradation is one of the early signs of osteoarthritis. It is not known how collagen degradation affects chondrocyte volume and morphology. Thus, the aim of this study was to investigate the effect of enzymatically induced collagen degradation on cell volume and shape changes in articular cartilage after a hypotonic challenge. Confocal laser scanning microscopy was used for imaging superficial zone chondrocytes in intact and degraded cartilage exposed to a hypotonic challenge. Fourier transform infrared microspectroscopy, polarized light microscopy, and mechanical testing were used to quantify differences in proteoglycan and collagen content, collagen orientation, and biomechanical properties, respectively, between the intact and degraded cartilage. Collagen content decreased and collagen orientation angle increased significantly (p < 0.05) in the superficial zone cartilage after collagenase treatment, and the instantaneous modulus of the samples was reduced significantly (p < 0.05). Normalized cell volume and height 20 min after the osmotic challenge (with respect to the original volume and height) were significantly (p < 0.001 and p < 0.01, respectively) larger in the intact compared to the degraded cartilage. These findings suggest that the mechanical environment of chondrocytes, specifically collagen content and orientation, affects cell volume and shape changes in the superficial zone articular cartilage when exposed to osmotic loading. This emphasizes the role of collagen in modulating cartilage mechanobiology in diseased tissue.

  10. Enzymatic digestion of articular cartilage results in viscoelasticity changes that are consistent with polymer dynamics mechanisms

    Directory of Open Access Journals (Sweden)

    June Ronald K


    Full Text Available Abstract Background Cartilage degeneration via osteoarthritis affects millions of elderly people worldwide, yet the specific contributions of matrix biopolymers toward cartilage viscoelastic properties remain unknown despite 30 years of research. Polymer dynamics theory may enable such an understanding, and predicts that cartilage stress-relaxation will proceed faster when the average polymer length is shortened. Methods This study tested whether the predictions of polymer dynamics were consistent with changes in cartilage mechanics caused by enzymatic digestion of specific cartilage extracellular matrix molecules. Bovine calf cartilage explants were cultured overnight before being immersed in type IV collagenase, bacterial hyaluronidase, or control solutions. Stress-relaxation and cyclical loading tests were performed after 0, 1, and 2 days of incubation. Results Stress-relaxation proceeded faster following enzymatic digestion by collagenase and bacterial hyaluronidase after 1 day of incubation (both p ≤ 0.01. The storage and loss moduli at frequencies of 1 Hz and above were smaller after 1 day of digestion by collagenase and bacterial hyaluronidase (all p ≤ 0.02. Conclusion These results demonstrate that enzymatic digestion alters cartilage viscoelastic properties in a manner consistent with polymer dynamics mechanisms. Future studies may expand the use of polymer dynamics as a microstructural model for understanding the contributions of specific matrix molecules toward tissue-level viscoelastic properties.

  11. bovine

    African Journals Online (AJOL)

    of various breeds under local conditions of management. (Hale, 1974b). AdditionaIly, this procedure has been used to assess the production of LH by the bovine anterior pituitary in vitro and to study the relationships between this production and the activity of the pineal- hypothalamic axis (Hayes, Knight & Symington, 1974;.

  12. Leptin produced by joint white adipose tissue induces cartilage degradation via upregulation and activation of matrix metalloproteinases. (United States)

    Hui, Wang; Litherland, Gary J; Elias, Martina S; Kitson, Gareth I; Cawston, Tim E; Rowan, Andrew D; Young, David A


    To investigate the effect of leptin on cartilage destruction. Collagen release was assessed in bovine cartilage explant cultures, while collagenolytic and gelatinolytic activities in culture supernatants were determined by bioassay and gelatin zymography. The expression of matrix metalloproteinases (MMP) was analysed by real-time RT-PCR. Signalling pathway activation was studied by immunoblotting. Leptin levels in cultured osteoarthritic joint infrapatellar fat pad or peri-enthesal deposit supernatants were measured by immunoassay. Leptin, either alone or in synergy with IL-1, significantly induced collagen release from bovine cartilage by upregulating collagenolytic and gelatinolytic activity. In chondrocytes, leptin induced MMP1 and MMP13 expression with a concomitant activation of STAT1, STAT3, STAT5, MAPK (JNK, Erk, p38), Akt and NF-κB signalling pathways. Selective inhibitor blockade of PI3K, p38, Erk and Akt pathways significantly reduced MMP1 and MMP13 expression in chondrocytes, and reduced cartilage collagen release induced by leptin or leptin plus IL-1. JNK inhibition had no effect on leptin-induced MMP13 expression or leptin plus IL-1-induced cartilage collagen release. Conditioned media from cultured white adipose tissue (WAT) from osteoarthritis knee joint fat pads contained leptin, induced cartilage collagen release and increased MMP1 and MMP13 expression in chondrocytes; the latter being partly blocked with an anti-leptin antibody. Leptin acts as a pro-inflammatory adipokine with a catabolic role on cartilage metabolism via the upregulation of proteolytic enzymes and acts synergistically with other pro-inflammatory stimuli. This suggests that the infrapatellar fat pad and other WAT in arthritic joints are local producers of leptin, which may contribute to the inflammatory and degenerative processes in cartilage catabolism, providing a mechanistic link between obesity and osteoarthritis.

  13. Platelet-rich plasma enhances the integration of bioengineered cartilage with native tissue in an in vitro model. (United States)

    Sermer, Corey; Kandel, Rita; Anderson, Jesse; Hurtig, Mark; Theodoropoulos, John


    Current therapies for cartilage repair can be limited by an inability of the repair tissue to integrate with host tissue. Thus, there is interest in developing approaches to enhance integration. We have previously shown that platelet-rich plasma (PRP) improves cartilage tissue formation. This raised the question as to whether PRP could promote cartilage integration. Chondrocytes were isolated from cartilage harvested from bovine joints, seeded on a porous bone substitute and grown in vitro to form an osteochondral-like implant. After 7 days, the biphasic construct was soaked in PRP for 30 min before implantation into the core of a donut-shaped biphasic explant of native cartilage and bone. Controls were not soaked in PRP. The implant-explant construct was cultured for 2-4 weeks. PRP-soaked bioengineered implants integrated with host tissue in 73% of samples, whereas controls only integrated in 19% of samples. The integration strength, as determined by a push-out test, was significantly increased in the PRP-soaked implant group (219 ± 35.4 kPa) compared with controls (72.0 ± 28.5 kPa). This correlated with an increase in glycosaminoglycan and collagen accumulation in the region of integration in the PRP-treated implant group, compared with untreated controls. Immunohistochemical studies revealed that the integration zone contained collagen type II and aggrecan. The cells at the zone of integration in the PRP-soaked group had a 3.5-fold increase in matrix metalloproteinase-13 gene expression compared with controls. These results suggest that PRP-soaked bioengineered cartilage implants may be a better approach for cartilage repair due to enhanced integration. Copyright © 2017 John Wiley & Sons, Ltd.

  14. Development of a Spring-Loaded Impact Device to Deliver Injurious Mechanical Impacts to the Articular Cartilage Surface (United States)

    Alexander, Peter G.; Song, Yingjie; Taboas, Juan M.; Chen, Faye H.; Melvin, Gary M.; Manner, Paul A.


    Objective: Traumatic impacts on the articular joint surface in vitro are known to lead to degeneration of the cartilage. The main objective of this study was to develop a spring-loaded impact device that can be used to deliver traumatic impacts of consistent magnitude and rate and to find whether impacts cause catabolic activities in articular cartilage consistent with other previously reported impact models and correlated with the development of osteoarthritic lesions. In developing the spring-loaded impactor, the operating hypothesis is that a single supraphysiologic impact to articular cartilage in vitro can affect cartilage integrity, cell viability, sulfated glycosaminoglycan and inflammatory mediator release in a dose-dependent manner. Design: Impacts of increasing force are delivered to adult bovine articular cartilage explants in confined compression. Impact parameters are correlated with tissue damage, cell viability, matrix and inflammatory mediator release, and gene expression 24 hours postimpact. Results: Nitric oxide release is first detected after 7.7 MPa impacts, whereas cell death, glycosaminoglycan release, and prostaglandin E2 release are first detected at 17 MPa. Catabolic markers increase linearly to maximal levels after ≥36 MPa impacts. Conclusions: A single supraphysiologic impact negatively affects cartilage integrity, cell viability, and GAG release in a dose-dependent manner. Our findings showed that 7 to 17 MPa impacts can induce cell death and catabolism without compromising the articular surface, whereas a 17 MPa impact is sufficient to induce increases in most common catabolic markers of osteoarthritic degeneration. PMID:26069650

  15. Effect of homologous synovial membrane on adult human articular cartilage in organ culture, and failure to influence it with D-penicillamine.


    Jacoby, R K


    Adult human articular cartilage has been maintained in organ culture for 8 days, and the culture medium, which was changed on alternate days, was pooled. Normal and rheumatoid cartilage was obtained from patients and 4 types of culture were prepared: (1) cartilage alone; (2) cartilage + D-penicillamine; (3) cartilage + homologous synovium; (4) cartilage, synovium, and D-penicillamine. The hexosamines and hexuronic acid were measured in the cartilage explants and in the medium. The quantity re...

  16. Cartilage Turnover Reflected by Metabolic Processing of Type II Collagen: A Novel Marker of Anabolic Function in Chondrocytes

    Directory of Open Access Journals (Sweden)

    Natasja Stæhr Gudmann


    Full Text Available The aim of this study was to enable measurement of cartilage formation by a novel biomarker of type II collagen formation. The competitive enzyme-linked immunosorbent assay (ELISA Pro-C2 was developed and characterized for assessment of the beta splice variant of type II procollagen (PIIBNP. This is expected to originate primarily from remodeling of hyaline cartilage. A mouse monoclonal antibody (Mab was raised in mouse, targeting specifically PIIBNP (QDVRQPG and used in development of the assay. The specificity, sensitivity, 4-parameter fit and stability of the assay were tested. Levels of PIIBNP were quantified in human serum (0.6–2.2 nM, human amniotic fluid (163–188 nM and sera from different animal species, e.g., fetal bovine serum (851–901 nM with general good linearity (100% (SD 7.6 recovery and good intra- and inter-assay variation (CV% < 10. Dose (0.1 to 100 ng/mL and time (7, 14 and 21 days dependent release of PIIBNP were evaluated in the conditioned medium from bovine cartilage explants (BEX and human cartilage explants (HEX upon stimulation with insulin-like growth factor (IGF-1, transforming growth factor (TGF-β1 and fibroblastic growth factor-2 (FGF-2. TGF-β1 and IGF-1 in concentrations of 10–100 ng/mL significantly (p < 0.05 induced release of PIIBNP in BEX compared to conditions without treatment (WO. In HEX, IGF-1 100 ng/mL was able to induce a significant increase of PIIBNP after one week compared to WO. FGF-2 did not induce a PIIBNP release in our models. To our knowledge this is the first assay, which is able to specifically evaluate PIIBNP excretion. The Pro-C2 assay seems to provide a promising and novel marker of type II collagen formation.

  17. Relative contribution of matrix metalloprotease and cysteine protease activities to cytokine-stimulated articular cartilage degradation. (United States)

    Sondergaard, B C; Henriksen, K; Wulf, H; Oestergaard, S; Schurigt, U; Bräuer, R; Danielsen, I; Christiansen, C; Qvist, P; Karsdal, M A


    Both matrix metalloprotease (MMP) activity and cathepsin K (CK) activity have been implicated in cartilage turnover. We investigated the relative contribution of MMP activity and CK activity in cartilage degradation using ex vivo and in vivo models. Bovine articular cartilage explants were stimulated with oncostatin M (OSM) 10 ng/ml and tumor necrosis factor-alpha (TNF-alpha) 20 ng/ml in the presence or absence of the broad-spectrum MMP inhibitor GM6001 and the cysteine protease inhibitor, E64. Cartilage degradation was evaluated in the conditioned medium by glycosaminoglycans (GAG), hydroxyproline, and cross-linked C-telopeptide fragments of type II collagen (CTX-II), which were compared to immunohistochemical evaluations of proteoglycans and CTX-II. We assessed MMP expression by gelatine zymography and CK expression by immunohistochemistry. In vivo, CTX-II release was measured from CK-deficient mice. OSM and TNF-alpha combined induced significant (Pdegradation products measured by hydroxyproline and CTX-II compared to vehicle control. The cytokines potently induced MMP expression, assessed by zymography, and CK expression investigated by immunohistochemistry. Inhibition of MMP activity completely abrogated hydroxyproline and CTX-II release (Pdegradation. In contrast, inhibition of cysteine proteases resulted in an increase rather than a decrease in MMP derived fragments of collagen type II degradation, CTX-II, suggesting altered collagen metabolism.

  18. Hernioplastia experimental em coelhos por meio de cartilagem auricular bovina conservada em glutaraldeído Rabbits experimental hernioplasty by means of bovine auricular cartilage preserved in glutaraldehyde

    Directory of Open Access Journals (Sweden)

    L.A.F. Silva


    , characterized by abscesses and fistulas. The histological section showed areas of inflammation and necrosis next to the periphery of the graft. In the animals euthanized after 30 days, there was no evidence of clinical alterations. Microscopic diagnosis of one of these animals showed intense fibroblastic proliferation, moderate neovascularization and inflammatory cells, predominantly mononuclear. One of the animals submitted to euthanasia at 45 days presented at necropsy adherence of bowel to the graft and impaired reconstitution of the parietal peritoneum. It is possible to infer that the grafted material presented satisfactory compatibility with the receptor tissue. Thus, it may be concluded that auricular bovine cartilage grafts preserved in 4% glutaraldehyde in experimental hernioplasty in rabbits presented evidence of good tissue integration and healing, with no elimination of the grafted material.

  19. Cartilage formation measured by a novel PIINP assay suggests that IGF-I does not stimulate but maintains cartilage formation ex vivo

    DEFF Research Database (Denmark)

    Madsen, S H; Sondergaard, B C; Jensen, Anne-Christine Bay


    OBJECTIVES: The aim of this study was to investigate the time-dependent effect of insulin-like growth factor-I (IGF)-I on cartilage, evaluated by a novel procollagen type II N-terminal propeptide (PIINP) formation assay. This was performed in a cartilage model. METHODS: Bovine articular cartilage...... and more than 3000% (p cartilage formation. The current developed...


    Klisch, Stephen M.; Asanbaeva, Anna; Oungoulian, Sevan R.; Masuda, Koichi; Thonar, Eugene J-MA; Davol, Andrew; Sah, Robert L.


    A cartilage growth mixture (CGM) model is proposed to address limitations of a model used in a previous study. New stress constitutive equations for the solid matrix are derived and collagen (COL) remodeling is incorporated into the CGM model by allowing the intrinsic COL material constants to evolve during growth. An analytical validation protocol based on experimental data from a recent in vitro growth study is developed. Available data included measurements of tissue volume, biochemical composition, and tensile modulus for bovine calf articular cartilage (AC) explants harvested at three depths and incubated for 13 days in 20% FBS and 20% FBS+β-aminopropionitrile. The proposed CGM model can match tissue biochemical content and volume exactly while predicting theoretical values of tensile moduli that do not significantly differ from experimental values. Also, theoretical values of a scalar COL remodeling factor are positively correlated with COL crosslink content, and mass growth functions are positively correlated with cell density. The results suggest that the CGM model may help to guide in vitro growth protocols for AC tissue via the a priori prediction of geometric and biomechanical properties. PMID:18532855

  1. Engineering Cartilage (United States)

    ... Research Matters NIH Research Matters March 3, 2014 Engineering Cartilage Artistic rendering of human stem cells on ... situations has been a major goal in tissue engineering. Cartilage contains water, collagen, proteoglycans, and chondrocytes. Collagens ...

  2. Inhibition of glycosaminoglycan incorporation influences collagen network formation during cartilage matrix production

    NARCIS (Netherlands)

    Bastiaansen-Jenniskens, Y.M.; Koevoet, W.; Jansen, K.M.B.; Verhaar, J.A.N.; Groot, J. de; Vanosch, G.J.V.M.


    To understand cartilage degenerative diseases and improve repair procedures, we investigate the influence of glycosaminoglycans (GAGs) on cartilage matrix biochemistry and functionality. Bovine articular chondrocytes were cultured in alginate beads with(out) para-nitrophenyl-beta-d-xyloside (PNPX)

  3. Ageing is associated with reduction of mechanically-induced activation of Smad2/3P signaling in articular cartilage

    NARCIS (Netherlands)

    Madej, W.M.; Caam, A.P.M. van; Blaney Davidson, E.N.; Hannink, G.J.; Buma, P.; Kraan, P.M. van der


    OBJECTIVE: Mechanical signals control key cellular processes in articular cartilage. Previously we have shown that mechanical compression is an important ALK5/Smad2/3P activator in cartilage explants. However, age-related changes in the cartilage are known to affect tissue mechanosensitivity and

  4. Design of porous scaffolds for cartilage tissue engineering using a three-dimensional fiber-deposition technique. (United States)

    Woodfield, T B F; Malda, J; de Wijn, J; Péters, F; Riesle, J; van Blitterswijk, C A


    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 homogeneous 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.

  5. Hypoxia inhibits hypertrophic differentiation and endochondral ossification in explanted tibiae.

    Directory of Open Access Journals (Sweden)

    Jeroen C H Leijten

    Full Text Available Hypertrophic differentiation of growth plate chondrocytes induces angiogenesis which alleviates hypoxia normally present in cartilage. In the current study, we aim to determine whether alleviation of hypoxia is merely a downstream effect of hypertrophic differentiation as previously described or whether alleviation of hypoxia and consequent changes in oxygen tension mediated signaling events also plays an active role in regulating the hypertrophic differentiation process itself.Fetal mouse tibiae (E17.5 explants were cultured up to 21 days under normoxic or hypoxic conditions (21% and 2.5% oxygen respectively. Tibiae were analyzed on growth kinetics, histology, gene expression and protein secretion.The oxygen level had a strong influence on the development of explanted fetal tibiae. Compared to hypoxia, normoxia increased the length of the tibiae, length of the hypertrophic zone, calcification of the cartilage and mRNA levels of hypertrophic differentiation-related genes e.g. MMP9, MMP13, RUNX2, COL10A1 and ALPL. Compared to normoxia, hypoxia increased the size of the cartilaginous epiphysis, length of the resting zone, calcification of the bone and mRNA levels of hyaline cartilage-related genes e.g. ACAN, COL2A1 and SOX9. Additionally, hypoxia enhanced the mRNA and protein expression of the secreted articular cartilage markers GREM1, FRZB and DKK1, which are able to inhibit hypertrophic differentiation.Collectively our data suggests that oxygen levels play an active role in the regulation of hypertrophic differentiation of hyaline chondrocytes. Normoxia stimulates hypertrophic differentiation evidenced by the expression of hypertrophic differentiation related genes. In contrast, hypoxia suppresses hypertrophic differentiation of chondrocytes, which might be at least partially explained by the induction of GREM1, FRZB and DKK1 expression.

  6. Ex vivo culture platform for assessment of cartilage repair treatment strategies. (United States)

    Schwab, Andrea; Meeuwsen, Annick; Ehlicke, Franziska; Hansmann, Jan; Mulder, Lars; Smits, Anthal; Walles, Heike; Kock, Linda


    There is a great need for valuable ex vivo models that allow for assessment of cartilage repair strategies to reduce the high number of animal experiments. In this paper we present three studies with our novel ex vivo osteochondral culture platform. It consists of two separated media compartments for cartilage and bone, which better represents the in vivo situation and enables supply of factors specific to the different needs of bone and cartilage. We investigated whether separation of the cartilage and bone compartments and/or culture media results in the maintenance of viability, structural and functional properties of cartilage tissue. Next, we evaluated for how long we can preserve cartilage matrix stability of osteochondral explants during long-term culture over 84 days. Finally, we determined the optimal defect size that does not show spontaneous self-healing in this culture system. It was demonstrated that separated compartments for cartilage and bone in combination with tissue-specific medium allow for long-term culture of osteochondral explants while maintaining cartilage viability, matrix tissue content, structure and mechanical properties for at least 56 days. Furthermore, we could create critical size cartilage defects of different sizes in the model. The osteochondral model represents a valuable preclinical ex vivo tool for studying clinically relevant cartilage therapies, such as cartilage biomaterials, for their regenerative potential, for evaluation of drug and cell therapies, or to study mechanisms of cartilage regeneration. It will undoubtedly reduce the number of animals needed for in vivo testing.

  7. Matrix accumulation and retention in embryonic cartilage and in vitro chondrogenesis. (United States)

    Maleski, M P; Knudson, C B


    Since hyaluronan anchors the proteoglycan-rich pericellular matrix to chondrocytes, hyaluronan-cell interactions may direct cartilage matrix assembly. To test this hypothesis, the competitive binding of hyaluronan hexasaccharides for native hyaluronan during matrix assembly, accumulation and retention in embryonic cartilage was studied. Chondrocytes released from explants with collagenase P retained pericellular matrices, but chondrocytes appeared "matrix-free" when released from hexasaccharide-treated explants. Decreased safranin O staining was also observed in the hexasaccharide-treated explants. This loss of proteoglycan retention was demonstrated quantitatively in the cartilage extracts and recovered in the media. The continual presence of hexasaccharides in micromass cultures resulted in decreased proteoglycan deposition. Increased proteoglycan retention, indicative of matrix repair, occurred following hexasaccharide wash-out. Thus, native hyaluronan-chondrocyte interactions are important for the assembly and maintenance of cartilage matrix.

  8. C2K77 ELISA detects cleavage of type II collagen by cathepsin K in equine articular cartilage. (United States)

    Noé, B; Poole, A R; Mort, J S; Richard, H; Beauchamp, G; Laverty, S


    Develop a species-specific ELISA for a neo-epitope generated by cathepsin K cleavage of equine type II collagen to: (1) measure cartilage type II collagen degradation by cathepsin K in vitro, (2) identify cytokines that upregulate cathepsin K expression and (3) compare cathepsin K with matrix metalloproteinase (MMP) collagenase activity in stimulated cartilage explants and freshly isolated normal and osteoarthritic (OA) articular cartilages. A new ELISA (C2K77) was developed and tested by measuring the activity of exogenous cathepsin K on equine articular cartilage explants. The ELISA was then employed to measure endogenous cathepsin K activity in cultured cartilage explants with or without stimulation by interleukin-1 beta (IL-1β), tumour necrosis-alpha (TNF-α), oncostatin M (OSM) and lipopolysaccharide (LPS). Cathepsin K activity in cartilage explants (control and osteoarthritic-OA) and freshly harvested cartilage (control and OA) was compared to that of MMPs employing C2K77 and C1,2C immunoassays. The addition of Cathepsin K to normal cartilage caused a significant increase (P K77 epitope release. Whereas the content of C1,2C, that reflects MMP collagenase activity, was increased in media by the addition to cartilage explants of TNF-α and OSM (P K77 which also unchanged in OA cartilages compared to normal. The ELISA C2K77 measured the activity of cathepsin K in equine cartilage which was unchanged in OA cartilage. Cytokines that upregulate MMP collagenase activity had no effect on endogenous cathepsin K activity, suggesting a different activation mechanism that requires further study. Copyright © 2017 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

  9. Effects of oral administration of phenylbutazone to horses on in vitro articular cartilage metabolism. (United States)

    Beluche, L A; Bertone, A L; Anderson, D E; Rohde, C


    To evaluate the effects of orally administered phenylbutazone on proteoglycan synthesis and chondrocyte inhibition by IL-1beta in articular cartilage explants of horses. 11 healthy 1- to 2-year-old horses. Horses were randomly assigned to the control (n = 5) or treated group (4.4 mg of phenylbutazone/kg of body weight, p.o., q 12 h; n = 6). Articular cartilage specimens were collected before treatment was initiated (day 0), after 14 days of treatment, and 2 weeks after cessation of treatment (day 30). Proteoglycan synthesis and stromelysin concentration in cartilage extracts were assessed after 72 hours of culture in medium alone or with recombinant human interleukin-1beta (IL-1beta; 0.1 ng/ml). On day 0, proteoglycan synthesis was significantly less in cartilage explants cultured in IL-1beta, compared with medium alone. Mean proteoglycan synthesis in explants collected on days 14 and 30 was significantly less in treated horses, compared with controls. However, incubation of explants from treated horses with IL-1beta did not result in a further decrease in proteoglycan synthesis. Significant differences in stromelysin concentration were not detected between or within groups. Oral administration of phenylbutazone for 14 days significantly decreased proteoglycan synthesis in articular culture explants from healthy horses to a degree similar to that induced by in vitro exposure to IL-1beta. Phenylbutazone should be used judiciously in athletic horses with osteoarthritis, because chronic administration may suppress proteoglycan synthesis and potentiate cartilage damage.

  10. Explant cultures of human colon

    DEFF Research Database (Denmark)

    Autrup, Herman; Barrett, L.A.; Jackson, F.E.


    . The ability to maintain colonic mucosa in culture was subject to both intra- and interindividual variation. Cultured human colonic mucosa also activated a chemical procarcinogen, benzo[a]pyrene, into metabolites which bound to cellular DNA. A 100-fold interindividual variation in this binding was observed.......Human colonic epithelium has been cultured as explants in a chemically defined medium for periods of 1 to 20 days. The viability of the explants was shown by the preservation of the ultrastructural features of the colonic epithelial cells and by active incorporation of radioactive precursors...... into cellular DNA and protein. A progressive decrease in the number of goblet cells, decrease in the depth of the crypts, and a change from a columnar to a cuboidal epithelium were observed. After 20 days in culture the colonic mucosa consisted of a single layer of cuboidal epithelial cells and a few glands...

  11. Type I collagen-based fibrous capsule enhances integration of tissue-engineered cartilage with native articular cartilage. (United States)

    Yang, Yueh-Hsun; Ard, Mary B; Halper, Jaroslava T; Barabino, Gilda A


    Successful integration of engineered constructs with host tissues is crucial for cartilage repair, yet achieving it remains challenging. A collagen I-based fibrous capsule characterized by increased cell density and decreased glycosaminoglycan deposition usually forms at the periphery of tissue-engineered cartilage. The current study aimed to evaluate the effects of a solid fibrous capsule on construct integration with native articular cartilage. To this end, capsule-containing (CC) and capsule-free (CF) constructs were grown by culturing chondrocyte-seeded scaffolds with insulin-like growth factor-1 and transforming growth factor-β1, respectively, in a wavy-walled bioreactor that imparts hydrodynamic forces for 4 weeks. The ability of harvested constructs to integrate with native cartilage was determined using a cartilage explant model. Our results revealed that adhesive stress between native cartilage and the CC constructs was 57% higher than that in the CF group, potentially due to the absence of glycosaminoglycans and increased cell density in the capsule region and deposition of denser and thicker collagen fibrils at the integration site. The present work demonstrates that the fibrous capsule can effectively enhance early integration of engineered and native cartilage tissues and thus suggests the need to include the capsule as a variable in the development of cartilage tissue engineering strategies.

  12. Unique Biology of Shoulder Cartilage in Comparison to the Cartilage Obtained from the Knee and Ankle Joints (United States)

    Chubinskaya, Susan; Meyer, Maximilian A.; Urita, Atsushi; Verma, Nikhil N.; Romeo, Anthony A.; Yanke, Adam Blair; Cole, Brian J.


    Objectives: Glenohumeral joint (GHJ) arthritis in relatively young active patients presents a considerable clinical challenge. Little is known regarding the biology and reparative capacity of GHJ articular cartilage and how it compares to other diarthrodial joints. The objectives of the current study were to 1) describe the histological and morphological appearance of human normal GHJ cartilage; and 2) investigate cellular responses of GHJ cartilage to interleukin-1β (IL-1β), in comparison to cartilage obtained from the knee and ankle of the same donors. Methods: GHJ, knee (femoral condyle) and ankle (talus) cartilage was obtained through the Gift of Hope Organ and Tissue Donor Network (Itasca, IL) from nine human donors with no documented history of joint diseases (58-75 yo, both genders). Gross morphology of each joint was assessed using Collins grading on a scale 0 to 4. Cartilage explants (3 mm diameter) were removed from each joint, cultured for 48 hours with or without interleukin-1β (IL-1β; 0.1ng/ml or 10ng/ml), and processed for histology with Safranin O, proteoglycan (PG) synthesis/content, and PCR for key extracellular matrix (ECM) genes: Col2, Agg, and SOX9. Results were compared between uncultured and cultured controls and across all three included joints. Results: Unlike grossly normal (Collins grades 0-1) knee and ankle cartilage, grossly normal GHJ cartilage with an intact surface displayed signs of subtle structural changes: loss of Safranin O in the upper layer and in the ECM and increased staining around chondrocytes suggesting elevated metabolic activity. Differences became more apparent with higher Collins grades or in the presence of IL-1β. Treatment with IL-1β (both doses) resulted in more than 2-fold inhibition of PG synthesis in GHJ cartilage (p<0.05), while only high dose IL-1β had the same effect on the knee or ankle cartilage. At the control level, expression of Col2 and Sox9 was comparable between all types of cartilages; Agg

  13. Electric Field Stimulation Enhances Healing of Post-Traumatic Osteoarthritic Cartilage (United States)


    development of more effective, long-term and even preventative treatments for osteoarthritis . Our results here, in developing a protocol in which stem cells ...utilize either endogenous canine chondrocytes or the clinically relevant canine cartilage stem cells called Synovium-Derived Stem Cells (SDSCs). Moreover...Apply EFs to ‘wounded’ canine cartilage explants in the presence of labeled synovium- derived stem cells (SDSCs). Major Task 2: Perform in vitro

  14. Suppression of human cartilage proteoglycan synthesis by rheumatoid synovial fluid mononuclear cells activated with mycobacterial 60-kd heat-shock protein

    NARCIS (Netherlands)

    Wilbrink, B.; Holewijn, M.; Bijlsma, J. W.; van Roy, J. L.; den Otter, W.; van Eden, W.


    To examine whether T cell reactivity toward heat-shock proteins (HSP) contributes to cartilage destruction in rheumatoid arthritis (RA). An in vitro system was used, in which human cartilage explants were cocultured with hsp60-activated synovial fluid mononuclear cells (SFMC) from patients with RA,

  15. Explant culture of rat colon: A model system for studying metabolism of chemical carcinogens

    DEFF Research Database (Denmark)

    Autrup, Herman; Stoner, G.D.; Jackson, F.


    . The explants were incubated at 30 degrees C. The viability of the tissue was measured both by incorporation of specific precursors into cellular macromolecules and by monitoring of tissue morphology with light and electron microscopy. Cultured rat colon was able to metabolize benzo[alpha]pyrene, 7.......5% bovine albumin or 5% fetal bovine serum. The dishes were placed in a controlled-atmosphere chamber which was gassed with 95% O2 and 5% CO2. The chamber then was placed on a rocker platform which rocked at 10 cycles per min causing the medium to flow intermittently over the epithelial surface...

  16. Causes of IOL explantation in Spain. (United States)

    Fernández-Buenaga, Roberto; Alio, Jorge L; Muñoz-Negrete, Francisco J; Barraquer Compte, Rafael I; Alio-Del Barrio, Jorge L


    To study the reasons and the demography of pseudophakic intraocular lens (IOL) explantation in Spain. In this observational multicenter retrospective study, the cases studied correspond to the 15 centers that constitute the Nodo Calidad Visual y Cirugia Refractiva of the Red Tematica de Investigacion Cooperativa (RETICS) sponsored by the Spanish Ministry of Health. Clinical data from all the patients who underwent explantation were assessed. The different reasons that caused the decision of explantation were analyzed. A total of 257 explanted pseudophakic IOLs have been studied. Patients' mean age when explantation occurred was 67.5 years (SD 13.5 [22-99]) and 135 were female (52.5%). The main causes for explantation were dislocation/decentration in 145 cases (56.3%), incorrect lens power in 33 cases (12.8%), IOL opacification in 29 eyes (11.3%), neuroadaptation failure in 16 cases (6.2%), pseudophakic bullous keratopathy in 6 eyes (2.3%), endophthalmitis in 5 cases (1.9%), and "other causes" in 23 eyes (8.9%). Treatment after explantation was posterior chamber IOL implantation in 149 eyes (58%), anterior chamber IOL implantation in 49 eyes (19.1%), aphakia in 39 eyes (15.2%), and missing information in 20 cases (7.8%). Mean time from implantation to explantation was 3.97 (SD 4.68 [0.005-21.1]) years. Dislocation/decentration (most of the time with lens in the bag) is the main cause for explantation in Spain, followed by incorrect lens power (which decreased greatly over past years) and IOL opacification. Posterior chamber IOL implantation is the most elected treatment after explantation.

  17. Cartilage immunoprivilege depends on donor source and lesion location. (United States)

    Arzi, B; DuRaine, G D; Lee, C A; Huey, D J; Borjesson, D L; Murphy, B G; Hu, J C Y; Baumgarth, N; Athanasiou, K A


    The ability to repair damaged cartilage is a major goal of musculoskeletal tissue engineering. Allogeneic (same species, different individual) or xenogeneic (different species) sources can provide an attractive source of chondrocytes for cartilage tissue engineering, since autologous (same individual) cells are scarce. Immune rejection of non-autologous hyaline articular cartilage has seldom been considered due to the popular notion of "cartilage immunoprivilege". The objective of this study was to determine the suitability of allogeneic and xenogeneic engineered neocartilage tissue for cartilage repair. To address this, scaffold-free tissue engineered articular cartilage of syngeneic (same genetic background), allogeneic, and xenogeneic origin were implanted into two different locations of the rabbit knee (n=3 per group/location). Xenogeneic engineered cartilage and control xenogeneic chondral explants provoked profound innate inflammatory and adaptive cellular responses, regardless of transplant location. Cytological quantification of immune cells showed that, while allogeneic neocartilage elicited an immune response in the patella, negligible responses were observed when implanted into the trochlea; instead the responses were comparable to microfracture-treated empty defect controls. Allogeneic neocartilage survived within the trochlea implant site and demonstrated graft integration into the underlying bone. In conclusion, the knee joint cartilage does not represent an immune privileged site, strongly rejecting xenogeneic but not allogeneic chondrocytes in a location-dependent fashion. This difference in location-dependent survival of allogeneic tissue may be associated with proximity to the synovium. Through a series of in vivo studies this research demonstrates that articular cartilage is not fully immunoprivileged. In addition, we now show that anatomical location of the defect, even within the same joint compartment, strongly influences the degree of the

  18. Simultaneous Magnetic Resonance Imaging and Consolidation Measurement of Articular Cartilage

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    Robert Mark Wellard


    Full Text Available Magnetic resonance imaging (MRI offers the opportunity to study biological tissues and processes in a non-disruptive manner. The technique shows promise for the study of the load-bearing performance (consolidation of articular cartilage and changes in articular cartilage accompanying osteoarthritis. Consolidation of articular cartilage involves the recording of two transient characteristics: the change over time of strain and the hydrostatic excess pore pressure (HEPP. MRI study of cartilage consolidation under mechanical load is limited by difficulties in measuring the HEPP in the presence of the strong magnetic fields associated with the MRI technique. Here we describe the use of MRI to image and characterize bovine articular cartilage deforming under load in an MRI compatible consolidometer while monitoring pressure with a Fabry-Perot interferometer-based fiber-optic pressure transducer.

  19. Lichen explants and natural occurrence of lichens

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    Kirschbaum, A.; Klee, R.


    Studies with lichen explants and with naturally occurring lichens, conducted in the Lower Main region in West Germany within the framework of an air hydgienic and meteorologic model study of that region, are described. Parmelia physodes explants from oak trees growing in nonpolluted areas were exposed in polluted areas, such as in an industrial area, an airport, a petroleum refinery, and near a large chemical plant. The degree of air pollution in the exposure site was evaluated by the degree of the lichen damage in seven grades. The large-scale average distribution of air pollution in the survey area was studied by surveying the natural occurrence of lichen species on 10 apple trees in area units of 6.25 sq km each. The lichen explant and lichen survey methods compared by the study of naturally occurring lichens were near the exposure site of lichen explants.

  20. Optical clearing of articular cartilage: a comparison of clearing agents (United States)

    Bykov, Alexander; Hautala, Tapio; Kinnunen, Matti; Popov, Alexey; Karhula, Sakari; Saarakkala, Simo; Nieminen, Miika T.; Tuchin, Valery


    Optical clearing technique was applied to the problem of OCT imaging of articular cartilage and subchondral bone. We show that optical clearing significantly enhances visualization of articular cartilage and cartilage-bone interface. The effect of different clearing agents was analyzed. For the clearing, iohexol solution and propylene glycol (PG) were used. Clearing was performed in vitro at room temperature by immersion method. Cylindrical osteochondral samples (d=4.8mm) were drilled from bovine lateral femur and stored in phosphate-buffered saline at -20°C until clearing. Monitoring of clearing process was performed using high-speed spectral-domain OCT system providing axial resolution of 5.8μm at 930nm. Total duration of experiment was 90-100min to ensure saturation of clearing. We have shown that iohexol solution and PG are capable to optically clear articular cartilage enabling reliable characterization of cartilagebone interface with OCT. Being a low osmolarity agent, iohexol provides minimal changes to the thickness of cartilage sample. Clearing saturation time for the cartilage sample with the thickness of 0.9 mm measured with OCT is of 50 min. However, less than 15 min is enough to reliably detect the rear cartilage boundary. Alternatively, PG significantly (60%) reduces the cartilage thickness enabling better visualization of subchondral bone. It was observed that PG has higher clearing rate. The clearing saturation time is of 30 min, however less than 5 min is enough to detect cartilage-bone interface. We conclude that iohexol solution is superior for OCT imaging of cartilage and cartilage-bone interface, while PG suits better for subhondral bone visualization.

  1. Suppression of Cartilage Degradation by Zingerone Involving the p38 and JNK MAPK Signaling Pathway. (United States)

    Ruangsuriya, Jetsada; Budprom, Piyaporn; Viriyakhasem, Nawarat; Kongdang, Patiwat; Chokchaitaweesuk, Chatchadawalai; Sirikaew, Nutnicha; Chomdej, Siriwadee; Nganvongpanit, Korakot; Ongchai, Siriwan


    Zingerone, an active compound that is present in cooked ginger, has been claimed to be a bioactive ingredient that holds the potential of preventing and/or treating diseases involving inflammation. In this study, zingerone was used to discover its properties against joint inflammation using interleukin-1β-induced osteoarthritis in cartilage explant and cell culture models. Zingerone was supplemented into the cartilage explant and cell culture media at different concentrations along with the presence of interleukin-1β, an inducer of osteoarthritis. Markers indicating cartilage degradation, inflammation, and the signaling molecules involved in the inflammatory induction were investigated. Diacerien, an anti-osteoarthritic drug, was used as a positive control. Zingerone at a concentration of 40 µM reduced the level of matrix metalloproteinase-13 to about 31.95 ± 4.33 % compared with the interleukin-1β-treated group and halted cartilage explant degradation as indicated by reducing the accumulative release of sulfated glycosaminoglycans by falling to the control concomitantly with an elevation of the remaining contents of uronic acid and collagen in the explant tissues when zingerone was added. In the SW1353 cell line model, zingerone efficiently suppressed the expression of TNF-α, interleukin-6, and interleukin-8 mRNA levels and tended to reduce the levels of both p38 and c-Jun N-terminal kinase phosphorylation. From the results of this study, it can be concluded that zingerone potentially reduced cartilage degradation, which is partially involved in p38 and c-Jun N-terminal kinases of the mitogen activator protein kinase signaling pathway leading to the reduction of proinflammatory cytokine amplification effects and cartilage-degrading enzyme syntheses. This finding supports the contention that ginger holds positive pharmaceutical effects against osteoarthritis. Georg Thieme Verlag KG Stuttgart · New York.

  2. Oxidant conditioning protects cartilage from mechanically induced damage. (United States)

    Ramakrishnan, Prem; Hecht, Benjamin A; Pedersen, Douglas R; Lavery, Matthew R; Maynard, Jerry; Buckwalter, Joseph A; Martin, James A


    Articular cartilage degeneration in osteoarthritis has been linked to abnormal mechanical stresses that are known to cause chondrocyte apoptosis and metabolic derangement in in vitro models. Evidence implicating oxidative damage as the immediate cause of these harmful effects suggests that the antioxidant defenses of chondrocytes might influence their tolerance for mechanical injury. Based on evidence that antioxidant defenses in many cell types are stimulated by moderate oxidant exposure, we hypothesized that oxidant preconditioning would reduce acute chondrocyte death and proteoglycan depletion in cartilage explants after exposure to abnormal mechanical stresses. Porcine cartilage explants were treated every 48 h with tert-butyl hydrogen peroxide (tBHP) at nonlethal concentrations (25, 100, 250, and 500 microM) for a varying number of times (one, two, or four) prior to a bout of unconfined axial compression (5 MPa, 1 Hz, 1800 cycles). When compared with untreated controls, tBHP had significant positive effects on post-compression viability, lactate production, and proteoglycan losses. Overall, the most effective regime was 100 microM tBHP applied four times. RNA analysis revealed significant effects of 100 microM tBHP on gene expression. Catalase, hypoxia-inducible factor-1alpha (HIF-1alpha), and glyceraldehyde 6-phosphate dehydrogenase (GAPDH) were significantly increased relative to untreated controls in explants treated four times with 100 microM tBHP, a regime that also resulted in a significant decrease in matrix metalloproteinase-3 (MMP-3) expression. These findings demonstrate that repeated exposure of cartilage to sublethal concentrations of peroxide can moderate the acute effects of mechanical stress, a conclusion supported by evidence of peroxide-induced changes in gene expression that could render chondrocytes more resistant to oxidative damage. (c) 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  3. Comparative digital cartilage histology for human and common osteoarthritis models

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    Pedersen DR


    Full Text Available Douglas R Pedersen, Jessica E Goetz, Gail L Kurriger, James A MartinDepartment of Orthopaedics and Rehabilitation, University of Iowa, Iowa City, IA, USAPurpose: This study addresses the species-specific and site-specific details of weight-bearing articular cartilage zone depths and chondrocyte distributions among humans and common osteoarthritis (OA animal models using contemporary digital imaging tools. Histological analysis is the gold-standard research tool for evaluating cartilage health, OA severity, and treatment efficacy. Historically, evaluations were made by expert analysts. However, state-of-the-art tools have been developed that allow for digitization of entire histological sections for computer-aided analysis. Large volumes of common digital cartilage metrics directly complement elucidation of trends in OA inducement and concomitant potential treatments.Materials and methods: Sixteen fresh human knees, 26 adult New Zealand rabbit stifles, and 104 bovine lateral plateaus were measured for four cartilage zones and the cell densities within each zone. Each knee was divided into four weight-bearing sites: the medial and lateral plateaus and femoral condyles.Results: One-way analysis of variance followed by pairwise multiple comparisons (Holm–Sidak method at a significance of 0.05 clearly confirmed the variability between cartilage depths at each site, between sites in the same species, and between weight-bearing articular cartilage definitions in different species.Conclusion: The present study clearly demonstrates multisite, multispecies differences in normal weight-bearing articular cartilage, which can be objectively quantified by a common digital histology imaging technique. The clear site-specific differences in normal cartilage must be taken into consideration when characterizing the pathoetiology of OA models. Together, these provide a path to consistently analyze the volume and variety of histologic slides necessarily generated

  4. Characterization of Myelomonocytoid Progenitor Cells with Mesenchymal Differentiation Potential Obtained by Outgrowth from Pancreas Explants

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    Marc-Estienne Roehrich


    Full Text Available Progenitor cells can be obtained by outgrowth from tissue explants during primary ex vivo tissue culture. We have isolated and characterized cells outgrown from neonatal mouse pancreatic explants. A relatively uniform population of cells showing a distinctive morphology emerged over time in culture. This population expressed monocyte/macrophage and hematopoietic markers (CD11b+ and CD45+, and some stromal-related markers (CD44+ and CD29+, but not mesenchymal stem cell (MSC-defining markers (CD90− and CD105− nor endothelial (CD31− or stem cell-associated markers (CD133− and stem cell antigen-1; Sca-1−. Cells could be maintained in culture as a plastic-adherent monolayer in culture medium (MesenCult MSC for more than 1 year. Cells spontaneously formed sphere clusters “pancreatospheres” which, however, were nonclonal. When cultured in appropriate media, cells differentiated into multiple mesenchymal lineages (fat, cartilage, and bone. Positive dithizone staining suggested that a subset of cells differentiated into insulin-producing cells. However, further studies are needed to characterize the endocrine potential of these cells. These findings indicate that a myelomonocytoid population from pancreatic explant outgrowths has mesenchymal differentiation potential. These results are in line with recent data onmonocyte-derivedmesenchymal progenitors (MOMPs.

  5. Articulation of Native Cartilage Against Different Femoral Component Materials. Oxidized Zirconium Damages Cartilage Less Than Cobalt-Chrome. (United States)

    Vanlommel, Jan; De Corte, Ronny; Luyckx, Jean Philippe; Anderson, Melissa; Labey, Luc; Bellemans, Johan


    Oxidized zirconium (OxZr) is produced by thermally driven oxidization creating an oxidized surface with the properties of a ceramic at the top of the Zr metal substrate. OxZr is much harder and has a lower coefficient of friction than cobalt-chrome (CoCr), both leading to better wear characteristics. We evaluated and compared damage to the cartilage of porcine patella plugs, articulating against OxZr vs CoCr. Our hypothesis was that, owing to its better wear properties, OxZr would damage cartilage less than CoCr. If this is true, OxZr might be a better material for the femoral component during total knee arthroplasty if the patella is not resurfaced. Twenty-one plugs from porcine patellae were prepared and tested in a reciprocating pin-on-disk machine while lubricated with bovine serum and under a constant load. Three different configurations were tested: cartilage-cartilage as the control group, cartilage-OxZr, and cartilage-CoCr. Macroscopic appearance, cartilage thickness, and the modified Mankin score were evaluated after 400,000 wear cycles. The control group showed statistically significant less damage than plugs articulating against both other materials. Cartilage plugs articulating against OxZr were statistically significantly less damaged than those articulating against CoCr. Although replacing cartilage by an implant always leads to deterioration of the cartilage counterface, OxZr results in less damage than CoCr. The use of OxZr might thus be preferable to CoCr in case of total knee arthroplasty without patella resurfacing. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Cartilage Engineering and Microgravity (United States)

    Toffanin, R.; Bader, A.; Cogoli, A.; Carda, C.; Fantazzini, P.; Garrido, L.; Gomez, S.; Hall, L.; Martin, I.; Murano, E.; Poncelet, D.; Pörtner, R.; Hoffmann, F.; Roekaerts, D.; Ronney, P.; Triebel, W.; Tummers, M.


    The complex effects of mechanical forces and growth factors on articular cartilage development still need to be investigated in order to identify optimal conditions for articular cartilage repair. Strictly controlled in vitro studies under modelled or space microgravity conditions can improve our understanding of the fundamental role of gravity in articular cartilage development. The main objective of this Topical Team is to use modelled microgravity as a tool to elucidate the fundamental science of cartilage regeneration. Particular attention is, therefore, given to the effects of physical forces under altered gravitational conditions, applied using controlled bioreactor systems, on cell metabolism, cell differentiation and tissue development. Specific attention is also directed toward the potential advantages of using magnetic resonance methods for the non-destructive characterisation of scaffolds, chondrocytes-polymer constructs and tissue engineered cartilage.

  7. Cartilage stem/progenitor cells are activated in osteoarthritis via interleukin-1β/nerve growth factor signaling. (United States)

    Jiang, Yangzi; Hu, Changchang; Yu, Shuting; Yan, Junwei; Peng, Hsuan; Ouyang, Hong Wei; Tuan, Rocky S


    Interleukin-1β (IL-1β) and nerve growth factor (NGF) are key regulators in the pathogenesis of inflammatory arthritis; specifically, IL-1β is involved in tissue degeneration and NGF is involved in joint pain. However, the cellular and molecular interactions between IL-1β and NGF in articular cartilage are not known. Cartilage stem/progenitor cells (CSPCs) have recently been identified in osteoarthritic (OA) cartilage on the basis of their migratory properties. Here we hypothesize that IL-1β/NGF signaling is involved in OA cartilage degeneration by targeting CSPCs. NGF and NGF receptor (NGFR: TrkA and p75NTR) expression in healthy and OA human articular cartilage and isolated chondrocytes was determined by immunostaining, qRT-PCR, flow cytometry and western blot. Articular cartilage derived stem/progenitor cells were collected and identified by stem/progenitor cell characteristics. 3D-cultured CSPC pellets and cartilage explants were treated with NGF and NGF neutralizing antibody, and extracellular matrix changes were examined by sulfated glycosaminoglycan (GAG) release and MMP expression and activity. Expression of NGF, TrkA and p75NTR was found to be elevated in human OA cartilage. Cellular changes upon IL-1β and/or NGF treatment were then examined. NGF mRNA and NGFR proteins levels were upregulated in cultured chondrocytes exposed to IL-1β. NGF was chemotactic for cells isolated from OA cartilage. Cells isolated on the basis of their chemotactic migration towards NGF demonstrated stem/progenitor cell characteristics, including colony-forming ability, multi-lineage differentiation potential, and stem cell surface markers. The effects of NGF perturbation in cartilage explants and 3D-cultured CSPCs were next analyzed. NGF treatment resulted in extracellular matrix catabolism indicated by increased sGAG release and MMP expression and activity; conversely, treatment with NGF neutralizing antibody inhibited increased MMP levels, and enhanced tissue inhibitor of

  8. Characterising the effects of in vitro mechanical stimulation on morphogenesis of developing limb explants. (United States)

    Chandaria, Vikesh V; McGinty, James; Nowlan, Niamh C


    Mechanical forces due to fetal movements play an important role in joint shape morphogenesis, and abnormalities of the joints relating to abnormal fetal movements can have long-term health implications. While mechanical stimulation during development has been shown to be important for joint shape, the relationship between the quantity of mechanical stimulation and the growth and shape change of developing cartilage has not been quantified. In this study, we culture embryonic chick limb explants in vitro in order to reveal how the magnitude of applied movement affects key aspects of the developing joint shape. We hypothesise that joint shape is affected by movement magnitude in a dose-dependent manner, and that a movement regime most representative of physiological fetal movements will promote characteristics of normal shape development. Chick hindlimbs harvested at seven days of incubation were cultured for six days, under either static conditions or one of three different dynamic movement regimes, then assessed for joint shape, cell survival and proliferation. We demonstrate that a physiological magnitude of movement in vitro promotes the most normal progression of joint morphogenesis, and that either under-stimulation or over-stimulation has detrimental effects. Providing insight into the optimal level of mechanical stimulation for cartilage growth and morphogenesis is pertinent to gaining a greater understanding of the etiology of conditions such as developmental dysplasia of the hip, and is also valuable for cartilage tissue engineering. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

  9. Dynamic compressive properties of bovine knee layered tissue (United States)

    Nishida, Masahiro; Hino, Yuki; Todo, Mitsugu


    In Japan, the most common articular disease is knee osteoarthritis. Among many treatment methodologies, tissue engineering and regenerative medicine have recently received a lot of attention. In this field, cells and scaffolds are important, both ex vivo and in vivo. From the viewpoint of effective treatment, in addition to histological features, the compatibility of mechanical properties is also important. In this study, the dynamic and static compressive properties of bovine articular cartilage-cancellous bone layered tissue were measured using a universal testing machine and a split Hopkinson pressure bar method. The compressive behaviors of bovine articular cartilage-cancellous bone layered tissue were examined. The effects of strain rate on the maximum stress and the slope of stress-strain curves of the bovine articular cartilage-cancellous bone layered tissue were discussed.

  10. Prostaglandin E2 role in inhibition of joint cartilage collagen destruction in patients with osteoarthritis

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    E V Chetina


    Full Text Available Prostaglandin E2 role in inhibition of articular cartilage collagen degradation in patients with osteoarthritis. Objective. To assess prostaglandin E2 (PGE2 role in inhibition of type II collagen digestion in explants of articular cartilage of pts with osteoarthritis (OA. Material and methods. Explants of articular cartilage of pts with OA were cultured with PGE2 1pg to 10 ng/ml. Type II collagen digestion was assessed with immuno-enzyme assay. Gene expression was evaluated with PCR in real time. Results. PGE2 10 pg/ml as well as transforming growth factor β2 (TGFβ2 suppressed type II collagen digestion in explants of articular cartilage of pts with OA. This concentration of PGE2 did not suppress proteoglycan (aggrecan degradation. Gene expression analysis in 5 OA pts showed that PGE2 10 pg/ml suppressed metallomonooxigenase (MMP-13, MMP-1 and marker of chondrocyte hypertrophy type X collagen (COL10A1 as well as proinflammatory cytokines interleukine (IL-1β and tumor necrosis factor (TNFα. Naproxen, nonselective cyclooxygenase(COX-2 and 1 inhibitor concentration from 5 to 30 mcg/ml blocked TGFβ2 induced collagen digestion inhibition proving that PGE2 mediate influence of this growth factor. Naproxen concentration 5 mcg/ml increased collagen degradation. Conclusion. The study showed that PGE2 is a chondroprotector because it is able to suppress selectively OA pts cartilage collagen degradation. Beside that cartilage chondrocyte hypertrophy in OA connected functionally with increased collagen digestion is also regulated by low concentrations of PGE2

  11. Investigation of polarization-sensitive optical coherence tomography towards the study of microstructure of articular cartilage (United States)

    Kasaragod, Deepa; Lu, Zenghai; Le Maitre, Christine; Wilkinson, J. Mark; Matcher, Stephen


    This paper highlights the extended Jones matrix calculus based multi-angle study carried out to understand the depth dependent structural orientation of the collagen fibers in articular cartilage using polarization-sensitive optical coherence tomography (PS-OCT). A 3D lamellar model for the collagen fiber orientation, with a quadratic profile for the arching of the collagen fibers in transitional zone which points towards an ordered arrangement of fibers in that zone is the basis of the organization architecture of collagen fibers in articular cartilage. Experimental data for both ex-vivo bovine fetlock and human patellar cartilage samples are compared with theoretical predictions, with a good quantitative agreement for bovine and a reasonable qualitative agreement for human articular cartilage samples being obtained

  12. In Vitro Engineering of High Modulus Cartilage-Like Constructs. (United States)

    Finlay, Scott; Seedhom, Bahaa B; Carey, Duane O; Bulpitt, Andy J; Treanor, Darren E; Kirkham, Jennifer


    To date, the outcomes of cartilage repair have been inconsistent and have frequently yielded mechanically inferior fibrocartilage, thereby increasing the chances of damage recurrence. Implantation of constructs with biochemical composition and mechanical properties comparable to natural cartilage could be advantageous for long-term repair. This study attempted to create such constructs, in vitro, using tissue engineering principles. Bovine synoviocytes were seeded on nonwoven polyethylene terephthalate fiber scaffolds and cultured in chondrogenic medium for 4 weeks, after which uniaxial compressive loading was applied using an in-house bioreactor for 1 h per day, at a frequency of 1 Hz, for a further 84 days. The initial loading conditions, determined from the mechanical properties of the immature constructs after 4 weeks in chondrogenic culture, were strains ranging between 13% and 23%. After 56 days (sustained at 84 days) of loading, the constructs were stained homogenously with Alcian blue and for type-II collagen. Dynamic compressive moduli were comparable to the high end values for native cartilage and proportional to Alcian blue staining intensity. We suggest that these high moduli values were attributable to the bioreactor setup, which caused the loading regime to change as the constructs developed, that is, the applied stress and strain increased with construct thickness and stiffness, providing continued sufficient cell stimulation as further matrix was deposited. Constructs containing cartilage-like matrix with response to load similar to that of native cartilage could produce long-term effective cartilage repair when implanted.

  13. Contrast Agent-Enhanced Computed Tomography of Articular Cartilage: Association with Tissue Composition and Properties

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    Silvast, T.S.; Jurvelin, J.S.; Aula, A.S.; Lammi, M.J.; Toeyraes, J. (Dept. of Clinical Neurophysiology, Kuopio Univ. Hospital, Kuopio (Finland))


    Background: Contrast agent-enhanced computed tomography may enable the noninvasive quantification of glycosaminoglycan (GAG) content of articular cartilage. It has been reported that penetration of the negatively charged contrast agent ioxaglate (Hexabrix) increases significantly after enzymatic degradation of GAGs. However, it is not known whether spontaneous degradation of articular cartilage can be quantitatively detected with this technique. Purpose: To investigate the diagnostic potential of contrast agent-enhanced cartilage tomography (CECT) in quantification of GAG concentration in normal and spontaneously degenerated articular cartilage by means of clinical peripheral quantitative computed tomography (pQCT). Material and Methods: In this in vitro study, normal and spontaneously degenerated adult bovine cartilage (n=32) was used. Bovine patellar cartilage samples were immersed in 21 mM contrast agent (Hexabrix) solution for 24 hours at room temperature. After immersion, the samples were scanned with a clinical pQCT instrument. From pQCT images, the contrast agent concentration in superficial as well as in full-thickness cartilage was calculated. Histological and functional integrity of the samples was quantified with histochemical and mechanical reference measurements extracted from our earlier study. Results: Full diffusion of contrast agent into the deep cartilage was found to take over 8 hours. As compared to normal cartilage, a significant increase (11%, P<0.05) in contrast agent concentration was seen in the superficial layer of spontaneously degenerated samples. Significant negative correlations were revealed between the contrast agent concentration and the superficial or full-thickness GAG content of tissue (|R|>0.5, P<0.01). Further, pQCT could be used to measure the thickness of patellar cartilage. Conclusion: The present results suggest that CECT can be used to diagnose proteoglycan depletion in spontaneously degenerated articular cartilage with a

  14. Tissue engineering of cartilage in space (United States)

    Freed, Lisa E.; Langer, Robert; Martin, Ivan; Pellis, Neal R.; Vunjak-Novakovic, Gordana


    Tissue engineering of cartilage, i.e., the in vitro cultivation of cartilage cells on synthetic polymer scaffolds, was studied on the Mir Space Station and on Earth. Specifically, three-dimensional cell-polymer constructs consisting of bovine articular chondrocytes and polyglycolic acid scaffolds were grown in rotating bioreactors, first for 3 months on Earth and then for an additional 4 months on either Mir (10−4–10−6 g) or Earth (1 g). This mission provided a unique opportunity to study the feasibility of long-term cell culture flight experiments and to assess the effects of spaceflight on the growth and function of a model musculoskeletal tissue. Both environments yielded cartilaginous constructs, each weighing between 0.3 and 0.4 g and consisting of viable, differentiated cells that synthesized proteoglycan and type II collagen. Compared with the Earth group, Mir-grown constructs were more spherical, smaller, and mechanically inferior. The same bioreactor system can be used for a variety of controlled microgravity studies of cartilage and other tissues. These results may have implications for human spaceflight, e.g., a Mars mission, and clinical medicine, e.g., improved understanding of the effects of pseudo-weightlessness in prolonged immobilization, hydrotherapy, and intrauterine development. PMID:9391122

  15. Interleukin-29 Enhances Synovial Inflammation and Cartilage Degradation in Osteoarthritis

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    Lingxiao Xu


    Full Text Available We have recently shown that IL-29 was an important proinflammatory cytokine in pathogenesis of rheumatoid arthritis (RA. Inflammation also contributes to the pathogenesis of osteoarthritis (OA. The aim of this study was to investigate the effect and mechanism of IL-29 on cytokine production and cartilage degradation in OA. The mRNA levels of IL-29 and its specific receptor IL-28Ra in peripheral blood mononuclear cells (PBMCs were significantly increased in OA patients when compared to healthy controls (HC. In the serum, IL-29 protein levels were higher in OA patients than those in HC. Immunohistochemistry revealed that both IL-29 and IL-28Ra were dramatically elevated in OA synovium compared to HC; synovial fibroblasts (FLS and macrophages were the main IL-29-producing cells in OA synovium. Furthermore, recombinant IL-29 augmented the mRNA expression of IL-1β, IL-6, IL-8, and matrix-metalloproteinase-3 (MMP-3 in OA FLS and increased cartilage degradation when ex vivo OA cartilage explant was coincubated with OA FLS. Finally, in OA FLS, IL-29 dominantly activated MAPK and nuclear factor-κB (NF-κB, but not Jak-STAT and AKT signaling pathway as examined by western blot. In conclusion, IL-29 stimulates inflammation and cartilage degradation by OA FLS, indicating that this cytokine is likely involved in the pathogenesis of OA.

  16. Effects of glucosamine on proteoglycan loss by tendon, ligament and joint capsule explant cultures. (United States)

    Ilic, M Z; Martinac, B; Samiric, T; Handley, C J


    To investigate the effect of glucosamine on the loss of newly synthesized radiolabeled large and small proteoglycans by bovine tendon, ligament and joint capsule. The kinetics of loss of (35)S-labeled large and small proteoglycans from explant cultures of tendon, ligament and joint capsule treated with 10mM glucosamine was investigated over a 10-day culture period. The kinetics of loss of (35)S-labeled small proteoglycans and the formation of free [(35)S]sulfate were determined for the last 10 days of a 15-day culture period. The proteoglycan core proteins were analyzed by gel electrophoresis followed by fluorography. The metabolism of tendon, ligament and joint capsule explants exposed to 10mM glucosamine was evaluated by incorporation of [(3)H]serine and [(35)S]sulfate into protein and glycosaminoglycans, respectively. Glucosamine at 10mM stimulated the loss of small proteoglycans from ligament explant cultures. This was due to the increased loss of both macromolecular and free [(35)S]sulfate to the medium indicating that glucosamine affected the release of small proteoglycans as well as their intracellular degradation. The degradation pattern of small proteoglycans in ligament was not affected by glucosamine. In contrast, glucosamine did not have an effect on the loss of large or small proteoglycans from tendon and joint capsule or large proteoglycans from ligament explant cultures. The metabolism of cells in tendon, ligament and joint capsule was not impaired by the presence of 10mM glucosamine. Glucosamine stimulated the loss of small proteoglycans from ligament but did not have an effect on small proteoglycan catabolism in joint capsule and tendon or large proteoglycan catabolism in ligament, tendon or synovial capsule. The consequences of glucosamine therapy at clinically relevant concentrations on proteoglycan catabolism in joint fibrous connective tissues need to be further assessed in an animal model.

  17. Gelatin Scaffolds with Controlled Pore Structure and Mechanical Property for Cartilage Tissue Engineering. (United States)

    Chen, Shangwu; Zhang, Qin; Nakamoto, Tomoko; Kawazoe, Naoki; Chen, Guoping


    Engineering of cartilage tissue in vitro using porous scaffolds and chondrocytes provides a promising approach for cartilage repair. However, nonuniform cell distribution and heterogeneous tissue formation together with weak mechanical property of in vitro engineered cartilage limit their clinical application. In this study, gelatin porous scaffolds with homogeneous and open pores were prepared using ice particulates and freeze-drying. The scaffolds were used to culture bovine articular chondrocytes to engineer cartilage tissue in vitro. The pore structure and mechanical property of gelatin scaffolds could be well controlled by using different ratios of ice particulates to gelatin solution and different concentrations of gelatin. Gelatin scaffolds prepared from ≥70% ice particulates enabled homogeneous seeding of bovine articular chondrocytes throughout the scaffolds and formation of homogeneous cartilage extracellular matrix. While soft scaffolds underwent cellular contraction, stiff scaffolds resisted cellular contraction and had significantly higher cell proliferation and synthesis of sulfated glycosaminoglycan. Compared with the gelatin scaffolds prepared without ice particulates, the gelatin scaffolds prepared with ice particulates facilitated formation of homogeneous cartilage tissue with significantly higher compressive modulus. The gelatin scaffolds with highly open pore structure and good mechanical property can be used to improve in vitro tissue-engineered cartilage.

  18. Mechanical characterization of articular cartilage by combining magnetic resonance imaging and finite-element analysis-a potential functional imaging technique

    Energy Technology Data Exchange (ETDEWEB)

    Julkunen, P; Korhonen, R K; Nissi, M J; Jurvelin, J S [Department of Physics, University of Kuopio, PO Box 1627, FI-70211 Kuopio (Finland)], E-mail:


    Magnetic resonance imaging (MRI) provides a method for non-invasive characterization of cartilage composition and structure. We aimed to see whether T{sub 1} and T{sub 2} relaxation times are related to proteoglycan (PG) and collagen-specific mechanical properties of articular cartilage. Specifically, we analyzed whether variations in the depthwise collagen orientation, as assessed by the laminae obtained from T{sub 2} profiles, affect the mechanical characteristics of cartilage. After MRI and unconfined compression tests of human and bovine patellar cartilage samples, fibril-reinforced poroviscoelastic finite-element models (FEM), with depthwise collagen orientations implemented from quantitative T{sub 2} maps (3 laminae for human, 3-7 laminae for bovine), were constructed to analyze the non-fibrillar matrix modulus (PG specific), fibril modulus (collagen specific) and permeability of the samples. In bovine cartilage, the non-fibrillar matrix modulus (R = -0.64, p < 0.05) as well as the initial permeability (R = 0.70, p < 0.05) correlated with T{sub 1}. In bovine cartilage, T{sub 2} correlated positively with the initial fibril modulus (R = 0.62, p = 0.05). In human cartilage, the initial fibril modulus correlated negatively (R = -0.61, p < 0.05) with T{sub 2}. Based on the simulations, cartilage with a complex collagen architecture (5 or 7 laminae), leading to high bulk T{sub 2} due to magic angle effects, provided higher compressive stiffness than tissue with a simple collagen architecture (3 laminae). Our results suggest that T{sub 1} reflects PG-specific mechanical properties of cartilage. High T{sub 2} is characteristic to soft cartilage with a classical collagen architecture. Contradictorily, high bulk T{sub 2} can also be found in stiff cartilage with a multilaminar collagen fibril network. By emerging MRI and FEM, the present study establishes a step toward functional imaging of articular cartilage.

  19. Boundary conditions for cartilage regeneration

    NARCIS (Netherlands)

    Auw Yang, K.G.


    Cartilage defects generally do not heal and may result in osteoarthritis (OA) development. Unfortunately, current treatment strategies result in repair tissue with insufficient structural and mechanical properties as compared to native cartilage, and, therefore, are thought to provide merely a

  20. Stabilization of gene expression and cell morphology after explant recycling during fin explant culture in goldfish

    Energy Technology Data Exchange (ETDEWEB)

    Chenais, Nathalie; Lareyre, Jean-Jacques; Le Bail, Pierre-Yves; Labbe, Catherine, E-mail:


    The development of fin primary cell cultures for in vitro cellular and physiological studies is hampered by slow cell outgrowth, low proliferation rate, poor viability, and sparse cell characterization. Here, we investigated whether the recycling of fresh explants after a first conventional culture could improve physiological stability and sustainability of the culture. The recycled explants were able to give a supplementary cell culture showing faster outgrowth, cleaner cell layers and higher net cell production. The cells exhibited a highly stabilized profile for marker gene expression including a low cytokeratin 49 (epithelial marker) and a high collagen 1a1 (mesenchymal marker) expression. Added to the cell spindle-shaped morphology, motility behavior, and actin organization, this suggests that the cells bore stable mesenchymal characteristics. This contrast with the time-evolving expression pattern observed in the control fresh explants during the first 2 weeks of culture: a sharp decrease in cytokeratin 49 expression was concomitant with a gradual increase in col1a1. We surmise that such loss of epithelial features for the benefit of mesenchymal ones was triggered by an epithelial to mesenchymal transition (EMT) process or by way of a progressive population replacement process. Overall, our findings provide a comprehensive characterization of this new primary culture model bearing mesenchymal features and whose stability over culture time makes those cells good candidates for cell reprogramming prior to nuclear transfer, in a context of fish genome preservation. - Highlights: • Recycled fin explants outgrow cells bearing stable mesenchymal traits. • Cell production and quality is enhanced in the recycled explant culture system. • Fresh fin primary culture is highly variable and loose epithelial traits over time.

  1. Principles of cartilage repair

    CERN Document Server

    Erggelet, Christoph; Mandelbaum, Bert R


    Cartilage defects affect patients of all age groups. Surgeons, teamdoctors, general practitioners and physiotherapists alike are expected to provide adequate care. Only individual treatment plans combining a well balanced choice of various options will be successful. Background knowledge, operative and non-operative therapies are described in concise chapters: Articular cartilage biology - Diagnostics - Surgical techniques - Symptomatic and alternative medications - Physiotherapy. Diagnostic findings and surgical procedures are generously illustrated by aquarelles and colour photographs. Recommendations for additional reading, description of important clinical scoring systems and a listing of analytic tools are added for further information.

  2. Sprifermin (rhFGF18) modulates extracellular matrix turnover in cartilage explants ex vivo

    DEFF Research Database (Denmark)

    Reker, Ditte; Kjelgaard-Petersen, Cecilie Freja; Siebuhr, Anne Sofie


    or placebo at weekly intervals, similar to the dosing regimen used in clinical trials. Pre-culturing with oncostatin M and tumour necrosis factor-a, was also used to induce an inflammatory state before treatment. Metabolic activity was measured using AlamarBlue, and chondrocyte proliferation was visualized...

  3. Biochemical composition of the superficial layer of articular cartilage. (United States)

    Crockett, R; Grubelnik, A; Roos, S; Dora, C; Born, W; Troxler, H


    To gain more information on the mechanism of lubrication in articular joints, the superficial layer of bovine articular cartilage was mechanically removed in a sheet of ice that formed on freezing the cartilage. Freeze-dried samples contained low concentrations of chondroitin sulphate and protein. Analysis of the protein by SDS PAGE showed that the composition of the sample was comparable to that of synovial fluid (SF). Attenuated total reflection infrared (ATR-IR) spectroscopy of the dried residue indicated that the sample contained mostly hyaluronan. Moreover, ATR-IR spectroscopy of the upper layer of the superficial layer, adsorbed onto silicon, showed the presence of phospholipids. A gel could be formed by mixing hyaluronan and phosphatidylcholine in water with mechanical properties similar to those of the superficial layer on cartilage. Much like the superficial layer of natural cartilage, the surface of this gel became hydrophobic on drying out. Thus, it is proposed that the superficial layer forms from hyaluronan and phospholipids, which associate by hydrophobic interactions between the alkyl chains of the phospholipids and the hydrophobic faces of the disaccharide units in hyaluronan. This layer is permeable to material from the SF and the cartilage, as shown by the presence of SF proteins and chondroitin sulphate. As the cartilage dries out after removal from the joint, the phospholipids migrate towards the surface of the superficial layer to reduce the surface tension. It is also proposed that the highly efficient lubrication in articular joints can, at least in part, be attributed to the ability of the superficial layer to adsorb and hold water on the cartilage surface, thus creating a highly viscous boundary protection. Copyright 2007 Wiley Periodicals, Inc.

  4. In vitro shoot regeneration from preconditioned explants of chickpea ...

    African Journals Online (AJOL)

    The present study reports the successful shoot regeneration of preconditioned mature embryo and embryonic axis explants of chickpea cv. Gokce. Explants were preconditioned with 10 mgl benzylaminopurine (BA) for 7 days followed by culture on Murashige and Skoog (MS) medium containing 0.25, 0.50, 1.00 and 2.00 ...

  5. In vitro plant regeneration from different explants of Cardiospermum ...

    African Journals Online (AJOL)

    A rapid and efficient protocol was developed for inducing indirect organogenesis using stem and leaf explants of Cardiospermum halicacabum L. Explants were cultured on MS medium supplemented with different concentrations of IAA, NAA and 2, 4-D (1.0 to 2.0 mg/l) combined with 0.5 mg/l BAP for callus induction.

  6. Effect of explant age, hormones on somatic embryogenesis and ...

    African Journals Online (AJOL)



    Feb 18, 2009 ... from Solanum melongena L. (eggplant) cotyledon explants is associated with hormone-modulated enhancement of polyamine biosynthesis and conjugation. Protoplasma. 211:51-63. Sharma P, Rajam MV (1995). GenoType, explant and position effects on organogenesis and somatic embryogenesis in ...

  7. Cytokine manipulation of explanted Dupuytren's affected human palmar fascia. (United States)

    Kuhn, M A; Payne, W G; Kierney, P C; Pu, L L; Smith, P D; Siegler, K; Ko, F; Wang, X; Robson, M C


    Dupuytren's disease plagues human hands and digits producing fibrotic nodules and fascial cords with resultant debilitating flexion contracture deformities. Interest in this condition is great but because the disease is specific to humans and study has been hampered by the lack of an in vivo model. By utilizing an in vivo "nude" rat model it is possible to maintain and study explanted Dupuytren's contracted palmar fascia for prolonged periods of time. Human specimens were divided into four, one for in vitro analysis, and three for model explantation. The explanted tissue was perfused with either transforming growth factor beta-2 (TGFbeta2), its antibody, or a control vehicle. Explant biopsies were obtained at 30 and 60 days and compared to tissue prior to explantation. Immunohistochemistry of collagen I and III, DNA synthesis, protein production, and fibroblast kinetics were serially determined. Perfusion of explanted Dupuytren's tissue by TGFbeta2 upregulated collagen I and III from biopsies obtained from the explants at 30 days when compared to vehicle control (P < 0.001). Perfusion with antibody prevented this upregulation when compared to vehicle control (P < 0.001). Cell cultures derived from fibroblasts obtained from biopsies of the explants perfused with TGFbeta2 increased DNA synthesis, protein production and fibroblast kinetics. These findings paralleled those from other fibroproliferative disorders suggesting a role for TGFbeta2 in the pathogenesis of Dupuytren's contracture as well as possible novel treatment approaches.

  8. Adventitious shoot regeneration from leaf explants of the valuable ...

    African Journals Online (AJOL)

    The objective of this study was to develop an efficient protocol for adventitious shoot regeneration for Plectranthus barbatus Andrews using leaf explants. The explants were cultured on MS (Murashige and Skoog, 1962) medium containing various concentration of kinetin (KN), 6-benzylaminopurine (BAP) and thidiazuron ...

  9. Effect of season, explants, growth regulators and sugar level on ...

    African Journals Online (AJOL)

    The effects of antioxidant treatments, 2,4-D concentrations and sucrose level were examined in order to optimize the induction and long term maintenance of callus cultures of Ficus religiosa L. from different explants including nodal segments, inter-nodal segments and shoot apices. The explants subjected to incubation for ...

  10. Efficient regeneration of plants from shoot tip explants of ...

    African Journals Online (AJOL)

    Dendrobium densiflorum Lindl. is one of the horticulturally important orchids of Nepal due to its beautiful yellowish flower and medicinal properties. The present study was carried out for plant regeneration from shoot tip explants of D. densiflorum by tissue culture technique. The shoot tip explants of this species, obtained ...

  11. Strategies for the selection of uncontaminated Eucalyptus explants ...

    African Journals Online (AJOL)

    The potential high yields of Eucalyptus shoot multiplication achieved with a temporary immersion culture system such as RITA® are compromised by losses caused by microbial contamination particularly bacteria, characteristic of the explants used to initiate the cultures. Disinfection of the explants through antibiotic ...

  12. Optimization of explants surface sterilization condition for field grown ...

    African Journals Online (AJOL)

    The aim of this work was to sterilize nodal explants, so as to mitigate microbial contamination in peach micropropagation. The nodal explants were treated with three concentrations levels (0.15, 0.25 and 0.5% (w/v) active ingredient of chlorine) of locally produced bleach, sodium hypochlorite (NaOCl) for varying exposure ...

  13. Cartilage imaging in sports medicine. (United States)

    Black, Brandon R; Chong, Le Roy; Potter, Hollis G


    Magnetic resonance imaging (MRI) using cartilage-sensitive sequences has been shown to be an accurate, noninvasive method by which to detect articular cartilage injury and early degeneration. These are important management considerations in an athletic population. The advantages of MRI include the lack of ionizing radiation, direct multiplanar capabilities and high-contrast resolution of articular soft tissue structures. The present review details imaging strategies for assessing cartilage in the athletic population, defines the normal MRI appearance of articular cartilage, and illustrates the spectrum of articular cartilage lesions seen in various joints of the body.

  14. Auxin Transport in Explants of Coleus 1 (United States)

    Gorter, Chr. J.; Veen, H.


    α-Naphthaleneacetic acid-C14, labeled in the carboxyl group, was applied in blocks of agar to the distal and to the proximal (either apical or basal) ends of explants of Coleus. The radioactivity in receiver blocks at the opposite ends was measured. Acropetal transport was slight, only 4% of the basipetal transport. Translocation of NAA-C14 was polar in basipetal direction. Only 1.4% of the radioactivity lost from donor blocks at the apical position reached the receiver blocks; the greatest part remained in the tissue and was immobilized there. All activity found in receiver blocks at the basal end appeared to be still in the form of NAA. There were no differences between petiole tissue and stem tissue, so far as the transport of NAA is concerned. PMID:16656237


    Directory of Open Access Journals (Sweden)

    Ariana Barlič


    Surveys show that the most frequently used surgical methods are mosaicplasty and bonemarrow stimulation with microfracturing. The efficacy of the autologous chondrocyte implantationmethod should be superior to microfracturing on a long run. Especially when(regeneration of the hyaline cartilage instead of fibrous tissue (fibrocartilage is concerned.However, it has not been scientifically proved yet

  16. In-vitro and in-vivo imaging of MMP activity in cartilage and joint injury

    Energy Technology Data Exchange (ETDEWEB)

    Fukui, Tomoaki; Tenborg, Elizabeth; Yik, Jasper H.N.; Haudenschild, Dominik R., E-mail:


    Non-destructive detection of cartilage-degrading activities represents an advance in osteoarthritis (OA) research, with implications in studies of OA pathogenesis, progression, and intervention strategies. Matrix metalloproteinases (MMPs) are principal cartilage degrading enzymes that contribute to OA pathogenesis. MMPSense750 is an in-vivo fluorimetric imaging probe with the potential to continuously and non-invasively trace real-time MMP activities, but its use in OA-related research has not been reported. Our objective is to detect and characterize the early degradation activities shortly after cartilage or joint injury with MMPSense750. We determined the appropriate concentration, assay time, and linear range using various concentrations of recombinant MMPs as standards. We then quantified MMP activity from cartilage explants subjected to either mechanical injury or inflammatory cytokine treatment in-vitro. Finally, we performed in-vivo MMP imaging of a mouse model of post-traumatic OA. Our in-vitro results showed that the optimal assay time was highly dependent on the MMP enzyme. In cartilage explant culture media, mechanical impact or cytokine treatment increased MMP activity. Injured knees of mice showed significantly higher fluorescent signal than uninjured knees. We conclude that MMPSense750 detects human MMP activities and can be used for in-vitro study with cartilage, as well as in-vivo studies of knee injury, and can offering real-time insight into the degradative processes that occurring within the joint before structural changes become evident radiographically. - Highlights: • MMPSense750 is near-infrared fluorescent probe which can detect MMP activity. • MMPSense750 can detect human MMP-3, -9, and -13. • The reaction kinetics with MMPSense750 were different for the three MMPs. • MMPSense750 can visualized real time MMP activity in mouse injured knees. • MMPSense750 is convenient tool to evaluate real-time MMP activity non-invasively.

  17. Stabilization of gene expression and cell morphology after explant recycling during fin explant culture in goldfish. (United States)

    Chenais, Nathalie; Lareyre, Jean-Jacques; Le Bail, Pierre-Yves; Labbe, Catherine


    The development of fin primary cell cultures for in vitro cellular and physiological studies is hampered by slow cell outgrowth, low proliferation rate, poor viability, and sparse cell characterization. Here, we investigated whether the recycling of fresh explants after a first conventional culture could improve physiological stability and sustainability of the culture. The recycled explants were able to give a supplementary cell culture showing faster outgrowth, cleaner cell layers and higher net cell production. The cells exhibited a highly stabilized profile for marker gene expression including a low cytokeratin 49 (epithelial marker) and a high collagen 1a1 (mesenchymal marker) expression. Added to the cell spindle-shaped morphology, motility behavior, and actin organization, this suggests that the cells bore stable mesenchymal characteristics. This contrast with the time-evolving expression pattern observed in the control fresh explants during the first 2 weeks of culture: a sharp decrease in cytokeratin 49 expression was concomitant with a gradual increase in col1a1. We surmise that such loss of epithelial features for the benefit of mesenchymal ones was triggered by an epithelial to mesenchymal transition (EMT) process or by way of a progressive population replacement process. Overall, our findings provide a comprehensive characterization of this new primary culture model bearing mesenchymal features and whose stability over culture time makes those cells good candidates for cell reprogramming prior to nuclear transfer, in a context of fish genome preservation. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Cartilage extracellular matrix as a biomaterial for cartilage regeneration. (United States)

    Kiyotake, Emi A; Beck, Emily C; Detamore, Michael S


    The extracellular matrix (ECM) of various tissues possesses the model characteristics that biomaterials for tissue engineering strive to mimic; however, owing to the intricate hierarchical nature of the ECM, it has yet to be fully characterized and synthetically fabricated. Cartilage repair remains a challenge because the intrinsic properties that enable its durability and long-lasting function also impede regeneration. In the last decade, cartilage ECM has emerged as a promising biomaterial for regenerating cartilage, partly because of its potentially chondroinductive nature. As this research area of cartilage matrix-based biomaterials emerged, investigators facing similar challenges consequently developed convergent solutions in constructing robust and bioactive scaffolds. This review discusses the challenges, emerging trends, and future directions of cartilage ECM scaffolds, including a comparison between two different forms of cartilage matrix: decellularized cartilage (DCC) and devitalized cartilage (DVC). To overcome the low permeability of cartilage matrix, physical fragmentation greatly enhances decellularization, although the process itself may reduce the chondroinductivity of fabricated scaffolds. The less complex processing of a scaffold composed of DVC, which has not been decellularized, appears to have translational advantages and potential chondroinductive and mechanical advantages over DCC, without detrimental immunogenicity, to ultimately enhance cartilage repair in a clinically relevant way. © 2016 New York Academy of Sciences.

  19. Studies of mineralization in tissue culture: optimal conditions for cartilage calcification (United States)

    Boskey, A. L.; Stiner, D.; Doty, S. B.; Binderman, I.; Leboy, P.


    The optimal conditions for obtaining a calcified cartilage matrix approximating that which exists in situ were established in a differentiating chick limb bud mesenchymal cell culture system. Using cells from stage 21-24 embryos in a micro-mass culture, at an optimal density of 0.5 million cells/20 microliters spot, the deposition of small crystals of hydroxyapatite on a collagenous matrix and matrix vesicles was detected by day 21 using X-ray diffraction, FT-IR microscopy, and electron microscopy. Optimal media, containing 1.1 mM Ca, 4 mM P, 25 micrograms/ml vitamin C, 0.3 mg/ml glutamine, no Hepes buffer, and 10% fetal bovine serum, produced matrix resembling the calcifying cartilage matrix of fetal chick long bones. Interestingly, higher concentrations of fetal bovine serum had an inhibitory effect on calcification. The cartilage phenotype was confirmed based on the cellular expression of cartilage collagen and proteoglycan mRNAs, the presence of type II and type X collagen, and cartilage type proteoglycan at the light microscopic level, and the presence of chondrocytes and matrix vesicles at the EM level. The system is proposed as a model for evaluating the events in cell mediated cartilage calcification.

  20. Tribology approach to the engineering and study of articular cartilage. (United States)

    Wimmer, Markus A; Grad, Sibylle; Kaup, Thomas; Hänni, Markus; Schneider, Erich; Gogolewski, Sylwester; Alini, Mauro


    This study has been based on the assumption that articular motion is an important aspect of mechanotransduction in synovial joints. For this reason a new bioreactor concept, able to reproduce joint kinematics more closely, has been designed. The prototype consists of a rotating scaffold and/or cartilage pin, which is pressed onto an orthogonally rotating ball. By oscillating pin and ball in phase difference, elliptical displacement trajectories are generated that are similar to the motion paths occurring in vivo. Simultaneously, dynamic compression may be applied with a linear actuator, while two-step-motors generate the rotation of pin and ball. The whole apparatus is placed in an incubator. The control station is located outside. Preliminary investigations at the gene expression level demonstrated promising results. Compared with free-swelling control and/or simply compression-loaded samples, chondrocyte-seeded scaffolds as well as nasal cartilage explants exposed to interface motion both showed elevated levels of cartilage oligomeric matrix protein mRNA. The final design of the bioreactor will include four individual stations in line, which will facilitate the investigation of motion-initiated effects at the contacting surfaces in more detail.

  1. Advances in cartilage tissue engineering : in vitro

    NARCIS (Netherlands)

    E.W. Mandl (Erik)


    textabstractWithin the body three subtypes of cartilage can be distinguished: hyaline cartilage, elastic cartilage and fibrocartilage. Hyaline cartilage is the predominant subtype and is mainly located in articular joints and in less extent in the nasal septum and cricoid. Elastic cartilage can be

  2. Explantation of the novel Ahmed glaucoma valve M4 implant. (United States)

    Hu, Wanda D; Pro, Michael J; Fudemberg, Scott J; Moster, Marlene R


    To report a series of cases involving Ahmed Glaucoma Valve M4 (AGV) explantation and to discuss the surgical technique to remove the drainage device. Four cases were identified that presented with AGV M4 postoperative complications necessitating tube shunt removal. Three patients presented with conjunctival erosion and 1 patient with persistent diplopia. AGV M4 implants were removed successfully between 1.5 and 9 months after implantation. Successful explantation of the AGV M4 novel implant was achieved in all cases without intraoperative or postoperative complications. If necessary, AGV M4 explantation can be successfully performed in the early postoperative period.

  3. MRI based knee cartilage assessment (United States)

    Kroon, Dirk-Jan; Kowalski, Przemyslaw; Tekieli, Wojciech; Reeuwijk, Els; Saris, Daniel; Slump, Cornelis H.


    Osteoarthritis is one of the leading causes of pain and disability worldwide and a major health problem in developed countries due to the gradually aging population. Though the symptoms are easily recognized and described by a patient, it is difficult to assess the level of damage or loss of articular cartilage quantitatively. We present a novel method for fully automated knee cartilage thickness measurement and subsequent assessment of the knee joint. First, the point correspondence between a pre-segmented training bone model is obtained with use of Shape Context based non-rigid surface registration. Then, a single Active Shape Model (ASM) is used to segment both Femur and Tibia bone. The surfaces obtained are processed to extract the Bone-Cartilage Interface (BCI) points, where the proper segmentation of cartilage begins. For this purpose, the cartilage ASM is trained with cartilage edge positions expressed in 1D coordinates at the normals in the BCI points. The whole cartilage model is then constructed from the segmentations obtained in the previous step. An absolute thickness of the segmented cartilage is measured and compared to the mean of all training datasets, giving as a result the relative thickness value. The resulting cartilage structure is visualized and related to the segmented bone. In this way the condition of the cartilage is assessed over the surface. The quality of bone and cartilage segmentation is validated and the Dice's coefficients 0.92 and 0.86 for Femur and Tibia bones and 0.45 and 0.34 for respective cartilages are obtained. The clinical diagnostic relevance of the obtained thickness mapping is being evaluated retrospectively. We hope to validate it prospectively for prediction of clinical outcome the methods require improvements in accuracy and robustness.

  4. Mechanical Testing of Cartilage Constructs. (United States)

    Olvera, Dinorath; Daly, Andrew; Kelly, Daniel John


    A key goal of functional cartilage tissue engineering is to develop constructs with mechanical properties approaching those of the native tissue. Herein we describe a number of tests to characterize the mechanical properties of tissue engineered cartilage. Specifically, methods to determine the equilibrium confined compressive (or aggregate) modulus, the equilibrium unconfined compressive (or Young's) modulus, and the dynamic modulus of tissue engineered cartilaginous constructs are described. As these measurements are commonly used in both the articular cartilage mechanics literature and the cartilage tissue engineering literature to describe the mechanical functionality of cartilaginous constructs, they facilitate comparisons to be made between the properties of native and engineered tissues.

  5. Making post-mortem implantable cardioverter defibrillator explantation safe

    DEFF Research Database (Denmark)

    Räder, Sune B E W; Zeijlemaker, Volkert; Pehrson, Steen


    that the resting voltage over the operating person would not exceed 50 V. CONCLUSION: The use of intact medical gloves made of latex, neoprene, or plastic eliminates the potential electrical risk during explantation of an ICD. Two gloves on each hand offer sufficient protection. We will recommend the use......AIMS: The aim of this study is to investigate whether protection with rubber or plastic gloves during post-mortem explantation of an implantable cardioverter defibrillator (ICD) offers enough protection for the explanting operator during a worst-case scenario (i.e. ICD shock). METHODS AND RESULTS......: We investigated the insulating properties of rubber and plastic gloves (double layer) within the first 60 min exposure (mimicking the maximum time of an explantation procedure) to saline (simulating the effects of body fluids on the gloves). For latex gloves, we measured an increase in voltage up...

  6. Using magnetic resonance elastography to assess the dynamic mechanical properties of cartilage (United States)

    Lopez, Orlando; Amrami, Kimberly; Rossman, Phillip; Ehman, Richard L.


    This work explored the feasibility of using Magnetic Resonance Elastography (MRE) technology to enable in vitro quantification of dynamic mechanical behavior of cartilage through its thickness. A customized system for MRE of cartilage was designed to include components for adequate generation and detection of high frequency mechanical shear waves within small and stiff materials. The system included components for mechanical excitation, motion encoding, and imaging of small samples. Limitations in sensitivity to motion encoding of high frequency propagating mechanical waves using a whole body coil (i.e. Gmax = 2.2 G/cm) required the design of a local gradient coil system to achieve a gain in gradient strength of at least 5 times. The performance of the new system was tested using various cartilage-mimicking phantom materials. MRE of a stiff 5% agar gelatin phantom demonstrated gains in sensitivity to motion encoding of high frequency mechanical waves in cartilage like materials. MRE of fetal bovine cartilage samples yielded a distribution of shear stiffness within the thickness of the cartilage similar to values found in the literature, hence, suggesting the feasibility of using MRE to non-invasively and directly assess the dynamic mechanical properties of cartilage.

  7. Genipin crosslinking of cartilage enhances resistance to biochemical degradation and mechanical wear. (United States)

    McGann, Megan E; Bonitsky, Craig M; Jackson, Mariah L; Ovaert, Timothy C; Trippel, Stephen B; Wagner, Diane R


    Collagen crosslinking enhances many beneficial properties of articular cartilage, including resistance to chemical degradation and mechanical wear, but many crosslinking agents are cytotoxic. The purpose of this study was to evaluate the effectiveness of genipin, a crosslinking agent with favorable biocompatibility and cytotoxicity, as a potential treatment to prevent the degradation and wear of articular cartilage. First, the impact of genipin concentration and treatment duration on the viscoelastic properties of bovine articular cartilage was quantified. Next, two short-term (15 min) genipin crosslinking treatments were chosen, and the change in collagenase digestion, cartilage wear, and the friction coefficient of the tissue with these treatments was measured. Finally, chondrocyte viability after exposure to these genipin treatments was assessed. Genipin treatment increased the stiffness of healthy, intact cartilage in a dose-dependent manner. The 15-min crosslinking treatments improved cartilage's resistance to both chemical degradation, particularly at the articular surface, and to damage due to mechanical wear. These enhancements were achieved without sacrificing the low coefficient of friction of the tissue and at a genipin dose that preserved chondrocyte viability. The results of this study suggest that collagen crosslinking via genipin may be a promising preventative treatment to slow the degradation of cartilage. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  8. An ovine tracheal explant culture model for allergic airway inflammation

    Directory of Open Access Journals (Sweden)

    Abeynaike Latasha


    Full Text Available Abstract Background The airway epithelium is thought to play an important role in the pathogenesis of asthmatic disease. However, much of our understanding of airway epithelial cell function in asthma has been derived from in vitro studies that may not accurately reflect the interactive cellular and molecular pathways active between different tissue constituents in vivo. Methods Using a sheep model of allergic asthma, tracheal explants from normal sheep and allergic sheep exposed to house dust mite (HDM allergen were established to investigate airway mucosal responses ex vivo. Explants were cultured for up to 48 h and tissues were stained to identify apoptotic cells, goblet cells, mast cells and eosinophils. The release of cytokines (IL-1α, IL-6 and TNF-α by cultured tracheal explants, was assessed by ELISA. Results The general morphology and epithelial structure of the tracheal explants was well maintained in culture although evidence of advanced apoptosis within the mucosal layer was noted after culture for 48 h. The number of alcian blue/PAS positive mucus-secreting cells within the epithelial layer was reduced in all cultured explants compared with pre-cultured (0 h explants, but the loss of staining was most evident in allergic tissues. Mast cell and eosinophil numbers were elevated in the allergic tracheal tissues compared to naïve controls, and in the allergic tissues there was a significant decline in mast cells after 24 h culture in the presence or absence of HDM allergen. IL-6 was released by allergic tracheal explants in culture but was undetected in cultured control explants. Conclusions Sheep tracheal explants maintain characteristics of the airway mucosa that may not be replicated when studying isolated cell populations in vitro. There were key differences identified in explants from allergic compared to control airways and in their responses in culture for 24 h. Importantly, this study establishes the potential for the

  9. Linking Cellular and Mechanical Processes in Articular Cartilage Lesion Formation: A Mathematical Model

    Directory of Open Access Journals (Sweden)

    Georgi I Kapitanov


    Full Text Available Post-traumatic osteoarthritis affects almost 20% of the adult US population. An injurious impact applies a significant amount of physical stress on articular cartilage and can initiate a cascade of biochemical reactions that can lead to the development of osteoarthritis. In our effort to understand the underlying biochemical mechanisms of this debilitating disease, we have constructed a multiscale mathematical model of the process with three components: cellular, chemical, and mechanical. The cellular component describes the different chondrocyte states according to the chemicals these cells release. The chemical component models the change in concentrations of those chemicals. The mechanical component contains a simulation of a blunt impact applied onto a cartilage explant and the resulting strains that initiate the biochemical processes. The scales are modeled through a system of partial-differential equations and solved numerically. The results of the model qualitatively capture the results of laboratory experiments of drop-tower impacts on cartilage explants. The model creates a framework for incorporating explicit mechanics, simulated by finite element analysis, into a theoretical biology framework. The effort is a step toward a complete virtual platform for modeling the development of post-traumatic osteoarthritis, which will be used to inform biomedical researchers on possible non-invasive strategies for mitigating the disease.

  10. Growth factors and cartilage repair.

    NARCIS (Netherlands)

    Berg, W.B. van den; Kraan, P.M. van der; Scharstuhl, A.; Beuningen, H.M. van


    Growth factors are obvious tools to enhance cartilage repair. Understanding of reactivities in normal and arthritic cartilage and potential side effects on other compartments in the joint will help to identify possibilities and limitations. Growth factor responses have been evaluated in normal and

  11. Regulators of articular cartilage homeostasis

    NARCIS (Netherlands)

    Leijten, Jeroen Christianus Hermanus


    Prevention of hypertrophic differentiation is essential for successful cartilage repair strategies. Although this process is essential for longitudinal growth, it also is part of degenerative cartilage diseases such as osteoarthiritis. Moreover, it limits the use of cell types prone to this process

  12. Cartilage oligomeric matrix protein neoepitope in the synovial fluid of horses with acute lameness: A new biomarker for the early stages of osteoarthritis. (United States)

    Skiöldebrand, E; Ekman, S; Mattsson Hultén, L; Svala, E; Björkman, K; Lindahl, A; Lundqvist, A; Önnerfjord, P; Sihlbom, C; Rüetschi, U


    Clinical tools to diagnose the early changes of osteoarthritis (OA) that occur in the articular cartilage are lacking. We sought to identify and quantify a novel cartilage oligomeric matrix protein (COMP) neoepitope in the synovial fluid from the joints of healthy horses and those with different stages of OA. In vitro quantitative proteomics and assay development with application in synovial fluids samples obtained from biobanks of well-characterised horses. Articular cartilage explants were incubated with or without interleukin-1β for 25 days. Media were analysed via quantitative proteomics. Synovial fluid was obtained from either normal joints (n = 15) or joints causing lameness (n = 17) or with structural OA lesions (n = 7) and analysed for concentrations of the COMP neoepitope using a custom-developed inhibition enzyme-linked immunosorbent assay (ELISA). Explants were immunostained with polyclonal antibodies against COMP and the COMP neoepitopes. Semitryptic COMP peptides were identified and quantified in cell culture media from cartilage explants. A rabbit polyclonal antibody was raised against the neoepitope of the N-terminal portion of one COMP fragment (sequence SGPTHEGVC). An inhibition ELISA was developed to quantify the COMP neoepitope in synovial fluid. The mean concentration of the COMP neoepitope significantly increased in the synovial fluid from the joints responsible for acute lameness compared with normal joints and the joints of chronically lame horses and in joints with chronic structural OA. Immunolabelling for the COMP neoepitope revealed a pericellular staining in the interleukin-1β-stimulated explants. The ELISA is based on polyclonal antisera rather than a monoclonal antibody. The increase in the COMP neoepitope in the synovial fluid from horses with acute lameness suggests that this neoepitope has the potential to be a unique candidate biomarker for the early molecular changes in articular cartilage associated with OA. © 2017 The Authors

  13. [Bionic design of articular cartilage]. (United States)

    Qin, Jun; Zhang, Wenguang; Wu, Gang; Wang, Chengtao


    Natural articular cartilage is well known as a special connective tissue with multiple effects and functions, which are important and irreplaceable, in human synovial joints. Biomedical, histological and pathological characteristics of articular cartilage, as well as biomaterial, biomechanical and bio-tribological properties thereof, are summarized from a novel aspect of bionics. Bionic design of articualr cartilage at macro-level and micro-level is carried out from three aspects, i.e., structure, material, and function; and a bionic design model of articular cartilage is set up. As a result, this basic research would be helpful to providing theoretical and practical basis for innovational design and manufacturing of new-style artificial joint with "soft-cushion bearing", and of bionic artificial cartilage.

  14. Articular cartilage bioreactors and bioprocesses. (United States)

    Darling, Eric M; Athanasiou, Kyriacos A


    This review summarizes the major approaches for developing articular cartilage, using bioreactors and mechanical stimuli. Cartilage cells live in an environment heavily influenced by mechanical forces. The development of cartilaginous tissue is dependent on the environment that surrounds it, both in vivo and in vitro. Chondrocytes must be cultured in a way that gives them the proper concentration of nutrients and oxygen while removing wastes. A mechanical force must also be applied during the culturing process to produce a phenotypically correct tissue. Four main types of forces are currently used in cartilage-culturing processes: hydrostatic pressure, direct compression, "high"-shear fluid environments, and "low"-shear fluid environments. All these forces have been integrated into culturing devices that serve as bioreactors for articular cartilage. The strengths and weaknesses of each device and stimulus are explored, as is the future of cartilage bioreactors.

  15. Characterization of an Ex vivo Femoral Head Model Assessed by Markers of Bone and Cartilage Turnover (United States)

    Madsen, Suzi Hoegh; Goettrup, Anne Sofie; Thomsen, Gedske; Christensen, Søren Tvorup; Schultz, Nikolaj; Henriksen, Kim; Bay-Jensen, Anne-Christine; Karsdal, Morten Asser


    Objective: The pathophysiology of osteoarthritis involves the whole joint and is characterized by cartilage degradation and altered subchondral bone turnover. At present, there is a need for biological models that allow investigation of the interactions between the key cellular players in bone/cartilage: osteoblasts, osteoclasts, and chondrocytes. Methods: Femoral heads from 3-, 6-, 9-, and 12-week-old female mice were isolated and cultured for 10 days in serum-free media in the absence or presence of IGF-I (100 nM) (anabolic stimulation) or OSM (10 ng/mL) + TNF-α (20 ng/mL) (catabolic stimulation). Histology on femoral heads before and after culture was performed, and the growth plate size was examined to evaluate the effects on cell metabolism. The conditioned medium was examined for biochemical markers of bone and cartilage degradation/formation. Results: Each age group represented a unique system regarding the interest of bone or cartilage metabolism. Stimulation over 10 days with OSM + TNF-α resulted in depletion of proteoglycans from the cartilage surface in all ages. Furthermore, OSM + TNF-α decreased growth plate size, whereas IGF-I increased the size. Measurements from the conditioned media showed that OSM + TNF-α increased the number of osteoclasts by approximately 80% and induced bone and cartilage degradation by approximately 1200% and approximately 2600%, respectively. Stimulation with IGF-I decreased the osteoclast number and increased cartilage formation by approximately 30%. Conclusion: Biochemical markers and histology together showed that the catabolic stimulation induced degradation and the anabolic stimulation induced formation in the femoral heads. We propose that we have established an explant whole-tissue model for investigating cell-cell interactions, reflecting parts of the processes in the pathogenesis of joint degenerative diseases. PMID:26069585

  16. 77 FR 29914 - Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products (United States)


    ... RIN 0579-AC68 Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products AGENCY... live bovines and products derived from bovines with regard to bovine spongiform encephalopathy. This... products to revise the conditions for the importation of live bovines and products derived from bovines...

  17. Boswellia frereana (frankincense) suppresses cytokine-induced matrix metalloproteinase expression and production of pro-inflammatory molecules in articular cartilage. (United States)

    Blain, Emma J; Ali, Ahmed Y; Duance, Victor C


    The aim of this study was to assess the anti-inflammatory efficacy of Boswellia frereana extracts in an in vitro model of cartilage degeneration and determine its potential as a therapy for treating osteoarthritis. Cartilage degradation was induced in vitro by treating explants with 5 ng/ml interleukin1alpha (IL-1alpha) and 10 ng/ml oncostatin M (OSM) over a 28-day period, in the presence or absence of 100 microg/ml B. frereana. Treatment of IL-1alpha/OSM stimulated cartilage explants with B. frereana inhibited the breakdown of the collagenous matrix. B. frereana reduced MMP9 and MMP13 mRNA levels, inhibited MMP9 expression and activation, and significantly reduced the production of nitrite (stable end product of nitric oxide), prostaglandin E2 and cycloxygenase-2. Epi-lupeol was identified as the principal constituent of B. frereana. This is the first report on the novel anti-inflammatory properties of Boswellia frereana in an in vitro model of cartilage degradation. We have demonstrated that B. frereana prevents collagen degradation, and inhibits the production of pro-inflammatory mediators and MMPs. Due to its efficacy we propose that B. frereana should be examined further as a potential therapeutic agent for treating inflammatory symptoms associated with arthritis. (c) 2009 John Wiley & Sons, Ltd.

  18. Anomalous NMR Relaxation in Cartilage Matrix Components and Native Cartilage: Fractional-Order Models (United States)

    Magin, Richard L.; Li, Weiguo; Velasco, M. Pilar; Trujillo, Juan; Reiter, David A.; Morgenstern, Ashley; Spencer, Richard G.


    We present a fractional-order extension of the Bloch equations to describe anomalous NMR relaxation phenomena (T1 and T2). The model has solutions in the form of Mittag-Leffler and stretched exponential functions that generalize conventional exponential relaxation. Such functions have been shown by others to be useful for describing dielectric and viscoelastic relaxation in complex, heterogeneous materials. Here, we apply these fractional-order T1 and T2 relaxation models to experiments performed at 9.4 and 11.7 Tesla on type I collagen gels, chondroitin sulfate mixtures, and to bovine nasal cartilage (BNC), a largely isotropic and homogeneous form of cartilage. The results show that the fractional-order analysis captures important features of NMR relaxation that are typically described by multi-exponential decay models. We find that the T2 relaxation of BNC can be described in a unique way by a single fractional-order parameter (α), in contrast to the lack of uniqueness of multi-exponential fits in the realistic setting of a finite signal-to-noise ratio. No anomalous behavior of T1 was observed in BNC. In the single-component gels, for T2 measurements, increasing the concentration of the largest components of cartilage matrix, collagen and chondroitin sulfate, results in a decrease in α, reflecting a more restricted aqueous environment. The quality of the curve fits obtained using Mittag-Leffler and stretched exponential functions are in some cases superior to those obtained using mono- and bi-exponential models. In both gels and BNC, α appears to account for microstructural complexity in the setting of an altered distribution of relaxation times. This work suggests the utility of fractional-order models to describe T2 NMR relaxation processes in biological tissues. PMID:21498095

  19. Regional Articular Cartilage Abnormalities of the Hip. (United States)

    Link, Thomas M; Schwaiger, Benedikt J; Zhang, Alan L


    Imaging of hip cartilage is challenging because of its limited thickness and complex geometry and therefore requires advanced MRI techniques. However, cartilage abnormalities are found in a number of disease entities, and their diagnosis may impact patient management. This article will provide pertinent information about the motivation to image hip cartilage, which imaging techniques to use, and how to analyze cartilage; finally, we will discuss disease entities with regional cartilage lesions, including the typical MRI findings. Because the detection and quantification of regional cartilage abnormalities are critical for guidance of operative and nonoperative management of hip disorders, radiologists should be familiar with imaging and analysis techniques for assessing hip cartilage.

  20. Citrus tissue culture employing vegetative explants. (United States)

    Chaturvedi, H C; Singh, S K; Sharma, A K; Agnihotri, S


    Citrus being a number one fruit of the world due to its high nutritional value, huge production of fruits and fruit products, the citrus industry may be considered a major fruit industry. Though citrus orchard area in India is comparable to USA, the produce is far less, while its export is nil. Biotechnology has played an outstanding role in boosting the citrus industry, e.g., in Spain, which is now the biggest exporter of citrus fruit with the application of micrografting. Amongst the fruit trees, perhaps the maximum tissue culture research has been done in citrus during the past four decades, however, the results of practical value are meagre. The shortfalls in citrus tissue culture research and some advancements made in this direction along with bright prospects are highlighted, restricting the review to vegetative explants only. Whilst utilization of nucellar embryogenesis is limited to rootstocks, the other aspects, like, regeneration and proliferation of shoot meristems measuring 200 microm in length--a global breakthrough--of two commercially important scion species, Citrus aurantifolia and C. sinensis and an important rootstock, C. limonia, improvement of micrografting technique, cloning of the same two scion species as well as some Indian rootstock species, employing nodal stem segments of mature trees, of immense practical value have been elaborated. A rare phenomenon of shift in the morphogenetic pattern of differentiation from shoot bud differentiation to embryoid formation occurred during the long-term culture of stem callus of C. grandis. Stem callus-regenerated plants of C. aurantifolia, C. sinensis and C. grandis showed variation in their ploidy levels and a somaclonal variant of C. sinensis, which produced seedless fruits was isolated. Tailoring of rooting in microshoots to a tap root-like system by changing the inorganic salt composition of the rooting medium, resulting in 100% transplant success, and germplasm preservation through normal growth

  1. Antiviral effects of bovine interferons on bovine respiratory tract viruses.


    Fulton, R W; Downing, M M; Cummins, J M


    The antiviral effects of bovine interferons on the replication of bovine respiratory tract viruses were studied. Bovine turbinate monolayer cultures were treated with bovine interferons and challenged with several bovine herpesvirus 1 strains, bovine viral diarrhea virus, parainfluenza type 3 virus, goat respiratory syncytial virus, bovine respiratory syncytial virus, bovine adenovirus type 7, or vesicular stomatitis virus. Treatment with bovine interferons reduced viral yield for each of the...

  2. Origin and function of cartilage stem/progenitor cells in osteoarthritis. (United States)

    Jiang, Yangzi; Tuan, Rocky S


    Articular cartilage is a physiologically non-self-renewing avascular tissue with a singular cell type, the chondrocyte, which functions as the load-bearing surface of the arthrodial joint. Injury to cartilage often progresses spatiotemporally from the articular surface to the subchondral bone, leading to development of degenerative joint diseases such as osteoarthritis (OA). Although lacking intrinsic reparative ability, articular cartilage has been shown to contain a population of stem cells or progenitor cells, similar to those found in many other adult tissues, that are thought to be involved in the maintenance of tissue homeostasis. These so-called cartilage-derived stem/progenitor cells (CSPCs) have been observed in human, equine and bovine articular cartilage, and have been identified, isolated and characterized on the basis of expression of stem-cell-related surface markers, clonogenicity and multilineage differentiation ability. However, the origin and functions of CSPCs are incompletely understood. We review here the current status of CSPC research and discuss the possible origin of these cells, what role they might have in cartilage repair, and their therapeutic potential in OA.

  3. Contrast agent enhanced pQCT of articular cartilage

    Energy Technology Data Exchange (ETDEWEB)

    Kallioniemi, A S [Department of Physics, University of Kuopio, POB 1627, 70211 Kuopio (Finland); Jurvelin, J S [Department of Physics, University of Kuopio, POB 1627, 70211 Kuopio (Finland); Nieminen, M T [Department of Diagnostic Radiology, POB 50, 90029 OYS, Oulu University Hospital, Oulu (Finland); Lammi, M J [Department of Anatomy, Institute of Biomedicine, University of Kuopio, POB 1627, 70211 Kuopio (Finland); Toeyraes, J [Department of Physics, University of Kuopio, POB 1627, 70211 Kuopio (Finland)


    The delayed gadolinium enhanced MRI of cartilage (dGEMRIC) technique is the only non-invasive means to estimate proteoglycan (PG) content in articular cartilage. In dGEMRIC, the anionic paramagnetic contrast agent gadopentetate distributes in inverse relation to negatively charged PGs, leading to a linear relation between T{sub 1,Gd} and spatial PG content in tissue. In the present study, for the first time, contrast agent enhanced peripheral quantitative computed tomography (pQCT) was applied, analogously to dGEMRIC, for the quantitative detection of spatial PG content in cartilage. The suitability of two anionic radiographic contrast agents, gadopentetate and ioxaglate, to detect enzymatically induced PG depletion in articular cartilage was investigated. First, the interrelationships of x-ray absorption, as measured with pQCT, and the contrast agent solution concentration were investigated. Optimal contrast agent concentrations for the following experiments were selected. Second, diffusion rates for both contrast agents were investigated in intact (n = 3) and trypsin-degraded (n 3) bovine patellar cartilage. The contrast agent concentration of the cartilaginous layer was measured prior to and 2-27 h after immersion. Optimal immersion time for the further experiments was selected. Third, the suitability of gadopentetate and ioxaglate enhanced pQCT to detect the enzymatically induced specific PG depletion was investigated by determining the contrast agent concentrations and uronic acid and water contents in digested and intact osteochondral samples (n = 16). After trypsin-induced PG loss (-70%, p < 0.05) the penetration of gadopentetate and ioxaglate increased (p < 0.05) by 34% and 48%, respectively. Gadopentetate and ioxaglate concentrations both showed strong correlation (r = -0.95, r -0.94, p < 0.01, respectively) with the uronic acid content. To conclude, contrast agent enhanced pQCT provides a technique to quantify PG content in normal and experimentally

  4. The slice culture method for following development of tooth germs in explant culture. (United States)

    Alfaqeeh, Sarah A; Tucker, Abigail S


    Explant culture allows manipulation of developing organs at specific time points and is therefore an important method for the developmental biologist. For many organs it is difficult to access developing tissue to allow monitoring during ex vivo culture. The slice culture method allows access to tissue so that morphogenetic movements can be followed and specific cell populations can be targeted for manipulation or lineage tracing. In this paper we describe a method of slice culture that has been very successful for culture of tooth germs in a range of species. The method provides excellent access to the tooth germs, which develop at a similar rate to that observed in vivo, surrounded by the other jaw tissues. This allows tissue interactions between the tooth and surrounding tissue to be monitored. Although this paper concentrates on tooth germs, the same protocol can be applied to follow development of a number of other organs, such as salivary glands, Meckel's cartilage, nasal glands, tongue, and ear.

  5. Correlated response of in vitro regeneration capacity from different source of explants inCucumis melo. (United States)

    Molina, R V; Nuez, F


    The variation among and within different populations of the regeneration ability from leaf, cotyledon and hypocotyl explants has been studied. A control population and two lines selected by their regeneration capacity from leaf explants were used. Significant differences among the plants of the control population,for the organogenic response, were detected. The regeneration capacity varies depending on the type of explant. Selection in order to improve the regeneration frequency from leaf explants also raises the organogenic response in the other explant types. This result suggests the presence of a partial common genetic system controlling the regeneration frequency of the diverse types of explants.

  6. Hypoxia Inhibits Hypertrophic Differentiation and Endochondral Ossification in Explanted Tibiae

    NARCIS (Netherlands)

    Leijten, Jeroen Christianus Hermanus; Moreira Teixeira, Liliana; Landman, Ellie; van Blitterswijk, Clemens; Karperien, Hermanus Bernardus Johannes


    Purpose: Hypertrophic differentiation of growth plate chondrocytes induces angiogenesis which alleviates hypoxia normally present in cartilage. In the current study, we aim to determine whether alleviation of hypoxia is merely a downstream effect of hypertrophic differentiation as previously

  7. A dual flow bioreactor with controlled mechanical stimulation for cartilage tissue engineering. (United States)

    Spitters, Tim W G M; Leijten, Jeroen C H; Deus, Filipe D; Costa, Ines B F; van Apeldoorn, Aart A; van Blitterswijk, Clemens A; Karperien, Marcel


    In cartilage, tissue engineering bioreactors can create a controlled environment to study chondrocyte behavior under mechanical stimulation or produce chondrogenic grafts of clinically relevant size. Here we present a novel bioreactor that combines mechanical stimulation with a two compartment system through which nutrients can be supplied solely by diffusion from opposite sides of a tissue-engineered construct. This design is based on the hypothesis that creating gradients of nutrients, growth factors, and growth factor antagonists can aid in the generation of zonal tissue-engineered cartilage. Computational modeling predicted that the design facilitates the creation of a biologically relevant glucose gradient. This was confirmed by quantitative glucose measurements in cartilage explants. In this system, it is not only possible to create gradients of nutrients, but also of anabolic or catabolic factors. Therefore, the bioreactor design allows control over nutrient supply and mechanical stimulation useful for in vitro generation of cartilage constructs that can be used for the resurfacing of articulated joints or as a model for studying osteoarthritis disease progression.

  8. Strain-rate-dependent non-linear tensile properties of the superficial zone of articular cartilage. (United States)

    Ahsanizadeh, Sahand; Li, LePing


    The tensile properties of articular cartilage play an important role in the compressive behavior and integrity of the tissue. The stress-strain relationship of cartilage in compression was observed previously to depend on the strain-rate. This strain-rate dependence has been thought to originate mainly from fluid pressurization. However, it was not clear to what extent the tensile properties of cartilage contribute to the strain-rate dependence in compressive behavior of cartilage. The aim of the present study was to quantify the strain-rate dependent stress-strain relationship and hysteresis of articular cartilage in tension. Uniaxial tensile tests were performed to examine the strain-rate dependent non-linear tensile properties of the superficial zone of bovine knee cartilage. Tensile specimens were oriented in the fiber direction indicated by the India ink method. Seven strain-rates were used in the measurement ranging from 0.1 to 80%/s, which corresponded to nearly static to impact joint loadings. The experimental data showed substantial strain-rate and strain-magnitude dependent load response: for a given strain-magnitude, the tensile stress could vary by a factor of 1.95 while the modulus by a factor of 1.58 with strain-rate; for a given strain-rate, the modulus at 15% strain could be over four times the initial modulus at no strain. The energy loss in cartilage tension upon unloading exhibited a complex variation with the strain-rate. The strain-rate dependence of cartilage in tension observed from the present study is relatively weaker than that in compression observed previously, but is considerable to contribute to the strain-rate dependent load response in compression.

  9. Role of Insulin-Transferrin-Selenium in Auricular Chondrocyte Proliferation and Engineered Cartilage Formation in Vitro

    Directory of Open Access Journals (Sweden)

    Xia Liu


    Full Text Available The goal of this study is to determine the effects of Insulin-Transferrin-Selenium (ITS on proliferation of auricular chondrocytes and formation of engineered cartilage in vitro. Pig auricular monolayer chondrocytes and chondrocyte pellets were cultured in media containing 1% ITS at different concentrations of fetal bovine serum (FBS, 10%, 6%, 2%, 0%, or 10% FBS alone as a control for four weeks. Parameters including cell proliferation in monolayer, wet weight, collagen type I/II/X (Col I, II, X and glycosaminoglycan (GAG expression, GAG content of pellets and gene expression associated with cartilage formation/dedifferentiation (lost cartilage phenotype/hypertrophy within the chondrocyte pellets were assessed. The results showed that chondrocytes proliferation rates increased when FBS concentrations increased (2%, 6%, 10% FBS in ITS supplemented groups. In addition, 1% ITS plus 10% FBS significantly promoted cell proliferation than 10% FBS alone. No chondrocytes grew in ITS alone medium. 1% ITS plus 10% FBS enhanced cartilage formation in terms of size, wet weight, cartilage specific matrices, and homogeneity, compared to 10% FBS alone group. Furthermore, ITS prevented engineered cartilage from dedifferentiation (i.e., higher index of Col II/Col I mRNA expression and expression of aggrecan and hypertrophy (i.e., lower mRNA expression of Col X and MMP13. In conclusion, our results indicated that ITS efficiently enhanced auricular chondrocytes proliferation, retained chondrogenic phenotypes, and promoted engineered cartilage formation when combined with FBS, which is potentially used as key supplementation in auricular chondrocytes and engineered cartilage culture.

  10. Contrast enhanced cartilage imaging: Comparison of ionic and non-ionic contrast agents

    Energy Technology Data Exchange (ETDEWEB)

    Wiener, Edzard [Department of Radiology, Technical University Munich, Ismaninger Str. 22, D-81675 Munich (Germany)]. E-mail:; Woertler, Klaus [Department of Radiology, Technical University Munich, Ismaninger Str. 22, D-81675 Munich (Germany); Weirich, Gregor [Institute of Pathology, Technical University Munich, Troger Str. 18, D-81675 Munich (Germany); Rummeny, Ernst J. [Department of Radiology, Technical University Munich, Ismaninger Str. 22, D-81675 Munich (Germany); Settles, Marcus [Department of Radiology, Technical University Munich, Ismaninger Str. 22, D-81675 Munich (Germany)


    Our objective was to compare relaxation effects, dynamics and spatial distributions of ionic and non-ionic contrast agents in articular cartilage at concentrations typically used for direct MR arthrography at 1.5 T. Dynamic MR-studies over 11 h were performed in 15 bovine patella specimens. For each of the contrast agents gadopentetate dimeglumine, gadobenate dimeglumine, gadoteridol and mangafodipir trinatrium three patellae were placed in 2.5 mmol/L contrast solution. Simultaneous measurements of T {sub 1} and T {sub 2} were performed every 30 min using a high-spatial-resolution 'MIX'-sequence. T {sub 1}, T {sub 2} and {delta}R {sub 1}, {delta}R {sub 2} profile plots across cartilage thickness were calculated to demonstrate the spatial and temporal distributions. The charge is one of the main factors which controls the amount of the contrast media diffusing into intact cartilage, but independent of the charge, the spatial distribution across cartilage thickness remains highly inhomogeneous even after 11 h of diffusion. The absolute {delta}R {sub 2}-effect in cartilage is at least as large as the {delta}R {sub 1}-effect for all contrast agents. Maximum changes were 5-12 s{sup -1} for {delta}R {sub 1} and 8-15 s{sup -1} for {delta}R {sub 2}. This study indicates that for morphologically intact cartilage only the amount of contrast agents within cartilage is determined by the charge but not the spatial distribution across cartilage thickness. In addition, {delta}R {sub 2} can be considered for quantification of contrast agent concentrations, since it is of the same magnitude and less time consuming to measure than {delta}R {sub 1}.

  11. Explant age, auxin concentrations and media type affect callus ...

    African Journals Online (AJOL)

    The effects of explant age of oil palm (Elaeis huineensis) embryo axes, 15 and 18 weeks after anthesis (WAA), media type (Eeuwens and Murashige and Skoog) supplemented with various concentrations of 2,4-D on callus production employing standard in vitro techniques were investigated. The results of the study showed ...

  12. Usual interstitial pneumonia: histologic study of biopsy and explant specimens. (United States)

    Katzenstein, Anna-Luise A; Zisman, David A; Litzky, Leslie A; Nguyen, Binh T; Kotloff, Robert M


    The pathologic findings in biopsy and subsequent explant specimens from 20 patients with usual interstitial pneumonia (UIP) were reviewed to refine histologic criteria for diagnosis, to identify factors that may confound diagnosis, and to assess the relationship of UIP and nonspecific interstitial pneumonia (NSIP). One case of NSIP was also identified and included for comparison. Surgical biopsies from 15 of the 20 UIP cases were diagnosed as UIP, whereas 5 showed only nondiagnostic changes. An important new observation is that areas resembling nonspecific interstitial pneumonia (NSIP-like areas) are present in the majority of UIP cases in both biopsy and explant specimens, and they are extensive in some. Ten of the 15 UIP biopsies were considered straightforward, with typical patchy interstitial fibrosis, honeycomb change, and fibroblast foci. Five cases were considered difficult because of prominent NSIP-like areas in two, extensive honeycomb change in one, superimposed diffuse alveolar damage in one, and superimposed bronchiolitis obliterans-organizing pneumonia in one. The most helpful feature for diagnosing UIP in difficult cases was the presence of a distinct patchwork appearance to the characteristic uneven or variegated parenchymal involvement along with evidence of architectural derangement. No explant showing UIP was preceded by biopsy findings of NSIP, and the one NSIP case appeared similar at biopsy and explant. NSIP or NSIP-like areas and UIP may reflect different mechanisms of fibrosis related either to different severity of injury or to different injuries.

  13. High frequency plant regeneration from shoot tip explants of ...

    African Journals Online (AJOL)

    A high frequency and rapid regeneration protocol was developed from shoot tip explants of Citrullus colocynthis on Murashige and Skoog (MS) medium supplemented with N6-benzylamino-purine (BAP, 0.5 mg/l) and α-naphthalene acetic acid (NAA, 0.5 mg/l). Highest number of shoots (23.0 ± 0.567) was obtained on MS ...

  14. Adventitious shoot regeneration from in vitro stem explants of ...

    African Journals Online (AJOL)

    An efficient in vitro plant regeneration system from stem explants was established in Phellodendron amurense. Factors influencing shoot regeneration from stems including culture medium type, combinations of plant growth regulators and carbon source in the medium were investigated. Adventitious shoot regeneration was ...

  15. Plant regeneration via somatic embryogenesis from root explants of ...

    African Journals Online (AJOL)

    A system for induction of callus and plant regeneration via somatic embryogenesis from root explants of Hevea brasiliensis Muell. Arg. clone Reyan 87-6-62 was evaluated. The influence of plant growth regulators (PGRs) including 2,4-dichlorophenoxyacetic acid (2,4-D), 6-benzylaminopurine (6-BA) and kinetin (KT) on ...

  16. Somatic embryogenesis from leaf explants of hermaphrodite Carica ...

    African Journals Online (AJOL)

    In culture medium supplemented with 2,4-dichlorophenoxyacetic, FEC overgrew into a yellowish friable mass that fully covered the leaf explants. The somatic embryogenesis process occurred asynchronously, with new globular embryos continuously forming from the FEC. Torpedo and early cotyledonary somatic embryos ...

  17. Direct and indirect plant regeneration from various explants of ...

    African Journals Online (AJOL)



    Apr 18, 2011 ... 1700s from North America. In Turkey, it is an exotic species of poplar and was introduced in the late 1960s from the USA. ..... Balkan H, Tanrıyar H, Calikoglu M, Ogras T, Ozden O, Tulukcu M,. Tank T (1998). Genotype differencies in direct plant regeneration from stem explants of Populus tremula in Turkey.

  18. Influence of explanting season on in vitro multiplication of the ...

    African Journals Online (AJOL)



    Jul 20, 2009 ... and multiple shoot formation was induced between July and September on MS medium supplemented with BAP ... Table 1. Response of apical bud and nodal explants of T. procumbens cultured on MS medium supplemented with different ... through TLC on silica gel-G plates using the method of Holloway.

  19. Effects of genotype, explant type and nutrient medium components ...

    African Journals Online (AJOL)

    The objective of the study was to develop an efficient method for shoot regeneration of canola (Brassica napus L.) and to compare the regeneration capacity of different explants on MS medium with several combinations of plant growth regulators. The experiments showed that the morphogenetical potential of canola ...

  20. Explants, hormones and sucrose influence in vitro shoot ...

    African Journals Online (AJOL)

    Zantedeschia is an important and rapidly expanding cut flower in Kenya today. A protocol for in vitro shoot regeneration of Zantedeschia, using tuber, leaf and shoot primordium explants on Murashige and Skoog (MS) (1962) basal salts, supplemented with 6-benzyalamino purine (BAP) or Kinetin is described. Of the four ...

  1. Effect of plant growth regulators, explants type and efficient plantlet ...

    African Journals Online (AJOL)



    Dec 5, 2011 ... nodal segment and shoot tip) and different concentrations of plant growth regulators. Callus formation and shoot differentiation ... Key words: Naringi crenulata, callus, regeneration, leaf explants, peroxidase, total soluble protein. INTRODUCTION ... primary health care (Mousumi et al., 2007). The success.

  2. The effect of plant growth regulators, explants and cultivars on ...

    African Journals Online (AJOL)

    Spinach (Spinacia oleracea L.) is an important vegetable crop of which dioecy in nature has made cultivar improvement difficult using traditional breeding methods; therefore, production of high amount of disease free spinach is critical. To achieve the best explants and media for spinach tissue culture, the effects of two ...

  3. Simple, effective and economical explant-surface sterilization ...

    African Journals Online (AJOL)



    Oct 19, 2009 ... Three different surface sterilization methods were evaluated using seeds and excised embryos of cowpea, rice and sorghum as explants: Method 1: Ethanol alone in concentrations of 95, 90, 85 and 70% at different time intervals and observed at different days. Method 2: Locally produced bleaching.

  4. AAV Serotype Testing on Cultured Human Donor Retinal Explants

    NARCIS (Netherlands)

    Buck, Thilo M; Pellissier, Lucie P; Vos, Rogier M; van Dijk, Elon H C; Boon, Camiel J F; Wijnholds, J.


    This protocol details on a screening method for infectivity and tropism of different serotypes of adeno-associated viruses (AAVs) on human retinal explants with cell-type specific or ubiquitous green fluorescent protein (GFP) expression vectors. Eyes from deceased adult human donors are enucleated

  5. Somatic embryogenesis and plant regeneration from leaf explants of ...

    African Journals Online (AJOL)

    An attempt was made to study the somatic embryogenesis and plant regeneration from the in vitro leaf explants of Rumex vesicarius L. a renowned medicinal plant, which belongs to polygonaceae family. Effective in vitro regeneration of R. vesicarius was achieved via young leaf derived somatic embryo cultures.

  6. Efficient plant regeneration from leaf explants of Solanum americanum

    African Journals Online (AJOL)

    A very efficient system for direct plant regeneration from in vitro–derived leaf explants of Solanum americanum was developed. S. americanum is a tropical plant with important medical properties. The in vitro procedure that was established consists of (i) induction of shoots from leaf tissue, (ii) elongation of shoots, and (iii) ...

  7. Regeneration potential of seedling explants of chilli ( Capsicum ...

    African Journals Online (AJOL)

    A study was conducted with hypocotyl, cotyledon and shoot tip of chilli as explants for regeneration on MS medium supplemented with different concentrations and combinations of auxins and cytokinins. Regeneration potential was determined by two ways. One is regeneration of shoot via callus formation from hypocotyls ...

  8. High-frequency shoot regeneration of nodal explants from ...

    African Journals Online (AJOL)

    This paper describes the shoot regeneration of nodal segments from a medicinal plant, Tetrastigma hemsleyanum Diels et Gilg (Vitaceae). The highest number of shoots (7.27 shoots per explant) was observed in MS medium supplemented with 4 mg/l BA after six weeks of inoculation. 2 mg/l BA in combination with 0.1 mg/l ...

  9. callus induction from epicotyl and hypocotyl explants of

    African Journals Online (AJOL)

    Department of Botany, Obafemi Awolowo University, Ile-Ife. Nigeria. (Submitted: 31 May 2004; Accepted: 31 October 2004). Abstract. Epicotyl and hypocotyl explants of Parkia biglobosa (Locust bean) were cultured in vitro to investigate their callogenic capacity. Established cultures were obtained and maintained on MS ...

  10. High-frequency shoot regeneration of nodal explants from ...

    African Journals Online (AJOL)



    Jun 29, 2011 ... develop a rapid and efficient in vitro multiplication and regeneration system using nodal explants. MATERIALS AND METHODS. Plant material and initiation of in vitro shoot cultures. Young in vivo shoots with six to eight nodes of T. hemsleyanum were collected from wild population in Zhejiang Province, ...

  11. Regeneration of plantlets from nodal and shoot tip explants of ...

    African Journals Online (AJOL)

    Anoectochilus elatus Lindley is an endangered terrestrial orchid. A procedure for the regeneration of complete plantlets of A. elatus Lindley through node and shoot tip explants resulted directly in shoots when cultured on a full strength Murashige and Skoog (1962) medium supplemented with cytokinins at different ...

  12. In vitro propagation of Alstroemeria using rhizome explants derived ...

    African Journals Online (AJOL)



    Sep 19, 2007 ... For surface sterilization of rhizome, those obtained from pot plants, (3 cm in length) were excised and washed thoroughly under running tap water for 10 min. Thereafter, the rhizome explants sterilized by immersion for 35 min in 40% (v/v) commercial bleach (containing 5.54% sodium hypochlorite) and.

  13. In vitro shoot regeneration from preconditioned explants of chickpea ...

    African Journals Online (AJOL)



    Mar 14, 2011 ... cultivated edible legumes used as vegetable. Chickpeas are grown in the .... the culture medium had negative effect on shoot length and resulted in smaller shoots on both ... Effect of pre-conditioning on number of shoots and shoot length of mature embryo and embryonic axis explant of chickpea cv. Gokce.

  14. Adventitious shoot regeneration from cultured leaf explants of Petunia

    African Journals Online (AJOL)



    Jun 26, 2012 ... the first bedding plant; in addition Petunia is the most ... leaf explants of Petunia using MS medium containing. 2 mgL .... suggested that differences in BA uptake and metabolism ... cation might have an inhibitory effect on shoot.

  15. Design and Fabrication of Anatomical Bioreactor Systems Containing Alginate Scaffolds for Cartilage Tissue Engineering


    Gharravi, Anneh Mohammad; Orazizadeh, Mahmoud; Ansari-Asl, Karim; Banoni, Salem; Izadi, Sina; Hashemitabar, Mahmoud


    The aim of the present study was to develop a tissue-engineering approach through alginate gel molding to mimic cartilage tissue in a three-dimensional culture system. The perfusion biomimetic bioreactor was designed to mimic natural joint. The shear stresses exerting on the bioreactor chamber were calculated by Computational Fluid Dynamic (CFD). Several alginate/bovine chondrocyte constructs were prepared, and were cultured in the bioreactor. Histochemical and immunohistochemical staining me...

  16. Light Absorptive Properties of Articular Cartilage, ECM Molecules, Synovial Fluid, and Photoinitiators as Potential Barriers to Light-Initiated Polymer Scaffolding Procedures. (United States)

    Finch, Anthony J; Benson, Jamie M; Donnelly, Patrick E; Torzilli, Peter A


    Objective Many in vivo procedures to repair chondral defects use ultraviolet (UV)-photoinitiated in situ polymerization within the cartilage matrix. Chemical species that absorb UV light might reduce the effectiveness of these procedures by acting as light absorption barriers. This study evaluated whether any of the individual native biochemical components in cartilage and synovial fluid interfered with the absorption of light by common scaffolding photosensitizers. Materials UV-visible spectroscopy was performed on each major component of cartilage in solution, on bovine synovial fluid, and on four photosensitizers, riboflavin, Irgacure 2959, quinine, and riboflavin-5'-phosphate. Molar extinction and absorption coefficients were calculated at wavelengths of maximum absorbance and 365 nm. Intact articular cartilage was also examined. Results The individual major biochemical components of cartilage, Irgacure 2959, and quinine did not exhibit a significant absorption at 365 nm. Riboflavin and riboflavin-5'-phosphate were more effectual light absorbers at 365 nm, compared with the individual native species. Intact cartilage absorbed a significantly greater amount of UV light in comparison with the native species. Conclusion Our results indicate that none of the individual native species in cartilage will interfere with the absorption of UV light at 365 nm by these commonly used photoinitiators. Intact cartilage slices exhibited significant light absorption at 365 nm, while also having distinct absorbance peaks at wavelengths less than 300 nm. Determining the UV absorptive properties of the biomolecules native to articular cartilage and synovial fluid will aid in optimizing scaffolding procedures to ensure sufficient scaffold polymerization at a minimum UV intensity.

  17. The Removal of Hydrogel Explants: An Analysis of 467 Consecutive Cases

    NARCIS (Netherlands)

    Crama, N.; Klevering, B.J.


    PURPOSE: To describe the complications associated with hydrogel explants and to describe the indications, surgical technique, and risks involved in the removal of a hydrogel explant. DESIGN: Single-center, retrospective interventional case series. PARTICIPANTS: Patients who underwent surgical

  18. Particulate cartilage under bioreactor-induced compression and shear. (United States)

    Wang, Ning; Grad, Sibylle; Stoddart, Martin J; Niemeyer, Philipp; Reising, Kilian; Schmal, Hagen; Südkamp, Norbert P; Alini, Mauro; Salzmann, Gian M


    Our aim was to explore the effect of varying in vitro culture conditions on general chondrogenesis of minced cartilage (MC) fragments. Minced, fibrin-associated, bovine articular cartilage fragments were cultured in vitro within polyurethane scaffold rings. Constructs were maintained either free swelling for two or four weeks (control), underwent direct mechanical knee-joint-specific bioreactor-induced dynamic compression and shear, or they were maintained free swelling for two weeks followed by two weeks of bioreactor stimulation. Samples were collected for glycosaminoglycan (GAG)/DNA quantification; collagen type I, collagen type II, aggrecan, cartilage oligomeric matrix protein (COMP), proteoglycan-4 (PRG-4) messenger RNA (mRNA) analysis; histology and immunohistochemistry. Cellular outgrowth and neomatrix formation was successfully accomplished among all groups. GAG/DNA and collagen type I mRNA were not different between groups; chondrogenic genes collagen type II, aggrecan and COMP revealed a significant downregulation among free-swelling constructs over time (week two through week four). Mechanical loading was able to maintain chondrogenic expression with significantly stronger expression at long-term time points (four weeks) in comparison with four-week control. Histology and immunohistochemistry revealed that bioreactor culture induced stronger cellular outgrowth than free-swelling constructs. However, weaker collagen type II and aggrecan expression with an increased collagen type I expression was noted among this outgrowth neotissue. The method of MC culture is feasible under in vitro free-swelling and dynamic loading conditions, simulating in vivo posttransplantation. Mechanical stimulation significantly provokes cellular outgrowth and long-term chondrogenic maturation at the mRNA level, whereas histology depicts immature neotissue where typical cartilage matrix is expected.

  19. Morphologic differences observed by scanning electron microscopy according to the reason for pseudophakic IOL explantation

    DEFF Research Database (Denmark)

    Fernandez-Buenaga, Roberto; Alio, Jorge L.; Ramirez, Jose M.


    Purpose To compare variations in surface morphology, as studied by scanning electron microscopy (SEM), of explanted intraocular lenses (IOLs) concerning the cause leading to the explantation surgery. Methods In this prospective multicenter study, explanted IOLs were analyzed by SEM and energy-dis...

  20. Effect of storage media and time on fin explants culture in the ...

    African Journals Online (AJOL)

    On the outgrowth of cells, fin explants stored for seven days before culturing showed significantly higher growth (P<0.05) as observed on the fin explants stored in Dulbecco's modified Eagles medium (DMEM, 84.44%), phosphate buffered saline (PBS, 62.42%) and in control/fresh fin explants (100%), compared with the ...

  1. Absorção de macronutrientes por explantes de bananeira in vitro Macronutrient absorption by banana explants in vitro

    Directory of Open Access Journals (Sweden)

    Josefa Diva Nogueira Diniz


    Full Text Available Com o objetivo de estudar a absorção de macronutrientes (N, P, K, Ca, Mg e S em explantes de bananeira cv. Prata Anã, foram utilizados explantes de plantas estabelecidas in vitro, inoculados em meio básico de Murashige & Skoog (1962 contendo sacarose (30 g/L, e BAP (3,5 mg/L com sete tratamentos, representados pelos períodos de 0, 10, 20, 30, 40, 50 e 60 dias de cultivo e três repetições. As quantidades de macronutrientes totais absorvidas pelos explantes seguiram a ordem: K > N > Ca > ou = P > Mg @ S. O P foi o nutriente absorvido mais rapidamente pelos explantes, com 75% extraído do meio de cultivo nos primeiros 30 dias, cessando sua absorção aos 50 dias, restando ainda 9% no meio de cultivo. A absorção do S cessou também aos 50 dias, quando 66% deste nutriente ainda permanecia no meio de cultivo. Este resultado sugere haver uma relação, quanto à absorção, entre esses dois nutrientes. As maiores taxas de absorção de todos os nutrientes foram verificadas nos primeiros 20 dias. O rizoma, o pseudocaule e as folhas, se diferenciaram quanto à concentração e extração ou acúmulo de nutrientes.The absorption of the nutrients (N, P, K, Ca, Mg and S by banana (Musa sp. cv. Prata Anã explants on the basic medium of Murashige & Skoog (1962 supplemented with sucrose (30 g/L and BAP (3.5 mg/L were evaluated at 0, 10, 20, 30, 40, 50 and 60 days after inoculation. The seven treatments were arranged on a completely randomized design with three replicates. The sequence of nutrient absorption by the explants was K > N > Ca > or = P > Mg @ S. The P was the nutrient with the fastest absorption rate and at the 30th day the explants had already absorbed 75% of the P from the medium. The P absorption stopped by the 50th day. The S absorption stopped at the 50th day with 66% of it remaining in the medium. The results suggested a close relationship between these two nutrients. The highest rates of nutrient absorption were observed during the

  2. Cyclin-Dependent Kinase 9 inhibition protects cartilage from the catabolic effects of pro-inflammatory cytokines (United States)

    Yik, Jasper H. N.; Hu, Zi’ang; Kumari, Ratna; Christiansen, Blaine A.; Haudenschild, Dominik R.


    Objective CDK9 controls the activation of primary inflammatory response genes. We determined whether CDK9 inhibition protects cartilage from the catabolic effects of pro-inflammatory cytokines. Methods Human chondrocytes were challenged with different pro-inflammatory stimuli (IL-1β, lipopolysaccharides, and TNFα), in the presence or absence of the CDK9 inhibitor Flavopiridol, or siRNA. The mRNA expression of inflammatory mediators, catabolic, and anabolic genes were determined by real-time PCR. Cartilage explants were incubated with IL-1β, with or without Flavopiridol, for 6 days. Cartilage matrix degradation was assessed by the release of glycosaminoglycan (GAG) and cleaved Type II collagen (Col2a) peptides. Results CDK9 inhibition by Flavopiridol, or knockdown by siRNA, effectively suppressed iNOS mRNA induction by all three pro-inflammatory stimuli. Results from NFkB-targets PCR array showed that Flavopiridol suppressed the induction of a broad range of inflammatory mediator genes (59 out of 67 tested) by IL-1β. CDK9 inhibition also suppressed induction of catabolic genes MMP 1, 3, 9, 13, and ADAMTS4, 5; but did not affect the basal expression of anabolic genes such as Col2a, aggrecan, and COMP, and housekeeping genes. Flavopiridol had no apparent short-term cytotoxicity as assessed by glucose-6-phosphate dehydrogenase activity. Finally, in IL-1β-treated cartilage explants, Flavopiridol reduced the release of matrix degradation products GAG and cleaved Col2a peptides, but did not affect long-term chondrocyte viability. Conclusion CDK9 activity is required for the primary inflammatory response in chondrocytes. Flavopiridol suppresses the induction of inflammatory mediators and catabolic genes to protect cartilage from the deleterious effects of pro-inflammatory cytokines, without impacting cell viability and functions. PMID:24470357

  3. Diffusion of Paramagnetically Labeled Proteins in Cartilage: Enhancement of the 1-D NMR Imaging Technique (United States)

    Foy, Brent D.; Blake, Joseph


    Quantifying the diffusive transport of large molecules in avascular cartilage tissue is important both for planning potential pharamacological treatments and for gaining insight into the molecular-scale structure of cartilage. In this work, the diffusion coefficients of gadolinium-DTPA and Gd-labeled versions of four proteins-lysozyme, trypsinogen, ovalbumin, and bovine serum albumin (BSA) with molecular weights of 14,300, 24,000, 45,000, and 67,000, respectively-have been measured in healthy and degraded calf cartilage. The experimental technique relies on the effect of the paramagnetic on the relaxation properties of the surrounding water, combined with the time course of a 1-dimensional spatial profile of the water signal in the cartilage sample. The enhanced technique presented here does not require a prior measurement of the relaxivity of the paramagnetic compound in the sample of interest. The data are expressed as the ratio of the diffusion coefficient of a compound in cartilage to its diffusion coefficient in water. For healthy cartilage, this ratio was 0.34 ± 0.07 for Gd-DTPA, the smallest compound, and fell to 0.3 ± 0.1 for Gd-lysozyme, 0.08 ± 0.04 for Gd-trypsinogen, and 0.07 ± 0.04 for Gd-ovalbumin. Gd-BSA did not appear to enter healthy cartilage tissue beyond a surface layer. After the cartilage had been degraded by 24-h trypsinization, these ratios were 0.60 ± 0.03 for Gd-DTPA, 0.40 ± 0.08 for Gd-lysozyme, 0.42 ± 0.09 for Gd-trypsinogen, 0.16 ± 0.14 for Gd-ovalbumin, and 0.11 ± 0.05 for Gd-BSA. Thus, degradation of the cartilage led to increases in the diffusion coefficient of up to fivefold for the Gd-labeled proteins. These basic transport parameters yield insights on the nature of pore sizes and chemical-matrix interactions in the cartilage tissue and may prove diagnostically useful for identifying the degree and nature of damage to cartilage.


    African Journals Online (AJOL)

    BACKGROUND: Tuberculosis is caused by a number of Mycobacterium species, of which Mycobacterium bovis, causing 'bovine tuberculosis' is ... KEY WORDS: Mycobacterium bovis, Zoonosis, Holeta, Ethiopia causing 'bovine tuberculosis ..... isolation of infected animals in which communal grazing and watering practiced.

  5. Transcriptomic profiling of cartilage ageing

    Directory of Open Access Journals (Sweden)

    Mandy Jayne Peffers


    Full Text Available The musculoskeletal system is severely affected by the ageing process, with many tissues undergoing changes that lead to loss of function and frailty. Articular cartilage is susceptible to age related diseases, such as osteoarthritis. Applying RNA-Seq to young and old equine cartilage, we identified an over-representation of genes with reduced expression relating to extracellular matrix, degradative proteases, matrix synthetic enzymes, cytokines and growth factors in cartilage from older donors. Here we describe the contents and quality controls in detail for the gene expression and related results published by Peffers and colleagues in Arthritis Research and Therapy 2013 associated with the data uploaded to ArrayExpress (E-MTAB-1386.

  6. Microwave treatment of xenogeneic cartilage transplants

    NARCIS (Netherlands)

    Visser, C. E.; Boon, M. E.; Visser, P. E.; Kok, L. P.


    Human rib cartilage was irradiated with microwaves according to six different methods and transplanted into rabbits. Untreated rib cartilage preserved in Cialit served as a control. After 12 and 40 wk of implantation, the microscopic appearance of these xenogeneic cartilage transplants was given a

  7. An enzyme-sensitive PEG hydrogel based on aggrecan catabolism for cartilage tissue engineering. (United States)

    Skaalure, Stacey C; Chu, Stanley; Bryant, Stephanie J


    A new cartilage-specific degradable hydrogel based on photoclickable thiol-ene poly(ethylene glycol) (PEG) hydrogels is presented. The hydrogel crosslinks are composed of the peptide, CRDTEGE-ARGSVIDRC, derived from the aggrecanase-cleavable site in aggrecan. This new hydrogel is evaluated for use in cartilage tissue engineering by encapsulating bovine chondrocytes from different cell sources (skeletally immature (juvenile) and mature (adult) donors and adult cells stimulated with proinflammatory lipopolysaccharide (LPS)) and culturing for 12 weeks. Regardless of cell source, a twofold decrease in compressive modulus is observed by 12 weeks, but without significant hydrogel swelling indicating limited bulk degradation. For juvenile cells, a connected matrix rich in aggrecan and collagen II, but minimal collagens I and X is observed. For adult cells, less matrix, but similar quality, is deposited. Aggrecanase activity is elevated, although without accelerating bulk hydrogel degradation. LPS further decreases matrix production, but does not affect aggrecanase activity. In contrast, matrix deposition in the nondegradable hydrogels consists of aggrecan and collagens I, II, and X, indicative of hypertrophic cartilage. Lastly, no inflammatory response in chondrocytes is observed by the aggrecanase-sensitive hydrogels. Overall, it is demonstrated that this new aggrecanase-sensitive hydrogel, which is degradable by chondrocytes and promotes a hyaline-like engineered cartilage, is promising for cartilage regeneration. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Wavy-walled bioreactor supports increased cell proliferation and matrix deposition in engineered cartilage constructs. (United States)

    Bueno, Ericka M; Bilgen, Bahar; Barabino, Gilda A


    Hydrodynamic forces in bioreactors can decisively influence extracellular matrix deposition in engineered cartilage constructs. In the present study, the reduced fluid shear, high-axial mixing environment provided by a wavy-walled bioreactor was exploited in the cultivation of cartilage constructs using polyglycolic acid scaffolds seeded with bovine articular chondrocytes. Increased growth as defined by weight, cell proliferation and extracellular matrix deposition was observed in cartilage constructs from wavy-walled bioreactors in comparison with those from spinner flasks cultured under the same conditions. The wet weight composition of 4-week constructs from the wavy-walled bioreactor was similar to that of spinner flask constructs, but the former were 60% heavier due to equally higher incorporation of extracellular matrix and 30% higher cell population. It is most likely that increased construct matrix incorporation was a result of increased mitotic activity of chondrocytes cultured in the environment of the wavy-walled bioreactor. A layer of elongated cells embedded in type I collagen formed at the periphery of wavy-walled bioreactor and spinner flask constructs, possibly as a response to local shear forces. On the basis of the robustness and reproducibility of the extracellular matrix composition of cartilage constructs, the wavy-walled bioreactor demonstrated promise as an experimental cartilage tissue-engineering vessel. Increased construct growth in the wavy-walled bioreactor may lead to enhanced mechanical properties and expedited in vitro cultivation.

  9. Valgus Stress Radiographs Predict Lateral-Compartment Cartilage Thickness but Not Cartilage Degeneration in Varus Osteoarthritis. (United States)

    Waldstein, Wenzel; Schmidt-Braekling, Tom; Perino, Giorgio; Kasparek, Maximilian F; Windhager, Reinhard; Boettner, Friedrich


    Intact cartilage in the lateral compartment is an important requirement for medial unicompartmental knee arthroplasty. This study sought to determine how measurements of joint space width in the lateral compartment on valgus stress radiographs compare to cartilage thickness as measured with a precise needle test, and whether cartilage thickness is a predictor of cartilage degeneration. A consecutive series of 100 knees undergoing total knee arthroplasty for end-stage varus osteoarthritis was studied. Twenty-eight knees were retrospectively excluded because not all data were available, leaving 72 knees (61 patients; mean age, 67 years [49-87]). On calibrated valgus stress radiographs, lateral-compartment joint space width was measured. During surgery, osteochondral samples of the distal lateral femur and the lateral tibia plateau were harvested. Cartilage thickness and histology were assessed. Cartilage thickness of tibia and femur was defined as lateral-compartment cartilage thickness. Lateral-compartment joint space width on valgus stress radiographs and lateral-compartment cartilage thickness correlated well (rs = 0.671, P cartilage histology according to the osteoarthritis cartilage histopathology assessment system, and cartilage thickness on the lateral tibia plateau (rs = -0.060, P = .614) and cartilage thickness on the distal lateral femur (rs = -0.128, P = .282) was observed. Valgus stress radiographs can assess combined cartilage thickness in the lateral compartment of the knee. Cartilage thickness, however, is a poor predictor of cartilage degeneration. Copyright © 2016 Elsevier Inc. All rights reserved.

  10. Laboratory analyses of two explanted hydrophobic acrylic intraocular lenses

    Directory of Open Access Journals (Sweden)

    Yunhai Dai


    Full Text Available Two three-piece hydrophobic acrylic intraocular lenses (IOLs were explanted from two patients at 7 and 9 years, respectively, after implantation, because of poor fundus visualisation and/or a clinically significant decrease in visual acuity related to their opacified IOLs. In addition to light microscopy, scanning electron microscopy and energy dispersive X-ray spectroscopy, confocal laser scanning microscopy was used for the first time to observe the explanted IOLs. The clinical aspect seemed to correspond to the phenomenon of surface light scattering, while laboratory analyses showed dense glistenings in the central layer of the IOL optic, which had no change next to the surface. Further studies on these phenomena are needed.

  11. Age-related alterations in TGF beta signaling as a causal factor of cartilage degeneration in osteoarthritis. (United States)

    van der Kraan, Peter M


    Age is the most important risk factor for primary osteoarthritis (OA). Members of the TGF-β superfamily play a crucial role in chondrocyte differentiation and maintenance of healthy articular cartilage. We have investigated whether age-related changes in TGF-β superfamily signaling components play a role in the relationship between OA-related cartilage degradation and aging. The relationship between age, OA and TGF-β superfamily signaling was studied using murine experimental OA models, aging mice, bovine articular cartilage and human OA cartilage. The effects of TGF-β on cartilage homeostasis was studied with immunohistochemistry, Q-RT-PCR and signaling pathway analysis with Western blotting and the application of specific TGF-β inhibitors. We have found that TGF-β loses its protective effects in old cartilage. Moreover, we found that on chondrocytes, TGF-β not only signals via the canonical type I receptor ALK5 (TGFBR1) but also via the ALK1 (ACVRL1) receptor. Remarkably, signaling via ALK5 (Smad2/3 route) results in protective while ALK1 signaling (Smad1/5/8 route) results in deleterious responses in articular chondrocytes. In cartilage of aging mice it was detected that the ALK1/ALK5 ratio is significantly increased, favoring TGF-β signaling via the Smad1/5/8 route, inducing changes in chondrocyte differentiation and matrix metalloproteinase-13 (MMP-13) expression. Moreover, human OA cartilage showed a significant correlation between ALK1 and MMP-13 expression. Since in mice aging and OA in often goes hand in hand, we also analyzed age-related expression of TGF-β superfamily related signaling molecules in healthy bovine cartilage in an age range from 6 months to 14 years. In this cohort of aging cartilage, we found that mainly signaling receptors determining the Smad2/3 pathway were decreased with age while Smad1/5/8-related signaling molecules did not alter, confirming our findings in aging mice. Old cartilage appears to be less protected by TGF

  12. Callus induction from epicotyl and hypocotyl explants of Parkia ...

    African Journals Online (AJOL)

    Established cultures were obtained and maintained on MS medium supplemented with either 2,4-D or NAA, each of concentration range of 0.4 – 1.0 mg/L. In general, while higher concentrations of 0.8 and 1.0 mg/L NAA appeared to have favoured callus production from these explants, the same concentrations of 2, 4-D ...

  13. Diverse response of tomato fruit explants to high temperature

    Directory of Open Access Journals (Sweden)

    Zofia Starck


    Full Text Available Tomato explants (fruit with a pedicel and a piece of peduncle, with fruit growth stimulated by treating the flowers with NOA + GA3 (NG-series were used as a model system for studying the effect of high temperature on C-sucrose uptake, its distribution and Ca retranslocation. Two cultivars with contrasting responses to high temperature were compared. In sensitive cv. Roma heat stress during 22h (40oC for 10h and 30oC for 12h, drastically depressed the uptake of 14C-sucrose coinciding with diminished fruit 14C-supply. It also decreased the specific activity of soluble acid invertase and the calcium content. All these strong negative responses to high temperature were markedly reduced in the NG-treated series involving remobilization of Ca to the fruits and a higher stability of the invertase activity. This indicates the indirect role of flower treatment with NG in addaptation to heat stress. In tolerant cv. Robin even higher temperatures (42oC for 10h and 34oC for 12h were not stressful. They did not affect the 14C-sucrose uptake and stimulated 14C-supply to the fruit. Increased specific activity of acid invertase and a higher calcium content were also recorded but only in the control explants. In contrast to cv. Roma elevated temperature was slightly stressful for cv. Robin explants of NG-series. The differences in response of both cultivar explants to elevated temperature, based on unequal fruit supply with 14C-sucrose, seem to be causaly connected with two factors: the invertase activity being more or less sensitive to the heat stress, the ability to translocate Ca to the heated fruits.

  14. Regeneration potential of seedling explants of chilli (Capsicum ...

    African Journals Online (AJOL)



    Feb 18, 2009 ... gation techniques for mass multiplication have become imperative. Conventional ..... the shooting media. For direct shoot regeneration, shoot tip explants were cultured onto MS medium supplemented with 5.0 mg L-1. BAP + 3.0 mg L-1 AgN03, 5.0 mg L-1 BAP + 0.1 mg L-1. NAA. It might be concluded from ...

  15. Shoot Regeneration from Leaf Explants of Withania somnifera (L. Dunal

    Directory of Open Access Journals (Sweden)

    Aruna Girish JOSHI


    Full Text Available Regeneration from leaf explants of Withania somnifera (L. for mass propagation was studied on Murashige and Skoog�s medium supplemented with Kinetin (Kn and 6-benzylaminopurine (BAP alone or in combination. Shoot buds were induced from the midrib on the abaxial side in presence of Kn and BAP (4 �M. These shoot buds developed into shoots on the same medium. Rooting of these shoots was achieved in 0.5 �M of IBA.

  16. Shoot Regeneration from Leaf Explants of Withania somnifera (L.) Dunal


    Aruna Girish JOSHI; Mainavati A. PADHYA


    Regeneration from leaf explants of Withania somnifera (L.) for mass propagation was studied on Murashige and Skoog�s medium supplemented with Kinetin (Kn) and 6-benzylaminopurine (BAP) alone or in combination. Shoot buds were induced from the midrib on the abaxial side in presence of Kn and BAP (4 �M). These shoot buds developed into shoots on the same medium. Rooting of these shoots was achieved in 0.5 �M of IBA.

  17. Shoot Regeneration from Leaf Explants of Withania somnifera (L. Dunal

    Directory of Open Access Journals (Sweden)

    Aruna Girish JOSHI


    Full Text Available Regeneration from leaf explants of Withania somnifera (L. for mass propagation was studied on Murashige and Skoogs medium supplemented with Kinetin (Kn and 6-benzylaminopurine (BAP alone or in combination. Shoot buds were induced from the midrib on the abaxial side in presence of Kn and BAP (4 M. These shoot buds developed into shoots on the same medium. Rooting of these shoots was achieved in 0.5 M of IBA.

  18. callus induction and proliferation from cotyledon explants in ...

    African Journals Online (AJOL)



    amélioration de la qualité des graines est fortement souhaitable. Toutefois, cela exige au préalable l'existence de systèmes efficaces de régénération de plantes. L'induction de cals à partir des explants cotylédon a été étudiée chez ...

  19. An efficient plant regeneration protocol from petiole explants of ...

    African Journals Online (AJOL)

    The highest percentage of shoot buds induction (64.0%) was observed on MS medium supplemented with 0.52 mgL-1 TDZ with organic additives; adenine sulphate (50 mgL-1) + glutamine (100 mgL-1) + L-arginine (25 mgL-1) + citric acid (0.0025%) + ascorbic acid (0.005%). A maximum of six shoots per explant were ...

  20. In vitro regeneration from internodal explants of bitter melon ...

    African Journals Online (AJOL)

    Adventitious shoots were produced from organogenic callus when it was transferred to MS medium supplemented with 4.0 μM TDZ, 1.5 μM 2,4-D and 0.07 mM L-glutamine with shoot induction frequency of 96.5% and regeneration of adventitious shoots from callus (48 shoots per explant). Shoot proliferation occurred when ...

  1. Tropic responses of potato single-node explant cultures


    Vinterhalter D.; Vinterhalter Branka


    A special in-vitro protocol was elaborated enabling the production of potato single-node explant plantlets that can be used as objects for tropic studies. In light-grown plantlets, achievement of a full (90°) phototropic (PT) curvature required 75 to 120 min of continuous unilateral blue light irradiation or 120-135 min of gravitropic stimulation (GT). Time-lapse photography revealed that the curves describing PT and GT bending have a sigmoid shape. Continuous BL irradiation was necessa...

  2. Materials characterization of explanted polypropylene hernia mesh: Patient factor correlation. (United States)

    Smith, Sarah E; Cozad, Matthew J; Grant, David A; Ramshaw, Bruce J; Grant, Sheila A


    This study quantitatively assessed polypropylene (PP) hernia mesh degradation and its correlation with patient factors including body mass index, tobacco use, and diabetes status with the goal of improving hernia repair outcomes through patient-matched mesh. Thirty PP hernia mesh explants were subjected to a tissue removal process followed by assessment of their in vivo degradation using Fourier transform infrared, differential scanning calorimetry, and thermogravimetric analysis analyses. Results were then analyzed with respect to patient factors (body mass index, tobacco use, and diabetes status) to determine their influence on in vivo hernia mesh oxidation and degradation. Twenty of the explants show significant surface oxidation. Tobacco use exhibits a positive correlation with modulated differential scanning calorimetry melt temperature and exhibits significantly lower TGA decomposition temperatures than non-/past users. Chemical and thermal characterization of the explanted meshes indicate measurable degradation while in vivo regardless of the patient population; however, tobacco use is correlated with less oxidation and degradation of the polymeric mesh possibly due to a reduced inflammatory response. © The Author(s) 2015.

  3. Comparative experimental infection of Listeria monocytogenes and Listeria ivanovii in bovine trophoblasts. (United States)

    Rocha, Cláudia E; Mol, Juliana P S; Garcia, Luize N N; Costa, Luciana F; Santos, Renato L; Paixão, Tatiane A


    Listeria monocytogenes is a Gram-positive, facultative intracellular and invasive bacterium that has tropism to the placenta, and causes fetal morbidity and mortality in several mammalian species. While infection with L. monocytogenes and L. ivanovii are known as important causes of abortion and reproductive failure in cattle, the pathogenesis of maternal-fetal listeriosis in this species is poorly known. This study used the bovine chorioallantoic membrane explant model to investigate the kinetics of L. monocytogenes, L. ivanovii, and L. innocua infections in bovine trophoblastic cells for up to 8 h post infection. L. monocytogenes and L. ivanovii were able to invade and multiply in trophoblastic cells without causing cell death or inducing expression of pro-inflammatory genes. Although L. innocua was unable to multiply in bovine trophoblastic cells, it induced transcription of the pro-inflammatory mediator CXCL6. This study demonstrated for the first time the susceptibility of bovine trophoblastic cells to L. monocytogenes and L. ivanovii infection.

  4. Requirement for protein kinase R in interleukin-1alpha-stimulated effects in cartilage. (United States)

    Tam, Christine L; Hofbauer, Maria; Towle, Christine A


    Interleukin-1 (IL-1) has pleiotropic effects in cartilage. The interferon-induced, double stranded RNA-activated protein kinase PKR that phosphorylates eukaryotic initiation factor 2 (eIF2) alpha has been implicated in cytokine effects in chondrocytes. A compound was recently identified that potently suppresses PKR autophosphorylation (IC50 approximately 200 etaM) and partially restores PKR-inhibited translation in a cell-free system with significant effect in the nanomolar range. The objectives of this study were to exploit this potent PKR inhibitor to assess whether PKR kinase activity is required for catabolic and proinflammatory effects of IL-1alpha in cartilage and to determine whether IL-1alpha causes an increase in eIF2alpha phosphorylation that is antagonized by the PKR inhibitor. Cartilage explants were incubated with the PKR inhibitor and IL-1alpha. Culture media were assessed for sulfated glycosaminoglycan as an indicator of proteoglycan degradation and for prostaglandin E(2). Cartilage extracts were analyzed by Western blot for cyclooxygenase-2 and phosphorylated signaling molecules. Nanomolar concentrations of the PKR inhibitor suppressed proteoglycan degradation and cyclooxygenase-2 accumulation in IL-1alpha-activated cartilage. The PKR inhibitor stimulated or inhibited PGE(2) production with a biphasic dose response relationship. IL-1alpha increased the phosphorylation of both PKR and eIF2alpha, and nanomolar concentrations of PKR inhibitor suppressed the IL-1alpha-induced changes in phosphorylation. The results strongly support PKR involvement in pathways activated by IL-1alpha in chondrocytes.

  5. Strategies for Stratified Cartilage Bioprinting

    NARCIS (Netherlands)

    Schuurman, W.


    Multiple materials, cells and growth factors can be combined into one construct by the use of a state–of-the-art bioprinter. This technique may in the future make the fabrication of complete tissues or organs possible. In this thesis the feasibility of the bioprinting of cartilage and the

  6. Postnatal development of articular cartilage

    NARCIS (Netherlands)

    Turnhout, van M.C.


    Articular cartilage (AC) is the thin layer of tissue that covers the ends of the bones in the synovial joints in mammals. Functional adult AC has depth-dependent mechanical properties that are not yet present at birth. These depth-dependent mechanical properties in adult life are the result of a

  7. Femoral head cartilage disarticulation disorder (United States)

    Femoral head cartilage disarticulation disorder and necrosis is a major skeletal problem in broiler breeders since they are maintained for a long time in the farm. The etiology of this disease is not well understood. A field study was conducted to understand the basis of this metabolic disease. Six ...

  8. Cartilage-selective genes identified in genome-scale analysis of non-cartilage and cartilage gene expression

    Directory of Open Access Journals (Sweden)

    Cohn Zachary A


    Full Text Available Abstract Background Cartilage plays a fundamental role in the development of the human skeleton. Early in embryogenesis, mesenchymal cells condense and differentiate into chondrocytes to shape the early skeleton. Subsequently, the cartilage anlagen differentiate to form the growth plates, which are responsible for linear bone growth, and the articular chondrocytes, which facilitate joint function. However, despite the multiplicity of roles of cartilage during human fetal life, surprisingly little is known about its transcriptome. To address this, a whole genome microarray expression profile was generated using RNA isolated from 18–22 week human distal femur fetal cartilage and compared with a database of control normal human tissues aggregated at UCLA, termed Celsius. Results 161 cartilage-selective genes were identified, defined as genes significantly expressed in cartilage with low expression and little variation across a panel of 34 non-cartilage tissues. Among these 161 genes were cartilage-specific genes such as cartilage collagen genes and 25 genes which have been associated with skeletal phenotypes in humans and/or mice. Many of the other cartilage-selective genes do not have established roles in cartilage or are novel, unannotated genes. Quantitative RT-PCR confirmed the unique pattern of gene expression observed by microarray analysis. Conclusion Defining the gene expression pattern for cartilage has identified new genes that may contribute to human skeletogenesis as well as provided further candidate genes for skeletal dysplasias. The data suggest that fetal cartilage is a complex and transcriptionally active tissue and demonstrate that the set of genes selectively expressed in the tissue has been greatly underestimated.

  9. Enhanced micropropagation and tiller formation in sugarcane through pretreatment of explants with thidiazuron (TDZ). (United States)

    Kumari, Kavita; Lal, Madan; Saxena, Sangeeta


    An efficient, simple and commercially applicable protocol for rapid micropropagation of sugarcane has been designed using variety Co 05011. Pretreatment of shoot tip explants with thidiazuron (TDZ) induced high frequency regeneration of shoot cultures with improved multiplication ratio. The highest frequency (80%) of shoot initiation in explants pretreated with 10 mg/l of TDZ was obtained during the study. Maximum 65% shoot cultures could be established from the explants pretreated with TDZ as compared to minimum 40% establishment in explants without pretreatment. The explants pretreated with 10 mg/l of TDZ required minimum 40 days for the establishment of shoot cultures as compared to untreated explants which required 60 days. The highest average number of shoots per culture (19.1) could be obtained from the explants pretreated with 10 mg/l of TDZ, indicating the highest multiplication ratio (1:6). Highest rooting (over 94%) was obtained in shoots regenerated from pretreated explants on ½ strength MS medium containing 5.0 mg/l of NAA and 50 g/l of sucrose within 15 days. Higher number of tillers/clump (15.3) could be counted in plants regenerated from pretreated explants than untreated ones (10.9 tillers/clump) in field condition, three months after transplantation. Molecular analysis using RAPD and DAMD markers suggested that the pretreatment of explants with TDZ did not adversely affect the genetic stability of regenerated plants and maintained high clonal purity.

  10. Applied osmotic loading for promoting development of engineered cartilage. (United States)

    Sampat, Sonal R; Dermksian, Matthew V; Oungoulian, Sevan R; Winchester, Robert J; Bulinski, J Chloë; Ateshian, Gerard A; Hung, Clark T


    This study investigated the potential use of static osmotic loading as a cartilage tissue engineering strategy for growing clinically relevant grafts from either synovium-derived stem cells (SDSCs) or chondrocytes. Bovine SDSCs and chondrocytes were individually encapsulated in 2% w/v agarose and divided into chondrogenic media of osmolarities 300 (hypotonic), 330 (isotonic), and 400 (hypertonic, physiologic) mOsM for up to 7 weeks. The application of hypertonic media to constructs comprised of SDSCs or chondrocytes led to increased mechanical properties as compared to hypotonic (300mOsM) or isotonic (330mOsM) media (pBVH) law. Results confirmed that chondrocytes behave as perfect osmometers; however SDSCs deviated from the BVH relation. © 2013 Elsevier Ltd. All rights reserved.

  11. Optimization of Brassica napus (canola) explant regeneration for genetic transformation. (United States)

    Maheshwari, Priti; Selvaraj, Gopalan; Kovalchuk, Igor


    Brassica napus (canola) is the second largest oilseed crop in the world. It is among the first crops to be genetically transformed, and genetically modified cultivars are in commercial production at very significant levels. Despite the early lead with respect to transgenesis, there remain cultivars that are recalcitrant to transformation. To address this, we have conducted an elaborate investigation of the conditions for regenerating shoots from hypocotyl explants from four genetic lines: Invigor 5020, Westar and Topas as well as a microspore culture derived line of Topas (Line 4079). We analyzed the effect of hormonal combinations in regeneration medium, donor plant age and explant type on the regeneration capacity of these plants. The analysis showed that hypocotyls of eight-day-old seedlings grown on media supplemented with 1mg/L dinitrophenylhydrazine (2,4-D) produced the most shoots. Globular somatic embryos emerged following two weeks of 2,4-D treatment. When transferred to the medium containing 5mg/L benzyladenine (BA), approximately 82% of embryos produced shoots within six weeks. Invigor plants were shown to regenerate more efficiently than Topas; the number of plantlets regenerated from Invigor was approximately 40-50% more as compared to Topas or Line 4079. When hypocotyl explants were co-cultivated with the Agrobacterium strain GV3101 harboring a binary vector carrying a firefly luciferase reporter gene (LUC), significant numbers of plantlets were LUC-positive in a luciferase assay. Frequency of such plants were: Invigor 5020 (54.2 ± 2.5%), Westar (53.7 ± 5.3), Topas (16.0 ± 0.24) and Line 4079 (13.4 ± 4). Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Effect of disulfide bonding and multimerization on proteoglycan 4's cartilage boundary lubricating ability and adsorption. (United States)

    Abubacker, Saleem; Ponjevic, Dragana; Ham, Hyun O; Messersmith, Phillip B; Matyas, John R; Schmidt, Tannin A


    The objectives of this study were to assess the cartilage boundary lubricating ability of (1) nonreduced (NR) disulfide-bonded proteoglycan 4 (PRG4) multimers versus PRG4 monomers and (2) NR versus reduced and alkylated (R/A) PRG4 monomers and to assess (3) the ability of NR PRG4 multimers versus monomers to adsorb to an articular cartilage surface. PRG4 was separated into two preparations, PRG4 multimer enriched (PRG4Multi+) and PRG4 multimer deficient (PRG4Multi-), using size exclusion chromatography (SEC) and characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The cartilage boundary lubricating ability of PRG4Multi+ and PRG4Multi- was compared at a physiological concentration (450 μg/mL) and assessed over a range of concentrations (45, 150, and 450 μg/mL). R/A and NR PRG4Multi- were evaluated at 450 μg/mL. Immunohistochemistry with anti-PRG4 antibody 4D6 was performed to visualize the adsorption of PRG4 preparations to the surface of articular cartilage explants. Separation into enriched populations of PRG4Multi+ and PRG4Multi- was achieved using SEC and was confirmed by SDS-PAGE. PRG4Multi+ and PRG4Multi- both functioned as effective friction-reducing cartilage boundary lubricants at 450 μg/mL, with PRG4Multi+ being more effective than PRG4Multi-. PRG4Multi+ lubricated in a dose-dependent manner, however, PRG4Multi- did not. R/A PRG4Multi- lubricated similar to NR PRG4Multi-. PRG4-containing solutions showed 4D6 immunoreactivity at the articular surface; the immunoreactive intensity of PRG4Multi+ appeared to be similar to SF, whereas PRG4Multi- appeared to have less intensity. These results demonstrate that the intermolecular disulfide-bonded multimeric structure of PRG4 is important for its ability to adsorb to a cartilage surface and function as a boundary lubricant. These findings contribute to a greater understanding of the molecular basis of cartilage boundary lubrication of PRG4. Elucidating the PRG4 structure

  13. Characterization of Articular Cartilage Recovery and Its Correlation with Optical Response in the Near-Infrared Spectral Range. (United States)

    Afara, Isaac Oluwaseun; Singh, Sanjleena; Moody, Hayley; Zhang, Lihai; Oloyede, Adekunle


    In this study, we examine the capacity of a new parameter, based on the recovery response of articular cartilage, to distinguish between healthy and damaged tissues. We also investigate whether or not this new parameter correlates with the near-infrared (NIR) optical response of articular cartilage. Normal and artificially degenerated (proteoglycan-depleted) bovine cartilage samples were nondestructively probed using NIR spectroscopy. Subsequently they were subjected to a load and unloading protocol, and the recovery response was logged during unloading. The recovery parameter, elastic rebound ( ER), is based on the strain energy released as the samples underwent instantaneous elastic recovery. Our results reveal positive relationship between the rebound parameter and cartilage proteoglycan content (normal samples: 2.20 ± 0.10 N mm; proteoglycan-depleted samples: 0.50 ± 0.04 N mm for 1 hour of enzymatic treatment and 0.13 ± 0.02 N mm for 4 hours of enzymatic treatment). In addition, multivariate analysis using partial least squares regression was employed to investigate the relationship between ER and NIR spectral data. The results reveal significantly high correlation ( R(2)cal = 98.35% and R(2)val = 79.87%; P cartilage in the combined NIR regions 5,450 to 6,100 cm(-1) and 7,500 to 12,500 cm(-1). We conclude that ER can indicate the mechanical condition and state of health of articular cartilage. The correlation of ER with cartilage optical response in the NIR range could facilitate real-time evaluation of the tissue's integrity during arthroscopic surgery and could also provide an important tool for cartilage assessment in tissue engineering and regeneration research.

  14. Leptin alone and in combination with interleukin-1-beta induced cartilage degradation potentially inhibited by EPA and DHA. (United States)

    Phitak, Thanyaluck; Boonmaleerat, Kanchanit; Pothacharoen, Peraphan; Pruksakorn, Dumnoensun; Kongtawelert, Prachya


    Osteoarthritis (OA) is the most common form of arthritis. Obesity has been believed to be an important risk factor for OA development and the progression of not only load-bearing joints, but low-load-bearing joints as well. Increased leptin has been the focus of a link between obesity and OA. In this study, the effects of pathological (100ng/ml) or supra-pathological (10μg/ml) concentrations of leptin alone or in combination with IL1β on cartilage metabolisms were studied in porcine cartilage explant. The involved mechanisms were examined in human articular chondrocytes (HACs). Moreover, the protective effect of omega-3 polyunsaturated acids, eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA) was also investigated. Leptin (10μg/ml) alone or in combination with IL1β could induce cartilage destruction, although lower concentrations had no effect. Leptin activated NFκB, ERK, JNK and p38 in HACs, which led to the induction of MMP3, MMP13 and ADAMTS4 secretions. The combined effect could further induce those enzymes through the additive effect on activation of NFκB and JNK. Interestingly, both EPA and DHA could inhibit cartilage damage induced by leptin plus IL1β by reducing the activation of NFκB and JNK, which led to the decrease of ADAMTS4 secretion. Altogether, only a supra-pathological concentration of leptin alone or in combination with IL1β could induce cartilage destruction, whereas a pathological one could not. This effect could be inhibited by EPA and DHA. To gain greater understanding of the link between leptin and OA, the effect of different levels of leptin on several states of OA cartilage requires further investigation.

  15. Preclinical Studies for Cartilage Repair (United States)

    Hurtig, Mark B.; Buschmann, Michael D.; Fortier, Lisa A.; Hoemann, Caroline D.; Hunziker, Ernst B.; Jurvelin, Jukka S.; Mainil-Varlet, Pierre; McIlwraith, C. Wayne; Sah, Robert L.; Whiteside, Robert A.


    Investigational devices for articular cartilage repair or replacement are considered to be significant risk devices by regulatory bodies. Therefore animal models are needed to provide proof of efficacy and safety prior to clinical testing. The financial commitment and regulatory steps needed to bring a new technology to clinical use can be major obstacles, so the implementation of highly predictive animal models is a pressing issue. Until recently, a reductionist approach using acute chondral defects in immature laboratory species, particularly the rabbit, was considered adequate; however, if successful and timely translation from animal models to regulatory approval and clinical use is the goal, a step-wise development using laboratory animals for screening and early development work followed by larger species such as the goat, sheep and horse for late development and pivotal studies is recommended. Such animals must have fully organized and mature cartilage. Both acute and chronic chondral defects can be used but the later are more like the lesions found in patients and may be more predictive. Quantitative and qualitative outcome measures such as macroscopic appearance, histology, biochemistry, functional imaging, and biomechanical testing of cartilage, provide reliable data to support investment decisions and subsequent applications to regulatory bodies for clinical trials. No one model or species can be considered ideal for pivotal studies, but the larger animal species are recommended for pivotal studies. Larger species such as the horse, goat and pig also allow arthroscopic delivery, and press-fit or sutured implant fixation in thick cartilage as well as second look arthroscopies and biopsy procedures. PMID:26069576

  16. Supporting Biomaterials for Articular Cartilage Repair (United States)

    Duarte Campos, Daniela Filipa; Drescher, Wolf; Rath, Björn; Tingart, Markus


    Orthopedic surgeons and researchers worldwide are continuously faced with the challenge of regenerating articular cartilage defects. However, until now, it has not been possible to completely mimic the biological and biochemical properties of articular cartilage using current research and development approaches. In this review, biomaterials previously used for articular cartilage repair research are addressed. Furthermore, a brief discussion of the state of the art of current cell printing procedures mimicking native cartilage is offered in light of their use as future alternatives for cartilage tissue engineering. Inkjet cell printing, controlled deposition cell printing tools, and laser cell printing are cutting-edge techniques in this context. The development of mimetic hydrogels with specific biological properties relevant to articular cartilage native tissue will support the development of improved, functional, and novel engineered tissue for clinical application. PMID:26069634

  17. Izogenic cartilage transfer in rhinoplasty procedure. (United States)

    Yigit, Baris; Bicer, Ahmet; Aytop, Derya


    Cartilage is commonly grafted during primary and secondary rhinoplasties as a means of addressing both functional and esthetic issues. Generally, such grafts are taken from the nasal septum, but auricular conchae or ribs may serve as donor sites if needed. However, the latter often entail considerable morbidity and graft mismatch. To circumvent these drawbacks, use of implants or processed cartilage (allogenic or xenogenic in origin) has been proposed. Herein, the isogenic transfer of nasal septal cartilage between identical twins is reported.

  18. Can Glucosamine Supplements Protect My Knee Cartilage from Osteoarthritis? (United States)

    ... Can glucosamine supplements protect my knee cartilage from osteoarthritis? Answers from Brent A. Bauer, M.D. Study results on this question have ... build cartilage. The most common type of arthritis, osteoarthritis wears away the slick cartilage that covers the ...

  19. Absorção de micronutrientes por explantes de bananeira in vitro Micronutrient absorption by banana explants in vitro

    Directory of Open Access Journals (Sweden)

    Josefa Diva Nogueira Diniz


    Full Text Available Foi estudada a absorção dos micronutrientes B, Cu, Fe, Mn e Zn em explantes de bananeira (Musa sp., cultivar Prata Anã em meio básico de Murashige & Skoog suplementado com 30g/L de sacarose e 3,5mg/L de BAP. O experimento foi realizado em delineamento completamente casualizado, com três repetições. Na matéria seca dos propágulos inteiros, rizomas, pseudocaules e folhas foram avaliadas a concentração e extração de nutrientes, e, no meio de cultivo, a quantidade remanescente aos 0, 10, 20, 30, 40, 50 e 60 dias. A maior quantidade de micronutrientes extraída pelos propágulos foi observada nos primeiros 20 dias, exceto no tocante ao Mn, que foi aproximadamente constante durante todo o período. O Fe e o Cu foram os micronutrientes absorvidos em maior e menor quantidade, respectivamente. As concentrações de B, Zn, Mn, e Cu remanescentes no meio de cultivo aos 60 dias foram de 52, 61, 77 e 78%, respectivamente, o que sugere que podem ser reduzidas no meio básico MS para o cultivo de explantes de bananeira.The absorption of the micronutrients B, Cu, Fe, Mn and Zn by banana (Musa sp. cultivar "Prata Anã" explants on the basic medium of Murashige & Skoog supplemented with BAP (3.5 mg/L and sucrose (30 g/L were evaluated at 0, 10, 20, 30, 40, 50 and 60 days after the inoculation. The experiment was arranged on a completely randomized design with three replications. Concentration of the micronutrients in the medium and in dry matter of the whole propagule and in the rhizome, pseudostem and leaves was also evaluated. Absorption of Mn was approximately constant during all the period of the experiment, while the other micronutrients had their higher absorption observed on the first 20 days. At the end of the experiment concentrations of B, Zn, Mn and Cu in the medium were 52, 61, 77 and 78%, respectively. These results point out that it is possible to reduce the concentration of these micronutrients on the basic medium MS for banana

  20. MRI of Native Knee Cartilage Delamination Injuries. (United States)

    White, Candace L; Chauvin, Nancy A; Waryasz, Gregory R; March, Bradford T; Francavilla, Michael L


    The purpose of this article is to describe the normal imaging appearance of cartilage and the pathophysiologic findings, imaging appearance, and surgical management of cartilage delamination. Delamination injuries of knee cartilage signify surgical lesions that can lead to significant morbidity without treatment. These injuries may present with clinical symptoms identical to those associated with meniscal injury, and arthroscopic identification can be difficult, thereby creating a role for imaging diagnosis. A low sensitivity of imaging identification of delamination injury of the knee is reported in the available literature, although vast improvements in MRI of cartilage have since been introduced.

  1. Cartilage thickness measurements from optical coherence tomography (United States)

    Rogowska, Jadwiga; Bryant, Clifford M.; Brezinski, Mark E.


    We describe a new semiautomatic image processing method for detecting the cartilage boundaries in optical coherence tomography (OCT). In particular, we focus on rabbit cartilage since this is an important animal model for testing both chondroprotective agents and cartilage repair techniques. The novel boundary-detection system presented here consists of (1) an adaptive filtering technique for image enhancement and speckle reduction, (2) edge detection, and (3) edge linking by graph searching. The procedure requires several steps and can be automated. The quantitative measurements of cartilage thickness on OCT images correlated well with measurements from histology.

  2. Advances in treatment of articular cartilage injuries

    Directory of Open Access Journals (Sweden)

    Yuan-cheng LI


    Full Text Available Cartilage is a kind of terminally differentiated tissue devoid of vessel or nerve, and it is difficult to repair by itself after damage. Many studies for the treatment of cartilage injuries were performed in recent years aiming at repair of the structure and restoration of its function for injured joint. This article reviews the traditional methods of treatment for cartilage injuries, such as joint lavage with the aid of arthroscope, abrasion chondroplasty, laser abrasion and chondroplasty, and drilling of the subchondral bone-marrow space. The research advances in treatment of articular cartilage injuries with tissue engineering were summarized.

  3. Cellular and Acellular Approaches for Cartilage Repair (United States)


    There are several choices of cells to use for cartilage repair. Cells are used as internal or external sources and sometimes in combination. In this article, an analysis of the different cell choices and their use and potential is provided. Embryonic cartilage formation is of importance when finding more about how to be able to perfect cartilage repair. Some suggestions for near future research based on up-to-date knowledge on chondrogenic cells are given to hopefully stimulate more studies on the final goal of cartilage regeneration. PMID:27340516

  4. Microscopic and spectroscopic investigation of an explanted opacified intraocular lens

    Energy Technology Data Exchange (ETDEWEB)

    Simon, V., E-mail: [Babeş-Bolyai University, Faculty of Physics and Interdisciplinary Research Institute on Bio-Nano-Sciences, 400084 Cluj-Napoca (Romania); Radu, T.; Vulpoi, A. [Babeş-Bolyai University, Faculty of Physics and Interdisciplinary Research Institute on Bio-Nano-Sciences, 400084 Cluj-Napoca (Romania); Rosca, C. [Optilens Clinic of Ophthalmology, 400604 Cluj-Napoca (Romania); Eniu, D. [Iuliu Haţieganu University of Medicine and Pharmacy, Department of Molecular Sciences, 400349 Cluj-Napoca (Romania)


    Highlights: • Changes on intraocular lens (IOL) surface after implantation. • Partial opacification of IOL central area. • Elemental composition on IOL surface prior to and after implantation. • First XPS depth profiling examination of the opacifying deposits. • Cell-mediated hydroxyapatite structuring. - Abstract: The investigated polymethylmethacrylate intraocular lens explanted an year after implantation presented a fine granularity consisting of ring-like grains of about 15 μm in diameter. In order to evidence the changes occurred on intraocular lens relative to morphology, elemental composition and atomic environments, microscopic and spectroscopic analyses were carried out using scanning electron microscopy (SEM), Fourier transform infrared (FTIR), energy-dispersive X-ray (EDS), and X-ray photoelectron (XPS) spectroscopies. The results revealed that the grains contain hydroxyapatite mineral phase. A protein layer covers the lens both in opacified and transparent zones. The amide II band is like in basal epithelial cells. The shape and size of the grains, and the XPS depth profiling results indicate the possibility of a cell-mediated process involving lens epithelial cells which fagocitated apoptotic epithelial cells, and in which the debris derived from cell necrosis were calcified. To the best of our knowledge, this is the first investigation on explanted intraocular lenses using XPS depth profiling in order to examine the inside of the opacifying deposits.

  5. Micropropagation of Cyrtopodium paludicolum (Orchidaceae from root tip explants

    Directory of Open Access Journals (Sweden)

    Dayana Rotili Nunes Picolotto


    Full Text Available An efficient protocol for in vitro plant propagation of Cyrtopodium paludicolum has been developed using root tips dissected from well-developed seedlings. Root tips were cultured on Knudson medium supplemented with α-naphthaleneacetic acid (NAA, and/or thidiazuron (TDZ. TDZ did not induce protocorm-like bodies (PLBs in the NAA absence, indicating phytoregulators synergistic effect. Medium supplemented with 1.34 μM NAA and 2.27 μM TDZ resulted in better response on PBLs, and subsequent shoot differentiation (55.25 shoots per explant, and in better rooting number and root length responses, favoring acclimatization with 90% of survived plants. However, the medium supplemented with only NAA (1.34 μM resulted in 33.50 shoots per explant. Histological sections confirmed that only one PLB was induced per responsive root tip, and it showed numerous dispersed and extended meristemoids, or division centers that originated new PBLs. Additionally, this protocol could be an excellent model to study molecular aspects of root to shoot conversion.

  6. 78 FR 73993 - Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products (United States)


    ... Health Inspection Service 9 CFR Parts 92, 93, 94, 95, 96, and 98 RIN 0579-AC68 Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products Corrections In rule document 2013-28228 appearing on...

  7. 77 FR 20319 - Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products (United States)


    ...; ] DEPARTMENT OF AGRICULTURE Animal and Plant Health Inspection Service 9 CFR Part 93 RIN 0579-AC68 Bovine Spongiform Encephalopathy; Importation of Bovines and Bovine Products Correction In proposed rule document...

  8. Infrared microspectroscopic determination of collagen cross-links in articular cartilage (United States)

    Rieppo, Lassi; Kokkonen, Harri T.; Kulmala, Katariina A. M.; Kovanen, Vuokko; Lammi, Mikko J.; Töyräs, Juha; Saarakkala, Simo


    Collagen forms an organized network in articular cartilage to give tensile stiffness to the tissue. Due to its long half-life, collagen is susceptible to cross-links caused by advanced glycation end-products. The current standard method for determination of cross-link concentrations in tissues is the destructive high-performance liquid chromatography (HPLC). The aim of this study was to analyze the cross-link concentrations nondestructively from standard unstained histological articular cartilage sections by using Fourier transform infrared (FTIR) microspectroscopy. Half of the bovine articular cartilage samples (n=27) were treated with threose to increase the collagen cross-linking while the other half (n=27) served as a control group. Partial least squares (PLS) regression with variable selection algorithms was used to predict the cross-link concentrations from the measured average FTIR spectra of the samples, and HPLC was used as the reference method for cross-link concentrations. The correlation coefficients between the PLS regression models and the biochemical reference values were r=0.84 (p<0.001), r=0.87 (p<0.001) and r=0.92 (p<0.001) for hydroxylysyl pyridinoline (HP), lysyl pyridinoline (LP), and pentosidine (Pent) cross-links, respectively. The study demonstrated that FTIR microspectroscopy is a feasible method for investigating cross-link concentrations in articular cartilage.

  9. Cloning and sequence analysis of a partial cDNA for chicken cartilage proteoglycan core protein. (United States)

    Sai, S; Tanaka, T; Kosher, R A; Tanzer, M L


    A chicken embryo sternal cartilage cDNA library, created in the plasmid expression vector pUC9, was screened for sequences coding for immunologically detectable core protein of the large, major proteoglycan of cartilage. A 1229-base-pair cDNA clone was isolated that contained only one extended open reading frame, which had sequences coding for a polypeptide of 379 amino acid residues. These deduced sequences corresponded to those anticipated from current models of proteoglycan structure; a deduced sequence encompassing 21 amino acids was almost identical to a known sequence of bovine nasal cartilage proteoglycan. Significant homology was found between the deduced amino acid sequence of the proteoglycan and two regions of a chicken hepatic lectin. Immunoprecipitation of the products of cell-free translation yielded a component of about 340 kDa, and transfer blot hybridization of sternal cartilage RNA showed a single mRNA of about 8.1 kilobases. Hybridizable mRNA sequences were readily detectable by dot-blot analyses of the cytoplasm of cartilaginous tissues of the chicken embryo, whereas similar analyses of prechondrogenic limb mesenchymal cells did not demonstrate such hybridizable mRNA signals. Images PMID:3460082

  10. Viability of human chondrocytes in an ex vivo model in relation to temperature and cartilage depth. (United States)

    Drobnic, M; Mars, T; Alibegović, A; Bole, V; Balazic, J; Grubic, Z; Brecelj, J


    Chondrocytes in human articular cartilage remain viable post-mortem. It has however not been established yet how the storage temperature affects their survival, which is essential information when post-mortem cartilage is used for toxicologic studies. Our aim was to construct a simple model of explanted knee cartilage and to test the influences of time and temperature on the viability of chondrocytes in the ex vivo conditions. Osteochondral cylinders were procured from the cadaveric femoral condyles. The cylinders were embedded in water-tight rubber tubes, which formed separate chondral and osteal compartments. Tubes were filled with normal saline, without additives, to keep chondrocytes under close-to-normal conditions. The samples were divided into two groups stored at 4 degrees C and 35 degrees C, respectively. Three samples of each of these two groups were analysed at the time of removal, and then three and nine days later. Images of Live-Dead staining were scanned by a confocal laser microscope. Count of viable chondrocytes in four regions, from surface to bone, was obtained using image analysis software. The regression model revealed that the number of viable chondrocytes decreased every day by 19% and that an increase in temperature by 1 degree C decreased their viability by 5.8%. The temperature effect fell by 0.2 percentage points for every 100 microm from the surface to the bone. Herein we demonstrate that chondrocytes remain viable in the ex vivo model of human knee cartilage long enough to be able to serve as a model for toxicologic studies. Their viability is, however, significantly influenced by time and temperature.

  11. Diagnosis of bovine neosporosis. (United States)

    Dubey, J P; Schares, G


    The protozoan parasite Neospora caninum is a major cause of abortion in cattle. The diagnosis of neosporosis-associated mortality and abortion in cattle is difficult. In the present paper we review histologic, serologic, immunohistochemical, and molecular methods for dignosis of bovine neosporosis. Although not a routine method of diagnosis, methods to isolate viable N. caninum from bovine tissues are also reviewed.

  12. Proinflammatory and Anabolic Gene Expression Effects of Platelet-Rich Gel Supernatants on Equine Synovial Membrane Explants Challenged with Lipopolysaccharide. (United States)

    Carmona, Jorge U; Ríos, Diana L; López, Catalina; Álvarez, María E; Pérez, Jorge E


    Platelet-rich plasma (PRP) preparations are used in horses with osteoarthritis (OA). However, some controversies remain regarding the ideal concentration of platelets and leukocytes to produce an adequate anti-inflammatory and anabolic response in the synovial membrane. The aims of this study were to study the influence of leukoconcentrated platelet-rich gel (Lc-PRG) and leukoreduced platelet-rich gel (Lr-PRG) supernatants on the quantitative expression of some proinflammatory and anabolic genes in equine synovial membrane explants (SMEs) challenged with lipopolysaccharide (LPS). SMEs from six horses were cultured over 96 h. Then, SMEs were harvested for RNA extraction and quantitative gene expression analysis by RT-qPCR for nuclear factor kappa B (NFκB), matrix metalloproteinase 13 (MMP-13), a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS-4), collagen type I alpha 1 (COL1A1), collagen type II alpha 1 (COL2A1), and cartilage oligomeric matrix protein (COMP). The 25% and 50% Lc-PRG supernatants led to downregulation of NFκB, MMP-13, ADAMTS-4, COL1A1, COL2A1, and COMP in SMEs. Lr-PRG supernatants (particularly at the 50% concentration) induced downregulation of NFκB, MMP-13, ADAMTS-4, and COL1A1 and upregulation of COL2A1 and COMP. Lr-PRG supernatants should be used for the treatment of inflammatory arthropathies in horses because they have anti-inflammatory and anabolic effects in the synovial membrane.

  13. Predicting knee cartilage loss using adaptive partitioning of cartilage thickness maps

    DEFF Research Database (Denmark)

    Jørgensen, Dan Richter; Dam, Erik B.; Lillholm, Martin


    This study investigates whether measures of knee cartilage thickness can predict future loss of knee cartilage. A slow and a rapid progressor group was determined using longitudinal data, and anatomically aligned cartilage thickness maps were extracted from MRI at baseline. A novel machine learning...... framework was then trained using these maps. Compared to measures of mean cartilage plate thickness, group separation was increased by focusing on local cartilage differences. This result is central for clinical trials where inclusion of rapid progressors may help reduce the period needed to study effects...

  14. Deferoxamine Suppresses Collagen Cleavage and Protease, Cytokine, and COL10A1 Expression and Upregulates AMPK and Krebs Cycle Genes in Human Osteoarthritic Cartilage. (United States)

    Tchetina, Elena V; Markova, Galina A; Poole, A Robin; Zukor, David J; Antoniou, John; Makarov, Sergey A; Kuzin, Aleksandr N


    This study reports the effects of the iron chelator deferoxamine (DFO) on collagen cleavage, inflammation, and chondrocyte hypertrophy in relation to energy metabolism-related gene expression in osteoarthritic (OA) articular cartilage. Full-depth explants of human OA knee articular cartilage from arthroplasty were cultured with exogenous DFO (1-50 μM). Type II collagen cleavage and phospho-adenosine monophosphate-activated protein kinase (pAMPK) concentrations were measured using ELISAs. Gene expression studies employed real-time PCR and included AMPK analyses in PBMCs. In OA explants collagen cleavage was frequently downregulated by 10-50 μM DFO. PCR analysis of 7 OA patient cartilages revealed that 10 μM DFO suppressed expression of MMP-1, MMP-13, IL-1β, and TNFα and a marker of chondrocyte hypertrophy, COL10A1. No changes were observed in the expression of glycolysis-related genes. In contrast, expressions of genes associated with the mitochondrial Krebs cycle (TCA), AMPK, HIF1α, and COL2A1 were upregulated. AMPK gene expression was reduced in OA cartilage and increased in PBMCs from the same patients compared to healthy controls. Our studies demonstrate that DFO is capable of suppressing excessive collagenase-mediated type II collagen cleavage in OA cartilage and reversing phenotypic changes. The concomitant upregulation of proanabolic TCA-related gene expressions points to a potential for availability of energy generating substrates required for matrix repair by end-stage OA chondrocytes. This might normally be prevented by high whole-body energy requirements indicated by elevated AMPK expression in PBMCs of OA patients.

  15. Deferoxamine Suppresses Collagen Cleavage and Protease, Cytokine, and COL10A1 Expression and Upregulates AMPK and Krebs Cycle Genes in Human Osteoarthritic Cartilage

    Directory of Open Access Journals (Sweden)

    Elena V. Tchetina


    Full Text Available This study reports the effects of the iron chelator deferoxamine (DFO on collagen cleavage, inflammation, and chondrocyte hypertrophy in relation to energy metabolism-related gene expression in osteoarthritic (OA articular cartilage. Full-depth explants of human OA knee articular cartilage from arthroplasty were cultured with exogenous DFO (1–50 μM. Type II collagen cleavage and phospho-adenosine monophosphate-activated protein kinase (pAMPK concentrations were measured using ELISAs. Gene expression studies employed real-time PCR and included AMPK analyses in PBMCs. In OA explants collagen cleavage was frequently downregulated by 10–50 μM DFO. PCR analysis of 7 OA patient cartilages revealed that 10 μM DFO suppressed expression of MMP-1, MMP-13, IL-1β, and TNFα and a marker of chondrocyte hypertrophy, COL10A1. No changes were observed in the expression of glycolysis-related genes. In contrast, expressions of genes associated with the mitochondrial Krebs cycle (TCA, AMPK, HIF1α, and COL2A1 were upregulated. AMPK gene expression was reduced in OA cartilage and increased in PBMCs from the same patients compared to healthy controls. Our studies demonstrate that DFO is capable of suppressing excessive collagenase-mediated type II collagen cleavage in OA cartilage and reversing phenotypic changes. The concomitant upregulation of proanabolic TCA-related gene expressions points to a potential for availability of energy generating substrates required for matrix repair by end-stage OA chondrocytes. This might normally be prevented by high whole-body energy requirements indicated by elevated AMPK expression in PBMCs of OA patients.

  16. Effects of different omega-3 sources, fish oil, krill oil, and green-lipped mussel against cytokine-mediated canine cartilage degradation. (United States)

    Buddhachat, Kittisak; Siengdee, Puntita; Chomdej, Siriwadee; Soontornvipart, Kumpanart; Nganvongpanit, Korakot


    Our purpose was to evaluate the protective effect of three marine omega-3 sources, fish oil (FO), krill oil (KO), and green-lipped mussel (GLM) against cartilage degradation. Canine cartilage explants were stimulated with either 10 ng/mL interleukin-1β (IL-1β) or IL-1β/oncostatin M (10 ng/mL each) and then treated with various concentrations of docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA; 3 and 30 μg/mL), FO, KO, or GLM (250, 500, and 1000 μg/mL) for 28 days. Gene expression was then investigated in primary canine chondrocytes. Our results showed that DHA and EPA as well as omega-3 sources could suppress matrix degradation in cytokine-induced cartilage explants by significantly reducing the increase of sulfated glycosaminoglycans (s-GAGs) and preserving uronic acid and hydroxyproline content (except GLM). These agents were not able to reduce IL-1β-induced IL1B and TNFA expression but were able to down-regulate the expression of the catabolic genes MMP1, MMP3, and MMP13 and up-regulate the anabolic genes AGG and COL2A1; FO and KO were especially effective. Our findings indicated that FO and KO were superior to GLM for their protective effect against proteoglycan and collagen degradation. Hence, FO and KO could serve as promising sources of chondroprotective agents.

  17. Dynamic mechanical loading enhances functional properties of tissue-engineered cartilage using mature canine chondrocytes. (United States)

    Bian, Liming; Fong, Jason V; Lima, Eric G; Stoker, Aaron M; Ateshian, Gerard A; Cook, James L; Hung, Clark T


    The concept of cartilage functional tissue engineering (FTE) has promoted the use of physiologic loading bioreactor systems to cultivate engineered tissues with load-bearing properties. Prior studies have demonstrated that culturing agarose constructs seeded with primary bovine chondrocytes from immature joints, and subjected to dynamic deformation, produced equilibrium compressive properties and proteoglycan content matching the native tissue. In the process of translating these results to an adult canine animal model, it was found that protocols previously successful with immature bovine primary chondrocytes did not produce the same successful outcome when using adult canine primary chondrocytes. The objective of this study was to assess the efficacy of a modified FTE protocol using adult canine chondrocytes seeded in agarose hydrogel and subjected to dynamic loading. Two modes of dynamic loading were applied to constructs using custom bioreactors: unconfined axial compressive deformational loading (DL; 1 Hz, 10% deformation) or sliding contact loading (Slide; 0.5 Hz, 10% deformation). Loading for 3 h daily was initiated on day 0, 14, or 28 (DL0, DL14, DL28, and Slide14). Constructs with applied loading (both DL and Slide) exhibited significant increases in Young's modulus compared with free-swelling control as early as day 28 in culture (p engineered constructs compare favorably with (and exceed in some cases) those of native canine knee (patella groove and condyle) cartilage. Our findings successfully demonstrate an FTE strategy incorporating clinically relevant, adult chondrocytes and gel scaffold for engineering cartilage replacement tissue. These results, using continuous growth factor supplementation, are in contrast to our previously reported studies with immature chondrocytes where the sequential application of dynamic loading after transient transforming growth factor-beta3 application was found to be a superior culture protocol. Sliding, which simulates

  18. Biomechanical evaluation of suture holding properties of native and tissue engineered articular cartilage (United States)

    DuRaine, GD; Arzi, B; Lee, JK; Lee, CA; Responte, DJ; Hu, JC; Athanasiou, KA


    Objective The purpose of this study was to determine suture-holding properties of tissue engineered neocartilage relative to native articular cartilage. To this end, suture pull-out strength was quantified for native articular cartilage and for neocartilages possessing various mechanical properties. Methods Suture holding properties were examined in vitro and in vivo. Neocartilage from bovine chondrocytes was engineered using two sets of exogenous stimuli resulting in neotissue of different biochemical compositions. Compressive and tensile properties and glycosaminoglycan, collagen, and pyridinoline cross-link contents were assayed (study 1). Suture pull-out strength was compared between neocartilage constructs, and bovine and leporine native cartilage. Uniaxial pull-out test until failure was performed after passing 6-0 Vicryl through each tissue (study 2). Subsequently, neocartilage was implanted into a rabbit model to examine short-term suture holding ability in vivo (study 3). Results Neocartilage glycosaminoglycan and collagen content per wet weight reached 4.55% ± 1.62% and 4.21 ± 0.77%, respectively. Tensile properties for neocartilage constructs reached 2.6 ± 0.77 MPa for Young’s modulus and 1.39 ± 0.63 MPa for ultimate tensile strength. Neocartilage reached ~33% of suture pull-out strength of native articular cartilage. Neocartilage cross-link content reached 50% of native values, and suture pull-out strength correlated positively with cross-link content (R2=0.74). Neocartilage sutured into rabbit osteochondral defects was successfully maintained for 3 weeks. Conclusion This study shows that pyridinoline cross-links in neocartilage may be vital in controlling suture pull-out strength. Neocartilage produced in vitro with one-third of native tissue pull-out strength appears sufficient for construct suturing and retention in vivo. PMID:24848644

  19. High throughput generated micro-aggregates of chondrocytes stimulate cartilage formation in vitro and in vivo. (United States)

    Moreira Teixeira, L S; Leijten, J C H; Sobral, J; Jin, R; van Apeldoorn, A A; Feijen, J; van Blitterswijk, C; Dijkstra, P J; Karperien, M


    Cell-based cartilage repair strategies such as matrix-induced autologous chondrocyte implantation (MACI) could be improved by enhancing cell performance. We hypothesised that micro-aggregates of chondrocytes generated in high-throughput prior to implantation in a defect could stimulate cartilaginous matrix deposition and remodelling. To address this issue, we designed a micro-mould to enable controlled high-throughput formation of micro-aggregates. Morphology, stability, gene expression profiles and chondrogenic potential of micro-aggregates of human and bovine chondrocytes were evaluated and compared to single-cells cultured in micro-wells and in 3D after encapsulation in Dextran-Tyramine (Dex-TA) hydrogels in vitro and in vivo. We successfully formed micro-aggregates of human and bovine chondrocytes with highly controlled size, stability and viability within 24 hours. Micro-aggregates of 100 cells presented a superior balance in Collagen type I and Collagen type II gene expression over single cells and micro-aggregates of 50 and 200 cells. Matrix metalloproteinases 1, 9 and 13 mRNA levels were decreased in micro-aggregates compared to single-cells. Histological and biochemical analysis demonstrated enhanced matrix deposition in constructs seeded with micro-aggregates cultured in vitro and in vivo, compared to single-cell seeded constructs. Whole genome microarray analysis and single gene expression profiles using human chondrocytes confirmed increased expression of cartilage-related genes when chondrocytes were cultured in micro-aggregates. In conclusion, we succeeded in controlled high-throughput formation of micro-aggregates of chondrocytes. Compared to single cell-seeded constructs, seeding of constructs with micro-aggregates greatly improved neo-cartilage formation. Therefore, micro-aggregation prior to chondrocyte implantation in current MACI procedures, may effectively accelerate hyaline cartilage formation.

  20. Injectable hydrogels for cartilage tissue engineering

    NARCIS (Netherlands)

    Jin, R.


    Tissue engineering is a promising method for the regeneration of cartilage defects. This approach generally involves the use of a three-dimensional scaffold which can act as a temporary artificial extracellular matrix (ECM) for healthy cartilage cells, chondrocytes. Hydrogels represent a class of

  1. Magnetic Resonance Imaging of Cartilage Repair (United States)

    Trattnig, Siegfried; Winalski, Carl S.; Marlovits, Stephan; Jurvelin, Jukka S.; Welsch, Goetz H.; Potter, Hollis G.


    Articular cartilage lesions are a common pathology of the knee joint, and many patients may benefit from cartilage repair surgeries that offer the chance to avoid the development of osteoarthritis or delay its progression. Cartilage repair surgery, no matter the technique, requires a noninvasive, standardized, and high-quality longitudinal method to assess the structure of the repair tissue. This goal is best fulfilled by magnetic resonance imaging (MRI). The present article provides an overview of the current state of the art of MRI of cartilage repair. In the first 2 sections, preclinical and clinical MRI of cartilage repair tissue are described with a focus on morphological depiction of cartilage and the use of functional (biochemical) MR methodologies for the visualization of the ultrastructure of cartilage repair. In the third section, a short overview is provided on the regulatory issues of the United States Food and Drug Administration (FDA) and the European Medicines Agency (EMEA) regarding MR follow-up studies of patients after cartilage repair surgeries. PMID:26069565

  2. Effect of media composition and explant type on the regeneration of ...

    African Journals Online (AJOL)

    Two as well as three way interactions of three eggplant genotypes, media compositions and explants (hypocotyl, cotyledon and leaf) showed significant differences for plant regeneration. Among three explants, hypocotyl induced highest percent callusing, but cotyledon showed best results for somatic embryogenesis on all ...

  3. Effect of 2,4-D, explants type and cultivar on the callogenesis ...

    African Journals Online (AJOL)

    Although, statistically insignificant, the 8 mg/l 2,4-D concentration was visually the best in callus formation from explants both within and across the cassava cultivars. This study had shown that different cassava explants respond differently to tissue culture conditions established for callus culture formation. Key words: Auxin ...

  4. Influence of genotype and age of explant source on the capacity for ...

    African Journals Online (AJOL)



    Apr 12, 2010 ... The embryogenic capacities of flower explants from one- and two-week-old male inflorescence buds from Musa acuminata Cavendish, AAA, genotypes 'Williams' and 'Grand Naine' were investigated. Explants of hands with immature flowers were excised and induced for embryogenesis. Highly significant ...

  5. In vitro morphogenic events in culture of Lotus corniculatus L. seedling root explants

    Directory of Open Access Journals (Sweden)

    Jan J. Rybczyński


    Full Text Available The experiments were carried out on Lotus corniculatus (L. seedling root explants of the cultivar varieties Skrzeszowicka, Caroll A10 and strain 175. Callus formation and shoot regeneration were the major explant response depended mainly on of the studied genotype and used plant growth regulators (PGRs. Primary cortex of proximal and distal end of explant was the most active tissue for callus proliferation. For shoot primordia differentiation deeper zones of cortex took a part. The process of meristematic centre initiation was not uniform and various level of shoot differentiation events were observed not earlier than 3 weeks of culture. Usually, the shoot primordia regeneration began on proximal rather than distal end of the explant. BAP rather than urea derivatives stimulated shoot proliferation in extended cultures. Increasing of BAP and TDZ concentrations brought about the explant polarity and expansion of the meristematic zones. The explant position in root did not have significant influence on the number of regenerated shoots. The cultures only had better bud formation by TDZ when compared to BAP. BAP stimulated bud formation and development of the shoots from them. Short term of TDZ treatment of explants stimulated meristem formation which developed into buds and shoots. CPPU stimulated callus proliferation and bud formation when explants pretreatment was prolonged from 12 to 36 hrs.


    NARCIS (Netherlands)


    A simple method is presented for selective cell culture of human mesangial cells using explanatation of mesangial cell hillocks. Glomeruli which had been incubated with collagenase were explanted on plastic tissue culture flasks. Three to 6 weeks after explantation, a rapidly growing multilayer of

  7. High Throughput and Mechano-Active Platforms to Promote Cartilage Regeneration and Repair (United States)

    Mohanraj, Bhavana

    Traumatic joint injuries initiate acute degenerative changes in articular cartilage that can lead to progressive loss of load-bearing function. As a result, patients often develop post-traumatic osteoarthritis (PTOA), a condition for which there currently exists no biologic interventions. To address this need, tissue engineering aims to mimic the structure and function of healthy, native counterparts. These constructs can be used to not only replace degenerated tissue, but also build in vitro, pre-clinical models of disease. Towards this latter goal, this thesis focuses on the design of a high throughput system to screen new therapeutics in a micro-engineered model of PTOA, and the development of a mechanically-responsive drug delivery system to augment tissue-engineered approaches for cartilage repair. High throughput screening is a powerful tool for drug discovery that can be adapted to include 3D tissue constructs. To facilitate this process for cartilage repair, we built a high throughput mechanical injury platform to create an engineered cartilage model of PTOA. Compressive injury of functionally mature constructs increased cell death and proteoglycan loss, two hallmarks of injury observed in vivo. Comparison of this response to that of native cartilage explants, and evaluation of putative therapeutics, validated this model for subsequent use in small molecule screens. A primary screen of 118 compounds identified a number of 'hits' and relevant pathways that may modulate pathologic signaling post-injury. To complement this process of therapeutic discovery, a stimuli-responsive delivery system was designed that used mechanical inputs as the 'trigger' mechanism for controlled release. The failure thresholds of these mechanically-activated microcapsules (MAMCs) were influenced by physical properties and composition, as well as matrix mechanical properties in 3D environments. TGF-beta released from the system upon mechano-activation stimulated stem cell

  8. Cartilage tissue engineering for degenerative joint disease. (United States)

    Nesic, Dobrila; Whiteside, Robert; Brittberg, Mats; Wendt, David; Martin, Ivan; Mainil-Varlet, Pierre


    Pain in the joint is often due to cartilage degeneration and represents a serious medical problem affecting people of all ages. Although many, mostly surgical techniques, are currently employed to treat cartilage lesions, none has given satisfactory results in the long term. Recent advances in biology and material science have brought tissue engineering to the forefront of new cartilage repair techniques. The combination of autologous cells, specifically designed scaffolds, bioreactors, mechanical stimulations and growth factors together with the knowledge that underlies the principles of cell biology offers promising avenues for cartilage tissue regeneration. The present review explores basic biology mechanisms for cartilage reconstruction and summarizes the advances in the tissue engineering approaches. Furthermore, the limits of the new methods and their potential application in the osteoarthritic conditions are discussed.

  9. Effect of low oxygen tension on tissue-engineered cartilage construct development in the concentric cylinder bioreactor. (United States)

    Saini, Sunil; Wick, Timothy M


    Cartilage is exposed to low oxygen tension in vivo, suggesting culture in a low-oxygen environment as a strategy to enhance matrix deposition in tissue-engineered cartilage in vitro. To assess the effects of oxygen tension on cartilage matrix accumulation, porous polylactic acid constructs were dynamically seeded in a concentric cylinder bioreactor with bovine chondrocytes and cultured for 3 weeks at either 20 or 5% oxygen tension. Robust chondrocyte proliferation and matrix deposition were achieved. After 22 days in culture, constructs from bioreactors operated at either 20 or 5% oxygen saturation had similar chondrocyte densities and collagen content. During the first 12 days of culture, the matrix glycosaminoglycan (GAG) deposition rate was 19.5 x 10(-9) mg/cell per day at 5% oxygen tension and 65% greater than the matrix GAG deposition rate at 20% oxygen tension. After 22 days of bioreactor culture, constructs at 5% oxygen contained 4.5 +/- 0.3 mg of GAG per construct, nearly double the 2.5 +/- 0.2 mg of GAG per construct at 20% oxygen tension. These data demonstrate that culture in bioreactors at low oxygen tension favors the production and retention of GAG within cartilage matrix without adversely affecting chondrocyte proliferation or collagen deposition. Bioreactor studies such as these can identify conditions that enhance matrix accumulation and construct development for cartilage tissue engineering.

  10. A simple technique of intraocular lenses explantation for single-piece foldable lenses

    Directory of Open Access Journals (Sweden)

    Arup Bhaumik


    Full Text Available Foldable intraocular lenses (IOLs are most commonly used in modern-day cataract surgery. Explantation of these IOLs is not frequently encountered, but sometimes extreme situations may demand the same. Commonly explantation is achieved by bisecting the IOL inside the anterior chamber with a cutter and delivering the pieces out one by one. This may require corneal wound extension with associated damage and endothelial loss leading to visual deterioration. We devised a simple, innovative IOL explantation technique utilizing a modified Alcon A cartridge and snare. This can successfully refold the IOL to be explanted inside the eye and deliver it out through the same wound. The device has limitations with very thick optic lenses, multipiece, and silicon IOLs. In conclusion, we describe a simple, innovative, and reproducible technique to explant almost any single piece IOL without compromising the original surgery and yielding very satisfactory outcomes.

  11. Bovine Herpesvirus 4 infections and bovine mastitis

    NARCIS (Netherlands)

    Wellenberg, Gerardus Johannus


    Mastitis is an often occurring disease in dairy cattle with an enormous economic impact for milk producers worldwide. Despite intensive research, which is historically based on the detection of bacterial udder pathogens, still around 20-35% of clinical cases of bovine mastitis have an unknown

  12. Cartilage repair in the degenerative ageing knee (United States)

    Brittberg, Mats; Gomoll, Andreas H; Canseco, José A; Far, Jack; Lind, Martin; Hui, James


    Background and purpose Cartilage damage can develop due to trauma, resulting in focal chondral or osteochondral defects, or as more diffuse loss of cartilage in a generalized organ disease such as osteoarthritis. A loss of cartilage function and quality is also seen with increasing age. There is a spectrum of diseases ranging from focal cartilage defects with healthy surrounding cartilage to focal lesions in degenerative cartilage, to multiple and diffuse lesions in osteoarthritic cartilage. At the recent Aarhus Regenerative Orthopaedics Symposium (AROS) 2015, regenerative challenges in an ageing population were discussed by clinicians and basic scientists. A group of clinicians was given the task of discussing the role of tissue engineering in the treatment of degenerative cartilage lesions in ageing patients. We present the outcomes of our discussions on current treatment options for such lesions, with particular emphasis on different biological repair techniques and their supporting level of evidence. Results and interpretation Based on the studies on treatment of degenerative lesions and early OA, there is low-level evidence to suggest that cartilage repair is a possible treatment for such lesions, but there are conflicting results regarding the effect of advanced age on the outcome. We concluded that further improvements are needed for direct repair of focal, purely traumatic defects before we can routinely use such repair techniques for the more challenging degenerative lesions. Furthermore, we need to identify trigger mechanisms that start generalized loss of cartilage matrix, and induce subchondral bone changes and concomitant synovial pathology, to maximize our treatment methods for biological repair in degenerative ageing joints. PMID:27910738

  13. Integration of Stem Cell to Chondrocyte-Derived Cartilage Matrix in Healthy and Osteoarthritic States in the Presence of Hydroxyapatite Nanoparticles.

    Directory of Open Access Journals (Sweden)

    Rupak Dua

    Full Text Available We investigated the effectiveness of integrating tissue engineered cartilage derived from human bone marrow derived stem cells (HBMSCs to healthy as well as osteoarthritic cartilage mimics using hydroxyapatite (HA nanoparticles immersed within a hydrogel substrate. Healthy and diseased engineered cartilage from human chondrocytes (cultured in agar gels were integrated with human bone marrow stem cell (HBMSC-derived cartilaginous engineered matrix with and without HA, and evaluated after 28 days of growth. HBMSCs were seeded within photopolymerizable poly (ethylene glycol diacrylate (PEGDA hydrogels. In addition, we also conducted a preliminary in vivo evaluation of cartilage repair in rabbit knee chondral defects treated with subchondral bone microfracture and cell-free PEGDA with and without HA. Under in vitro conditions, the interfacial shear strength between tissue engineered cartilage derived from HBMSCs and osteoarthritic chondrocytes was significantly higher (p < 0.05 when HA nanoparticles were incorporated within the HBMSC culture system. Histological evidence confirmed a distinct spatial transition zone, rich in calcium phosphate deposits. Assessment of explanted rabbit knees by histology demonstrated that cellularity within the repair tissues that had filled the defects were of significantly higher number (p < 0.05 when HA was used. HA nanoparticles play an important role in treating chondral defects when osteoarthritis is a co-morbidity. We speculate that the calcified layer formation at the interface in the osteoarthritic environment in the presence of HA is likely to have attributed to higher interfacial strength found in vitro. From an in vivo standpoint, the presence of HA promoted cellularity in the tissues that subsequently filled the chondral defects. This higher presence of cells can be considered important in the context of accelerating long-term cartilage remodeling. We conclude that HA nanoparticles play an important role in

  14. Concentric cylinder bioreactor for production of tissue engineered cartilage: effect of seeding density and hydrodynamic loading on construct development. (United States)

    Saini, Sunil; Wick, Timothy M


    A concentric cylinder bioreactor has been developed to culture tissue engineered cartilage constructs under hydrodynamic loading. This bioreactor operates in a low shear stress environment, has a large growth area for construct production, allows for dynamic seeding of constructs, and provides for a uniform loading environment. Porous poly-lactic acid constructs, seeded dynamically in the bioreactor using isolated bovine chondrocytes, were cultured for 4 weeks at three seeding densities (60, 80, 100 x 10(6) cells per bioreactor) and three different shear stresses (imposed at 19, 38, and 76 rpm) to characterize the effect of chondrocyte density and hydrodynamic loading on construct growth. Construct seeding efficiency with chondrocytes is greater than 95% within 24 h. Extensive chondrocyte proliferation and matrix deposition are achieved so that after 28 days in culture, constructs from bioreactors seeded at the highest cell densities contain up to 15 x 10(6) cells, 2 mg GAG, and 3.5 mg collagen per construct and exhibit morphology similar to that of native cartilage. Bioreactors seeded with 60 million chondrocytes do not exhibit robust proliferation or matrix deposition and do not achieve morphology similar to that of native cartilage. In cultures under different steady hydrodynamic loading, the data demonstrate that higher shear stress suppresses matrix GAG deposition and encourages collagen incorporation. In contrast, under dynamic hydrodynamic loading conditions, cartilage constructs exhibit robust matrix collagen and GAG deposition. The data demonstrate that the concentric cylinder bioreactor provides a favorable hydrodynamic environment for cartilage construct growth and differentiation. Notably, construct matrix accumulation can be manipulated by hydrodynamic loading. This bioreactor is useful for fundamental studies of construct growth and to assess the significance of cell density, nutrients, and hydrodynamic loading on cartilage development. In addition

  15. Application of an acoustofluidic perfusion bioreactor for cartilage tissue engineering


    Li, Siwei; Glynne-Jones, Peter; Andriotis, Orestis G.; Ching, Kuan Y.; Jonnalagadda, Umesh S.; Oreffo, Richard O.C.; Hill, Martyn; Tare, Rahul S.


    Cartilage grafts generated using conventional static tissue engineering strategies are characterised by low cell viability, suboptimal hyaline cartilage formation and, critically, inferior mechanical competency, which limit their application for resurfacing articular cartilage defects. To address the limitations of conventional static cartilage bioengineering strategies and generate robust, scaffold-free neocartilage grafts of human articular chondrocytes, the present study utilised custom-bu...

  16. Correlation of Pre-Explant Lactate Dehydrogenase Concentrations and Findings During Post-Explant Pump Analysis of the HeartMate II Left Ventricular Assist Device. (United States)

    Sood, Vikram; Alam, Osman; Tchantchaleishvili, Vakhtang; Pagani, Francis D; Aaronson, Keith D; Haft, Jonathan; Joyce, David L; Joyce, Lyle D; Daly, Richard C; Maltais, Simon; Stulak, John M


    Analyses of the HeartMate II left ventricular assist device are routinely performed after explant if returned to the manufacturer. Findings from manufacturer-reported pump analyses have not been correlated with pre-explant serum lactate dehydrogenase (LDH) values. Between May 2004 and December 2014, 502 patients underwent primary HeartMate II implantation. Seventy pumps were explanted in 58 patients and returned to the manufacturer for pump analysis: 51 (73%) for suspected pump thrombosis, 12 (17%) for device-related infection, and 7 (13%) for percutaneous lead fracture. Median time from implant to explant was 12.4 months (range, 1 to 57 months). Pump thrombus was confirmed in 53 of 70 pumps (76%). Stratified by major clinical indications for explant of suspected pump thrombus or infection and lead fracture, the presence of pump thrombus was identified in 47 of 51 (92%) and 6 of 19 (32%), respectively. The median of 1) all LDH values, 2) maximum LDH values, and 3) interval change in LDH between 6 and 2 months before device explant were 1,061, 1,940, and -27 IU/L with thrombus present and 533, 504, and 13 IU/L in the absence of thrombus. Density estimation of LDH concentrations identified that a LDH value of 1,155 IU/L in the 6 months preceding pump thrombus had a 78% sensitivity and 90% specificity for findings of pump thrombus at the time of pump analysis. Pre-explant LDH concentrations significantly correlated with findings on manufacture-performed pump analysis. These data validate LDH surveillance as an important clinical tool for identification of pump thrombus. Copyright © 2017 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.

  17. The influence of collagen network integrity on the accumulation of gadolinium-based MR contrast agents in articular cartilage

    Energy Technology Data Exchange (ETDEWEB)

    Wiener, Edzard; Schmidt, C.; Diederichs, G. [Charite - Universitaetsmedizin Berlin (Germany). Inst. fuer Radiologie; Settles, M. [Klinikum rechts der Isar, Muenchen (Germany). Inst. fuer Roentgendiagnostik; Weirich, G. [Klinikum Rechts der Isar, Muenchen (Germany). Inst. fuer Pathologie und Pathologische Anatomie


    Delayed gadolinium-enhanced MR imaging of cartilage is used to quantify the proteoglycan loss in early osteoarthritis. It is assumed that T 1 after Gd-DTPA administration in the near equilibrium state reflects selective proteoglycan loss from cartilage. To investigate the influence of the collagen network integrity on contrast accumulation, the relaxation rates {delta}R1 and {delta}R2 were compared after Gd-DTPA administration in a well established model of osteoarthritis. Collagen or proteoglycan depletion was induced by the proteolytic enzymes papain and collagenase in healthy bovine patellar cartilage. Using a dedicated MRI sequence, T{sub 1} and T{sub 2} maps were simultaneously acquired before and 11 h after Gd-DTPA administration. Depth-dependent profiles of {delta}R1 and {delta}R2 were calculated in healthy, proteoglycan and collagen-depleted articular cartilage and the mean values of different cartilage layers were compared using the Mann-Whitney-U test. In superficial layers (1 mm) there was no significant difference (p > 0.05) in either {delta}R1 or {delta}R2 between proteoglycan-depleted (16.6 {+-} 1.2 s{sup -1}, 15.9 {+-} 1.0 s{sup -1}) and collagen-depleted articular cartilage (15.3 {+-} 0.9 s{sup -1}, 15.5 {+-} 0.9 s{sup -1}). In deep layers (3 mm) both parameters were significantly higher (p = 0.005, 0.03) in proteoglycan-depleted articular cartilage (12.3 {+-} 1.1 s{sup -1}, 9.8 {+-} 0.8 s{sup -1}) than in collagen-depleted articular cartilage (9.1 {+-} 1.1 s{sup -1}, 8.7 {+-} 0.7 s{sup -1}). Both proteoglycan loss and alterations in the collagen network influence the accumulation of Gd-DTPA in articular cartilage with significant differences between superficial and deep cartilage layers. (orig.)

  18. Cartilage tissue engineering: its potential and uses. (United States)

    Kuo, Catherine K; Li, Wan-Ju; Mauck, Robert L; Tuan, Rocky S


    The prevalent nature of osteoarthritis, a cartilage degenerative disease that results in the erosion of joint surfaces and loss of mobility, underscores the importance of developing functional articular cartilage replacement. Recent research efforts have focused on tissue engineering as a promising approach for cartilage regeneration and repair. Tissue engineering is a multidisciplinary research area that incorporates both biological and engineering principles for the purpose of generating new, living tissues to replace the diseased/damaged tissue and restore tissue/organ function. This review surveys and highlights the current concepts and recent progress in cartilage tissue engineering, and discusses the challenges and potential of this rapidly advancing field of biomedical research. Cartilage tissue engineering is critically dependent on selection of appropriate cells (differentiated or progenitor cells); fabrication and utilization of biocompatible and mechanically suitable scaffolds for cell delivery; stimulation with chondrogenically bioactive molecules introduced in the form of recombinant proteins or via gene transfer; and application of dynamic, mechanical loading regimens for conditioning of the engineered tissue constructs, including the design of specialized biomechanically active bioreactors. Cell selection, scaffold design and biological stimulation remain the challenges of function tissue engineering. Successful regeneration or replacement of damaged or diseased cartilage will depend on future advances in our understanding of the biology of cartilage and stem cells and technological development in engineering.

  19. Thickness Distribution of Glenohumeral Joint Cartilage. (United States)

    Schleich, Christoph; Bittersohl, Bernd; Antoch, Gerald; Krauspe, Rüdiger; Zilkens, Christoph; Kircher, Jörn


    High-resolution 3-dimensional cartilage-specific magnetic resonance imaging (MRI) was performed at 3 T to test the following hypotheses: (1) there is a nonuniform cartilage thickness distribution both on the proximal humerus and on the glenoid surface and (2) the glenohumeral joint as a combined system is congruent with the level of the joint cartilage surface without substantial radial mismatch. Inclusion of 38 volunteers (19 females, mean age 24.34 ± 2.22 years; range 21-29 years) in a prospective study. Measurements of: cartilage thickness in 3 regions and 3 zones; radius of both circles (glenoid and humeral cartilage) for congruency calculation using 3-T MRI with 3-dimensional dual-echo steady-state sequence with water excitation. A homogenous mean cartilage thickness (1.2-1.5 mm) and slightly higher values for the glenoidal articulating surface radii both in the mid-paracoronar section (2.4 vs. 2.1 cm, P cartilage changes at the shoulder for future studies.

  20. Does intraarticular inflammation predict biomechanical cartilage properties? (United States)

    Waldstein, Wenzel; Perino, Giorgio; Jawetz, Shari T; Gilbert, Susannah L; Boettner, Friedrich


    Intact cartilage in the lateral compartment is an important requirement for medial unicompartmental knee arthroplasty (UKA). Progression of cartilage degeneration in the lateral compartment is a common failure mode of medial UKA. Little is known about factors that influence the mechanical properties of lateral compartment cartilage. The purposes of this study were to answer the following questions: (1) Does the synovial fluid white blood cell count predict the biomechanical properties of macroscopically intact cartilage of the distal lateral femur? (2) Is there a correlation between MRI grading of synovitis and the biomechanical properties of macroscopically intact cartilage? (3) Is there a correlation between the histopathologic assessment of the synovium and the biomechanical properties of macroscopically intact cartilage? The study included 84 patients (100 knees) undergoing primary TKA for varus osteoarthritis between May 2010 and January 2012. All patients underwent preoperative MRI to assess the degree of synovitis. During surgery, the cartilage of the distal lateral femur was assessed macroscopically using the Outerbridge grading scale. In knees with an Outerbridge grade of 0 or 1, osteochondral plugs were harvested from the distal lateral femur for biomechanical and histologic assessment. The synovial fluid was collected to determine the white blood cell count. Synovial tissue was taken for histologic evaluation of the degree of synovitis. The mean aggregate modulus and the mean dynamic modulus were significantly greater in knees with 150 or less white blood cells/mL synovial fluid compared with knees with greater than 150 white blood cells/mL synovial fluid. There was no correlation among MRI synovitis grades, histopathologic synovitis grades, and biomechanical cartilage properties. The study suggests that lateral compartment cartilage in patients with elevated synovial fluid white blood cell counts has a reduced ability to withstand compressive loads

  1. Azithromycin is able to control Toxoplasma gondii infection in human villous explants (United States)


    Background Although Toxoplasma gondii infection is normally asymptomatic, severe cases of toxoplasmosis may occur in immunosuppressed patients or congenitally infected newborns. When a fetal infection is established, the recommended treatment is a combination of pyrimethamine, sulfadiazine and folinic acid (PSA). The aim of the present study was to evaluate the efficacy of azithromycin to control T. gondii infection in human villous explants. Methods Cultures of third trimester human villous explants were infected with T. gondii and simultaneously treated with either PSA or azithromycin. Proliferation of T. gondii, as well as production of cytokines and hormones by chorionic villous explants, was analyzed. Results Treatment with either azithromycin or PSA was able to control T. gondii infection in villous explants. After azithromycin or PSA treatment, TNF-α, IL-17A or TGF-β1 levels secreted by infected villous explants did not present significant differences. However, PSA-treated villous explants had decreased levels of IL-10 and increased IL-12 levels, while treatment with azithromycin increased production of IL-6. Additionally, T. gondii-infected villous explants increased secretion of estradiol, progesterone and HCG + β, while treatments with azithromycin or PSA reduced secretion of these hormones concurrently with decrease of parasite load. Conclusions In conclusion, these results suggest that azithromycin may be defined as an effective alternative drug to control T. gondii infection at the fetal-maternal interface. PMID:24885122


    Directory of Open Access Journals (Sweden)

    Yohana de Oliveira-Cauduro


    Full Text Available aims of this research were to evaluate different culture media for indirect organogenesis and shoot multiplication of Eucalyptus benthamii x Eucalyptus dunnii. For organogenesis, leaf explants were used to test the following treatments: two culture media (MS N/2 and JADS supplemented with 0.1 μM 1-naphthaleneacetic acid (NAA and thidiazuron (TDZ (0.1 or 0.5 μM, with or without PVP- 40 (250 mg L-1. The percentage of oxidized explants, callus forming explants, explants with anthocyanin, buds, shoots and the shoot number per explant were evaluated. In the multiplication experiment, isolated shoots were cultivated in MS, JADS and WPM media, all supplemented with 1.11 μM BAP. Four subcultures were carried out every 28 days. In every subculture the explant oxidation, partial or total leaf chlorosis, fresh mass and mean number of shoot per explant were evaluated. The MS N/2 medium supplemented with 0.1 μM NAA and 0.5 μM TDZ promoted the highest rate of organogenesis (8.3% and the culture media MS supplemented with 1.11 μM BAP the multiplication rate was higher than in the other media, in the first and the second subcultures (9.28 and 9.24, respectively, without differences between the three media in the following subcultures. 

  3. An ex vivo porcine nasal mucosa explants model to study MRSA colonization.

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    Pawel Tulinski

    Full Text Available Staphylococcus aureus is an opportunistic pathogen able to colonize the upper respiratory tract and skin surfaces in mammals. Methicillin-resistant S. aureus ST398 is prevalent in pigs in Europe and North America. However, the mechanism of successful pig colonization by MRSA ST398 is poorly understood. To study MRSA colonization in pigs, an ex vivo model consisting of porcine nasal mucosa explants cultured at an air-liquid interface was evaluated. In cultured mucosa explants from the surfaces of the ventral turbinates and septum of the pig nose no changes in cell morphology and viability were observed up to 72 h. MRSA colonization on the explants was evaluated followed for three MRSA ST398 isolates for 180 minutes. The explants were incubated with 3×10(8 CFU/ml in PBS for 2 h to allow bacteria to adhere to the explants surface. Next the explants were washed and in the first 30 minutes post adhering time, a decline in the number of CFU was observed for all MRSA. Subsequently, the isolates showed either: bacterial growth, no growth, or a further reduction in bacterial numbers. The MRSA were either localized as clusters between the cilia or as single bacteria on the cilia surface. No morphological changes in the epithelium layer were observed during the incubation with MRSA. We conclude that porcine nasal mucosa explants are a valuable ex vivo model to unravel the interaction of MRSA with nasal tissue.

  4. An equine joint friction test model using a cartilage-on-cartilage arrangement. (United States)

    Noble, Prisca; Collin, Bernard; Lecomte-Beckers, Jacqueline; Magnée, Adrien; Denoix, Jean M; Serteyn, Didier


    This study describes an equine joint friction test using a cartilage-on-cartilage arrangement and investigates the influence of age and load on the frictional response. Osteochondral plugs were extracted from equine shoulder joints (2-5 years, n=12; 10-14 years, n=15), and mounted in a pin-on-disc tribometer. The frictional response was then measured under constant conditions (2N; 20 degrees C; 5 mm/s), and with increasing load (2N, 5N, 10N). In all experiments, the friction coefficient of young cartilage was significantly (Plubrication remained stable, cartilage ageing may have been responsible for lubrication regime change. The cartilage-on-cartilage model could be used to better understand lubrication regime disturbances in healthy and diseased equine joints, and to test the efficacy of various bio-lubricant treatments. Copyright (c) 2008 Elsevier Ltd. All rights reserved.

  5. Genome-wide transcriptome and expression profile analysis of Phalaenopsis during explant browning.

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    Chuanjun Xu

    Full Text Available Explant browning presents a major problem for in vitro culture, and can lead to the death of the explant and failure of regeneration. Considerable work has examined the physiological mechanisms underlying Phalaenopsis leaf explant browning, but the molecular mechanisms of browning remain elusive. In this study, we used whole genome RNA sequencing to examine Phalaenopsis leaf explant browning at genome-wide level.We first used Illumina high-throughput technology to sequence the transcriptome of Phalaenopsis and then performed de novo transcriptome assembly. We assembled 79,434,350 clean reads into 31,708 isogenes and generated 26,565 annotated unigenes. We assigned Gene Ontology (GO terms, Kyoto Encyclopedia of Genes and Genomes (KEGG annotations, and potential Pfam domains to each transcript. Using the transcriptome data as a reference, we next analyzed the differential gene expression of explants cultured for 0, 3, and 6 d, respectively. We then identified differentially expressed genes (DEGs before and after Phalaenopsis explant browning. We also performed GO, KEGG functional enrichment and Pfam analysis of all DEGs. Finally, we selected 11 genes for quantitative real-time PCR (qPCR analysis to confirm the expression profile analysis.Here, we report the first comprehensive analysis of transcriptome and expression profiles during Phalaenopsis explant browning. Our results suggest that Phalaenopsis explant browning may be due in part to gene expression changes that affect the secondary metabolism, such as: phenylpropanoid pathway and flavonoid biosynthesis. Genes involved in photosynthesis and ATPase activity have been found to be changed at transcription level; these changes may perturb energy metabolism and thus lead to the decay of plant cells and tissues. This study provides comprehensive gene expression data for Phalaenopsis browning. Our data constitute an important resource for further functional studies to prevent explant browning.

  6. Reproducible simulation of respiratory motion in porcine lung explants

    Energy Technology Data Exchange (ETDEWEB)

    Biederer, J. [Dept. of Diagnostic Radiology, Univ. Hospital Schleswig-Holstein, Campus Kiel (Germany); Dept. of Radiology, German Cancer Research Center, Heidelberg (Germany); Plathow, C. [Dept. of Diagnostic Radiology, Eberhard-Karls-Univ. Tuebingen (Germany); Dept. of Radiology, German Cancer Research Center, Heidelberg (Germany); Schoebinger, M.; Meinzer, H.P. [Dept. of Medical and Biological Informatics, German Cancer Research Center, Heidelberg (Germany); Tetzlaff, R.; Puderbach, M.; Zaporozhan, J.; Kauczor, H.U. [Dept. of Radiology, German Cancer Research Center, Heidelberg (Germany); Bolte, H.; Heller, M. [Dept. of Diagnostic Radiology, Univ. Hospital Schleswig-Holstein, Campus Kiel (Germany)


    Purpose: To develop a model for exactly reproducible respiration motion simulations of animal lung explants inside an MR-compatible chest phantom. Materials and Methods: The materials included a piston pump and a flexible silicone reconstruction of a porcine diaphragm and were used in combination with an established MR-compatible chest phantom for porcine heart-lung preparations. The rhythmic inflation and deflation of the diaphragm at the bottom of the artificial thorax with water (1-1.5 L) induced lung tissue displacement resembling diaphragmatic breathing. This system was tested on five porcine heart-lung preparations using 1.5T MRI with transverse and coronal 3D-GRE (TR/TE=3.63/1.58, 256 x 256 matrix, 350 mm FOV, 4 mm slices) and half Fourier T2-FSE (TR/TE=545/29, 256 x 192, 350 mm, 6 mm) as well as multiple row detector CT (16 x 1 mm collimation, pitch 1.5, FOV 400 mm, 120 mAs) acquired at five fixed inspiration levels. Dynamic CT scans and coronal MRI with dynamic 2D-GRE and 2D-SS-GRE sequences (image frequencies of 10/sec and 3/sec, respectively) were acquired during continuous 'breathing' (7/minute). The position of the piston pump was visually correlated with the respiratory motion visible through the transparent wall of the phantom and with dynamic displays of CT and MR images. An elastic body splines analysis of the respiratory motion was performed using CT data. Results: Visual evaluation of MRI and CT showed three-dimensional movement of the lung tissue throughout the respiration cycle. Local tissue displacement inside the lung explants was documented with motion maps calculated from CT. The maximum displacement at the top of the diaphragm (mean 26.26 [SD 1.9] mm on CT and 27.16 [SD 1.5] mm on MRI, respectively [p=0.25; Wilcoxon test]) was in the range of tidal breathing in human patients. Conclusion: The chest phantom with a diaphragmatic pump is a promising platform for multi-modality imaging studies of the effects of respiratory lung

  7. Cartilage invasion patterns in laryngeal cancer. (United States)

    Gómez Serrano, Manuel; Iglesias Moreno, María Cruz; Gimeno Hernández, Jesús; Ortega Medina, Luis; Martín Villares, Cristina; Poch Broto, Joaquín


    The cartilaginous invasion determines the T and is one of the most common sources of mistake in tumor staging. Also it is of great importance when planning any therapeutic alternative. In the latest revision of the TNM classification a clear distinction is made between infiltration of cartilage without going through it, considered a T3 recently and that would be a T4 according to the previous classification, and those going through the cartilage, classified as T4a. While this classification makes the difference in depth of infiltration, it does not emphasize the extent of invasion. This paper provides a detailed description of the laryngeal cartilage tumor infiltration by whole organ serial section in which the invasion is considered both horizontal (transcartilaginous) and vertical (extent of invasion) and establishing patterns of three-dimensional infiltration of the cartilage. This is a cross-sectional study of prevalence. 275 records of patients treated for laryngeal squamous cell carcinoma between 1995 and 2000 were reviewed. The pathological processing of laryngectomy surgical specimens was performed following the method of whole organ serial section described by G. F. Tucker. The following patterns of cartilaginous infiltration were defined: (1) transcartilaginous infiltration; (2a) partial focal infiltration of the cartilage: infiltration not going through the cartilage but occupying one third or less of its extent; (2b) partial extensive infiltration of the cartilage: infiltration occupying two thirds or more of its length and (3) no cartilage infiltration: tumor in contact with the cartilage (paraglottic space) but without affecting it. 161 patients met the inclusion criteria. The most frequent tumor location was supraglottic (58 cases) followed by glottic (47). 109 patients (67.7 %) were treated with total laryngectomy. Partial surgical techniques were performed in the remaining cases. TNM tumor staging was performed according to the results of

  8. The rapid manufacture of uniform composite multicellular-biomaterial micropellets, their assembly into macroscopic organized tissues, and potential applications in cartilage tissue engineering. (United States)

    Babur, Betul Kul; Kabiri, Mahboubeh; Klein, Travis Jacob; Lott, William B; Doran, Michael Robert


    We and others have published on the rapid manufacture of micropellet tissues, typically formed from 100-500 cells each. The micropellet geometry enhances cellular biological properties, and in many cases the micropellets can subsequently be utilized as building blocks to assemble complex macrotissues. Generally, micropellets are formed from cells alone, however when replicating matrix-rich tissues such as cartilage it would be ideal if matrix or biomaterials supplements could be incorporated directly into the micropellet during the manufacturing process. Herein we describe a method to efficiently incorporate donor cartilage matrix into tissue engineered cartilage micropellets. We lyophilized bovine cartilage matrix, and then shattered it into microscopic pieces having average dimensions manufacture of thousands of replica composite micropellets, with each micropellet having a material/CD core and a cellular surface. This micropellet organization enabled the rapid bulking up of the micropellet core matrix content, and left an adhesive cellular outer surface. This morphological organization enabled the ready assembly of the composite micropellets into macroscopic tissues. Generically, this is a versatile method that enables the rapid and uniform integration of biomaterials into multicellular micropellets that can then be used as tissue building blocks. In this study, the addition of CD resulted in an approximate 8-fold volume increase in the micropellets, with the donor matrix functioning to contribute to an increase in total cartilage matrix content. Composite micropellets were readily assembled into macroscopic cartilage tissues; the incorporation of CD enhanced tissue size and matrix content, but did not enhance chondrogenic gene expression.

  9. Morphogenetic response of cotyledon and leaf explants of melon (Cucumis melo L. cv. Amarillo Oro

    Directory of Open Access Journals (Sweden)

    Fernanda Vidigal Duarte Souza


    Full Text Available Callus cultures from cotyledon and leaf explants of a Spanish cultivar of melon (Amarillo Oro were tested for growth and morphogenic capacity on several culture media with different concentrations of IAA (indole-3-acetic acid in combination with 1.0 mg.L-1 BA (6-benzylaminopurine or 6.0 mg.L-1 KIN (kinetin. The best results were achieved with cotyledon explants. The leaf explants presented low bud formation capacity. Variability of organogenic response on cotyledons of different age (7, 5, 3 and 1-day-old was evaluated. The age of explant had a significant influence on bud induction. Cotyledon explants from 7-day-old seedlings showed higher organogenic index and development of shoots when cultured onto MS medium supplemented with 1.5 mg.L-1 of IAA and 1.0 mg.L-1 of BA. The effect of cut type of cotyledonary explants on organogenic response was also investigated. Explants cut transversally showed the best results. The addition of copper sulfate in the culture medium promoted a qualitative improvement of the regenerated shoots.Explantes de cotilédones e folhas do cultivar de melão Amarillo Oro foram cultivados para avaliação do potencial morfogenético em diferentes meios de cultura com diversas concentrações de ácido indolacético (AIA, em combinação com 1,0 mg.L-1 benziladenina (BA e 6,0 mg.L-1 de cinetina (CIN. Os melhores resultados foram obtidos com explantes de cotilédones, sendo que explantes de folhas mostraram uma capacidade baixa na indução de gemas. A variação na resposta organogênica em cotilédones de diferentes idades fisiológicas (1, 3, 5 e 7 dias após a germinação mostraram que os melhores resultados foram obtidos com cotilédones de sete dias cultivados no meio MS suplementado com 1,5 mg.L-1 de AIA e 1,0 mg.L-1 de BA. O efeito do tipo de corte na resposta organogênica de explantes cotiledonares foi também avaliado. Os melhores resultados foram obtidos com explantes cortados transversalmente. Adi

  10. Chondrocalcin is internalized by chondrocytes and triggers cartilage destruction via an interleukin-1β-dependent pathway. (United States)

    Bantsimba-Malanda, Claudie; Cottet, Justine; Netter, Patrick; Dumas, Dominique; Mainard, Didier; Magdalou, Jacques; Vincourt, Jean-Baptiste


    Chondrocalcin is among the most highly synthesized polypeptides in cartilage. This protein is released from its parent molecule, type II pro-collagen, after secretion by chondrocytes. A participation of extracellular, isolated chondrocalcin in mineralization was proposed more than 25 years ago, but never demonstrated. Here, exogenous chondrocalcin was found to trigger MMP13 secretion and cartilage destruction ex vivo in human cartilage explants and did so by modulating the expression of interleukin-1β in primary chondrocyte cultures in vitro. Chondrocalcin was found internalized by chondrocytes. Uptake was found mediated by a single 18-mer peptide of chondrocalcin, which does not exhibit homology to any known cell-penetrating peptide. The isolated peptide, when artificially linked as a tetramer, inhibited gene expression regulation by chondrocalcin, suggesting a functional link between uptake and gene expression regulation. At the same time, the tetrameric peptide potentiated chondrocalcin uptake by chondrocytes, suggesting a cooperative mechanism of entry. The corresponding peptide from type I pro-collagen supported identical cell-penetration, suggesting that this property may be conserved among C-propeptides of fibrillar pro-collagens. Structural modeling localized this peptide to the tips of procollagen C-propeptide trimers. Our findings shed light on unexpected function and mechanism of action of these highly expressed proteins from vertebrates. © 2013. Published by Elsevier B.V. All rights reserved.

  11. Micropropagation Of Merremia Quinquefolia (L. Hallier F. From Nodal Explants

    Directory of Open Access Journals (Sweden)

    Kher Mafatlal M.


    Full Text Available Merremia quinquefolia, is an important medicinal plant of the family Convolvulaceae known for its vasoconstrictor, uterotonic, neurohormonic, sympathicolytic and sedative effects. In the present investigation effect of cytokinins 6-benzylaminopurine (BAP, kinetin (Kn and thidiazuron (TDZ, at concentrations 1.0, 2.0, 3.0, 4.0 and 5.0 mg·dm−3 on in vitro shoot multiplication from nodal explants of M. quinquefolia was evaluated. Bud breaking and emergence of shoots started within 10-15 days of inoculation in all media containing cytokinin. Murashige and Skoog (MS medium supplemented with 4.0 mg·dm−3 BAP resulted in maximum number of shoots from single node within 45 days. In vitro raised shoots were successfully rooted on ½ mineral salts of MS medium with 3% sucrose supplemented with 2.0 mg·dm−3 indole-3-butyric acid (IBA. This is the first report on in vitro propagation of Merremia quinquefolia. This study can be useful for development of micropropagation protocols for related taxa.

  12. Shoot meristem: an ideal explant for Zea mays L. transformation. (United States)

    Sairam, R V; Parani, M; Franklin, G; Lifeng, Z; Smith, B; MacDougall, J; Wilber, C; Sheikhi, H; Kashikar, N; Meeker, K; Al-Abed, D; Berry, K; Vierling, R; Goldman, S L


    We report on a rapid high-frequency somatic embryogenesis and plant regeneration protocol for Zea mays. Maize plants were regenerated from complete shoot meristem (3-4 mm) explants via organogenesis and somatic embryogenesis. In organogenesis, the shoot meristems were directly cultured on a high-cytokinin medium comprising 5-10 mg x L(-1) 6-benzylaminopurine (BAP). The number of multiple shoots produced per meristem varied from six to eight Plantlet regeneration through organogenesis resulted in just four weeks. Callus was induced in five days of incubation on an auxin-modified Murashige and Skoog (MS) medium. Prolific callus, with numerous somatic embryos, developed within 3-4 weeks when cultured on an auxin medium containing 5 mg 2,4-dichlorophenoxyacetic acid x L(-1). The number of multiple shoots varied from three to six per callus. Using R23 (Pioneer, Hi-Bred, Johnston, Iowa), the frequency of callus induction was consistently in excess of 80% and plant regeneration ranged between 47 and 64%. All regenerated plantlets survived in the greenhouse and produced normal plants. Each transgenic plant produced leaves, glumes, and anthers that uniformly expressed green fluorescent protein (GFP). The GFP gene segregated in the pollen. Based on this data it is concluded that the transgenics arose from single-cell somatic embryos. The rate of transfer DNA (T-DNA) transfer to complete shoot meristems of Zea mays was high on the auxin medium and was independent of using super-virulent strains of Agrobacterium.

  13. Tropic responses of potato single-node explant cultures

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    Vinterhalter D.


    Full Text Available A special in-vitro protocol was elaborated enabling the production of potato single-node explant plantlets that can be used as objects for tropic studies. In light-grown plantlets, achievement of a full (90° phototropic (PT curvature required 75 to 120 min of continuous unilateral blue light irradiation or 120-135 min of gravitropic stimulation (GT. Time-lapse photography revealed that the curves describing PT and GT bending have a sigmoid shape. Continuous BL irradiation was necessary for the induction of continuous PT bending. If the BL was turned off after 30-50 min of PT stimulation, the bending gradually decreased and stopped in darkness after 25.0 ± 2.0 min. Within this period, curvature increased by 15.5 ± 1.5°. When the BL was turned off upon completion of PT bending (when the plantlets reached an angle of 90°, the plantlets entered the phase of fast straightening. The 90° PT curvature was significantly exaggerated in darkness by turning the jars from a vertical to horizontal position providing 120.74 ± 2.5° as the final curvature angle after two more hours in darkness.

  14. Nanomechanics of the Cartilage Extracellular Matrix (United States)

    Han, Lin; Grodzinsky, Alan J.; Ortiz, Christine


    Cartilage is a hydrated biomacromolecular fiber composite located at the ends of long bones that enables proper joint lubrication, articulation, loading, and energy dissipation. Degradation of extracellular matrix molecular components and changes in their nanoscale structure greatly influence the macroscale behavior of the tissue and result in dysfunction with age, injury, and diseases such as osteoarthritis. Here, the application of the field of nanomechanics to cartilage is reviewed. Nanomechanics involves the measurement and prediction of nanoscale forces and displacements, intra- and intermolecular interactions, spatially varying mechanical properties, and other mechanical phenomena existing at small length scales. Experimental nanomechanics and theoretical nanomechanics have been applied to cartilage at varying levels of material complexity, e.g., nanoscale properties of intact tissue, the matrix associated with single cells, biomimetic molecular assemblies, and individual extracellular matrix biomolecules (such as aggrecan, collagen, and hyaluronan). These studies have contributed to establishing a fundamental mechanism-based understanding of native and engineered cartilage tissue function, quality, and pathology.

  15. PRP and Articular Cartilage: A Clinical Update

    National Research Council Canada - National Science Library

    Marmotti, Antonio; Rossi, Roberto; Castoldi, Filippo; Roveda, Eliana; Michielon, Gianni; Peretti, Giuseppe M


      The convincing background of the recent studies, investigating the different potentials of platelet-rich plasma, offers the clinician an appealing alternative for the treatment of cartilage lesions and osteoarthritis...

  16. Quantitative Imaging Biomarkers of Knee Cartilage Composition

    NARCIS (Netherlands)

    J. van Tiel (Jasper)


    markdownabstractFor a long time, radiography and subsequently conventional magnetic resonance imaging (MRI) were used as imaging biomarkers for evaluating cartilage morphological disease state in osteoarthritis (OA). Because research is switching its focus towards disease modification or even

  17. Cartilage Repair in Football (Soccer) Athletes (United States)

    Bekkers, J.E.J.; de Windt, Th.S.; Brittberg, M.


    The prevalence of focal articular cartilage lesions among athletes is higher than in the general population. Treatment goals differ considerably between the professional and recreational athlete. High financial stakes and the short duration of a professional career influence the treatment selection for the professional athlete, while such parameters weigh differently in recreational sports. This article describes our investigation of the relation between sports and a high prevalence of focal cartilage lesions. In addition, we provide a critical review of the best available evidence for cartilage surgery and treatment selection, evaluate specific patient profiles for professional and recreational athletes, and propose a treatment algorithm for the treatment of focal cartilage lesions in football (soccer) players. PMID:26069606

  18. The minor collagens in articular cartilage

    DEFF Research Database (Denmark)

    Luo, Yunyun


    Articular cartilage is a connective tissue consisting of a specialized extracellular matrix (ECM) that dominates the bulk of its wet and dry weight. Type II collagen and aggrecan are the main ECM proteins in cartilage. However, little attention has been paid to less abundant molecular components......, especially minor collagens, including type IV, VI, IX, X, XI, XII, XIII, and XIV, etc. Although accounting for only a small fraction of the mature matrix, these minor collagens not only play essential structural roles in the mechanical properties, organization, and shape of articular cartilage, but also...... fulfil specific biological functions. Genetic studies of these minor collagens have revealed that they are associated with multiple connective tissue diseases, especially degenerative joint disease. The progressive destruction of cartilage involves the degradation of matrix constituents including...

  19. Materials science: Like cartilage, but simpler

    DEFF Research Database (Denmark)

    Skov, Anne Ladegaard


    The properties of articular cartilage, which lines bones in joints, depend partlyon repulsion between components of the material. A new synthetic gel that mimics this feature has rare, direction-dependent properties.......The properties of articular cartilage, which lines bones in joints, depend partlyon repulsion between components of the material. A new synthetic gel that mimics this feature has rare, direction-dependent properties....

  20. Animal Models for Cartilage Regeneration and Repair (United States)

    Szczodry, Michal; Bruno, Stephen


    Articular cartilage injury and degeneration are leading causes of disability. Animal studies are critically important to developing effective treatments for cartilage injuries. This review focuses on the use of animal models for the study of the repair and regeneration of focal cartilage defects. Animals commonly used in cartilage repair studies include murine, lapine, canine, caprine, porcine, and equine models. There are advantages and disadvantages to each model. Small animal rodent and lapine models are cost effective, easy to house, and useful for pilot and proof-of-concept studies. The availability of transgenic and knockout mice provide opportunities for mechanistic in vivo study. Athymic mice and rats are additionally useful for evaluating the cartilage repair potential of human cells and tissues. Their small joint size, thin cartilage, and greater potential for intrinsic healing than humans, however, limit the translational value of small animal models. Large animal models with thicker articular cartilage permit study of both partial thickness and full thickness chondral repair, as well as osteochondral repair. Joint size and cartilage thickness for canine, caprine, and mini-pig models remain significantly smaller than that of humans. The repair and regeneration of chondral and osteochondral defects of size and volume comparable to that of clinically significant human lesions can be reliably studied primarily in equine models. While larger animals may more closely approximate the human clinical situation, they carry greater logistical, financial, and ethical considerations. A multifactorial analysis of each animal model should be carried out when planning in vivo studies. Ultimately, the scientific goals of the study will be critical in determining the appropriate animal model. PMID:19831641

  1. The comparison between the absorption rate of rib cartilage graft with conchal cartilage graft in rabbit

    Directory of Open Access Journals (Sweden)

    Soraya Shahrokh


    Full Text Available Background: Cartilage grafts is one integral component in the various fields of plastic surgery particular rhinoplasty. Surgeons usually use from various sources, including the septum of the nose, ears and rib. Complications such deformity and reabsorbtion may be created with use of the cartilage. Area of the removal of cartilage can prevent these complications. The aim of this study was to compare the absorption rate and viability of cartilage autograft between two common donor site, the rib and the concha. Methods: This experimental study was performed on October 2014 in animal laboratory of Hazrat Fatima Hospital, Tehran, Iran. In this study, 15 New Zealand white male rabbits, weighing 2000-2500 g, approximately 12 to 16 weeks of age were used. In each rabbit, a piece of one ear and one cartilage was excised. After careful weighting of grafts, we implanted the rib cartilage graft into the left pocket and the conchal cartilage graft into the right one. After 8 weeks, the grafts were removed and weighed precisely and photography was carried out. The specimens were fixed in 10% formalin solution for histologic examination was. An example of hematoxylin and eosin staining and cut (H&E were performed and samples of live chondrocytes and fibrosis were examined by a pathologist. Results: We lost 3 rabbits during our study. The results showed that the average weight of a graft from the ear within 2 months, but this increase was not statistically significant (P= 0.152. In the rib graft weight loss over 2 months, and this reduction was statistically significant (P= 0.009. The resorption between two group was not significant but the amount of fibrosis was more in conchal cartilage graft. Conclusion: According to the study it can be concluded that absorption rib cartilage is somewhat better results than the cartilage of the ear. More studies, in addition to cartilage implants longer human studies can contribute to more accurate conclusions.

  2. Inorganic polyphosphate stimulates cartilage tissue formation. (United States)

    St-Pierre, Jean-Philippe; Wang, Qishan; Li, Shu Qiu; Pilliar, Robert M; Kandel, Rita A


    Clinical utilization of tissue-engineered cartilage constructs has been limited by their inferior mechanical properties compared to native articular cartilage. A number of strategies have been investigated to increase the accumulation of major extracellular matrix components within in vitro-formed cartilage, including the administration of growth factors and mechanical stimulation. In this study, the anabolic effect of inorganic polyphosphates, a linear polymer of orthophosphate residues linked by phosphoanhydride bonds, was demonstrated in both chondrocyte cultures and native articular cartilage cultured ex vivo. Compared to untreated controls, polyphosphate treatment of three-dimensional primary chondrocyte cultures induced increased glycosaminoglycan and collagen accumulation in a concentration- and chain length-dependent manner. This effect was transient, because chondrocytes express exopolyphosphatases that hydrolyze polyphosphate. The anabolic effect of polyphosphates was accompanied by a lower rate of DNA increase within the chondrocyte cultures treated with inorganic polyphosphate. Inorganic polyphosphate enhances cartilage matrix accumulation and is a promising approach to improve the quality of tissue-engineered cartilage constructs.

  3. [Therapeutic algorithm for traumatic cartilage injuries]. (United States)

    Miltner, Oliver; Hagemann, Lars; Ristan, Steven; Siebert, Christian H


    Reports regarding sport injuries frequently pertain to the knee. Although ligament and meniscus damage are the most common, cartilage injuries are of great interest. Even with the great variety of treatment modalities available, the healing of these cartilage injuries remains problematic. Due to the complex structure of hyaline cartilage joint surface, repair has proven to be very difficult. The conservative treatment options range from orthotic devices and physical therapy to systemic and intraarticular medication. In case of failure, a wide variety of surgical interventions exist. Among these surgical treatment forms, one must differentiate between the repair and the reconstruction of hyaline joint surfaces. In the latter group only the osteochondral autologous transplantation procedures allow for the reconstruction of a cartilaginous lesion with hyaline cartilage as part of a single procedure. This paper will provide an overview of most common therapeutic approaches to cartilage injuries available today. Even with the ongoing discussions with regard to cartilage healing, the basics such as the ligamentous stability of the affected joint, the mechanical axis of the extremity and good neuromuscular control must always be part of the algorithm.

  4. Extracellular vesicles in cartilage homeostasis and osteoarthritis. (United States)

    Miyaki, Shigeru; Lotz, Martin K


    Extracellular vesicles carry bioactive molecules that can be transferred between cells and tissues. The purpose of this review is to describe how extracellular vesicles regulate functions of cells in cartilage and other joint tissues. The potential application of extracellular vesicles in the treatment of osteoarthritis and as biomarkers will also be discussed. Extracellular vesicles are found in synovial fluid, in articular cartilage and in the supernatants of synoviocytes and chondrocytes. Extracellular vesicles in cartilage have been proposed to be involved in cross talk between cells in joint tissues and to affect extracellular matrix turnover and inflammation. Extracellular vesicles from arthritic joints can promote abnormal gene expression and changes in cartilage extracellular matrix, including abnormal mineralization. Promising results were obtained in the therapeutic application of mesenchymal stem cell-derived extracellular vesicles for cartilage repair and experimental osteoarthritis. Extracellular vesicles have emerged as vehicles for the exchange of bioactive signaling molecules within cartilage and between joint tissues to promote joint homeostasis and arthritis pathogenesis. As the molecular content of extracellular vesicles can be customized, they offer utility in therapeutic applications.

  5. Swelling and curling behaviors of articular cartilage. (United States)

    Setton, L A; Tohyama, H; Mow, V C


    A new experimental method was developed to quantify parameters of swelling-induced shape change in articular cartilage. Full-thickness strips of cartilage were studied in free-swelling tests and the swelling-induced stretch, curvature, and areal change were measured. In general, swelling-induced stretch and curvature were found to increase in cartilage with decreasing ion concentration, reflecting an increasing tendency to swell and "curl" at higher swelling pressures. An exception was observed at the articular surface, which was inextensible for all ionic conditions. The swelling-induced residual strain at physiological ionic conditions was estimated from the swelling-induced stretch and found to be tensile and from 3-15 percent. Parameters of swelling were found to vary with sample orientation, reflecting a role for matrix anisotropy in controlling the swelling-induced residual strains. In addition, the surface zone was found to be a structurally important element, which greatly limits swelling of the entire cartilage layer. The findings of this study provide the first quantitative measures of swelling-induced residual strain in cartilage ex situ, and may be readily adapted to studies of cartilage swelling in situ.

  6. Design and fabrication of anatomical bioreactor systems containing alginate scaffolds for cartilage tissue engineering. (United States)

    Gharravi, Anneh Mohammad; Orazizadeh, Mahmoud; Ansari-Asl, Karim; Banoni, Salem; Izadi, Sina; Hashemitabar, Mahmoud


    The aim of the present study was to develop a tissue-engineering approach through alginate gel molding to mimic cartilage tissue in a three-dimensional culture system. The perfusion biomimetic bioreactor was designed to mimic natural joint. The shear stresses exerting on the bioreactor chamber were calculated by Computational Fluid Dynamic (CFD). Several alginate/bovine chondrocyte constructs were prepared, and were cultured in the bioreactor. Histochemical and immunohistochemical staining methods for the presence of glycosaminoglycan(GAG), overall matrix production and type II collagen protein were performed, respectively. The dynamic mechanical device applied a linear mechanical displacement of 2 mm to 10 mm. The CFD modeling indicated peak velocity and maximum wall shear stress were 1.706×10(-3)m/s and 0.02407 dyne/cm(2), respectively. Histochemical and immunohistochemical analysis revealed evidence of cartilage-like tissue with lacunas similar to those of natural cartilage and the production of sulfated GAG of matrix by the chondrons, metachromatic territorial matrix-surrounded cells and accumulation of type II collagen around the cells. The present study indicated that when chondrocytes were seeded in alginate hydrogel and cultured in biomimetic cell culture system, cells survived well and secreted newly synthesized matrix led to improvement of chondrogenesis.

  7. Comparison of photopolymerizable thiol-ene PEG and acrylate-based PEG hydrogels for cartilage development. (United States)

    Roberts, Justine J; Bryant, Stephanie J


    When designing hydrogels for tissue regeneration, differences in polymerization mechanism and network structure have the potential to impact cellular behavior. Poly(ethylene glycol) hydrogels were formed by free-radical photopolymerization of acrylates (chain-growth) or thiol-norbornenes (step-growth) to investigate the impact of hydrogel system (polymerization mechanism and network structure) on the development of engineered tissue. Bovine chondrocytes were encapsulated in hydrogels and cultured under free swelling or dynamic compressive loading. In the acrylate system immediately after encapsulation chondrocytes exhibited high levels of intracellular ROS concomitant with a reduction in hydrogel compressive modulus and higher variability in cell deformation upon compressive strain; findings that were not observed in the thiol-norbornene system. Long-term the quantity of sulfated glycosaminoglycans and total collagen was greater in the acrylate system, but the quality resembled that of hypertrophic cartilage with positive staining for aggrecan, collagens I, II, and X and collagen catabolism. The thiol-norbornene system led to hyaline-like cartilage production especially under mechanical loading with positive staining for aggrecan and collagen II and minimal staining for collagens I and X and collagen catabolism. Findings from this study confirm that the polymerization mechanism and network structure have long-term effects on the quality of engineered cartilage, especially under mechanical loading. Copyright © 2013 Elsevier Ltd. All rights reserved.

  8. Cultivar, explant type and culture medium influencing embryogenesis and organogenegenesis in Anthurium spp.

    Directory of Open Access Journals (Sweden)

    Yaowaphan Sontikun


    Full Text Available The effect of cultivars, culture media, explant type and wounding on culture response of anthurium were studied. Valantino gave the highest callus formation (83.73% significantly different from Sonat (78.73% and Plew Thien Phuket (45.66%. Valantino and Plew Thien Phuket gave meristematic nodular callus (MNC whereas Sonat produced embryogenic-like callus (ELC. Modified Murashige and Skoog (MMS medium gave the highest callus formation from both leaf (86.6% and node (100%. Callus obtained in Nitsch and Nitsch (NN and MMS was MNC while woody plant (WPM medium provided ELC. For explant types, internode gave the highest callus formation (72.63%. Nodal and internodal explant gave ELC while the leaf explant yielded MNC. Wounding leaf blades tended to promote more MNC.

  9. Organ explant culture of neonatal rat ventricles: a new model to study gene and cell therapy.

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    A Dénise den Haan

    Full Text Available Testing cardiac gene and cell therapies in vitro requires a tissue substrate that survives for several days in culture while maintaining its physiological properties. The purpose of this study was to test whether culture of intact cardiac tissue of neonatal rat ventricles (organ explant culture may be used as a model to study gene and cell therapy. We compared (immuno histology and electrophysiology of organ explant cultures to both freshly isolated neonatal rat ventricular tissue and monolayers. (Immuno histologic studies showed that organ explant cultures retained their fiber orientation, and that expression patterns of α-actinin, connexin-43, and α-smooth muscle actin did not change during culture. Intracellular voltage recordings showed that spontaneous beating was rare in organ explant cultures (20% and freshly isolated tissue (17%, but common (82% in monolayers. Accordingly, resting membrane potential was -83.9±4.4 mV in organ explant cultures, -80.5±3.5 mV in freshly isolated tissue, and -60.9±4.3 mV in monolayers. Conduction velocity, measured by optical mapping, was 18.2±1.0 cm/s in organ explant cultures, 18.0±1.2 cm/s in freshly isolated tissue, and 24.3±0.7 cm/s in monolayers. We found no differences in action potential duration (APD between organ explant cultures and freshly isolated tissue, while APD of monolayers was prolonged (APD at 70% repolarization 88.8±7.8, 79.1±2.9, and 134.0±4.5 ms, respectively. Organ explant cultures and freshly isolated tissue could be paced up to frequencies within the normal range for neonatal rat (CL 150 ms, while monolayers could not. Successful lentiviral (LV transduction was shown via Egfp gene transfer. Co-culture of organ explant cultures with spontaneously beating cardiomyocytes increased the occurrence of spontaneous beating activity of organ explant cultures to 86%. We conclude that organ explant cultures of neonatal rat ventricle are structurally and electrophysiologically similar

  10. Human rheumatoid arthritis tissue production of IL-17A drives matrix and cartilage degradation: synergy with tumour necrosis factor-alpha, Oncostatin M and response to biologic therapies.

    LENUS (Irish Health Repository)

    Moran, Ellen M


    INTRODUCTION: The aim of this study was to examine IL-17A in patients, following anti-TNF-alpha therapy and the effect of IL-17A on matrix turnover and cartilage degradation. METHODS: IL-17A expression was examined by ELISA and immunohistology in the rheumatoid arthritis (RA) joints. RA whole synovial tissue explant (RA ST), primary synovial fibroblasts (RASFC), human cartilage and chondrocyte cultures were stimulated with IL-17A +\\/- TNF-alpha and Oncostatin M (OSM). Matrix metalloproteinase (MMP) and tissue inhibitor (TIMP-1) were assessed by ELISA and zymography. Cartilage proteoglycan release was assessed histologically by Safranin-O staining. Clinical parameters, IL-17A, MMP\\/TIMP were assessed in patients pre\\/post biologic therapy. RESULTS: IL-17A levels were higher in RA vs osteoarthritis (OA)\\/normal joints (P < 0.05). IL-17A up-regulated MMP-1, -2, -9, and -13 in RA ST, RASFC, cartilage and chondrocyte cultures (P < 0.05). In combination with TNF-alpha and OSM, IL-17A shifted the MMP:TIMP-1 ratio in favor of matrix degradation (all P < 0.05). Cartilage proteoglycan depletion in response to IL-17A was mild; however, in combination with TNF-alpha or OSM showed almost complete proteoglycan depletion. Serum IL-17A was detected in 28% of patients commencing biologic therapy. IL-17A negative patients demonstrated reductions post therapy in serum MMP1\\/TIMP4, MMP3\\/TIMP1 and MMP3\\/TIMP4 ratios and an increase in CS846 (all P < 0.05). No significant changes were observed in IL-17A positive patients. CONCLUSIONS: IL-17A is produced locally in the inflamed RA joint. IL-17A promotes matrix turnover and cartilage destruction, especially in the presence of other cytokines, mimicking the joint environment. IL-17A levels are modulated in vivo, following anti-TNF therapy, and may reflect changes in matrix turnover.

  11. Enzootic bovine leucosis. (United States)

    Tyler, L


    Enzootic bovine leucosis is associated with infection by bovine leucosis virus. The incubation period is measured in years and a minority of infected animals develop clinical signs. The disease is widespread in Europe and elsewhere and can cause significant economic loss. The epidemiology is incompletely understood and findings from one cattle production system may not be directly applicable to another. Major control programmes exist in Denmark and West Germany and control schemes are being developed elsewhere. Eradication of enzootic bovine leucosis has been established as a goal in the EEC and research is revealing the ways in which this goal may be attained. To be effective, control and epidemiological monitoring must be interactive. Recently introduced serological tests, of improved sensitivity, provide a valuable tool.

  12. Immunogenicity of unprocessed and photooxidized bovine and human osteochondral grafts in collagen-sensitive mice

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    Lehmann Paul V


    Full Text Available Abstract Background Autologous and allogeneic osteochondral grafts have been used to repair damaged or diseased cartilage. There are drawbacks to both of these methods, however. Another possible source for osteochondral grafting is photooxidized xenograft scaffolds. The purpose of this study was to evaluate the adaptive immune response to unprocessed and photooxidized xenogeneic osteochondral grafts in a collagen-sensitive mouse model. Methods Unprocessed and photooxidized bovine and human osteochondral grafts were used. The grafts were implanted subcutaneously in collagen-sensitive DBA/1LacJ mice for four or twelve weeks. ELISPOT assays were conducted with spleen cells to evaluate the number of collagen-specific T cells that produce IL-2, IL-4, IL-5 or IFN-γ. Serum was collected and ELISA assays were performed to determine the titers of collagen-specific and total IgG, IgG1, IgG2a, or IgM antibodies. Histology was conducted on the retrieved osteochondral grafts. Results Results indicated that, with respect to adaptive T cell immunity, the photooxidized bovine grafts, unprocessed human grafts and photooxidized human grafts did not induce a significant response to collagen. The unprocessed bovine grafts, however, were slightly more immunogenic, inducing a weak immune response. With respect to antibody production, the bovine grafts were less immunogenic than the human grafts. Bovine collagen-specific IgG antibodies were not induced by these grafts, but production of IgM after twelve weeks was observed with both the unprocessed and photooxidized bovine grafts. In contrast, photooxidized human osteochondral grafts induced IgG1 and IgG2a antibodies, while the unprocessed human grafts did not. Pre-existing human collagen-specific IgM antibodies were present in all mice, including sham-operated negative controls that did not receive an implant. Histological analysis revealed some degree of fibrous encapsulation and inflammatory infiltrations in both

  13. Protection against LPS-induced cartilage inflammation and degradation provided by a biological extract of Mentha spicata (United States)


    Background A variety of mint [Mentha spicata] has been bred which over-expresses Rosmarinic acid (RA) by approximately 20-fold. RA has demonstrated significant anti-inflammatory activity in vitro and in small rodents; thus it was hypothesized that this plant would demonstrate significant anti-inflammatory activity in vitro. The objectives of this study were: a) to develop an in vitro extraction procedure which mimics digestion and hepatic metabolism, b) to compare anti-inflammatory properties of High-Rosmarinic-Acid Mentha spicata (HRAM) with wild-type control M. spicata (CM), and c) to quantify the relative contributions of RA and three of its hepatic metabolites [ferulic acid (FA), caffeic acid (CA), coumaric acid (CO)] to anti-inflammatory activity of HRAM. Methods HRAM and CM were incubated in simulated gastric and intestinal fluid, liver microsomes (from male rat) and NADPH. Concentrations of RA, CA, CO, and FA in simulated digest of HRAM (HRAMsim) and CM (CMsim) were determined (HPLC) and compared with concentrations in aqueous extracts of HRAM and CM. Cartilage explants (porcine) were cultured with LPS (0 or 3 μg/mL) and test article [HRAMsim (0, 8, 40, 80, 240, or 400 μg/mL), or CMsim (0, 1, 5 or 10 mg/mL), or RA (0.640 μg/mL), or CA (0.384 μg/mL), or CO (0.057 μg/mL) or FA (0.038 μg/mL)] for 96 h. Media samples were analyzed for prostaglandin E2 (PGE2), interleukin 1β (IL-1), glycosaminoglycan (GAG), nitric oxide (NO) and cell viability (differential live-dead cell staining). Results RA concentration of HRAMsim and CMsim was 49.3 and 0.4 μg/mL, respectively. CA, FA and CO were identified in HRAMsim but not in aqueous extract of HRAM. HRAMsim (≥ 8 μg/mL) inhibited LPS-induced PGE2 and NO; HRAMsim (≥ 80 μg/mL) inhibited LPS-induced GAG release. RA inhibited LPS-induced GAG release. No anti-inflammatory or chondroprotective effects of RA metabolites on cartilage explants were identified. Conclusions Our biological extraction procedure produces a

  14. Protection against LPS-induced cartilage inflammation and degradation provided by a biological extract of Mentha spicata

    Directory of Open Access Journals (Sweden)

    Kott Laima S


    Full Text Available Abstract Background A variety of mint [Mentha spicata] has been bred which over-expresses Rosmarinic acid (RA by approximately 20-fold. RA has demonstrated significant anti-inflammatory activity in vitro and in small rodents; thus it was hypothesized that this plant would demonstrate significant anti-inflammatory activity in vitro. The objectives of this study were: a to develop an in vitro extraction procedure which mimics digestion and hepatic metabolism, b to compare anti-inflammatory properties of High-Rosmarinic-Acid Mentha spicata (HRAM with wild-type control M. spicata (CM, and c to quantify the relative contributions of RA and three of its hepatic metabolites [ferulic acid (FA, caffeic acid (CA, coumaric acid (CO] to anti-inflammatory activity of HRAM. Methods HRAM and CM were incubated in simulated gastric and intestinal fluid, liver microsomes (from male rat and NADPH. Concentrations of RA, CA, CO, and FA in simulated digest of HRAM (HRAMsim and CM (CMsim were determined (HPLC and compared with concentrations in aqueous extracts of HRAM and CM. Cartilage explants (porcine were cultured with LPS (0 or 3 μg/mL and test article [HRAMsim (0, 8, 40, 80, 240, or 400 μg/mL, or CMsim (0, 1, 5 or 10 mg/mL, or RA (0.640 μg/mL, or CA (0.384 μg/mL, or CO (0.057 μg/mL or FA (0.038 μg/mL] for 96 h. Media samples were analyzed for prostaglandin E2 (PGE2, interleukin 1β (IL-1, glycosaminoglycan (GAG, nitric oxide (NO and cell viability (differential live-dead cell staining. Results RA concentration of HRAMsim and CMsim was 49.3 and 0.4 μg/mL, respectively. CA, FA and CO were identified in HRAMsim but not in aqueous extract of HRAM. HRAMsim (≥ 8 μg/mL inhibited LPS-induced PGE2 and NO; HRAMsim (≥ 80 μg/mL inhibited LPS-induced GAG release. RA inhibited LPS-induced GAG release. No anti-inflammatory or chondroprotective effects of RA metabolites on cartilage explants were identified. Conclusions Our biological extraction procedure produces

  15. Impact of mold infections in explanted lungs on outcomes of lung transplantation. (United States)

    Vadnerkar, Aniket; Clancy, Cornelius J; Celik, Umit; Yousem, Samuel A; Mitsani, Dimitra; Toyoda, Yoshiya; Nguyen, Minh-Ly; Kwak, Eun J; Pilewski, Joseph; Silveira, Fernanda P; Crespo, Maria; Nguyen, M Hong


    Little is known about the incidence or significance of mold infections in the explanted lungs of lung transplant recipients. We reviewed the histopathology of the explanted lungs from 304 patients who underwent lung transplantation at our institution from 2005 to 2007 and received alemtuzumab induction therapy and posttransplant voriconazole prophylaxis. Invasive mold infections were present in the explanted lungs of 5% (14 of 304) of patients, including chronic necrotizing pneumonias (n=7), mycetomas (n=4), and invasive fungal pneumonias (n=3). Only 21% (3 of 14) received immunosuppressive therapy within 1 year before lung transplantation, suggesting that lung damage itself predisposed patients to mold infections. The risk of mold infection was higher in patients with cystic fibrosis (11%, 4 of 35) than other underlying lung diseases (4%, 10 of 269). Pulmonary mold infections were not diagnosed or suspected in 57% (8 of 14) of patients. Despite secondary voriconazole prophylaxis, fungal infections developed in 43% (6 of 14) of patients with mold infections of the explanted lungs compared with 14% (42 of 290) of patients without mold infections (P=0.01). Three patients developed invasive fungal infections while on voriconazole prophylaxis and three developed fungal infections more than 8 months after the discontinuation of voriconazole. The mortality attributable to invasive fungal infections among patients with mold infections of the explanted lungs was 29% (4 of 14). Invasive mold infections in the explanted lungs are often not recognized before lung transplantation and are associated with poor outcomes.

  16. Cartilage Repair Surgery: Outcome Evaluation by Using Noninvasive Cartilage Biomarkers Based on Quantitative MRI Techniques?

    Directory of Open Access Journals (Sweden)

    Pia M. Jungmann


    Full Text Available Background. New quantitative magnetic resonance imaging (MRI techniques are increasingly applied as outcome measures after cartilage repair. Objective. To review the current literature on the use of quantitative MRI biomarkers for evaluation of cartilage repair at the knee and ankle. Methods. Using PubMed literature research, studies on biochemical, quantitative MR imaging of cartilage repair were identified and reviewed. Results. Quantitative MR biomarkers detect early degeneration of articular cartilage, mainly represented by an increasing water content, collagen disruption, and proteoglycan loss. Recently, feasibility of biochemical MR imaging of cartilage repair tissue and surrounding cartilage was demonstrated. Ultrastructural properties of the tissue after different repair procedures resulted in differences in imaging characteristics. T2 mapping, T1rho mapping, delayed gadolinium-enhanced MRI of cartilage (dGEMRIC, and diffusion weighted imaging (DWI are applicable on most clinical 1.5 T and 3 T MR scanners. Currently, a standard of reference is difficult to define and knowledge is limited concerning correlation of clinical and MR findings. The lack of histological correlations complicates the identification of the exact tissue composition. Conclusions. A multimodal approach combining several quantitative MRI techniques in addition to morphological and clinical evaluation might be promising. Further investigations are required to demonstrate the potential for outcome evaluation after cartilage repair.

  17. Osteoarthritic human cartilage is more sensitive to transforming growth factor beta than is normal cartilage

    NARCIS (Netherlands)

    Lafeber, F. P.; Vander Kraan, P. M.; Huber-Bruning, O.; Vanden Berg, W. B.; Bijlsma, J. W.


    Osteoarthritis is a degenerative joint disease, characterized by the destruction of the articular cartilage. One of the first changes in the osteoarthritic articular cartilage is a reduction in proteoglycan content. In this study we demonstrate that transforming growth factor beta (TGF beta), a

  18. Correction of Asian Short Nose with Lower Lateral Cartilage Repositioning and Ear Cartilage Grafting

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    Jin Suk Byun, MD, PhD


    Conclusions: LLC repositioning and ear cartilage grafting aid in the correction of short nose in Asians. With LLC repositioning and ear cartilage grafting, the nasal tip can be positioned in accordance with the patient’s anatomic limits. The entire nasal tip and columella can be lengthened, while the tip maintains its mobility.

  19. In end stage osteoarthritis, cartilage tissue pentosidine levels are inversely related to parameters of cartilage damage

    NARCIS (Netherlands)

    Vos, P.A.J.M.; Mastbergen, S.C.; Huisman, A.M.; Boer,; Groot,; Polak, A.A.; Lafeber, F.P.J.G.


    Objectives: Age is the most prominent predisposition for development of osteoarthritis (OA). Age-related changes of articular cartilage are likely to play a role. Advanced glycation endproducts (AGEs) accumulate in cartilage matrix with increasing age and adversely affect the biomechanical

  20. Cartilage Repair Surgery: Outcome Evaluation by Using Noninvasive Cartilage Biomarkers Based on Quantitative MRI Techniques? (United States)

    Jungmann, Pia M.; Baum, Thomas; Bauer, Jan S.; Karampinos, Dimitrios C.; Link, Thomas M.; Li, Xiaojuan; Trattnig, Siegfried; Rummeny, Ernst J.; Woertler, Klaus; Welsch, Goetz H.


    Background. New quantitative magnetic resonance imaging (MRI) techniques are increasingly applied as outcome measures after cartilage repair. Objective. To review the current literature on the use of quantitative MRI biomarkers for evaluation of cartilage repair at the knee and ankle. Methods. Using PubMed literature research, studies on biochemical, quantitative MR imaging of cartilage repair were identified and reviewed. Results. Quantitative MR biomarkers detect early degeneration of articular cartilage, mainly represented by an increasing water content, collagen disruption, and proteoglycan loss. Recently, feasibility of biochemical MR imaging of cartilage repair tissue and surrounding cartilage was demonstrated. Ultrastructural properties of the tissue after different repair procedures resulted in differences in imaging characteristics. T2 mapping, T1rho mapping, delayed gadolinium-enhanced MRI of cartilage (dGEMRIC), and diffusion weighted imaging (DWI) are applicable on most clinical 1.5 T and 3 T MR scanners. Currently, a standard of reference is difficult to define and knowledge is limited concerning correlation of clinical and MR findings. The lack of histological correlations complicates the identification of the exact tissue composition. Conclusions. A multimodal approach combining several quantitative MRI techniques in addition to morphological and clinical evaluation might be promising. Further investigations are required to demonstrate the potential for outcome evaluation after cartilage repair. PMID:24877139

  1. Free Diced Cartilage: A New Application of Diced Cartilage Grafts in Primary and Secondary Rhinoplasty. (United States)

    Kreutzer, Christian; Hoehne, Julius; Gubisch, Wolfgang; Rezaeian, Farid; Haack, Sebastian


    Irregularities or deformities of the nasal dorsum after hump reduction account for a significant number of revision rhinoplasties. The authors therefore developed a technique of meticulously dicing and exactly placing free diced cartilage grafts, harvested from septum, rib, or ear cartilage. The cartilage paste is used for smoothening, augmentation, or camouflaging of the nasal dorsum in primary or revision rhinoplasties. A retrospective analysis of multisurgeon consecutive open approach rhinoplasties from January to December of 2014 was conducted at a single center. The authors compared the outcome of three different techniques to augment or cover the nasal dorsum after an observation period of 7 months. In group I, 325 patients with free diced cartilage grafts as the only onlay were included. In group II, consisting of 73 patients, the dorsal onlay was either fascia alone or in combination with free diced cartilage grafts. Forty-eight patients in group III received a dorsal augmentation with the classic diced cartilage in fascia technique. Four hundred forty-six patients undergoing primary and secondary rhinoplasties in which one of the above-mentioned diced cartilage techniques was used were included in the study. The authors found revision rates for dorsal irregularities within the 7-month postoperative observation period of 5.2, 8.2, and 25 percent for groups I, II, and III, respectively. The authors' findings strongly support their clinical experience that the free diced cartilage graft technique presents an effective and easily reproducible method for camouflage and augmentation in aesthetic and reconstructive rhinoplasty.

  2. Non-invasive monitoring of cytokine-based regenerative treatment of cartilage by hyperspectral unmixing (Conference Presentation) (United States)

    Mahbub, Saabah B.; Succer, Peter; Gosnell, Martin E.; Anwaer, Ayad G.; Herbert, Benjamin; Vesey, Graham; Goldys, Ewa M.


    Extracting biochemical information from tissue autofluorescence is a promising approach to non-invasively monitor disease treatments at a cellular level, without using any external biomarkers. Our recently developed unsupervised hyperspectral unmixing by Dependent Component Analysis (DECA) provides robust and detailed metabolic information with proper account of intrinsic cellular heterogeneity. Moreover this method is compatible with established methods of fluorescent biomarker labelling. Recently adipose-derived stem cell (ADSC) - based therapies have been introduced for treating different diseases in animals and humans. ADSC have been shown promise in regenerative treatments for osteoarthritis and other bone and joint disorders. One of the mechanism of their action is their anti-inflammatory effects within osteoarthritic joints which aid the regeneration of cartilage. These therapeutic effects are known to be driven by secretions of different cytokines from the ADSCs. We have been using the hyperspectral unmixing techniques to study in-vitro the effects of ADSC-derived cytokine-rich secretions with the cartilage chip in both human and bovine samples. The study of metabolic effects of different cytokine treatment on different cartilage layers makes it possible to compare the merits of those treatments for repairing cartilage.

  3. Aggrecan structure in amphibian cartilage

    Directory of Open Access Journals (Sweden)

    Covizi D.Z.


    Full Text Available The structure of the large proteoglycan present in the bullfrog epiphyseal cartilage was studied by immunochemical and biochemical methods. The isolated monomer showed a polydisperse behavior on Sepharose CL2B, with a peak at Kav = 0.14. Chondroitin sulfate chains were identified by HPLC analysis of the products formed by chondroitinase digestion and mercuric acetate treatment. These chains have approximately 38 disaccharides, a Di45:Di68 ratio of 1.6 and GalNAc4S + GalNAc4,6S are the main non-reducing terminals. Keratan sulfate was identified by the use of two monoclonal antibodies in Western blots after chondroitinase ABC treatment. A keratan sulfate-rich region (~110 kDa was isolated by sequential treatment with chondroitinase ABC and proteases. We also employed antibodies in Western blotting experiments and showed that the full length deglycosylated core protein is about 300 kDa after SDS-PAGE. Domain-specific antibodies revealed the presence of immunoreactive sites corresponding to G1/G2 and G3 globular domains and the characterization of this large proteoglycan as aggrecan. The results indicate the high conservation of the aggrecan domain structure in this lower vertebrate.

  4. Structures of benzo(a)pyrene-nucleic acid adducts formed in human and bovine bronchial explants

    DEFF Research Database (Denmark)

    Jeffrey, A.M.; Weinstein, I.B.; Jenette, K.W.


    PUBLICATION by Sims et al. of evidence that the 7,8-dihydrodiol-9,10-oxide of benzo(a)pyrene (BP) is a metabolic intermediate in the covalent binding of this ubiquitous polycyclic aromatic hydrocarbon to DNA in hamster embryo cells1 was followed by many related publications2. Grover et al. 3 also...

  5. Ultrasound elastomicroscopy for articular cartilage: from static to transient and 1D to 2D (United States)

    Zheng, Yongping; Bridal, Sharon L.; Shi, Jun; Saied, Amena; Lu, Minghua; Jaffre, Britta; Mak, Arthur F. T.; Laugier, Pascal; Qin, Ling


    Articular cartilage (AC) is a biological weight-bearing tissue covering the ends of articulating bones within synovial joints. Its function very much depends on the unique multi-layered structure and the depth-dependent material properties, which have not been well invetigated nondestructively. In this study, transient depth-dependent material properties of bovine patella cartilage were measured using ultrasound elastomicroscopy methods. A 50 MHz focused ultrasound transducer was used to collect A-mode ultrasound echoes from the articular cartilage during the compression and subsequent force-relaxation. The transient displacements of the cartilage tissues at different depths were calculated from the ultrasound echoes using a cross-correlation technique. It was observed that the strains in the superficial zone were much larger than those in the middle and deep zones as the equilibrium state was approached. The tissues inside the AC layer continued to move during the force-relaxation phase after the compression was completed. This process has been predicted by a biphasic theory. In this study, it has been verified experimentally. It was also observed that the tissue deformations at different depths of AC were much more evenly distributed before force-relaxation. AC specimens were also investigated using a 2D ultrasound elastomicroscopy system that included a 3D translating system for moving the ultrasound transducer over the specimens. B-mode RF ultrasound signals were collected from the specimens under different loading levels applied with a specially designed compressor. Preliminary results demonstrated that the scanning was repeatable with high correlation of radio frequency signals obtained from the same site during different scans when compression level was unchanged (R2 > 0.97). Strains of the AC specimens were mapped using data collected with this ultrasound elastomicroscope. This system can also be potentially used for the assessment of other biological

  6. Intervet Symposium: bovine neosporosis

    NARCIS (Netherlands)

    Schetters, T.; Dubey, J.P.; Adrianarivo, A.; Frankena, K.; Romero, J.J.; Pérez, E.; Heuer, C.; Nicholson, C.; Russell, D.; Weston, J.


    This article summarises the most relevant data of presentations delivered at the 19th International Conference of the World Association for the Advancement of Veterinary Parasitology (WAAVP) held in New Orleans, LA, USA, from 10 to 14 August 2003) in a symposium session on bovine neosporosis. The

  7. Genotyping bovine coronaviruses. (United States)

    Bovine coronaviruses (BoCV) are enveloped, single-stranded, positive-sense RNA viruses of the Coronaviridae family. Infection is associated with enteritis and pneumonia in calves and Winter Dysentery in adult cattle. Strains, isolated more than 50 years ago, are used in vaccines and as laboratory ...

  8. Indução de calos embriogênicos em explantes de cupuaçuzeiro Induction of embryogenics calli in cupuassu explants

    Directory of Open Access Journals (Sweden)

    Maria das Graças Rodrigues Ferreira


    Full Text Available Objetivou-se a indução de calos embriogênicos em cupuaçuzeiro, em função do tipo de explante e meio de cultura. Foram testados como explantes, segmentos cotiledonares e eixos embrionários divididos em três partes: região da plúmula, radícula e hipocótilo. Os explantes foram cultivados em 2 diferentes meios de cultura: 1 MS suplementado com 2,4-D (1 mg L-1 e Cinetina (0,25 mg L-1; 2 MS acrescido de ANA (5 mg L-1 e Cinetina (0,25 mg L-1. Constatou-se que a região do hipocótilo foi a parte mais responsiva do eixo embrionário, formando calos com aspecto branco e friável. As auxinas testadas nos meios não estimularam o processo embriogênico em calos de cupuaçuzeiro.It was studied the induction of embryogenics calli in cupuassu, in function of kind of explant and culture medium. Cotyledons segments and embryonic axes were tested and divided in three parts: region of plumule, radicule and hypocotile. The explants were cultivated in two different culture medium: 1 MS supplemented with 2,4-D (1 mg L-1 and Kinetin (0,25 mg L-1; 2 MS supplemented with NAA (5 mg L-1 and Kinetin (0,25 mg L-1. The hypocotile region demonstrated to be more responsive segment of the embryonic axe, forming callus with white and friable aspect. No somatic embryogenesis was evidenced in callus of cupuassu with auxines testeds in the medium.

  9. Polarized IR microscopic imaging of articular cartilage

    Energy Technology Data Exchange (ETDEWEB)

    Ramakrishnan, Nagarajan; Xia Yang; Bidthanapally, Aruna [Department of Physics and Center for Biomedical Research, Oakland University, Rochester, MI 48309 (United States)


    The objective of this spectroscopic imaging study is to understand the anisotropic behavior of articular cartilage under polarized infrared radiation at 6.25 {mu}m pixel resolution. Paraffin embedded canine humeral cartilage-bone blocks were used to obtain 6 {mu}m thick tissue sections. Two wire grid polarizers were used to manipulate the polarization states of IR radiation by setting them for various polarizer/analyzer angles. The characteristics of the major chemical components (amide I, amide II, amide III and sugar) of articular cartilage were investigated using (a) a polarizer and (b) a combination of a polarizer and an analyzer. These results were compared to those obtained using only an analyzer. The infrared anisotropy (variation in infrared absorption as a function of polarization angles) of amide I, amide II and amide III bands correlates with the orientation of collagen fibrils along the tissue depth in different histological zones. An 'anisotropic flipping' region of amide profiles indicates the possibility of using Fourier transform infrared imaging (FTIRI) to determine the histological zones in cartilage. Cross-polarization experiment indicates the resolution of overlapping peaks of collagen triple helix and/or proteoglycan in articular cartilage.

  10. Repair of osteochondral defects in rabbits with ectopically produced cartilage

    NARCIS (Netherlands)

    Emans, PJ; Hulsbosch, M; Wetzels, GMR; Bulstra, SK; Kuijer, R


    Cartilage has poor regenerative capacity. Donor site morbidity and interference with joint homeostasis should be considered when applying the autologous chondrocyte transplantation technique. The use of ectopically produced cartilage, derived from periosteum, might be a novel method to heal

  11. Primary cilia expression in bone marrow in response to mechanical stimulation in explant bioreactor culture. (United States)

    Coughlin, T R; Schiavi, J; Alyssa Varsanik, M; Voisin, M; Birmingham, E; Haugh, M G; McNamara, L M; Niebur, G L


    Bone marrow contains a multitude of mechanically sensitive cells that may participate in mechanotransduction. Primary cilia are sensory organelles expressed on mesenchymal stem cells (MSCs), osteoblasts, osteocytes, and other cell types that sense fluid flow in monolayer culture. In marrow, cilia could similarly facilitate the sensation of relative motion between adjacent cells or interstitial fluid. The goal of this study was to determine the response of cilia to mechanical stimulation of the marrow. Bioreactors were used to supply trabecular bone explants with low magnitude mechanical stimulation (LMMS) of 0.3 ×g at 30 Hz for 1 h/d, 5 d/week, inducing shear stresses in the marrow. Four groups were studied: unstimulated (UNSTIM), stimulated (LMMS), and with and without chloral hydrate (UNSTIM+CH and LMMS+CH, respectively), which was used to disrupt cilia. After 19 days of culture, immunohistochemistry for acetylated α-tubulin revealed that more cells expressed cilia in culture compared to in vivo controls. Stimulation decreased the number of cells expressing cilia in untreated explants, but not in CH-treated explants. MSCs represented a greater fraction of marrow cells in the untreated explants than CH-treated explants. MSCs harvested from the stimulated groups were more proliferative than in the unstimulated explants, but this effect was absent from CH treated explants. In contrast to the marrow, neither LMMS nor CH treatment affected bone formation as measured by mineralising surface. Computational models indicated that LMMS does not induce bone strain, and the reported effects were thus attributed to shear stress in the marrow. From a clinical perspective, genetic or pharmaceutical alterations of cilia expression may affect marrow health and function.

  12. Efficient Regeneration of �Caralis� Alstroemeria Cultivar from Rhizome Explants

    Directory of Open Access Journals (Sweden)

    Amir Ghaffar SHAHRIARI


    Full Text Available In this paper, the effects of a number of growth regulators as well as supplements to the Murashige and Skoog (MS basal medium were evaluated on the regeneration of Alstroemeria rhizome explants. In the first experiment the effects of three cytokinins (BA, TDZ and 2IP each at 0.5, 1 and 2 mg/l in combination with NAA (0.2 mg/l, followed by another PGR combination of 2IP (at 0.5, 1 and 2 mg/l with NAA (0 and 0.2 mg/l, on regeneration of rhizome-derived explants, was investigated. Through the second experiment, the effects of a number of supplements, including glucose (30 g/l as the alternative for sucrose, casein hydrolysate (1 g/l, asparagine and glutamine, (each at 30 mg/l added to MS medium, containing 1 mg/l BA and 0.2 mg/l NAA, was examined on rhizome explants regeneration. Among the tested cytokinins, BA induced better regeneration of rhizome explants, resulting in a higher number of shoots compared to the other cytokinins. A medium supplemented with 1 mg/l BA and 0.2 mg/l NAA proved to be the most effective, with an average of 4.16 regenerated shoots per explant. In the second PGR combination, addition of NAA at 0.2 mg/l improved regeneration, compared to NAA-free treatments. In the second experiment, glucose substitution for sucrose improved regeneration with an average of 5.10 regenerated shoots per explant, compared to 4.16 shoots in sucrose-containing medium; whereas glutamine and asparagine (with 2.66 shoots and casein hydrolysate (with 3.80 shoots showed a negative influence on rhizome explants regeneration.

  13. Efficient culture protocol for plant regeneration from cotyledonary petiole explants of Jatropha curcas L.

    Directory of Open Access Journals (Sweden)

    Ying Liu


    Full Text Available A high-frequency and reproducible protocol for induction of adventitious shoot buds and plant regeneration from cotyledonary petiole explants of Jatropha curcas L. has been developed. The cotyledonary petiole explants of J. curcas cultured directly on medium supplemented with thidiazuron (TDZ induce regeneration of poor quality shoot buds that have a low regeneration frequency. However, treating the explants with high concentrations (10–60 mg/L of TDZ solution for certain time periods (5–80 min significantly increased the regeneration frequency and improved the quality of the regenerated shoot buds. The best shoot buds induction (88.42% and number of shoot buds (12.67 per explant were observed when in vitro explants were treated with 20 mg/L TDZ solution for 20 min before being transferred on hormone-free medium after 30 days. Regeneration was also influenced by the orientation (horizontal or vertical of the explants on the medium, and by the origin of the cotyledonary petioles (in vitro or in vivo used for the preparation of explants. We performed subsequent experiments for elongation and rooting of the regenerated shoot buds. Addition of L-arginine to the medium was conducive to the elongation of the shoot buds. A concentration of 7.5 mg/L L-arginine yielded the best results. The elongated shoots could initiate roots to become intact plantlets in half-strength Murashige and Skoog medium containing 0.1 mg/L indole-3-butyric acid. After acclimatization, these plantlets could be transplanted to the soil and the growth was normal. Therefore, application of the methods described here helped to increase plant regeneration efficiency.

  14. Induction of mesenchymal stem cell chondrogenic differentiation and functional cartilage microtissue formation for in vivo cartilage regeneration by cartilage extracellular matrix-derived particles. (United States)

    Yin, Heyong; Wang, Yu; Sun, Zhen; Sun, Xun; Xu, Yichi; Li, Pan; Meng, Haoye; Yu, Xiaoming; Xiao, Bo; Fan, Tian; Wang, Yiguo; Xu, Wenjing; Wang, Aiyuan; Guo, Quanyi; Peng, Jiang; Lu, Shibi


    We propose a method of preparing a novel cell carrier derived from natural cartilage extracellular matrix (ECM), designated cartilage ECM-derived particles (CEDPs). Through a series of processes involving pulverization, sieving, and decellularization, fresh cartilage was made into CEDPs with a median diameter of 263 ± 48 μm. Under microgravity culture conditions in a rotary cell culture system (RCCS), bone marrow stromal cells (BMSCs) can proliferate rapidly on the surface of CEDPs with high viability. Histological evaluation and gene expression analysis indicated that BMSCs were differentiated into mature chondrocytes after 21 days of culture without the use of exogenous growth factors. Functional cartilage microtissue aggregates of BMSC-laden CEDPs formed as time in culture increased. Further, the microtissue aggregates were directly implanted into trochlear cartilage defects in a rat model (CEDP+MSC group). Gait analysis and histological results indicated that the CEDP+MSC group obtained better and more rapid joint function recovery and superior cartilage repair compared to the control groups, in which defects were treated with CEDPs alone or only fibrin glue, at both 6 and 12 weeks after surgery. In conclusion, the innovative cell carrier derived from cartilage ECM could promote chondrogenic differentiation of BMSCs, and the direct use of functional cartilage microtissue facilitated cartilage regeneration. This strategy for cell culture, stem cell differentiation and one-step surgery using cartilage microtissue for cartilage repair provides novel prospects for cartilage tissue engineering and may have further broad clinical applications. We proposed a method to prepare a novel cell carrier derived from natural cartilage ECM, termed cartilage ECM-derived particles (CEDPs), which can support proliferation of MSCs and facilitate their chondrogenic differentiation. Further, the direct use of functional cartilage microtissue of MSC-laden CEDP aggregates for

  15. Human elastic cartilage engineering from cartilage progenitor cells using rotating wall vessel bioreactor. (United States)

    Takebe, T; Kobayashi, S; Kan, H; Suzuki, H; Yabuki, Y; Mizuno, M; Adegawa, T; Yoshioka, T; Tanaka, J; Maegawa, J; Taniguchi, H


    Transplantation of bioengineered elastic cartilage is considered to be a promising approach for patients with craniofacial defects. We have previously shown that human ear perichondrium harbors a population of cartilage progenitor cells (CPCs). The aim of this study was to examine the use of a rotating wall vessel (RWV) bioreactor for CPCs to engineer 3-D elastic cartilage in vitro. Human CPCs isolated from ear perichondrium were expanded and differentiated into chondrocytes under 2-D culture conditions. Fully differentiated CPCs were seeded into recently developed pC-HAp/ChS (porous material consisted of collagen, hydroxyapatite, and chondroitinsulfate) scaffolds and 3-D cultivated utilizing a RWV bioreactor. 3-D engineered constructs appeared shiny with a yellowish, cartilage-like morphology. The shape of the molded scaffold was maintained after RWV cultivation. Hematoxylin and eosin staining showed engraftment of CPCs inside pC-HAp/ChS. Alcian blue and Elastica Van Gieson staining showed of proteoglycan and elastic fibers, which are unique extracellular matrices of elastic cartilage. Thus, human CPCs formed elastic cartilage-like tissue after 3-D cultivation in a RWV bioreactor. These techniques may assist future efforts to reconstruct complicate structures composed of elastic cartilage in vitro. Copyright © 2012 Elsevier Inc. All rights reserved.

  16. Role of Chondrocytes in Cartilage Formation, Progression of Osteoarthritis and Cartilage Regeneration

    Directory of Open Access Journals (Sweden)

    Hemanth Akkiraju


    Full Text Available Articular cartilage (AC covers the diarthrodial joints and is responsible for the mechanical distribution of loads across the joints. The majority of its structure and function is controlled by chondrocytes that regulate Extracellular Matrix (ECM turnover and maintain tissue homeostasis. Imbalance in their function leads to degenerative diseases like Osteoarthritis (OA. OA is characterized by cartilage degradation, osteophyte formation and stiffening of joints. Cartilage degeneration is a consequence of chondrocyte hypertrophy along with the expression of proteolytic enzymes. Matrix Metalloproteinases (MMPs and A Disintegrin and Metalloproteinase with Thrombospondin Motifs (ADAMTS are an example of these enzymes that degrade the ECM. Signaling cascades involved in limb patterning and cartilage repair play a role in OA progression. However, the regulation of these remains to be elucidated. Further the role of stem cells and mature chondrocytes in OA progression is unclear. The progress in cell based therapies that utilize Mesenchymal Stem Cell (MSC infusion for cartilage repair may lead to new therapeutics in the long term. However, many questions are unanswered such as the efficacy of MSCs usage in therapy. This review focuses on the role of chondrocytes in cartilage formation and the progression of OA. Moreover, it summarizes possible alternative therapeutic approaches using MSC infusion for cartilage restoration.

  17. Plant regeneration from cotyledonary explants of Eucalyptus camaldulensis Regeneração de plantas de Eucalyptus camaldulensis a partir das explantes cotiledonares

    Directory of Open Access Journals (Sweden)

    Roberson Dibax


    Full Text Available Breeding methods based on genetic transformation techniques need to be implemented for Eucalyptus camaldulensis to shorten the long breeding cycles and avoid manipulation of adult trees; that requires the development of plant regeneration protocols enabling development of plants from transformed tissues. The present work aimed to optimise the regeneration process already established for the species. Cotyledonary leaves of E. camaldulensis were cultured in MS medium supplemented with naphthaleneacetic acid (NAA and 6-benzylaminopurine (BAP combinations. The most efficient treatment for bud indirect regeneration (2.7 µmol L-1 NAA and 4.44 µmol L-1 BAP was used for further experiments. When explants were kept in the dark during the first 30 days, the percentage of explants forming calluses increased and explant necrosis was reduced in comparison with light-cultured explants. Mineral medium modifications were compared and half-strength MS mineral medium turned out to be as efficient as full-strength medium, producing 54% and 47% of explants with buds, respectively. For shoot elongation, MS medium with half-strength nitrate and ammonium salts, and 0.2% activated charcoal yielded rooted shoots 1 to 8 cm high after one month. The procedure is an efficient protocol for E. camadulensis plant regeneration, reducing the stages necessary for the obtention of complete plants.A implementação, para espécies florestais, de técnicas de melhoramento baseadas em métodos de transformação genética, permitirá reduzir os longos ciclos de melhoramento e evitar a manipulação de árvores adultas. Isto implica dispor de um protocolo de regeneração que permita o desenvolvimento de plantas a partir de tecidos transformados. Este trabalho teve como objetivo otimizar este protocolo de regeneração para Eucalyptus camaldulensis. Folhas cotiledonares foram cultivadas em meio de cultura MS suplementado com combinações de ácido naftalenoacético (ANA e 6

  18. Changes in glycemic control and body weight after explantation of the duodenal-jejunal bypass liner. (United States)

    Betzel, Bark; Koehestanie, Parviez; Homan, Jens; Aarts, Edo O; Janssen, Ignace M C; de Boer, Hans; Wahab, Peter J; Groenen, Marcel J M; Berends, Frits J


    The duodenal-jejunal bypass liner (DJBL) is an endoscopic device that induces weight loss and improves glycemic control in patients with type 2 diabetes mellitus (T2DM). The aim of the current study was to assess the effects of DJBL explantation on glycemic control and body weight. This prospective, observational study included only patients with T2DM who had the DJBL implanted for at least 6 months and had a follow-up of at least 12 months after explantation. The primary endpoints were changes in glycosylated hemoglobin A1c (HbA1c) and body weight during the 12 months after explantation. Secondary endpoints were changes in fasting plasma glucose, blood pressure, and plasma lipid levels. In total, 59 patients completed the 12-month follow-up after explantation. During this period body weight increased by 5.6 (standard deviation, 6.4) kg (P body weight remained 8.0 (SD 8.6) kg (P body weight loss of 7.4% (SD 7.6) (P weight gain and worsening of glycemic control, although some beneficial effects remained detectable 12 months after explantation. A change in strategy is needed to preserve the beneficial effects of DJBL treatment. (Clinical trial registration number: 746∖100111.). Copyright © 2017 American Society for Gastrointestinal Endoscopy. Published by Elsevier Inc. All rights reserved.

  19. Equine oviduct explant culture: a basic model to decipher embryo-maternal communication. (United States)

    Nelis, Hilde; D'Herde, Katharina; Goossens, Karen; Vandenberghe, Lynn; Leemans, Bart; Forier, Katrien; Smits, Katrien; Braeckmans, Kevin; Peelman, Luc; Van Soom, Ann


    Equine embryos remain for 6 days in the oviduct and thus there is a need for an in vitro model to study embryo-oviductal interactions in the horse, since this subtle way of communication is very difficult to analyse in vivo. Until now, no equine oviduct explant culture model has been characterised both morphologically and functionally. Therefore, we established a culture system for equine oviduct explants that maintained epithelial morphology during 6 days of culture, as revealed by light microscopy and transmission electron microscopy. We demonstrated the presence of highly differentiated, tall columnar, pseudostratified epithelium with basal nuclei, numerous nucleoli, secretory granules and apical cilia, which is very similar to the in vivo situation. Both epithelium and stromal cells originating from the lamina propria are represented in the explants. Moreover, at least 98% of the cells remained membrane intact and fewer than 2% of the cells were apoptotic after 6 days of culture. Although dark-cell degeneration, which is a hypoxia-related type of cell death, was observed in the centre of the explants, quantitative real-time PCR failed to detect upregulation of the hypoxia-related marker genes HIF1A, VEGFA, uPA, GLUT1 and PAI1. Since the explants remained morphologically and functionally intact and since the system is easy to set up, it appears to be an excellent tool for proteome, transcriptome and miRNome analysis in order to unravel embryo-maternal interactions in the horse.

  20. Optical and atomic force microscopy of an explanted AcrySof intraocular lens with glistenings. (United States)

    Dogru, M; Tetsumoto, K; Tagami, Y; Kato, K; Nakamae, K


    To assess the surface morphology and cause of glistenings in an explanted AcrySof intraocular lens (IOL). Shakai Hoken Kobe Central Hospital, Kobe, Japan. A 63-year-old Japanese man had implantation of an AcrySof IOL in the capsular bag. One month postoperatively, he had a neodymium:YAG laser capsulotomy for posterior capsule opacification, which changed the IOL's position in the capsular bag. A few months later, the patient developed disabling night glare from intralenticular glistenings and progressive hyperopic refractive error. The IOL was explanted and then analyzed by optical microscopy and atomic force microscopy (AFM). Laboratory analysis of control AcrySof IOLs kept in a balanced salt solution at steady room and body temperature for 2 months was also performed to evaluate the cause of the glistenings observed clinically. Optical microscopy showed that the explanted AcrySof IOL had several microvacuoles; no abnormalities were observed in the control AcrySof IOLs before or after folding at the room and body temperatures. The AFM analysis showed a significant change in the surface morphology of the explanted IOL, including vacuolar formations in the posterior surface as well as numerous anterior surface irregularities. No microvacuoles or surface morphology alterations were observed in the control AcrySof IOLs by AFM analysis. The glistenings in the explanted AcrySof IOL were likely caused by temperature changes and not mechanical stress from folding.

  1. Ex Vivo Produced Oral Mucosa Equivalent by Using the Direct Explant Cell Culture Technique

    Directory of Open Access Journals (Sweden)

    Kamile Öztürk


    Full Text Available Objective: The aim of this study is the histological and immunohistochemical evaluation of ex vivo produced oral mucosal equivalents using keratinocytes cultured by direct explant technique.Material and Methods: Oral mucosa tissue samples were obtained from the keratinized gingival tissues of 14 healthy human subjects. Human oral mucosa keratinocytes from an oral mucosa biopsy specimen were dissociated by the explant technique. Once a sufficient population of keratinocytes was reached, they were seeded onto the type IV collagen coated “AlloDerm” and taken for histological and immunohistochemical examinations at 11 days postseeding of the keratinocytes on the cadaveric human dermal matrix.Results: Histopathologically and immunohistochemically, 12 out of 14 successful ex vivo produced oral mucosa equivalents (EVPOME that consisted of a stratified epidermis on a dermal matrix have been developed with keratinocytes cultured by the explant technique.Conclusion: The technical handling involved in the direct explant method at the beginning of the process has fewer steps than the enzymatic method and use of the direct explant technique protocol for culturing of human oral mucosa keratinocyte may be more adequate for EVPOME production.

  2. Body weight independently affects articular cartilage catabolism. (United States)

    Denning, W Matt; Winward, Jason G; Pardo, Michael Becker; Hopkins, J Ty; Seeley, Matthew K


    Although obesity is associated with osteoarthritis, it is unclear whether body weight (BW) independently affects articular cartilage catabolism (i.e., independent from physiological factors that also accompany obesity). The primary purpose of this study was to evaluate the independent effect of BW on articular cartilage catabolism associated with walking. A secondary purpose was to determine how decreased BW influenced cardiovascular response due to walking. Twelve able-bodied subjects walked for 30 minutes on a lower-body positive pressure treadmill during three sessions: control (unadjusted BW), +40%BW, and -40%BW. Serum cartilage oligomeric matrix protein (COMP) was measured immediately before (baseline) and after, and 15 and 30 minutes after the walk. Heart rate (HR) and rate of perceived exertion (RPE) were measured every three minutes during the walk. Relative to baseline, average serum COMP concentration was 13% and 5% greater immediately after and 15 minutes after the walk. Immediately after the walk, serum COMP concentration was 14% greater for the +40%BW session than for the -40%BW session. HR and RPE were greater for the +40%BW session than for the other two sessions, but did not differ between the control and -40%BW sessions. BW independently influences acute articular cartilage catabolism and cardiovascular response due to walking: as BW increases, so does acute articular cartilage catabolism and cardiovascular response. These results indicate that lower-body positive pressure walking may benefit certain individuals by reducing acute articular cartilage catabolism, due to walking, while maintaining cardiovascular response. Key pointsWalking for 30 minutes with adjustments in body weight (normal body weight, +40% and -40% body weight) significantly influences articular cartilage catabolism, measured via serum COMP concentration.Compared to baseline levels, walking with +40% body weight and normal body weight both elicited significant increases in

  3. Shear and Compression Bioreactor for Cartilage Synthesis. (United States)

    Shahin, Kifah; Doran, Pauline M


    Mechanical forces, including hydrodynamic shear, hydrostatic pressure, compression, tension, and friction, can have stimulatory effects on cartilage synthesis in tissue engineering systems. Bioreactors capable of exerting forces on cells and tissue constructs within a controlled culture environment are needed to provide appropriate mechanical stimuli. In this chapter, we describe the construction, assembly, and operation of a mechanobioreactor providing simultaneous dynamic shear and compressive loading on developing cartilage tissues to mimic the rolling and squeezing action of articular joints. The device is suitable for studying the effects of mechanical treatment on stem cells and chondrocytes seeded into three-dimensional scaffolds.

  4. Organogênese de explante foliar de clones de Eucalyptus grandis x E. urophylla Organogenesis of the leaf explant of Eucalyptus grandis x E. urophylla clones

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    Elisa Cristina Soares de Carvalho Alves


    Full Text Available O objetivo deste trabalho foi avaliar os efeitos dos reguladores de crescimento TDZ [1-fenil-3-(1,2,3-tia-diazol-5-iluréia], BAP (6-benzilaminopurina e ANA (ácido naftalenoacético no desempenho da propagação in vitro por organogênese de explante foliar de três clones híbridos de Eucalyptus grandis x Eucalyptus urophylla. Houve resposta diferenciada dos clones quanto a intensidade, textura e coloração dos calos, em razão dos tratamentos com os reguladores de crescimento. Os melhores resultados de calejamento dos três genótipos foram observados nos tratamentos com a combinação dos reguladores de crescimento TDZ (0,5 mg L-1 e ANA (0,1 mg L-1, obtendo-se 100% de calejamento no explante foliar. Os piores resultados de calejamento foram observados nos tratamentos com a combinação dos reguladores de crescimento BAP (0,1 mg L-1 e ANA (0,1 mg L-1. Em relação à regeneração, a melhor resposta foi obtida com 1,0 mg L-1 BAP em que 8% dos calos formados a partir de explantes foliares regeneraram gemas, com número médio destas formadas por calo igual a 4,2.The aim of this work was to evaluate the effects of growth regulators TDZ [1-phenil-3-(1,2,3-thiadiazol-5-yl urea], BAP (6-benzilaminopurine e NAA (Naphthalene acetic acid on the in vitro propagation by organogenesis from foliar explants of Eucalyptus grandis x E. urophylla. Depending on the clone used, there were singular responses to growth regulators treatment regarding callusing intensity, texture and color. The best results of the three genotypes used were observed with the TDZ (0.5 mg L-1 and NAA (0.1 mg L-1 treatment, where 100% of the foliar explants presented callus. The worst results were observed with the BAP (0.1 mg L-1 and NAA (0.1 mg L-1 treatment. Subsequently, considering the regeneration process, the best response was achieved with 1.0 mg L-1 BAP, in which 8% of the calli regenerated buds, with an average of 4.2 buds per explant.

  5. Bovine parainfluenza-3 virus. (United States)

    Ellis, John A


    Bovine parainfluenza-3 virus (bPI(3)V) is a long-recognized, currently underappreciated, endemic infection in cattle populations. Clinical disease is most common in calves with poor passive transfer or decayed maternal antibodies. It is usually mild, consisting of fever, nasal discharge, and dry cough. Caused at least partly by local immunosuppressive effects, bPI(3)V infection is often complicated by coinfection with other respiratory viruses and bacteria, and is therefore an important component of enzootic pneumonia in calves and bovine respiratory disease complex in feedlot cattle. Active infection can be diagnosed by virus isolation from nasal swabs, or IF testing on smears made from nasal swabs. Timing of sampling is critical in obtaining definitive diagnostic test results. Parenteral and intranasal modified live vaccine combination vaccines are available. Priming early in calfhood with intranasal vaccine, followed by boosting with parenteral vaccine, may be the best immunoprophylactic approach. Copyright © 2010 Elsevier Inc. All rights reserved.

  6. Camel and bovine chymosin

    DEFF Research Database (Denmark)

    Jensen, Jesper Langholm; Mølgaard, Anne; Poulsen, Jens-Christian Navarro


    Bovine and camel chymosin are aspartic peptidases that are used industrially in cheese production. They cleave the Phe105-Met106 bond of the milk protein κ-casein, releasing its predominantly negatively charged C-terminus, which leads to the separation of the milk into curds and whey. Despite...... chymosin. Both enzymes possess local positively charged patches on their surface that can play a role in interactions with the overall negatively charged C-terminus of κ-casein. Camel chymosin contains two additional positive patches that favour interaction with the substrate. The improved electrostatic...... interactions arising from variation in the surface charges and the greater malleability both in domain movements and substrate binding contribute to the better milk-clotting activity of camel chymosin towards bovine milk....

  7. Mycotic bovine nasal granuloma

    Directory of Open Access Journals (Sweden)

    Conti Díaz Ismael Alejandro


    Full Text Available A case of mycotic bovine nasal granuloma in a 10 year-old Jersey cow, produced by Drechslera halodes is presented. Histopathological sections showed abundant hyaline and pigmented extra and intracellular fungal structures together with a polymorphic cellular granuloma formed by neutrophils, lymphocytes, plasmocytes, histiocytes and giant cells of the Langhans type. It is the first case of mycotic bovine nasal granuloma recognized in Uruguay although this disease seems to be frequent according to the opinion of veterinarian specialists. Another similar clinical case also in a Jersey cow from the same dairy house with an intense cellular infiltrate rich in eosinophils without granulomatous image, together with extracellular hyaline and fuliginous fungal forms, is also referred for comparative purposes. Geotrichum sp. was isolated. The need of an early diagnosis and treatment of the disease is stressed.

  8. Explant culture of human peripheral lung. I. Metabolism of benzo[alpha]pyrene

    DEFF Research Database (Denmark)

    Stoner, G.D.; Harris, C.C.; Autrup, Herman


    hydroxylase activity and could metabolize BP into forms that were bound to cellular DNA and protein. Peripheral lung had significantly lower aryl hydrocarbon hydroxylase activity than cultured bronchus but both tissues had similar binding levels of BP to DNA. Radioautographic studies indicated that all cell......Human lung explants have been maintained in vitro for a period of 25 days. Autoradiographic studies indicated that the broncholar epithelial cells, type 2 alveolar epithelial cells, and stromal fibroblasts incorporated 3H-thymidine during the culture. After 7 to 10 days, type 2 cells were...... the predominant alveolar epithelial cell type. Lamellar inclusion bodies were released from the type 2 cells and accumulated in the alveolar spaces. The metabolism of benzo[alpha]pyrene (BP) in human lung explants cultured for up to 7 days was investigated. Human lung explants had measurable aryl hydrocarbon...

  9. Resolving browning during the establishment of explant cultures in Vicia faba L. for genetic transformation

    Directory of Open Access Journals (Sweden)

    Helena Klenotičová


    Full Text Available Optimisation of in vitro regeneration systems of two explant types for low-tannine cultivars of faba bean based on culturing of shoot apices and cotyledonary nodes were provided by usage of various antioxidants - ascorbic acid, citric acid, glutathione and activated charcoal. In subsequent testing, the combined effects of antioxidants with transformation co-cultivation compounds acetosyringone and L-cysteine was studied. The application of antioxidants lead to decreased callogenesis, citric acids treatments (50 mg.l−1 dramatically decreased necrotic response of explants. However, citric acid, used together with ascorbic acid completely inhibited shoot growth in shoot apex cultures. Glutathion evoked hyperhydricity of explants. Activated charcoal induced rooting on media which are commonly used for shoot proliferation. Combination of acetosyringone with antioxidants influenced shoot proliferation, except of variant with ascorbic acid. Citric acid was the best and universal antioxidant in faba bean in vitro cultures and its use is recommended for faba bean genetic transformation experiments.

  10. Viral infections and bovine mastitis: a review. (United States)

    Wellenberg, G J; van der Poel, W H M; Van Oirschot, J T


    This review deals with the role of viruses in the aetiology of bovine mastitis. Bovine herpesvirus 1, bovine herpesvirus 4, foot-and-mouth disease virus, and parainfluenza 3 virus have been isolated from milk from cows with clinical mastitis. Intramammary inoculations of bovine herpesvirus 1 or parainfluenza 3 virus-induced clinical mastitis, while an intramammary inoculation of foot-and-mouth disease virus resulted in necrosis of the mammary gland. Subclinical mastitis has been induced after a simultaneous intramammary and intranasal inoculation of lactating cows with bovine herpesvirus 4. Bovine leukaemia virus has been detected in mammary tissue of cows with subclinical mastitis, but whether this virus was able to induce bovine mastitis has not been reported. Bovine herpesvirus 2, vaccinia, cowpox, pseudocowpox, vesicular stomatitis, foot-and-mouth disease viruses, and bovine papillomaviruses can play an indirect role in the aetiology of bovine mastitis. These viruses can induce teat lesions, for instance in the ductus papillaris, which result in a reduction of the natural defence mechanisms of the udder and indirectly in bovine mastitis due to bacterial pathogens. Bovine herpesvirus 1, bovine viral diarrhoea virus, bovine immunodeficiency virus, and bovine leukaemia virus infections may play an indirect role in bovine mastitis, due to their immunosuppressive properties. But, more research is warranted to underline their indirect role in bovine mastitis. We conclude that viral infections can play a direct or indirect role in the aetiology of bovine mastitis; therefore, their importance in the aetiology of bovine mastitis and their economical impact needs further attention.

  11. Comparative effects of plant growth regulators on leaf and stem explants of Labisia pumila var. alata. (United States)

    Ling, Anna Pick Kiong; Tan, Kinn Poay; Hussein, Sobri


    Labisia pumila var. alata, commonly known as 'Kacip Fatimah' or 'Selusuh Fatimah' in Southeast Asia, is traditionally used by members of the Malay community because of its post-partum medicinal properties. Its various pharmaceutical applications cause an excessive harvesting and lead to serious shortage in natural habitat. Thus, this in vitro propagation study investigated the effects of different plant growth regulators (PGRs) on in vitro leaf and stem explants of L. pumila. The capabilities of callus, shoot, and root formation were evaluated by culturing both explants on Murashige and Skoog (MS) medium supplemented with various PGRs at the concentrations of 0, 1, 3, 5, and 7 mg/L. Medium supplemented with 3 mg/L indole-3-butyric acid (IBA) showed the optimal callogenesis from both leaf and stem explants with (72.34 ± 19.55)% and (70.40 ± 14.14)% efficacy, respectively. IBA was also found to be the most efficient PGR for root induction. A total of (50.00 ± 7.07)% and (77.78 ± 16.47)% of root formation were obtained from the in vitro stem and leaf explants after being cultured for (26.5 ± 5.0) and (30.0 ± 8.5) d in the medium supplemented with 1 and 3 mg/L of IBA, respectively. Shoot formation was only observed in stem explant, with the maximum percentage of formation ((100.00 ± 0.00)%) that was obtained in 1 mg/L zeatin after (11.0 ± 2.8) d of culture. Callus, roots, and shoots can be induced from in vitro leaf and stem explants of L. pumila through the manipulation of types and concentrations of PGRs.

  12. Prolactin expression and secretion by human breast glandular and adipose tissue explants. (United States)

    Zinger, Michael; McFarland, Molly; Ben-Jonathan, Nira


    Prolactin (PRL) is a 23-kDa hormone produced by the pituitary and extrapituitary sites. The main target of PRL is the breast, where it affects cellular growth, differentiation, and milk production. Recent evidence suggests that locally produced PRL plays a role in breast tumorigenesis. Our objective was to examine PRL synthesis/release in different tissues of the human breast and determine the effect of ovarian steroids. Breast tissue, obtained from women undergoing mastectomy or breast reduction, was separated into glandular (nonmalignant) and adipose explants and incubated for 10 d. Conditioned media were analyzed for PRL by a bioassay. PRL release from glandular explants decreased by 60% from d 1-3, followed by a 4-fold increase on d 10. PRL release from adipose explants was unchanged from d 1-3 and increased more than 10-fold by d 10. PRL gene expression, determined by RT-PCR, was low on d 0 and markedly increased on d 10 in both types of explants. De novo synthesis of PRL was confirmed by metabolic labeling. Progesterone suppressed PRL release from glandular explants without affecting adipose explants. Estradiol did not alter PRL release from either tissue. In conclusion, the human breast produces and releases bioactive PRL, with a higher release rate by adipose than glandular tissue. The time-dependent rise in PRL release suggests removal from inhibitory control. Progesterone may be one of the factors that suppresses PRL production in the glandular compartment, whereas the factor(s) that regulate adipose PRL are unknown. These data suggest an autocrine/paracrine role for PRL in human glandular and adipose breast tissue.

  13. Analysis of Explanted Magnetically Controlled Growing Rods From Seven UK Spinal Centers. (United States)

    Joyce, Thomas J; Smith, Simon L; Rushton, Paul R P; Bowey, Andrew J; Gibson, Michael J


    Analysis of explanted MAGnetic Expansion Control (MAGEC) growing rods. To analyze explanted MAGEC rods used in management of early onset scoliosis and identify the mode of failure in such cases. Magnetically controlled growing rods are increasingly used as the option of choice for early onset scoliosis. However, being more complex than conventional growing rods they are perhaps more likely to succumb to multifarious failure modes. In addition, metallosis has been reported around failed MAGEC rods. Explanted MAGEC rods from seven UK spinal centers were obtained for independent analysis. Thirty-four MAGEC rods, from 18 children, explanted for reasons including failure of rod lengthening and maximum rod distraction reached, were cut open to allow internal components to be evaluated and assessed. Externally, all MAGEC rods showed localized marks, which were termed "growth marks" as they indicated growth of the rod in vivo, on the extending bar component. After cutting open, titanium wear debris was found inside all 34 (100%) MAGEC rods. Ninety-one percent (31/34) of MAGEC rods showed measurable wear of the extending bar, towards the magnet end. Substantial damage to the radial bearing was seen inside 74% (25/34) of MAGEC rods while O-ring seal failure was seen in 53% (18/34) of cases. In 44% (15/34) of MAGEC rods the drive pin was fractured but this was felt to be an effect of rod failure, not a cause. The combination of high volumes of titanium wear debris alongside O-ring seal damage likely accounts for the metallosis reported clinically around some MAGEC rods. Based on this explant data, a failure mechanism in MAGEC rods due to the natural off axis loading in the spine was proposed. This is the largest data set reporting a complete analysis of explanted MAGEC rods to date. 4.

  14. The effect of glucocorticoids on tendon cell viability in human tendon explants (United States)

    Lui, Wai Ting; Chuen Fu, Sai; Man Lee, Kwong


    Background and purpose Previous studies on the culture of human tenocytes have shown that dexamethasone and triamcino-lone reduce cell viability, suppress cell proliferation, and reduce collagen synthesis. However, such cell cultures lack the extracellular matrix and three-dimensional structure of normal tendons, which affects their response to stimuli. We established a human tendon explant culture system and tested the effects of dexamethasone and triamcinolone on cell viability. Methods Primary human tendon explant cultures were prepared from healthy hamstring tendons. Tendon strips were harvested from hamstring tendons and cultured in 24-well plates in Dulbecco’s modification of Eagle’s Medium (DMEM) supplemented with 2% fetal calf serum. The tendon explants were treated with 0 μM (control), 10 μM, or 100 μM dexamethasone sodium phosphate or 0 μM (control), 10 μM, or 100 μM triamcinolone acetonide in DMEM for 96 h. Cell viability was measured by Alamar blue assay before and after glucocorticoid treatment. Results Incubation with 10 μM and 100 μM dexamethasone reduced cell viability in human tendon explants by 35% and 45%, respectively, as compared to a 6% increase in the controls (p = 0.01, mixed-effects ANOVA). Triamcinolone at 10 μM and 100 μM reduced cell viability by 33% and 36%, respectively, as compared to a 9% increase in the controls (p = 0.07, mixed-effects ANOVA). Interpretation Human tendon explant cultures can be used to study the effects of glucocorticoids on human tendon. Dexamethasone and triamcinolone suppress the cell viability of human tendon in its natural 3-dimensional environment with matrix anchorage. Human tendon explant cultures provide a species-specific model for further investigation of the effects of glucocorticoids on the metabolism of the extracellular matrix of human tendon, and on its mechanical properties. PMID:19421908

  15. Diagnostic imaging in bovine orthopedics. (United States)

    Kofler, Johann; Geissbühler, Urs; Steiner, Adrian


    Although a radiographic unit is not standard equipment for bovine practitioners in hospital or field situations, ultrasound machines with 7.5-MHz linear transducers have been used in bovine reproduction for many years, and are eminently suitable for evaluation of orthopedic disorders. The goal of this article is to encourage veterinarians to use radiology and ultrasonography for the evaluation of bovine orthopedic disorders. These diagnostic imaging techniques improve the likelihood of a definitive diagnosis in every bovine patient but especially in highly valuable cattle, whose owners demand increasingly more diagnostic and surgical interventions that require high-level specialized techniques. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. Advances and Prospects in Stem Cells for Cartilage Regeneration (United States)

    Wang, Mingjie; Yuan, Zhiguo; Ma, Ning; Hao, Chunxiang; Guo, Weimin; Zou, Gengyi; Zhang, Yu; Chen, Mingxue; Gao, Shuang; Wang, Aiyuan; Wang, Yu; Sui, Xiang; Xu, Wenjing; Lu, Shibi


    The histological features of cartilage call attention to the fact that cartilage has a little capacity to repair itself owing to the lack of a blood supply, nerves, or lymphangion. Stem cells have emerged as a promising option in the field of cartilage tissue engineering and regenerative medicine and could lead to cartilage repair. Much research has examined cartilage regeneration utilizing stem cells. However, both the potential and the limitations of this procedure remain controversial. This review presents a summary of emerging trends with regard to using stem cells in cartilage tissue engineering and regenerative medicine. In particular, it focuses on the characterization of cartilage stem cells, the chondrogenic differentiation of stem cells, and the various strategies and approaches involving stem cells that have been used in cartilage repair and clinical studies. Based on the research into chondrocyte and stem cell technologies, this review discusses the damage and repair of cartilage and the clinical application of stem cells, with a view to increasing our systematic understanding of the application of stem cells in cartilage regeneration; additionally, several advanced strategies for cartilage repair are discussed. PMID:28246531

  17. Advances and Prospects in Stem Cells for Cartilage Regeneration

    Directory of Open Access Journals (Sweden)

    Mingjie Wang


    Full Text Available The histological features of cartilage call attention to the fact that cartilage has a little capacity to repair itself owing to the lack of a blood supply, nerves, or lymphangion. Stem cells have emerged as a promising option in the field of cartilage tissue engineering and regenerative medicine and could lead to cartilage repair. Much research has examined cartilage regeneration utilizing stem cells. However, both the potential and the limitations of this procedure remain controversial. This review presents a summary of emerging trends with regard to using stem cells in cartilage tissue engineering and regenerative medicine. In particular, it focuses on the characterization of cartilage stem cells, the chondrogenic differentiation of stem cells, and the various strategies and approaches involving stem cells that have been used in cartilage repair and clinical studies. Based on the research into chondrocyte and stem cell technologies, this review discusses the damage and repair of cartilage and the clinical application of stem cells, with a view to increasing our systematic understanding of the application of stem cells in cartilage regeneration; additionally, several advanced strategies for cartilage repair are discussed.

  18. Automatic detection of diseased regions in knee cartilage (United States)

    Qazi, Arish A.; Dam, Erik B.; Olsen, Ole F.; Nielsen, Mads; Christiansen, Claus


    Osteoarthritis (OA) is a degenerative joint disease characterized by articular cartilage degradation. A central problem in clinical trials is quantification of progression and early detection of the disease. The accepted standard for evaluating OA progression is to measure the joint space width from radiographs however; there the cartilage is not visible. Recently cartilage volume and thickness measures from MRI are becoming popular, but these measures don't account for the biochemical changes undergoing in the cartilage before cartilage loss even occurs and therefore are not optimal for early detection of OA. As a first step, we quantify cartilage homogeneity (computed as the entropy of the MR intensities) from 114 automatically segmented medial compartments of tibial cartilage sheets from Turbo 3D T 1 sequences, from subjects with no, mild or severe OA symptoms. We show that homogeneity is a more sensitive technique than volume quantification for detecting early OA and for separating healthy individuals from diseased. During OA certain areas of the cartilage are affected more and it is believed that these are the load-bearing regions located at the center of the cartilage. Based on the homogeneity framework we present an automatic technique that partitions the region on the cartilage that contributes to maximum homogeneity discrimination. These regions however, are more towards the noncentral regions of the cartilage. Our observation will provide valuable clues to OA research and may lead to improving treatment efficacy.

  19. Spatially resolved elemental distributions in articular cartilage

    Energy Technology Data Exchange (ETDEWEB)

    Reinert, T. E-mail:; Reibetanz, U.; Vogt, J.; Butz, T.; Werner, A.; Gruender, W


    In this study, the nuclear microprobe technique is employed to analyse the chemistry of joint cartilage in order to correlate internal structures of the collagen network with the elemental distribution. The samples were taken from pig's knee joint. 30 {mu}m thick coronar cross-sections were prepared by means of cryosectioning and freeze-drying. We performed simultaneously particle induced X-ray emission (PIXE), Rutherford backscattering spectrometry (RBS) and elastic recoil detection analysis (ERDA). Thus we obtained spatially resolved distributions of the elements H, C, N, O, P, S, Cl, K and Ca. The main components of the organic matrix are H, C, N and O. It was shown that their relations vary with the cartilage structures. It could be shown that zones with aligned collagen fibrils contain less sulphur and potassium but more chlorine. The higher chlorine concentration is remarkable because newest biochemical studies found that hypochloric acid is involved in cartilage degradation. Furthermore, the calcium distribution is still of great interest. Its correlation to structural changes inside the cartilage is still being discussed. It could be disproved that zones of higher calcium concentration are related to the aligned structures of the collagen network.

  20. Spatially resolved elemental distributions in articular cartilage (United States)

    Reinert, T.; Reibetanz, U.; Vogt, J.; Butz, T.; Werner, A.; Gründer, W.


    In this study, the nuclear microprobe technique is employed to analyse the chemistry of joint cartilage in order to correlate internal structures of the collagen network with the elemental distribution. The samples were taken from pig's knee joint. 30 μm thick coronar cross-sections were prepared by means of cryosectioning and freeze-drying. We performed simultaneously particle induced X-ray emission (PIXE), Rutherford backscattering spectrometry (RBS) and elastic recoil detection analysis (ERDA). Thus we obtained spatially resolved distributions of the elements H, C, N, O, P, S, Cl, K and Ca. The main components of the organic matrix are H, C, N and O. It was shown that their relations vary with the cartilage structures. It could be shown that zones with aligned collagen fibrils contain less sulphur and potassium but more chlorine. The higher chlorine concentration is remarkable because newest biochemical studies found that hypochloric acid is involved in cartilage degradation. Furthermore, the calcium distribution is still of great interest. Its correlation to structural changes inside the cartilage is still being discussed. It could be disproved that zones of higher calcium concentration are related to the aligned structures of the collagen network.

  1. PRP and Articular Cartilage: A Clinical Update

    Directory of Open Access Journals (Sweden)

    Antonio Marmotti


    Full Text Available The convincing background of the recent studies, investigating the different potentials of platelet-rich plasma, offers the clinician an appealing alternative for the treatment of cartilage lesions and osteoarthritis. Recent evidences in literature have shown that PRP may be helpful both as an adjuvant for surgical treatment of cartilage defects and as a therapeutic tool by intra-articular injection in patients affected by osteoarthritis. In this review, the authors introduce the trophic and anti-inflammatory properties of PRP and the different products of the available platelet concentrates. Then, in a complex scenario made of a great number of clinical variables, they resume the current literature on the PRP applications in cartilage surgery as well as the use of intra-articular PRP injections for the conservative treatment of cartilage degenerative lesions and osteoarthritis in humans, available as both case series and comparative studies. The result of this review confirms the fascinating biological role of PRP, although many aspects yet remain to be clarified and the use of PRP in a clinical setting has to be considered still exploratory.

  2. Molecular modulation of articular cartilage degradation

    NARCIS (Netherlands)

    Landman, Ellie


    Cartilage homeostasis is maintained due to a balance between anabolic and catabolic processes, that are regulated by a complex network of signaling pathways. Disturbance of one or more of these pathways disrupts this balance, resulting in excessive breakdown of the extracellular matrix and

  3. Zn deposition at the bone-cartilage interface in equine articular cartilage

    Energy Technology Data Exchange (ETDEWEB)

    Bradley, D.A. [Department of Physics, University of Surrey, Guildford, GU2 7XH (United Kingdom)], E-mail:; Moger, C.J.; Winlove, C.P. [School of Physics, University of Exeter, Exeter, EX4 4QL (United Kingdom)


    In articular cartilage metalloproteinases, a family of enzymes whose function relies on the presence of divalent cations such as Zn and Ca plays a central role in the normal processes of growth and remodelling and in the degenerative and inflammatory processes of arthritis. Another important enzyme, alkaline phosphatase, involved in cartilage mineralisation also relies on metallic cofactors. The local concentration of divalent cations is therefore of considerable interest in cartilage pathophysiology and several authors have used synchrotron X-ray fluorescence (XRF) to map metal ion distributions in bone and cartilage. We report use of a bench-top XRF analytical microscope, providing spatial resolution of 10 {mu}m and applicable to histological sections, facilitating correlation of the distribution with structural features. The study seeks to establish the elemental distribution in normal tissue as a precursor to investigation of changes in disease. For six samples prepared from equine metacarpophalangeal joint, we observed increased concentration of Zn and Sr ions around the tidemark between normal and mineralised cartilage. This is believed to be an active site of remodelling but its composition has hitherto lacked detailed characterization. We also report preliminary results on two of the samples using Proton-Induced X-ray Emission (PIXE). This confirms our previous observations using synchrotron-based XRF of enhanced deposition of Sr and Zn at the surface of the subchondral bone and in articular cartilage.

  4. Equivalence and precision of knee cartilage morphometry between different segmentation teams, cartilage regions, and MR acquisitions (United States)

    Schneider, E; Nevitt, M; McCulloch, C; Cicuttini, FM; Duryea, J; Eckstein, F; Tamez-Pena, J


    Objective To compare precision and evaluate equivalence of femorotibial cartilage volume (VC) and mean cartilage thickness (ThCtAB.Me) from independent segmentation teams using identical MR images from three series: sagittal 3D Dual Echo in the Steady State (DESS), coronal multi-planar reformat (DESS-MPR) of DESS and coronal 3D Fast Low Angle SHot (FLASH). Design 19 subjects underwent test-retest MR imaging at 3 Tesla. Four teams segmented the cartilage using prospectively defined plate regions and rules. Mixed models analysis of the pooled data were used to evaluate the effect of acquisition, team and plate on precision and Pearson correlations and mixed models to evaluate equivalence. Results Segmentation team differences dominated measurement variability in most cartilage regions for all image series. Precision of VC and ThCtAB.Me differed significantly by team and cartilage plate, but not between FLASH and DESS. Mean values of VC and ThCtAB.Me differed by team (Pprecision, were generally equivalent, and may be combined for cross-sectional analyses if potential systematic offsets are accounted for. Data from different teams should not be pooled unless equivalence is demonstrated for cartilage metrics of interest. PMID:22521758

  5. Effect of explant type, medium and soil mixture content on Dianthus deltoides L. rooting and acclimatization

    Directory of Open Access Journals (Sweden)

    Marković Marija


    Full Text Available The possibility of developing roots and shoots on Dianthus deltoides nodal and apical cuttings on rooting media was investigated in vitro conditions. The explants developed roots and shoots successfully, and the best results were achieved on MS medium without plant hormones. Number of roots per shoot, length of roots and ramification of roots were not significantly dependent on the medium (NAA-naphthaleneacetic acid concentration, or on the type of explant used (nodal or apical cuttings. The acclimatization rate of microplants was influenced by root numbers and ramification. The transfer to soil was successful, maximum survival rates (96% were achieved on peat - sand (4:1 mixture.

  6. Adherence of Candida albicans to bladder mucosa: development and application of a tissue explant assay. (United States)

    Lyman, C A; Navarro, E; Garrett, K F; Roberts, D D; Pizzo, P A; Walsh, T J


    In order to study the interactions between Candida species and uroepithelial tissue, a tissue explant assay was developed using bladder mucosa harvested from New Zealand white rabbits. Blastoconidia of Candida albicans, Candida tropicalis and Candida glabrata attached to the uroepithelial tissue in similar quantities. However, there was significantly more adherence to the uroepithelium by pre-germinated C. albicans compared with C. albicans blastoconidia. Furthermore, the amount of uroepithelial tissue injury was directly related to the length of exposure of the tissue to Candida. Thus, this tissue explant assay may provide a useful method for investigating properties related to fungal adherence to transitional uroepithelium and organism-mediated tissue injury.

  7. Articular Cartilage Regeneration: An Update of Possible Treatment Approaches

    Directory of Open Access Journals (Sweden)

    Ray Marks


    Full Text Available BACKGROUND: Osteoarthritis, a widespread chronically disabling disorder primarily affecting articular cartilage is said to be irreversible. Researchers have however, been examining processes and methods of promoting articular cartilage repair for some time. QUESTIONS: Can a case be made for the possibility of restoring osteoarthritic cartilage? How advanced is this undertaking? What barriers exist in translating basic studies in the clinical realm? What physical modalities are deemed efficacious in promoting cartilage structure? METHODS: All relevant publications detailing articular cartilage repair themes in the leading databases were examined. Specific emphasis was placed on a broad array of efforts and observations concerning articular cartilage and its repair. Articles of historic significance and more current strategies designed to foster cartilage repair were focused on, and reported in narrative form. Ideas extracted from the voluminous literature were those that answered one or more of the key questions driving this research. RESULTS: Numerous attempts have been made over time to foster cartilage repair, using a variety of approaches such as creating artificial cartilage, and transplanting stem cells into damaged cartilage to promote repair. Most current strategies are forged in laboratories and do not always account for the complex disease process, and the importance mechanical and inflammatory determinants play in the disease. However, manipulating biophysical, and biomechanical stimuli favorably is likely to hold promise for attenuating destruction of/or for fostering cartilage viability and repair, even in the presence of adverse osteoarthritic cartilage tissue changes. CONCLUSION: More work is needed to examine the key upstream determinants leading to articular cartilage destruction, and to enhancing the viability of the tissue. Employing carefully construed therapeutic strategies known to impact articular cartilage homeostasis

  8. Unsaturated phosphatidylcholines lining on the surface of cartilage and its possible physiological roles

    Directory of Open Access Journals (Sweden)

    Crawford Ross W


    Full Text Available Abstract Background Evidence has strongly indicated that surface-active phospholipid (SAPL, or surfactant, lines the surface of cartilage and serves as a lubricating agent. Previous clinical study showed that a saturated phosphatidylcholine (SPC, dipalmitoyl-phosphatidylcholine (DPPC, was effective in the treatment of osteoarthritis, however recent studies suggested that the dominant SAPL species at some sites outside the lung are not SPC, rather, are unsaturated phosphatidylcholine (USPC. Some of these USPC have been proven to be good boundary lubricants by our previous study, implicating their possible important physiological roles in joint if their existence can be confirmed. So far, no study has been conducted to identify the whole molecule species of different phosphatidylcholine (PC classes on the surface of cartilage. In this study we identified the dominant PC molecule species on the surface of cartilage. We also confirmed that some of these PC species possess a property of semipermeability. Methods HPLC was used to analyse the PC profile of bovine cartilage samples and comparisons of DPPC and USPC were carried out through semipermeability tests. Results It was confirmed that USPC are the dominant SAPL species on the surface of cartilage. In particular, they are Dilinoleoyl-phosphatidylcholine (DLPC, Palmitoyl-linoleoyl-phosphatidylcholine, (PLPC, Palmitoyl-oleoyl-phosphatidylcholine (POPC and Stearoyl-linoleoyl-phosphatidylcholine (SLPC. The relative content of DPPC (a SPC was only 8%. Two USPC, PLPC and POPC, were capable of generating osmotic pressure that is equivalent to that by DPPC. Conclusion The results from the current study confirm vigorously that USPC is the endogenous species inside the joint as against DPPC thereby confirming once again that USPC, and not SPC, characterizes the PC species distribution at non-lung sites of the body. USPC not only has better anti-friction and lubrication properties than DPPC, they also

  9. Induction of bulb organogenesis inin vitrocultures of tarda tulip (Tulipa tardaStapf.) from seed-derived explants. (United States)

    Maślanka, Małgorzata; Bach, Anna


    A protocol for obtaining bulbs via in vitro organogenesis was developed for tarda tulip ( Tulipa tarda Stapf). Scale explants were obtained from bulbs formed at the base of seedlings or from adventitious bulbs that developed from callus tissue forming on stolons or on germinating seeds. Some explants were subjected to chilling at 5°C for 12 wk. The culture media contained 3 or 6% sucrose and was supplemented with either no growth regulators, either 0.5 μM 6-benzyl-aminopurine (BAP) or 18.9 or 94.6 μM abscisic acid (ABA). Cultures were maintained in the dark at 20°C. Callus tissue developed mainly on media without growth regulators or with BAP. Callus was formed from up to 96% of explants derived from non-chilled adventitious bulbs that were treated with 3% sucrose and 0.5 μM BAP. Less callus was formed from chilled explants compared with non-chilled explants. Newly formed adventitious bulbs appeared on the explants via direct and indirect organogenesis. The media with BAP promoted the formation of adventitious bulbs at a rate of 56-92% from non-chilled explants, whereas a maximum rate of 36% was observed from chilled explants. ABA inhibited the induction of adventitious bulbs and callus. The adventitious bulbs obtained in these experiments contained a meristem, which was evidence that they had developed properly.

  10. An in vitro model for detecting skin irritants: methyl green-pyronine staining of human skin explant cultures

    NARCIS (Netherlands)

    Jacobs, J. J. L.; Lehé, C.; Cammans, K. D. A.; Das, P. K.; Elliott, G. R.


    We evaluated the potential of human organotypic skin explant cultures (hOSECs) for screening skin irritants. Test chemicals were applied to the epidermis of the skin explants which were incubated for 4, 24 or 48 h in tissue culture medium. A decrease in epidermal RNA staining, visualised in frozen

  11. Medial Femoral Condyle Cartilage Defect Biomechanics: Effect of Obesity, Defect Size, and Cartilage Thickness. (United States)

    Lacy, Kyle W; Cracchiolo, Allison; Yu, Stephen; Goitz, Henry


    Medial femoral condyle (MFC) chondral defects cause knee pain. Clinical studies have shown worse functional outcomes and cartilage defect fill rates after microfracture in obese patients (BMI ≥30) and for defects with size ≥2 cm(2). To determine the effect of obesity, defect size, and cartilage thickness on the force sustained at the base of full-thickness MFC cartilage defects during weightbearing. Controlled laboratory study. Eight human cadaveric knees were loaded in 15° of flexion. A sensor measured force across the medial compartment. The area at the base of the defect protected from load, termed the "area of containment," was quantified, and loads simulating weightbearing for BMIs of 20, 30, and 40 were applied. A full-thickness cartilage defect was created on the MFC. Cycles of loads were applied for defect sizes with diameters of 6, 8, 10, 12, 14, 16, 18, and 20 mm. A second sensor recorded force at the base of the defect for defects with diameters of 14, 16, 18, and 20 mm. Loads simulating BMI ≥30 led to a decrease in the area of containment for all defects ≥14 mm in diameter (P ≤ .038). Base of defect force increased for defects ≥16 mm in diameter (area, ≥2 cm(2)) between loaded and unloaded states (P ≤ .042) and for loads simulating BMI ≥30 (P ≤ .045). Cartilage rim thickness biomechanically unfavorable environment after microfracture in these patient subsets. These biomechanical findings corroborate clinical studies that have noted worse outcomes after microfracture in patients with BMI ≥30 and cartilage defects of size ≥2 cm(2). Further clinical studies are needed to compare microfracture with other cartilage restoration procedures in these patient subsets. © 2015 The Author(s).

  12. Autologous cartilage implantation for full thickness articular cartilage defects of the knee. (United States)

    Wasiak, J; Villanueva, E


    A variety of strategies have been employed for managing articular cartilage defects of the knee, including drilling and abrasion arthroplasty. These treatments are not always effective and when they are, the benefits may only be transitory. Unsuccessfully treated cartilage damage may progress to degenerative disease states and result in the need for a total knee replacement. In recent years the surgical implantation of healthy cartilage cells (autologous cartilage implantation [ACI] ) into damaged areas has been seen as an alternative option and is currently under investigation as a potential improvement over the current strategies for the management and treatment of articular cartilage defects. To determine the effectiveness of ACI in patients with full thickness articular cartilage defects of the knee. We searched the Cochrane Musculoskeletal Injuries Group specialised register (May 2002), Cochrane Controlled Trials Register (The Cochrane Library, Issue 3, 2002), MEDLINE (1966 to June Week 4 2001), CINAHL (1982 to July Week 2 2001), EMBASE (1980 to 2001 Week 27), SPORTDiscus (1949 to June 2001), Current Contents (1993 Week 26 to 2001 Week 30) and the National Research Register (Issue 2, May 2002). Randomised and quasi-randomised trials comparing ACI with any other type of treatment (including no treatment or placebo) for symptomatic cartilage defects of the medial or lateral femoral condyle, trochlea or patella. Two independent reviewers applied the entry criteria to identified studies. No completed randomised controlled trials investigating this treatment were identified through the above searches. One possible trial has been placed in Studies Awaiting Assessment, awaiting translation of the full trial report. Ongoing trials currently underway will be incorporated in future updates of this review. No information is available from RCTs which can influence current practice. Therefore, since current evidence is subject to the inherent weaknesses of case series or

  13. Knee cartilage segmentation and thickness computation from ultrasound images. (United States)

    Faisal, Amir; Ng, Siew-Cheok; Goh, Siew-Li; Lai, Khin Wee


    Quantitative thickness computation of knee cartilage in ultrasound images requires segmentation of a monotonous hypoechoic band between the soft tissue-cartilage interface and the cartilage-bone interface. Speckle noise and intensity bias captured in the ultrasound images often complicates the segmentation task. This paper presents knee cartilage segmentation using locally statistical level set method (LSLSM) and thickness computation using normal distance. Comparison on several level set methods in the attempt of segmenting the knee cartilage shows that LSLSM yields a more satisfactory result. When LSLSM was applied to 80 datasets, the qualitative segmentation assessment indicates a substantial agreement with Cohen's κ coefficient of 0.73. The quantitative validation metrics of Dice similarity coefficient and Hausdorff distance have average values of 0.91 ± 0.01 and 6.21 ± 0.59 pixels, respectively. These satisfactory segmentation results are making the true thickness between two interfaces of the cartilage possible to be computed based on the segmented images. The measured cartilage thickness ranged from 1.35 to 2.42 mm with an average value of 1.97 ± 0.11 mm, reflecting the robustness of the segmentation algorithm to various cartilage thickness. These results indicate a potential application of the methods described for assessment of cartilage degeneration where changes in the cartilage thickness can be quantified over time by comparing the true thickness at a certain time interval.

  14. [Physiology and pathology of the epiphyseal cartilage (author's transl)]. (United States)

    Cotta, H; Rauterberg, K


    Knowledge of the physiology of the epiphyseal cartilage, respectively epiphyseal plate, is essential for an understanding of defective growth and abnormal modeling of the long bones. The epiphyseal cartilage develops from the embryonal, cartilaginous long bone structure. The histology of the epiphyseal cartilage is characterised by definable zones representing the individual differentiation steps from the reformation of cartilage to chondrolysis. Modeling of the ends of the long bones is also influenced by a transversal and longitudinal direction of growth in the epiphyseal cartilage. The intercellular substance mainly contains collagin, proteoglycanes and non-collagenic proteins. These macromolecules are compounded by means of physicochemical bonds and are responsible for the special mechanical qualities of the hyaline cartilage. The process of mineralisation at the base of the epiphyseal cartilage is an essential differentiating step for the ossification processes which take place in the metaphysis. Two pathogenetic principles at the epiphyseal cartilage appear to be important for the defective growth of the long bones. On the one hand, the flowing equilibrium between the differentiation steps of cartilage reformation, transformation of the hyaline cartilage into a mineralised cartilaginous tissue and chondrolysis is changed, whereas on the other hand the turnover of these differentiation steps is retarded or accelerated.

  15. The role of bioreactors in cartilage tissue engineering. (United States)

    Mabvuure, Nigel; Hindocha, Sandip; Khan, Wasim S


    Cartilage tissue engineering is concerned with developing in vitro cartilage implants that closely match the properties of native cartilage, for eventual implantation to replace damaged cartilage. The three components to cartilage tissue engineering are cell source, such as in vitro expanded autologous chondrocytes or mesenchymal progenitor cells, a scaffold onto which the cells are seeded and a bioreactor which attempts to recreate the in vivo physicochemical conditions in which cartilage develops. Although much progress has been made towards the goal of developing clinically useful cartilage constructs, current constructs have inferior physicochemical properties than native cartilage. One of the reasons for this is the neglect of mechanical forces in cartilage culture. Bioreactors have been defined as devices in which biological or biochemical processes can be re-enacted under controlled conditions e.g. pH, temperature, nutrient supply, O2 tension and waste removal. The purpose of this review is to detail the role of bioreactors in the engineering of cartilage, including a discussion of bioreactor designs, current state of the art and future perspectives.

  16. Callus induction and plant regeneration from different explant types of Miscanthus x ogiformis Honda 'Giganteus'

    DEFF Research Database (Denmark)

    Holme, Inger Bæksted; Petersen, Karen Koefoed


    Different explants of Miscanthus x ogiformis Honda 'Giganteus' were tested in order to develop an efficient tissue culture system. Shoot apices, leaf and root sections from in vitro-propagated plants, and leaf and immature inflorescence sections from 6-month-old greenhouse-grown plants were used....

  17. Hypoxia preferentially destroys GABAergic neurons in developing rat neocortex explants in culture

    NARCIS (Netherlands)

    Romijn, H. J.; Ruijter, J. M.; Wolters, P. S.


    The hypothesis that hypoxic ischemia before or during the human birth process preferentially destroys GABAergic nerve cells, particularly in the neocortex, was tested in a tissue culture model system. To that end, rat neocortex explants dissected from 6-day-old rat pups and cultured to a

  18. Influence of genotype and age of explant source on the capacity for ...

    African Journals Online (AJOL)

    Influence of genotype and age of explant source on the capacity for somatic embryogenesis of two Cavendish banana cultivars (Musa acuminata Colla, AAA). M Youssef, A James, A Mayo-Mosqueda, JR Ku-Cauich, R Grijalva-Arango, RM Escobedo-GM ...

  19. Effect of growth regulators and explant types on callus induction in ...

    African Journals Online (AJOL)

    Different concentrations of growth regulators and three types of explants were investigated for their efficiency on callus induction in Telfairia occidentalis with a view of providing baseline information for the development of a callus initiation protocol. Three concentrations of kinetin (KN) (0.1, 3.3 and 5.0 mg/L) in combination ...


    Directory of Open Access Journals (Sweden)



    Full Text Available Observou-se alta contaminação bacteriana nos explantes de mamoeiro introduzidos in vitro, a partir de plantas matrizes desenvolvidas no campo, independentemente da época do ano em que se realizaram as coletas. O uso de desinfestantes superficiais, como álcool e hipoclorito de sódio, garantiram níveis aceitáveis de controle apenas para fungos, não para bactérias. A rifampicina, por tratamento de imersão ou introdução em meio de cultura, controlou satisfatoriamente as contaminações de caráter endofítico, obtendo-se 70% de explantes sadios, sem sinais de fitotoxicidade.High contamination by bacteria was observed in papaya tissue cuttings introduced in vitro from plants grown in the field, independent of the period of the year that samples were collected. The use of alcohol and sodium hypoclorite did not guarantee good bacteria control. Rifampicin, added as an immersion solution treatment or in the culture media, controlled the internal contamination of explants, without damaging the cuttings. Up to 70% of healthy tissue explants were obtained by the use of rifampicin.

  1. How to Perform a Late Surgical Explantation of a CoreValve Aortic Bioprothesis. (United States)

    Hernandez-Vaquero, Daniel; Pascual, Isaac; Diaz, Rocío; Álvarez-Cabo, Rubén; Moris, César; Silva, Jacobo


    As transcatheter techniques expand to younger patients, cardiac surgeons need to know a safe surgical technique to extract these kinds of prostheses. We describe here an adequate surgical strategy for the explantation of a CoreValve prosthesis that was implanted more than 5 years previously. Copyright © 2017 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.

  2. Microbiology of Explanted Suture Segments from Infected and Noninfected Surgical Patients (United States)

    Krepel, Candace J.; Marks, Richard M.; Rossi, Peter J.; Sanger, James; Goldblatt, Matthew; Graham, Mary Beth; Rothenburger, Stephen; Collier, John; Seabrook, Gary R.


    Sutures under selective host/environmental factors can potentiate postoperative surgical site infection (SSI). The present investigation characterized microbial recovery and biofilm formation from explanted absorbable (AB) and nonabsorbable (NAB) sutures from infected and noninfected sites. AB and NAB sutures were harvested from noninfected (70.9%) and infected (29.1%) sites in 158 patients. At explantation, devices were sonicated and processed for qualitative/quantitative bacteriology; selective sutures were processed for scanning electron microscopy (SEM). Bacteria were recovered from 85 (53.8%) explanted sites; 39 sites were noninfected, and 46 were infected. Suture recovery ranged from 11.1 to 574.6 days postinsertion. A significant difference in mean microbial recovery between noninfected (1.2 isolates) and infected (2.7 isolates) devices (P sutures was noted. Biofilm was present in 100% and 66.6% of infected and noninfected devices, respectively (P sutures provide a hospitable surface for microbial adherence: (i) a significant difference in microbial recovery from infected and noninfected sutures was noted, (ii) infected sutures harbored a mixed flora, including multidrug-resistant health care-associated pathogens, and (iii) a significant difference in the presence or absence of a biofilm in infected versus noninfected explanted devices was noted. Further studies to document the benefit of focused risk reduction strategies to minimize suture contamination and biofilm formation postimplantation are warranted. PMID:23175247

  3. Embryogenic cells in Dactylis glomerata L. (Poaceae) explants identified by cell tracking and by SERK expression

    NARCIS (Netherlands)

    Somleva, M.N.; Schmidt, E.D.L.; Vries, de S.C.


    Single mesophyll cells in leaf explants of Dactylis glomerata L. (Dactylis) that were competent to form somatic embryos directly or through callus were identified by semi-automatic cell tracking. These competent cells were a subpopulation of small, isodiametric, cytoplasm-rich cells located close to

  4. Re-use of explanted DDD pacemakers as VDD- clinical utility and cost effectiveness. (United States)

    Namboodiri, K K N; Sharma, Y P; Bali, H K; Grover, A


    Re-use of DDD pulse generators explanted from patients died of unrelated causes is associated with an additional cost of two transvenous leads if implanted as DDD itself, and high rate of infection according to some studies. We studied the clinical and economical aspects of reutilization of explanted DDD pacemakers programmed to VDD mode. Out of 28 patients who received VDD pacemaker during the period, October 2000- September 2001 in the Department of Cardiology, PGIMER, Chandigarh, 5 poor patients were implanted with explanted DDD pulse generators programmed to VDD mode. Each implantation was planned and carried out according to a standard protocol. The age ranged from 45 to 75 (mean-61) years. The indications for pacing were complete heart block (4) and second degree AV block (1). The clinical profile, costs and complications, if any were noted and followed up at regular intervals. The results were compared with patients who received new DDD pulse generators during this period. The additional cost for the atrial lead was not required in these patients. None of these patients had any local site infection. Compared to the two-lead system, the single lead system provided more rapid implantation and minimized complications associated with placement of an atrial lead. The explanted DDD pacemaker can be safely reused as VDD mode with same efficacy in selected patient population. This is associated with lower cost and complications compared to reimplantation as DDD itself.

  5. Production of immunoglobulins in gingival tissue explant cultures from juvenile periodontitis patients

    Energy Technology Data Exchange (ETDEWEB)

    Hall, E.R.; Falkler, W.A. Jr.; Suzuki, J.B. (Univ. of Maryland Dental School, Baltimore (USA))


    B lymphocytes and plasma cells are histologically observed in granulomatous periodontal tissues of juvenile periodontitis (JP) patients. Local immune processes may participate in protective or immunopathologic roles in the pathogenesis of this disease. An in vitro explant culture system was utilized to demonstrate the production of immunoglobulins by diseased JP tissues. Immunodiffusion studies using goat anti-human gamma, alpha, or mu chain serum revealed IgG to be the major immunoglobulin present in 92% of the day 1 supernatant fluids (SF) of the 47 JP gingival tissue explant cultures. IgA was present in 15% of the SF; however, no IgM was detected. Staph Protein A isolated 14C-labeled IgG from the SF, when allowed to react with goat anti-human gamma chain serum, formed lines of precipitation. Positive autoradiographs confirmed the biosynthesis of IgG by the explant cultures. The in vitro gingival tissue explant culture system described provides a useful model for the study of localized immunoglobulins produced by diseased tissues of JP patients.

  6. Staphylococcus aureus induces hypoxia and cellular damage in porcine dermal explants (United States)

    Methicillin-resistant Staphylococcus aureus (MRSA) can infect wounds and produce difficult-to- treat biofilms. To determine the extent that MRSA biofilms can deplete oxygen, change pH and damage host tissue, we developed a porcine dermal explant model on which we cultured GFP-labeled MRSA biofilms. ...


    Directory of Open Access Journals (Sweden)

    Alireza Bonyanpour


    Full Text Available ABSTRACT In this investigation, leaf explants of a local cultivar of dwarf pomegranate were placed on Murashige and Skoog (1962 (MS medium supplemented with various concentrations of 6-benzyl adenin (BA and naphthalene acetic acid (NAA for callus induction. After 40 days, maximum callus induction was observed on a media containing 1 mg L-1 BA and 0.2 to 0.4 mg L-1 NAA. However, the highest callus growth was obtained on a medium containing 1 mg L-1 BA and 1 mg L-1 NAA. The highest number of shoots (7 shoots per explants was obtained by transferring the calli to the media containing 5 mg L-1 BA with 0.1 mg L-1 NAA. Maximum shoot proliferation was observed when shoots were cultured on woody plant medium (WPM supplemented with 5 mg L-1 kinetin (Kin. In this treatment, after 4 subcultures, 36 shoots were produced from one original explant. Among treatments used in rooting experiments, shoots cultured on WPM medium containing 0.2 mg L-1 indol butyric acid (IBA had the maximum root percentage (100% and good root growth (2.06 cm mean length and 2 roots in each explants. Rooted plantlets were cultured in a soil mixture containing vermiculite (60%, perlite (30% and coco peat (10% v/v. After 2 months, 80% of plants survived and transferred to the greenhouse.

  8. Investigating the Skoog-Miller Model for Organogenesis Using Sweet Potato Root Explants. (United States)

    Delany, William; And Others


    Describes an experiment in which groups of students in a plant tissue culture course worked together to test application of the Skoog-Miller model (developed by Skoog and Miller in regeneration of tobacco experiments to demonstrate organogenesis) to sweet potato root explants. (ZWH)

  9. Growth Response of Explants of Irvingia Gabonensis (O'rorke, Baill ...

    African Journals Online (AJOL)

    Growth response of explants of Irvingia gabonensis to in vitro treatment was investigated using full, half and one quarter strength mineral components based on Murashige and Skoog medium. Plant growth regulator (kinetin-Kin) with concentration levels of 0, 1, 2, 3, 4 and 5mg/l were used for shoots initiation, while axillary ...


    Expression of AhR and ARNT mRNA in cultured human endometrial explants exposed to TCDD.Pitt JA, Feng L, Abbott BD, Schmid J, Batt RE, Costich TG, Koury ST, Bofinger DP.Curriculum in Toxicology, University of North Carolina, Chapel Hill, NC 27599, USA.Endom...

  11. Adventitious shoot regeneration from leaf explants of southern highbush blueberry cultivars (United States)

    Protocols were developed to optimize adventitious shoot regeneration from four southern highbush blueberry cultivars. Leaf explants from six-week-old shoots of the four cultivars were excised and cultured on ten WPM (woody plant medium)-based regeneration media each containing thidiazuron (TDZ) (4.5...

  12. Local changes in proteoglycan synthesis during culture are different for normal and osteoarthritic cartilage

    NARCIS (Netherlands)

    Lafeber, F. P.; van der Kraan, P. M.; van Roy, H. L.; Vitters, E. L.; Huber-Bruning, O.; van den Berg, W. B.; Bijlsma, J. W.


    Proteoglycan synthesis of mild-to-moderate osteoarthritic human knee cartilage was compared with that of normal cartilage of the same donor. Immediately after cartilage was obtained, the synthesis rate of proteoglycans was higher for osteoarthritic cartilage than for normal cartilage. Proteoglycan

  13. Surgical Treatment of Articular Cartilage Defects in the Knee: Are We Winning?

    Directory of Open Access Journals (Sweden)

    A. R. Memon


    Full Text Available Articular cartilage (AC injury is a common disorder. Numerous techniques have been employed to repair or regenerate the cartilage defects with varying degrees of success. Three commonly performed techniques include bone marrow stimulation, cartilage repair, and cartilage regeneration. This paper focuses on current level of evidence paying particular attention to cartilage regeneration techniques.

  14. High frequency organogenesis in hypocotyl, cotyledon, leaf and petiole explants of broccoli (Brassica oleracea L. var. italica), an important vegetable crop

    National Research Council Canada - National Science Library

    Kumar, Pankaj; Srivastava, D K


    ... organogenesis from hypocotyl, cotyledon, leaf and petiole explants of broccoli (Brassica oleracea L. var. italica cv. Solan green head) has been developed. Hypocotyl and cotyledon explants were used from 10 to 12...

  15. The impact of dietary long-chain polyunsaturated fatty acids on bone and cartilage in gilts and sows. (United States)

    O'Connor-Robison, C I; Spencer, J D; Orth, M W


    Dietary long-chain PFO including arachidonic acid (ARA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) are precursors for several inflammatory mediators. The objective of this study was to characterize the effects of dietary PFO supplementation on bone, cartilage, and synovial fluid in 2 ages of pigs. Sows and gilts were fed either control corn/soybean meal based diets or the control diet supplemented with PFO from Gromega (PFO; JBS United, Sheridan, IN). Sows were fed their diets for 24.5 mo and slaughtered at 43 mo while gilts were fed their diets from weaning until slaughter at 111 kg. Cartilage was harvested from both humeroulnar joints of 14 sows (7/treatment) and 16 gilts (8/treatment) within 30 h of slaughter for fatty acid analysis and explant cultures. Synovial fluid was collected from the carpal joints of each pig postmortem. The right fused radius/ulna was collected for computed tomography (CT) analysis. Cortical width and density were determined and trabecular density was measured at the distal radius. Cartilage explants were allocated to 24-well culture plates with 2 discs per well and cultured over 72 h at 37°C in serum-free Dulbecco's modified Eagle's medium: nutrient mixture F-12 (Ham) medium. Six wells/pig were treated with 10 ng/mL of recombinant porcine interleukin-1 (rpIL-1). At 24, 48, and 72 h of culture, media were removed and reserved for analysis of proteoglycans, nitric oxide (NO), and PGE2 concentrations. The CT scans of the radius/ulna from gilts revealed no differences for cortical width and bone density. Sows fed PFO had greater cortical width of the proximal ulna (P Gilts fed PFO had increased DHA (P gilts and sows had no effect on the variables tested in vitro. Although the PFO diet increased omega-3 incorporation into chondrocytes, the biological significance is unclear since concentrations of ARA were at least 9-fold higher than EPA or DHA. Therefore, if omega-3 fatty acids can mitigate inflammation in joints, the

  16. Biomechanical and immunohistochemical properties of meniscal cartilage after high hydrostatic pressure treatment. (United States)

    Naal, Florian D; Schauwecker, Johannes; Steinhauser, Erwin; Milz, Stefan; von Knoch, Fabian; Mittelmeier, Wolfram; Diehl, Peter


    Meniscal allograft processing procedures, in particular gamma irradiation, deteriorate the biomechanical and biological properties of the transplanted tissue. High hydrostatic pressure (HHP) treatment, widely used in food technology to inactivate microorganisms while preserving natural compounds, might serve as a gentle alternative to gamma irradiation in the processing of meniscal allografts. We therefore investigated the effects of HHP treatment on the biomechanical and immunohistochemical properties of meniscal cartilage. Specimens of bovine menisci were treated with HHP for 10 min (20 degrees C) at 300 MPa and 600 MPa. Untreated control samples were left at room temperature and ambient pressure. We performed repetitive cycling indentation-tests to assess the biomechanical properties-in particular the viscoelastic behavior-of HHP treated and untreated meniscal specimens. Immunohistochemical analysis for collagens type I, II, and III and for the proteoglycans versican, aggrecan and for link-protein was performed by immunolabeling cross-sections of untreated and at 600 MPa HHP treated specimens. Comparing untreated and HHP treated meniscal specimens there were no significant differences for all tested biomechanical parameters. All cross-sections of untreated and HHP treated specimens stained positive for the collagens and proteoglycans. We demonstrated that meniscal cartilage can be treated by HHP at levels as high as 600 MPa without affection of the biomechanical and immunochistochemical properties. Therefore, HHP treatment might serve as a gentle alternative to gamma irradiation in the processing of meniscal allografts. Further research is necessary to verificate the present results in vivo. (c) 2008 Wiley Periodicals, Inc.

  17. Efficient Regeneration of �Caralis� Alstroemeria Cultivar from Rhizome Explants

    Directory of Open Access Journals (Sweden)

    Amir Ghaffar SHAHRIARI


    Full Text Available In this paper, the effects of a number of growth regulators as well as supplements to the Murashige and Skoog (MS basal medium were evaluated on the regeneration of Alstroemeria rhizome explants. In the first experiment the effects of three cytokinins (BA, TDZ and 2IP each at 0.5, 1 and 2 mg/l in combination with NAA (0.2 mg/l, followed by another PGR combination of 2IP (at 0.5, 1 and 2 mg/l with NAA (0 and 0.2 mg/l, on regeneration of rhizome-derived explants, was investigated. Through the second experiment, the effects of a number of supplements, including glucose (30 g/l as the alternative for sucrose, casein hydrolysate (1 g/l, asparagine and glutamine, (each at 30 mg/l added to MS medium, containing 1 mg/l BA and 0.2 mg/l NAA, was examined on rhizome explants� regeneration. Among the tested cytokinins, BA induced better regeneration of rhizome explants, resulting in a higher number of shoots compared to the other cytokinins. A medium supplemented with 1 mg/l BA and 0.2 mg/l NAA proved to be the most effective, with an average of 4.16 regenerated shoots per explant. In the second PGR combination, addition of NAA at 0.2 mg/l improved regeneration, compared to NAA-free treatments. In the second experiment, glucose substitution for sucrose improved regeneration with an average of 5.10 regenerated shoots per explant, compared to 4.16 shoots in sucrose-containing medium; whereas glutamine and asparagine (with 2.66 shoots and casein hydrolysate (with 3.80 shoots showed a negative influence on rhizome explants� regeneration.

  18. Milk Thistle Extract and Silymarin Inhibit Lipopolysaccharide Induced Lamellar Separation of Hoof Explants in Vitro

    Directory of Open Access Journals (Sweden)

    Nicole Reisinger


    Full Text Available The pathogenesis of laminitis is not completely identified and the role of endotoxins (lipopolysaccharides, LPS in this process remains unclear. Phytogenic substances, like milk thistle (MT and silymarin, are known for their anti-inflammatory and antioxidant properties and might therefore have the potential to counteract endotoxin induced effects on the hoof lamellar tissue. The aim of our study was to investigate the influence of endotoxins on lamellar tissue integrity and to test if MT and silymarin are capable of inhibiting LPS-induced effects in an in vitro/ex vivo model. In preliminary tests, LPS neutralization efficiency of these phytogenics was determined in an in vitro neutralization assay. Furthermore, tissue explants gained from hooves of slaughter horses were tested for lamellar separation after incubation with different concentrations of LPS. By combined incubation of explants with LPS and either Polymyxin B (PMB; positive control, MT or silymarin, the influence of these substances on LPS-induced effects was assessed. In the in vitro neutralization assay, MT and silymarin reduced LPS concentrations by 64% and 75%, respectively, in comparison PMB reduced 98% of the LPS concentration. In hoof explants, LPS led to a concentration dependent separation. Accordantly, separation force was significantly decreased by 10 µg/mL LPS. PMB, MT and silymarin could significantly improve tissue integrity of explants incubated with 10 µg/mL LPS. This study showed that LPS had a negative influence on the structure of hoof explants in vitro. MT and silymarin reduced endotoxin activity and inhibited LPS-induced effects on the lamellar tissue. Hence, MT and silymarin might be used to support the prevention of laminitis and should be further evaluated for this application.

  19. Cartilage Injuries in the Adult Knee (United States)

    Moyad, Thomas F.


    Cartilage injuries are frequently recognized as a source of significant morbidity and pain in patients with previous knee injuries. The majority of patients who undergo routine knee arthroscopy have evidence of a chondral defect. These injuries represent a continuum of pathology from small, asymptomatic lesions to large, disabling defects affecting a major portion of one or more compartments within the knee joint. In comparison to patients with osteoarthritis, individuals with isolated chondral surface damage are often younger, significantly more active, and usually less willing to accept limitations in activities that require higher impact. At the present time, a variety of surgical procedures exist, each with their unique indications. This heterogeneity of treatment options frequently leads to uncertainty regarding which techniques, if any, are most appropriate for patients. The purpose of this review is to describe the workup and discuss the management techniques for cartilage injuries within the adult knee. PMID:26069581

  20. Cartilage tissue engineering on the surface of a novel gelatin-calcium-phosphate biphasic scaffold in a double-chamber bioreactor. (United States)

    Chang, Chih-Hung; Lin, Feng-Huei; Lin, Chien-Cheng; Chou, Cheng-Hung; Liu, Hwa-Chang


    Tissue engineering is a new approach to articular cartilage repair; however, the integration of the engineered cartilage into the host subchondral bone is a major problem in osteochondral injury. The aim of the present work, therefore, was to make a tissue-engineered osteochondral construct from a novel biphasic scaffold in a newly designed double-chamber bioreactor. This bioreactor was designed to coculture chondrocytes and osteoblasts simultaneously. The aim of this study was to prove that engineered cartilage could be formed with the use of this biphasic scaffold. The scaffold was constructed from gelatin and a calcium-phosphate block made from calcined bovine bone. The cartilage part of the scaffold had a uniform pore size of about 180 microm and approximate porosity of 75%, with the trabecular pattern preserved in the bony part of the scaffold. The biphasic scaffolds were seeded with porcine chondrocytes and cultured in a double-chamber bioreactor for 2 or 4 weeks. The chondrocytes were homogeneously distributed in the gelatin part of the scaffold, and secretion of the extracellular matrix was demonstrated histologically. The chondrocytes retained their phenotype after 4 weeks of culture, as proven immunohistochemically. After 4 weeks of culture, hyaline-like cartilage with lacuna formation could be clearly seen in the gelatin scaffold on the surface of the calcium phosphate. The results show that this biphasic scaffold can support cartilage formation on a calcium-phosphate surface in a double-chamber bioreactor, and it seems reasonable to suggest that there is potential for further application in osteochondral tissue engineering. (c) 2004 Wiley Periodicals, Inc.

  1. Influence of extremely low frequency, low energy electromagnetic fields and combined mechanical stimulation on chondrocytes in 3-D constructs for cartilage tissue engineering. (United States)

    Hilz, Florian M; Ahrens, Philipp; Grad, Sibylle; Stoddart, Martin J; Dahmani, Chiheb; Wilken, Frauke L; Sauerschnig, Martin; Niemeyer, Philipp; Zwingmann, Jörn; Burgkart, Rainer; von Eisenhart-Rothe, Rüdiger; Südkamp, Norbert P; Weyh, Thomas; Imhoff, Andreas B; Alini, Mauro; Salzmann, Gian M


    Articular cartilage, once damaged, has very low regenerative potential. Various experimental approaches have been conducted to enhance chondrogenesis and cartilage maturation. Among those, non-invasive electromagnetic fields have shown their beneficial influence for cartilage regeneration and are widely used for the treatment of non-unions, fractures, avascular necrosis and osteoarthritis. One very well accepted way to promote cartilage maturation is physical stimulation through bioreactors. The aim of this study was the investigation of combined mechanical and electromagnetic stress affecting cartilage cells in vitro. Primary articular chondrocytes from bovine fetlock joints were seeded into three-dimensional (3-D) polyurethane scaffolds and distributed into seven stimulated experimental groups. They either underwent mechanical or electromagnetic stimulation (sinusoidal electromagnetic field of 1 mT, 2 mT, or 3 mT; 60 Hz) or both within a joint-specific bioreactor and a coil system. The scaffold-cell constructs were analyzed for glycosaminoglycan (GAG) and DNA content, histology, and gene expression of collagen-1, collagen-2, aggrecan, cartilage oligomeric matrix protein (COMP), Sox9, proteoglycan-4 (PRG-4), and matrix metalloproteinases (MMP-3 and -13). There were statistically significant differences in GAG/DNA content between the stimulated versus the control group with highest levels in the combined stimulation group. Gene expression was significantly higher for combined stimulation groups versus static control for collagen 2/collagen 1 ratio and lower for MMP-13. Amongst other genes, a more chondrogenic phenotype was noticed in expression patterns for the stimulated groups. To conclude, there is an effect of electromagnetic and mechanical stimulation on chondrocytes seeded in a 3-D scaffold, resulting in improved extracellular matrix production. © 2013 Wiley Periodicals, Inc.

  2. TAK1 mediates BMP signaling in cartilage


    Greenblatt, Matthew B.; Shim, Jae-Hyuck; Glimcher, Laurie H.


    Although many signals are capable of activating MAPK signaling cascades in chondrocytes in vitro, the function of these pathways remains unclear in vivo. Here we report the phenotype of mice with a conditional deletion of TGF-β-activated kinase 1 (TAK1), a MAP3K family member, in cartilage using the collagen 2α promoter. These mice display chondrodysplasia characterized by neonatal-onset runting, delayed formation of secondary ossification centers, and defects in formation of the elbow and ta...

  3. Serum Cartilage Biomarkers and Shoulder Instability. (United States)

    Owens, Brett D; Cameron, Kenneth L; Bokshan, Steven L; Clifton, Kari B; Svoboda, Steven J; Wolf, Jennifer Moriatis


    Differences in cartilage biomarkers have been noted in patients with anterior cruciate ligament tears, but little is known about any similar relationship with shoulder instability. This study evaluated the relationship between serum cartilage biomarkers and shoulder instability. The authors present a prospective cohort study of young athletes followed from 2006 to 2010. A nested case-control analysis was conducted within this cohort to evaluate the association between preinjury collagen type II cleavage (a marker for type II collagen cleavage) and procollagen II carboxy propeptide (a marker of cartilage synthesis) and the subsequent likelihood of shoulder instability during the 4-year follow-up period. Preinjury collagen type II cleavage and procollagen II carboxy propeptide levels in 51 subjects who had shoulder instability were compared with levels in 210 subjects without documented anterior cruciate ligament or shoulder instability (control group) with commercially available enzyme-linked immunosorbent assay kits. Mean preinjury collagen type II cleavage levels in patients who subsequently had shoulder instability were significantly lower than those in the control group (73.91 vs 79.24 pg/mL, P=.03). No significant difference was found in preinjury procollagen II carboxy propeptide levels compared with the control group (359.94 vs 396.37, P=.24). This study is the first to examine the relationship between baseline collagen biomarkers and subsequent shoulder instability. The finding of lower baseline collagen type II cleavage levels in patients with subsequent shoulder instability may represent a genetic predisposition or a compensatory mechanism by which cartilage degradation is decreased in those who are more likely to have instability. [Orthopedics. 2017; 40(1):34-36.]. Copyright 2016, SLACK Incorporated.

  4. Cartilage Tissue Engineering: What Have We Learned in Practice? (United States)

    Doran, Pauline M


    Many technologies that underpin tissue engineering as a research field were developed with the aim of producing functional human cartilage in vitro. Much of our practical experience with three-dimensional cultures, tissue bioreactors, scaffold materials, stem cells, and differentiation protocols was gained using cartilage as a model system. Despite these advances, however, generation of engineered cartilage matrix with the composition, structure, and mechanical properties of mature articular cartilage has not yet been achieved. Currently, the major obstacles to synthesis of clinically useful cartilage constructs are our inability to control differentiation to the extent needed, and the failure of engineered and host tissues to integrate after construct implantation. The aim of this chapter is to distil from the large available body of literature the seminal approaches and experimental techniques developed for cartilage tissue engineering and to identify those specific areas requiring further research effort.

  5. Cartilage-targeting drug delivery: can electrostatic interactions help? (United States)

    Bajpayee, Ambika G; Grodzinsky, Alan J


    Current intra-articular drug delivery methods do not guarantee sufficient drug penetration into cartilage tissue to reach cell and matrix targets at the concentrations necessary to elicit the desired biological response. Here, we provide our perspective on the utilization of charge-charge (electrostatic) interactions to enhance drug penetration and transport into cartilage, and to enable sustained binding of drugs within the tissue's highly negatively charged extracellular matrix. By coupling drugs to positively charged nanocarriers that have optimal size and charge, cartilage can be converted from a drug barrier into a drug reservoir for sustained intra-tissue delivery. Alternatively, a wide variety of drugs themselves can be made cartilage-penetrating by functionalizing them with specialized positively charged protein domains. Finally, we emphasize that appropriate animal models, with cartilage thickness similar to that of humans, must be used for the study of drug transport and retention in cartilage.

  6. Viral infections and bovine mastitis: a review

    NARCIS (Netherlands)

    Wellenberg, G.J.; Poel, van der W.H.M.; Oirschot, van J.T.


    This review deals with the role of viruses in the aetiology of bovine mastitis. Bovine herpesvirus 1, bovine herpesvirus 4, foot-and-mouth disease virus, and parainfluenza 3 virus have been isolated from milk from cows with clinical mastitis. Intramammary inoculations of bovine herpesvirus 1 or

  7. Tracheal restoration in dogs with umbilical cord membrane of bovine preserved in glicerin

    Directory of Open Access Journals (Sweden)

    Alexandre Mazzanty


    Full Text Available The viability of the use the bovine umbilical membrane preserved in 98% glycerin implanted in the cervical trachea was studied. Seven adult mongrel dogs, three males and four females weighting 6 to 14 kg, were used. After the usual anesthesia protocol and asseptic technique, three tracheal rings were partially removed for implantation of a segment of the umbilical membrane. The animals were observed during 30 days and then reoperated for macroscopic observations and for fragment collection for histological evaluation. It occurred a repair of the tracheal lesion, with formation of granulation tissue rich in collagen fibers linking the extremities of the tracheal cartilages. Epithelial migration over the tracheal surface was also seen. It is concluded that the segment of the bovine umbilical cord preserved in 98% glycerin can be used in the repair of tracheal defects. It offers a temporary support for granulation tissue formation and epithelization in the implanted area.

  8. Mesenchymal Stem Cells for Cartilage Regeneration of TMJ Osteoarthritis


    Dixin Cui; Hongyu Li; Xin Xu; Ling Ye; Xuedong Zhou; Liwei Zheng; Yachuan Zhou


    Temporomandibular joint osteoarthritis (TMJ OA) is a degenerative disease, characterized by progressive cartilage degradation, subchondral bone remodeling, synovitis, and chronic pain. Due to the limited self-healing capacity in condylar cartilage, traditional clinical treatments have limited symptom-modifying and structure-modifying effects to restore impaired cartilage as well as other TMJ tissues. In recent years, stem cell-based therapy has raised much attention as an alternative approach...

  9. [Research progress of bioreactor biophysical factors in cartilage tissue engineering]. (United States)

    Ye, Gang; Zhang, Fangbiao; Shi, Hongcan


    To review the recent research progress of the bioreactor biophysical factors in cartilage tissue engineering. The related literature concerning the biophysical factors of bioreactor in cartilage tissue engineering was reviewed, analyzed, and summarized. Oxygen concentration, hydrostatic pressure, compressive force, and shear load in the bioreactor system have no unified standard parameters. Hydrostatic pressure and shear load have been in controversy, which restricts the application of bioreactors. The biophysical factors of broreactor in cartilage tissue engineering have to be studied deeply.

  10. Stress relaxation and cartilage shaping under laser radiation (United States)

    Sobol, Emil N.; Sviridov, Alexander P.; Bagratashvili, Victor N.; Omelchenko, Alexander I.; Ovchinnikov, Yuriy M.; Shekhter, Anatoliy B.; Downes, S.; Howdle, Steven; Jones, Nicholas; Lowe, J.


    The problem of a purposeful change of the shape of cartilage is of great importance for otolaryngology, orthopaedics, and plastic surgery. In 1992 we have found a possibility of controlled shaping of cartilage under moderate laser heating. This paper presents new results in studies of that phenomenon. We have measured temperature and stress in a tissue undergoing to irradiation with a Holmium laser. Study of cartilage structure allowed us to find conditions for laser shaping without pronounced alterations in the structure of matrix.

  11. Evaluation of early changes of cartilage biomarkers following arthroscopic meniscectomy in young Egyptian adults

    Directory of Open Access Journals (Sweden)

    Hamdy Khamis Koryem


    Conclusion: Cartilage volume loss by MRI combined with changes in cartilage matrix turnover detected by molecular biomarkers may reflect the initial changes associated with cartilage degeneration that account for early OA.

  12. Near Infrared Spectroscopic Mapping of Functional Properties of Equine Articular Cartilage

    NARCIS (Netherlands)

    Sarin, Jaakko K; Amissah, Michael; Brommer, Harold; Argüelles, David; Töyräs, Juha; Afara, Isaac O


    Mechanical properties of articular cartilage are vital for normal joint function, which can be severely compromised by injuries. Quantitative characterization of cartilage injuries, and evaluation of cartilage stiffness and thickness by means of conventional arthroscopy is poorly reproducible or

  13. Delayed Gadolinium Enhanced MRI of Cartilage (dGEMRIC) can be effectively applied for longitudinal cohort evaluation of articular cartilage regeneration

    NARCIS (Netherlands)

    Bekkers, J.E.J.; Lambertus, W.B.; Benink, R.J.; Tsuchida, A.I.; Vincken, K.L.; Dhert, W.J.A.; Creemers, L.B.; Saris, Daniël B.F.


    Objective Delayed gadolinium enhanced MRI of cartilage (dGEMRIC) facilitates non-invasive evaluation of the glycosaminoglycan content in articular cartilage. The primary aim of this study was to show that the dGEMRIC technique is able to monitor cartilage repair following regenerative cartilage

  14. Influencing micropropagation in Clitoria ternatea L. through the manipulation of TDZ levels and use of different explant types. (United States)

    Mukhtar, Seemab; Ahmad, Naseem; Khan, Md Imran; Anis, Mohammad; Aref, Ibrahim M


    A comparative performance of two explants types (CN and Nodal) for their efficiency to induce multiple shoot regeneration in Clitoria ternatea has been carried out. Thidiazuron (TDZ) in different concentrations (0.05-2.5 μM) was used as a supplement to the Murashige and Skoog's (MS) basal media. Explant type apart, two factors viz. concentration and exposure duration to TDZ played an important role in affecting multiple shoot regeneration. Cotyledonary node explants produced the best results at 0.1 μM TDZ, while in nodal explants the highest rate of shoot formation was achieved on MS medium supplemented with 1.0 μM TDZ. In both the explants, shoot multiplication increased when the regenerated shoots were subcultured on hormone free MS medium after 4 weeks of exposure to TDZ. Among the two, cotyledonary node explants produced considerably higher number of shoots at a comparatively lower concentration of TDZ than nodal explants. The regenerated shoots rooted best on MS medium containing 1.0 μM indole-3-butyric acid (IBA) and were successfully established in pots containing garden soil with 88 % survival rate. All the regenerated plants showed normal morphology and growth characteristics.

  15. Secondary cartilage revealed in a non-avian dinosaur embryo.

    Directory of Open Access Journals (Sweden)

    Alida M Bailleul

    Full Text Available The skull and jaws of extant birds possess secondary cartilage, a tissue that arises after bone formation during embryonic development at articulations, ligamentous and muscular insertions. Using histological analysis, we discovered secondary cartilage in a non-avian dinosaur embryo, Hypacrosaurus stebingeri (Ornithischia, Lambeosaurinae. This finding extends our previous report of secondary cartilage in post-hatching specimens of the same dinosaur species. It provides the first information on the ontogeny of avian and dinosaurian secondary cartilages, and further stresses their developmental similarities. Secondary cartilage was found in an embryonic dentary within a tooth socket where it is hypothesized to have arisen due to mechanical stresses generated during tooth formation. Two patterns were discerned: secondary cartilage is more restricted in location in this Hypacrosaurus embryo, than it is in Hypacrosaurus post-hatchlings; secondary cartilage occurs at far more sites in bird embryos and nestlings than in Hypacrosaurus. This suggests an increase in the number of sites of secondary cartilage during the evolution of birds. We hypothesize that secondary cartilage provided advantages in the fine manipulation of food and was selected over other types of tissues/articulations during the evolution of the highly specialized avian beak from the jaws of their dinosaurian ancestors.

  16. Mesenchymal Stem Cells for Treating Articular Cartilage Defects and Osteoarthritis. (United States)

    Wang, Yu; Yuan, Mei; Guo, Quan-yi; Lu, Shi-bi; Peng, Jiang


    Articular cartilage damage and osteoarthritis are the most common joint diseases. Joints are prone to damage caused by sports injuries or aging, and such damage regularly progresses to more serious joint disorders, including osteoarthritis, which is a degenerative disease characterized by the thinning and eventual wearing out of articular cartilage, ultimately leading to joint destruction. Osteoarthritis affects millions of people worldwide. Current approaches to repair of articular cartilage damage include mosaicplasty, microfracture, and injection of autologous chondrocytes. These treatments relieve pain and improve joint function, but the long-term results are unsatisfactory. The long-term success of cartilage repair depends on development of regenerative methodologies that restore articular cartilage to a near-native state. Two promising approaches are (i) implantation of engineered constructs of mesenchymal stem cell (MSC)-seeded scaffolds, and (ii) delivery of an appropriate population of MSCs by direct intra-articular injection. MSCs may be used as trophic producers of bioactive factors initiating regenerative activities in a defective joint. Current challenges in MSC therapy are the need to overcome current limitations in cartilage cell purity and to in vitro engineer tissue structures exhibiting the required biomechanical properties. This review outlines the current status of MSCs used in cartilage tissue engineering and in cell therapy seeking to repair articular cartilage defects and related problems. MSC-based technologies show promise when used to repair cartilage defects in joints.

  17. [Cartilage reshaping by laser in stomatology and maxillofacial surgery]. (United States)

    Mordon, S


    The restoration of congenital and traumatic malformations of the head and neck, together with the defects resulting from the trauma of ablative surgery, continue to pose significant problems to surgeons. The post-operative results are not always satisfactory because of the difficulty of shaping the cartilage and because of the tendency of cartilage to return to its original shape. Better understanding of laser-cartilage interaction and the development of a specific instrumentation Lasers (CO2, Nd: YAG, Ho: YAG) has enabled ex situ and in situ cartilage reshaping. A recent clinical study has demonstrated that nondestructive laser irradiation can reshape septal deviations

  18. From gristle to chondrocyte transplantation: treatment of cartilage injuries. (United States)

    Lindahl, Anders


    This review addresses the progress in cartilage repair technology over the decades with an emphasis on cartilage regeneration with cell therapy. The most abundant cartilage is the hyaline cartilage that covers the surface of our joints and, due to avascularity, this tissue is unable to repair itself. The cartilage degeneration seen in osteoarthritis causes patient suffering and is a huge burden to society. The surgical approach to cartilage repair was non-existing until the 1950s when new surgical techniques emerged. The use of cultured cells for cell therapy started as experimental studies in the 1970s that developed over the years to a clinical application in 1994 with the introduction of the autologous chondrocyte transplantation technique (ACT). The technology is now spread worldwide and has been further refined by combining arthroscopic techniques with cells cultured on matrix (MACI technology). The non-regenerating hypothesis of cartilage has been revisited and we are now able to demonstrate cell divisions and presence of stem-cell niches in the joint. Furthermore, cartilage derived from human embryonic stem cells and induced pluripotent stem cells could be the base for new broader cell treatments for cartilage injuries and the future technology base for prevention and cure of osteoarthritis. © 2015 The Author(s).

  19. Cutaneous Squamous Cell Carcinoma with Invasion through Ear Cartilage

    Directory of Open Access Journals (Sweden)

    Julie Boisen


    Full Text Available Cutaneous squamous cell carcinoma of the ear represents a high-risk tumor location with an increased risk of metastasis and local tissue invasion. However, it is uncommon for these cancers to invade through nearby cartilage. Cartilage invasion is facilitated by matrix metalloproteases, specifically collagenase 3. We present the unusual case of a 76-year-old man with an auricular squamous cell carcinoma that exhibited full-thickness perforation of the scapha cartilage. Permanent sections through the eroded cartilage confirmed tumor invasion extending to the posterior ear skin.

  20. Treatment of knee cartilage defect in 2010. (United States)

    Versier, G; Dubrana, F


    Treatment of knee cartilage defect, a true challenge, should not only reconstruct hyaline cartilage on a long-term basis, but also be able to prevent osteoarthritis. Osteochondral knee lesions occur in either traumatic lesions or in osteochondritis dissecans (OCD). These lesions can involve all the articular surfaces of the knee in its three compartments. In principle, this review article covers symptomatic ICRS grade C or D lesions, depth III and IV, excluding management of superficial lesions, asymptomatic lesions that are often discovered unexpectedly, and kissing lesions, which arise prior to or during osteoarthritis. For clarity sake, the international classifications used are reviewed, for both functional assessment (ICRS and functional IKDC for osteochondral fractures, Hughston for osteochondritis) and morphological lesion evaluations (the ICRS macroscopic evaluation for fractures, the Bedouelle or SOFCOT for osteochondritis, and MOCART for MRI). The therapeutic armamentarium to treat these lesions is vast, but accessibility varies greatly depending on the country and the legislation in effect. Many comparative studies have been conducted, but they are rarely of high scientific quality; the center effect is nearly constant because patients are often referred to certain centers for an expert opinion. The indications defined herein use algorithms that take into account the size of the cartilage defect and the patient's functional needs for cases of fracture and the vitality, stability, and size of the fragment for cases of osteochondritis dissecans. Fractures measuring less than 2 cm(2) are treated with either microfracturing or mosaic osteochondral grafting, between 2 and 4 cm(2) with microfractures covered with a membrane or a culture of second- or third-generation chondrocytes, and beyond this size, giant lesions are subject to an exceptional allografting procedure, harvesting from the posterior condyle, or chondrocyte culture on a 3D matrix to restore

  1. Inhibition of TGFβ cell signaling for limbal explant culture in serumless, defined xeno-free conditions. (United States)

    Zamudio, Aldo; Wang, Zheng; Chung, So-Hyang; Wolosin, J Mario


    Outgrowths of limbal epithelium by explant culture are used to treat limbal stem cell deficiency (LSCD). The explant culture medium is always complemented with serum, a complex solution which includes TGFβ. Since TGFβ is a cytostatic effector for epithelial proliferation we examined its effect on these cultures. Limbal biopsies were set on explant culture in DMEM/F12 with 5 ng/ml EGF and cholera toxin (ChT), ITS, and 5% FBS, henceforth SHEM or a) SHEMSB=SHEM plus SB431542 an inhibitor of TGFβ signaling; b) sfSHEM = SHEM with FBS replaced by 0.05% Albumax II; and c) sfSHEMSB and sfSHEMA83 = sfSHEM plus, respectively, SB431542 or A-83-01, another TGFβ inhibitor. After the initial outgrowths reached 3 cm in diameter, the limbal biopsies were serially transferred up to six times onto new inserts. Biopsy explant outgrowths were trypsinized and cell yield, morphology and stem-cell related JC-1 exclusion (IOVS, 52:4330) were determined by flow cytometry. Cells we plated at low density seeding to compare relative clonal proliferative activity. The expression of three proteins whose levels are associated with growth and differentiation states, Krt3, connexin 43 and p63 were determined by immunohistology and/or Western blot. Cell yield in rabbit, relative to SHEM (in %) were, SHEMSB, 104 ± 13 (p > 0.95); sfSHEM: 5 ± 3; and sfSHEMSB, 94 ± 18 (p > 0.95). Cell size and morphology, JC1 dye exclusion, Krt3, p63 and connexin 43 content, proliferation efficiency and the preservation of extended proliferative potential of the serially cultured biopsies were similar for SHEM, SHEMSB and sfSHEMSB. The only differences observed where reduced expression of Krt3 and increased preservation of p63 in the FBS-free medium. Removal of EGF from sfSHEMSB reduced yield by 92 ± 6% (p free medium caused a small, non-statistical decrease in growth rates. Equivalent results were observed in a preliminary experiment in human. These results suggest that in the absence serum

  2. A Novel 3D Skin Explant Model to Study Anaerobic Bacterial Infection

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    Grazieli Maboni


    Full Text Available Skin infection studies are often limited by financial and ethical constraints, and alternatives, such as monolayer cell culture, do not reflect many cellular processes limiting their application. For a more functional replacement, 3D skin culture models offer many advantages such as the maintenance of the tissue structure and the cell types present in the host environment. A 3D skin culture model can be set up using tissues acquired from surgical procedures or post slaughter, making it a cost effective and attractive alternative to animal experimentation. The majority of 3D culture models have been established for aerobic pathogens, but currently there are no models for anaerobic skin infections. Footrot is an anaerobic bacterial infection which affects the ovine interdigital skin causing a substantial animal welfare and financial impact worldwide. Dichelobacter nodosus is a Gram-negative anaerobic bacterium and the causative agent of footrot. The mechanism of infection and host immune response to D. nodosus is poorly understood. Here we present a novel 3D skin ex vivo model to study anaerobic bacterial infections using ovine skin explants infected with D. nodosus. Our results demonstrate that D. nodosus can invade the skin explant, and that altered expression of key inflammatory markers could be quantified in the culture media. The viability of explants was assessed by tissue integrity (histopathological features and cell death (DNA fragmentation over 76 h showing the model was stable for 28 h. D. nodosus was quantified in all infected skin explants by qPCR and the bacterium was visualized invading the epidermis by Fluorescent in situ Hybridization. Measurement of pro-inflammatory cytokines/chemokines in the culture media revealed that the explants released IL1β in response to bacteria. In contrast, levels of CXCL8 production were no different to mock-infected explants. The 3D skin model realistically simulates the interdigital skin and has

  3. Pathogenesis of bovine neosporosis. (United States)

    Dubey, J P; Buxton, D; Wouda, W


    The protozoan parasite Neospora caninum is a major pathogen of cattle and dogs, being a significant cause of abortion in cattle in many countries. It is one of the most efficiently transmitted parasites, with up to 90% of cattle infected in some herds. The pathogenesis of abortion due to Neospora is complex and only partially understood. Losses occur after a primary infection during pregnancy but more commonly as the result of recrudescence of a persistent infection during pregnancy. Parasitaemia is followed by invasion of the placenta and fetus. It is suggested that abortion occurs when primary parasite-induced placental damage jeopardises fetal survival directly or causes release of maternal prostaglandins that in turn cause luteolysis and abortion. Fetal damage may also occur due to primary tissue damage caused by the multiplication of N. caninum in the fetus or due to insufficient oxygen/nutrition, secondary to placental damage. In addition, maternal immune expulsion of the fetus may occur associated with maternal placental inflammation and the release of maternal pro-inflammatory cytokines in the placenta. Thus N. caninum is a primary pathogen capable of causing abortion either through maternal placental inflammation, maternal and fetal placental necrosis, fetal damage, or a combination of all three. The question of how N. caninum kills the fetus exposes the complex and finely balanced biological processes that have evolved to permit bovine and other mammalian pregnancies to occur. Defining these immunological mechanisms will shed light on potential methods of control of bovine neosporosis and enrich our understanding of the continuity of mammalian and protozoal survival.

  4. Enzymatic digestion of articular cartilage results in viscoelasticity changes that are consistent with polymer dynamics mechanisms

    National Research Council Canada - National Science Library

    June, Ronald K; Fyhrie, David P


    .... This study tested whether the predictions of polymer dynamics were consistent with changes in cartilage mechanics caused by enzymatic digestion of specific cartilage extracellular matrix molecules...

  5. One-stage explant-implant procedure of exposed porous orbital implants

    DEFF Research Database (Denmark)

    Toft, Peter B; Rasmussen, Marie L Roed; Prause, Jan Ulrik


    Purpose:  To investigate the risks of implant exposure after a combined explant-implant procedure in patients with an exposed porous orbital implant. Methods:  Twenty-four consecutive patients who had a combined explant-implant procedure of an exposed hydroxyapatite (21) or porous polyethylene (3......) orbital implant from January 2000 to February 2009 were included. The patient records were reviewed; patients were interviewed by telephone and invited for a clinical examination. Histopathological examination was carried out on the removed implants. Main outcome measures were: presence of exposure...... of the new implant or not, patient graded satisfaction with the cosmetic result, and presence of poor motility. Results:  None of the new implants became exposed or infected in the follow-up period of 25 [3-94] months (median [range]). The patients scored their satisfaction with the cosmetic result...

  6. Incubation under fluid dynamic conditions markedly improves the structural preservation in vitro of explanted skeletal muscles. (United States)

    Carton, Flavia; Calderan, Laura; Malatesta, Manuela


    Explanted organs and tissues represent suitable experimental systems mimicking the functional and structural complexity of the living organism, with positive ethical and economic impact on research activities. However, their preservation in culture is generally limited, thus hindering their application as experimental models for biomedical research. In the present study, we investigated the potential of an innovative fluid dynamic culture system to improve the structural preservation in vitro of explanted mouse skeletal muscles (soleus). We used light and transmission electron microscopy to compare the morphological features of muscles maintained either in multiwell plates under conventional conditions or in a bioreactor mimicking the flow of physiological fluids. Our results demonstrate that fluid dynamic conditions markedly slowed the progressive structural deterioration of the muscle tissue occurring during the permanence in the culture medium, prolonging the preservation of some organelles such as mitochondria up to 48 h.

  7. Effect of explant origin and different growth regulators on micropropagation of Pistacia atlantica ssp. mutica

    Directory of Open Access Journals (Sweden)

    Ali-Ashraf Mehrabi


    Full Text Available Propagation of wild pistachio as a multipurpose woody species is a hard and tedious task. In this research, an effective in vitro protocol was developed for rapid proliferation of wild pistachio (Pistacia atlantica ssp. mutica in MS medium supplemented with B5 vitamins and different growth regulators. Rooting of plantlets was tested by two treatments containing Rhizopon and IBA in ex vitro. With respect to the results, the nodal segments explants, produced the highest shoot frequency, leaf frequency and the tallest shoots. On the other hand, the tallest shoots were generated from shoot tip explant and medium containing of TDZ plus IAA. Both treatments (Rhizopon and IBA led to a remarkable increase in the number of roots, root length and rooting percentage compared to the control. These results may be applied for rapid proliferation to spread the pistachio trees and shrubs that are difficult and time consuming.

  8. Acetylcholine causes rooting in leaf explants of in vitro raised tomato (Lycopersicon esculentum Miller) seedlings. (United States)

    Bamel, Kiran; Gupta, Shrish Chandra; Gupta, Rajendra


    The animal neurotransmitter acetylcholine (ACh) induces rooting and promotes secondary root formation in leaf explants of tomato (Lycopersicon esculentum Miller var. Pusa Ruby), cultured in vitro on Murashige and Skoog's medium. The roots originate from the midrib of leaf explants and resemble taproot. ACh at 10(-5) M was found to be the optimum over a wide range of effective concentrations between 10(-7) and 10(-3) M. The breakdown products, choline and acetate were ineffective even at 10(-3) M concentration. ACh appears to have a natural role in tomato rhizogenesis because exogenous application of neostigmine, an inhibitor of ACh hydrolysis, could mimic the effect of ACh. Neostigmine, if applied in combination with ACh, potentiated the ACh effect.

  9. Ultrastructural analysis of initial stages of dedifferentiation of root explants of Gentiana cruciata seedlings

    Directory of Open Access Journals (Sweden)

    Anna Mikuła


    Full Text Available The studies were carried out on isolated roots of 10-day old seedlings of Gentiana cruciata, which were placed and cultured on induction medium of Murashige and Skoog (1962 supplemented with 1.0 mg/dm3 dicamba + 0.l mg/dm3 NAA + 2.00 mg/dm3 BAP + 80.0 mg/dm3 adenine sulphate. Changes in explants from the 3rd to the l lth day of culture with the help of light and electron microscope were observed. Observations showed gradual dedifferentiation of root tissues, which was seen earliest in cortex at the proximal end of the explant and shifted gradually inwards the root and towards distal parts of its elongation zone. The most intensive callus formation appeared at cut surface of explant, where proliferation of cells in both cortex and axial cylinder was recognised. In the distal part of the elongation zone, cell divisions occurred only in endoderm and in axial cylinder. The meristematic part of the root was inactive. Finally, the following areas were distinguished in the explant: (I an area of intensive cell divisions, i.e., the elongation zone; (II an area of cell dispersion; and (III the inactive meristem. The ultrastructure brought evidences of cell reorganisation as the meaning of cell readiness to the division. Observations showed an increased activity of mitochondria and Golgi structures, thickening of walls and disappearance of plasmodesmal connections. Amyloplasts and lipid bodies in tissues in which they had been scarce or had not appeared before founding. Intensively dividing cells showed features of meristematic cells. They had dense cytoplasm with numerous organelles, large centrally located nuclei, and "nucleolar vacuoles" inside nucleoli. Cortex-derived callus formed aggregates. Both pericycle and endoderm produced callus of characteristic dense structure and regular type of divisions.

  10. Effect of explant density and medium culture volumes on cassava micropropagation in Temporal Immersion System


    Milagros Basail; Victor Medero; Marilyn Martínez; José de la C. Ventura; Jorge López; Magaly García; Manuel Cabrera; Arletys Santos; Aymé Rayas; Carmen Pons; Maricel Bauta; Miguel Álvarez; Jesús García


    Due to the need of producing high quality planting material available to cassava growers, it has been necessary to look for alternatives in order to increase the efficiancy of in vitro propagation methods and their automation, such as the use of the Temporal Immersion Systems (RITA®). This work was carried out to increase the multiplication coefficient for cassava mass propagation through out Temporal Immersion Systems. The clone ‘CMC-40’ was used. Different medium volumes per explant, and ma...

  11. The Impact of Carbon Source, Explants and Growth Regulators on Callogenesis and Organogenesis of Artemisia annua

    Directory of Open Access Journals (Sweden)



    Full Text Available Artemisinin, a sesquiterpene lactone isolated from Artemisia annua L. plant is known for its antimalarial activity. The low content of artemisinin has stimulated researchers to enhance its production through biotechnological approaches such as tissue culture. The present study was initiated to study the effect of some important factors alone and in combination, on the callogenesis and organogenesis of Artemisia annua. The type of carbon source had a significant effect on NAA efficiency to callogenesis of A. annua, whereas the best callogenesis of A. absinthium was observed at 2 mg/L BAP + 30 g/L sucrose and in the absence of NAA, with root explants. Presence of BAP also had an important effect on callogenesis, especially in high concentrations. A suitable suspension culture was obtained in the MS basal medium containing 0.5 mg/L NAA and BAP, with 30 g/L glucose. Artemisinin was naturally production was at least 0.03 mg/g (w.dt at the first day and peaked on the 16th day with 0.31 mg/g (w.dt in the cell culture of A. annua. The maximum number of shoots (2.167 ± 1.484 was induced at 0.5 mg/L BAP + 0.1 mg/L NAA + glucose (30 g/L with leaf explants. However, treatments containing glucose did not show a good shoot induction. Longer shoots were induced in the medium containing either 0.5 mg/l NAA + 0.5 mg/l BAP + sucrose with leaf explants (1.493 cm ± 0.342 or 0.5 mg/l NAA + sucrose with stem explants (0.697 cm ± 0.930. Medium containing 0.5 or 2 mg/l NAA and sucrose (without BAP induced more roots though.

  12. Adventitious shoot regeneration from leaf explants of eastern cottonwood (Populus deltoides) cultured under photoautotrophic conditions. (United States)

    Mingozzi, Marco; Montello, Paul; Merkle, Scott


    Effects of photoautotrophic and photomixotrophic growth conditions on adventitious shoot regeneration from leaf explants of eastern cottonwood (Populus deltoides Bartr. ex Marsh.) were investigated. Rooting and proliferating shoot cultures (Stage I) were grown in either an elevated (1500 ppm) CO(2) concentration ([CO(2)]) at high photosynthetic photon flux (PPF; ~ 150 micromol m(-2) s(-1)) (photoautotrophic condition) with 0, 10 or 30 g l(-1) sucrose or under standard conditions (ambient (360 ppm) [CO(2)] at low PPF (~ 60 micromol m(-2) s(-1)) with 30 g l(-1) sucrose). Leaves harvested from these cultures were analyzed for soluble sugars and were used as explants for adventitious shoot regeneration (Stage II), which was also carried out under photoautotrophic and standard conditions. Photoautotrophic conditions during Stage I promoted growth of rooting shoots but inhibited axillary shoot proliferation. Photoautotrophic conditions during Stage II suppressed callus and adventitious bud production from leaf explants compared with standard conditions. The regeneration environment appeared to be more important in controlling bud formation than the conditions under which the donor shoots were grown. Regardless of Stage I treatment, bud production was up to 100-fold higher for leaves cultured under standard conditions than under photoautotrophic conditions. Once adventitious buds were differentiated from the leaf tissues, however, their elongation was faster under photoautotrophic conditions than that under standard conditions, with some shoots reaching 10 mm in length on leaf explants cultured under photoautotrophic conditions. Because total leaf soluble sugar concentration was always lowest in shoots under standard conditions, which also yielded the highest bud production, the results suggest that endogenous starvation enhanced shoot production.

  13. Characteristic of c-Kit+ progenitor cells in explanted human hearts


    Matuszczak, Sybilla; Czapla, Justyna; Jarosz-Biej, Magdalena; Wiśniewska, Ewa; Cichoń, Tomasz; Smolarczyk, Ryszard; Kobusińska, Magdalena; Gajda, Karolina; Wilczek, Piotr; Śliwka, Joanna; Zembala, Michał; Zembala, Marian; Szala, Stanisław


    According to literature data, self-renewing, multipotent, and clonogenic cardiac c-Kit+ progenitor cells occur within human myocardium. The aim of this study was to isolate and characterize c-Kit+ progenitor cells from explanted human hearts. Experimental material was obtained from 19 adult and 7 pediatric patients. Successful isolation and culture was achieved for 95 samples (84.1 %) derived from five different regions of the heart: right and left ventricles, atrium, intraventricular septum,...

  14. Successful Consecutive Expansion of Limbal Explants Using a Biosafe Culture Medium under Feeder Layer-Free Conditions. (United States)

    López-Paniagua, Marina; Nieto-Miguel, Teresa; de la Mata, Ana; Galindo, Sara; Herreras, José M; Corrales, Rosa M; Calonge, Margarita


    Transplantation of in vitro cultured limbal epithelial stem cells (LESCs) is a treatment widely used for LESC deficiency. However, the number of limbal tissue donors is limited, and protocols for LESC cultivation often include compounds and/or feeder layers that can induce side effects and/or increase the cost of the culture procedure. We investigated the feasibility of obtaining more than one limbal primary culture (LPC) from the same biopsy using a culture medium in which several potentially harmful compounds were replaced at the same time by biosafe supplements, allowing the LESC cultivation without feeder layers. We established feeder layer-free LPCs with three culture media: (1) a modified supplemental hormonal epithelial medium, containing potential harmful components (cholera toxin, dimethylsulfoxide, and fetal bovine serum [FBS]), (2) IOBA-FBS, a medium with FBS but with no other harmful supplements, and (3) IOBA-HS, similar to IOBA-FBS but with human serum instead of FBS. Additionally, the same limbal explant was consecutively cultured with IOBA-HS producing three cultures. LPCs were characterized by real-time reverse transcription polymerase chain reaction and/or immunofluorescence. LPCs cultured with the three media under feeder layer-free conditions showed cuboidal cells and no significant differences in the percentage of positive cells for limbal (ABCG2, p63, and K14) and corneal (K3, K12) proteins. Except for ABCG2, the relative mRNA expression of the LESC markers was significantly higher when IOBA-FBS or IOBA-HS was used. LPC1 showed characteristics similar to LPC0, while LPC2 cell morphology became elongated and the expression of some LESC markers was diminished. IOBA-HS enables the culturing of up to two biosafe homologous LPCs from one limbal tissue under feeder layer-free conditions. The routine use of this culture medium could improve both the biosafety and the number of available LPCs for potential clinical transplantation, as well as decrease

  15. Explant culture: An advantageous method for isolation of mesenchymal stem cells from human tissues. (United States)

    Hendijani, Fatemeh


    Mesenchymal stem cell (MSC) research progressively moves towards clinical phases. Accordingly, a wide range of different procedures were presented in the literature for MSC isolation from human tissues; however, there is not yet any close focus on the details to offer precise information for best method selection. Choosing a proper isolation method is a critical step in obtaining cells with optimal quality and yield in companion with clinical and economical considerations. In this concern, current review widely discusses advantages of omitting proteolysis step in isolation process and presence of tissue pieces in primary culture of MSCs, including removal of lytic stress on cells, reduction of in vivo to in vitro transition stress for migrated/isolated cells, reduction of price, processing time and labour, removal of viral contamination risk, and addition of supporting functions of extracellular matrix and released growth factors from tissue explant. In next sections, it provides an overall report of technical highlights and molecular events of explant culture method for isolation of MSCs from human tissues including adipose tissue, bone marrow, dental pulp, hair follicle, cornea, umbilical cord and placenta. Focusing on informative collection of molecular and methodological data about explant methods can make it easy for researchers to choose an optimal method for their experiments/clinical studies and also stimulate them to investigate and optimize more efficient procedures according to clinical and economical benefits. © 2017 John Wiley & Sons Ltd.

  16. Bioassay chamber for angiogenesis with perfused explanted arteries and electrospun scaffolding. (United States)

    Rubenstein, David; Han, Dong; Goldgraben, Sara; El-Gendi, Hebah; Gouma, Pelagia-Irene; Frame, Mary D


    The purpose of this study was to test the hypothesis that explanted perfused arteries can serve as the initial endothelial cell culture source to evaluate the onset of angiogenesis in a cellulose acetate electrospun scaffold. Electrospun scaffolds with fiber diameters roughly controlled in three broad ranges: 0.01 to 0.2, 0.2 to 1, and 1 to 5 microm (Nanomed Nanotechnol Biol Med 2:37-41, 2006), were used in cell culture to determine which provides the best culture topology. This scaffold was then tested in a bioassay chamber whose cellular source was an explanted abdominal aorta from donated euthanized mice. Scaffolds were draped over a cannulated vessel perfused for 24 h. Cell viability, density, and morphology were quantified. The largest fiber diameter group provided the best culture topology for human umbilical vein endothelial cells, showing high cell viability and density, and enhanced elongated cell morphology. Addition of single-walled carbon nanotubes decreased cell density significantly but chitosan heightened cell density and promoted spontaneous capillary tube like structure. Viability of endothelial cells increased with higher flow in the bioassay chamber. Endothelial cells showed a growth preference towards larger diameter fibers. Addition of chitosan improved culture conditions. Thus, this study provides a proof of principle for the possibility of co-culturing tissue engineered vascular networks from a perfused explant.

  17. Pistacia lentiscus fruit oil reduces oxidative stress in human skin explants caused by hydrogen peroxide. (United States)

    Ben Khedir, S; Moalla, D; Jardak, N; Mzid, M; Sahnoun, Z; Rebai, T


    We investigated the efficacy of Pistacia lentiscus fruit oil (PLFO) for protecting human skin from damage due to oxidative stress. PLFO contains natural antioxidants including polyphenols, sterols and tocopherols. We compared the antioxidant potential of PLFO with extra virgin olive oil (EVOO). Explants of healthy adult human skin were grown in culture with either PLFO or EVOO before adding hydrogen peroxide (H2O2). We also used cultured skin explants to investigate the effects of PLFO on lipid oxidation and depletion of endogenous antioxidant defense enzymes including glutathione peroxidase (GPx), superoxide dismutase (SOD) and catalase (CAT) one day after 2 h exposure to H2O2. We found that PLFO scavenged radicals and protected skin against oxidative injury. PLFO exhibited greater antioxidant and free radical scavenging activity than EVOO. Skin explants treated with PLFO inhibited H2O2 induced MDA formation by inhibition of lipid oxidation. In addition, the oil inhibited H2O2 induced depletion of antioxidant defense enzymes including GPx, SOD and CAT. We found that treatment with PLFO repaired skin damage owing to its antioxidant properties.

  18. In vitro regeneration from petiole explants of non-toxic Jatropha curcas

    KAUST Repository

    Kumar, Nitish


    Jatropha curcas, a multipurpose shrub has acquired significant economic potential as biodiesel plant. The seeds or pressed cake is toxic due to the presence of toxic substances and is not useful as food/fodder despite having the best protein composition. A simple, efficient, and reproducible method for plant regeneration through direct organogenesis from petiole explants of non-toxic J. curcas was developed using Murashige and Skoog (MS) medium supplemented with different concentrations of thidiazuron (TDZ). The best induction of shoot buds (57.61%), and number of shoot buds (4.98) per explant were obtained when in vitro petiole explants were placed horizontally on MS medium supplemented with 2.27 mu M TDZ. The Induced shoot buds were transferred to MS medium containing 10 mu M kinetin (Kn), 4.5 mu M 6-benzyl aminopurine (BA), and 5.5 mu M alpha-naphthaleneacetic acid (NAA) for shoot proliferation and subsequent elongation was achieved on MS medium supplemented with 2.25 mu M BA and 8.5 mu M IAA. The elongated shoots could be rooted on half-strength MS medium with 15 mu M IBA, 11.4 mu M IAA and 5.5 mu M NAA with more than 90% survival rate. (C) 2010 Elsevier B.V. All rights reserved.

  19. Pericardium Plug in the Repair of the Corneoscleral Fistula After Ahmed Glaucoma Valve Explantation (United States)

    Yoo, Chungkwon; Kwon, Sung Wook


    We report four cases in which a pericardium (Tutoplast®) plug was used to repair a corneoscleral fistula after Ahmed Glaucoma Valve (AGV) explantation. In four cases in which the AGV tube had been exposed, AGV explantation was performed using a pericardium (Tutoplast®) plug to seal the defect previously occupied by the tube. After debridement of the fistula, a piece of processed pericardium (Tutoplast®), measured 1 mm in width, was plugged into the fistula and secured with two interrupted 10-0 nylon sutures. To control intraocular pressure, a new AGV was implanted elsewhere in case 1, phaco-trabeculectomy was performed concurrently in case 2, cyclophotocoagulation was performed postoperatively in case 3 and anti-glaucomatous medication was added in case 4. No complication related to the fistula developed at the latest follow-up (range: 12~26 months). The pericardium (Tutoplast®) plug seems to be an effective method in the repair of corneoscleral fistulas resulting from explantation of glaucoma drainage implants. PMID:19096247

  20. Endogenous cytokinins in Cocos nucifera L. in vitro cultures obtained from plumular explants. (United States)

    Sáenz, L; Azpeitia, A; Oropeza, C; Jones, L H; Fuchsova, K; Spichal, L; Strnad, M


    Auxin induces in vitro somatic embryogenesis in coconut plumular explants through callus formation. Embryogenic calli and non-embryogenic calli can be formed from the initial calli. Analysis of endogenous cytokinins showed the occurrence of cytokinins with aromatic and aliphatic side chains. Fourteen aliphatic cytokinins and four aromatic cytokinins were analysed in the three types of calli and all the cytokinins were found in each type, although some in larger proportions than others. The most abundant cytokinins in each type of callus were isopentenyladenine-9-glucoside, zeatin-9-glucoside, zeatin riboside, isopentenyladenine riboside, dihydrozeatin and dihydrozeatin riboside in decreasing order. Total cytokinin content was compared between the three types of calli, and it was found to be lower in embryogenic calli compared to non-embryogenic calli or initial calli. The same pattern was observed for individual cytokinins. When explants were cultured in media containing exogenously added cytokinins, the formation of embryogenic calli in the explants was reduced. When 8-azaadenine (an anticytokinin) was added the formation of embryogenic calli and somatic embryos was increased. These results suggest that the difference in somatic embryo formation capacity observed between embryogenic calli and non-embryogenic calli is related to their endogenous cytokinin contents.

  1. Somatic embryogenesis from bud and leaf explants of date palm (Phoenix dactylifera L.) cv. Najda. (United States)

    Mazri, Mouaad Amine; Belkoura, Ilham; Meziani, Reda; Mokhless, Boutaïna; Nour, Souad


    An efficient regeneration system through somatic embryogenesis was developed for date palm cv. Najda. Adventitious bud and proximal leaf segments cultured on Murashige and Skoog (MS) medium supplemented with various combinations of auxins and cytokinins induced embryogenesis after at least 6 months of culture. Somatic embryogenesis induction seemed correlated with the type of the explant, the induction period and the auxin used. The highest rate of somatic embryogenesis (86.0%) was obtained on bud explants cultured on MS medium supplemented with 45.0 µM 2,4-dichlorophenoxyacetic acid (2,4-D), and 4.5 µM kinetin or 4.5 µM 6-(dimethylallylamino) purine (2iP). Whereas, low levels of embryogenesis were obtained on media supplemented with 1-naphthalene acetic acid (NAA) or 2-naphthoxyacetic acid (NOA). Proximal leaf segments showed somatic embryogenesis only when cultured on media supplemented with 2,4-D or picloram. Statistical analysis revealed significant effects of explant type and plant growth regulators (PGRs) combination on somatic embryogenesis. Somatic embryos were germinated successfully on PGR-free MS medium with or without activated charcoal (50.0-60.0 and 26.6-36.6%, respectively), and 80.0% of plantlets survived after transferring to a glasshouse for 6 months. Our results will be useful for large-scale propagation of date palm cv. Najda, characterized by high fruit quality and bayoud disease resistance.

  2. Callogenesis in root explants of four species of the family Solanaceae after inducing by Agrobacterium rhizogenes

    Directory of Open Access Journals (Sweden)

    Zahra Shakeran


    Full Text Available Studying explants affected by Agrobacterium rhizogenes shows that in addition to possible formation of hairy roots, it is likely that callogenesis can be induced in these tissues. The T-DNA region of A. rhizogenes codes enzymes that participate in biosynthesis of plants growth hormones. These hormones also affect callogenesis, hence, the formation of various calluses with different morphological properties are possible. It is very likely that the level of biosynthetic growth hormone, the plasmid carried by each bacteria strain, the position of T-DNA, and the level of gene expression contribute to this morphologic variation. In this study, the root explants of four species of the family Solanaceae namely Atropa belladonna, Datura metel, D. stramonium and Hyoscyamus niger were induced by using different strains of A. rhizogenes (A4, A7, AR15834, AR318, AR9402 and AR9543. Some of these explants entered callus phase and formed various calluses with different colors and shapes. Moreover, in some callus samples hairy roots were also appeared. These variations were probably caused by variations in the levels and ratios of auxin and cytokinine hormons after the induction. As shown in previous studies, the amount of secondary metabolites is reduced due to undifferentiated tissue produced in the callogenesis process.

  3. Bystander-induced differentiation: A major response to targeted irradiation of a urothelial explant model

    Energy Technology Data Exchange (ETDEWEB)

    Belyakov, Oleg V. [Gray Cancer Institute, P.O. Box 100, Mount Vernon Hospital, Northwood, Middlesex HA6 2JR (United Kingdom) and Radiation and Environmental Science Centre, Dublin Institute of Technology, Focas Institute, Kevin St., Dublin 9 (Ireland)]. E-mail:; Folkard, Melvyn [Gray Cancer Institute, P.O. Box 100, Mount Vernon Hospital, Northwood, Middlesex HA6 2JR (United Kingdom); Mothersill, Carmel [Radiation and Environmental Science Centre, Dublin Institute of Technology, Focas Institute, Kevin St., Dublin 9 (Ireland); Prise, Kevin M. [Gray Cancer Institute, P.O. Box 100, Mount Vernon Hospital, Northwood, Middlesex HA6 2JR (United Kingdom); Michael, Barry D. [Gray Cancer Institute, P.O. Box 100, Mount Vernon Hospital, Northwood, Middlesex HA6 2JR (United Kingdom)


    A ureter primary explant technique, using porcine tissue sections was developed to study bystander effects under in vivo like conditions where dividing and differentiated cells are present. Targeted irradiations of ureter tissue fragments were performed with the Gray Cancer Institute charged particle microbeam at a single location (2 {mu}m precision) with 10 {sup 3}He{sup 2+} particles (5 MeV; LET 70 keV/{mu}m). After irradiation the ureter tissue section was incubated for 7 days allowing explant outgrowth to be formed. Differentiation was estimated using antibodies to Uroplakin III, a specific marker of terminal urothelial differentiation. Even although only a single region of the tissue section was targeted, thousands of additional cells were found to undergo bystander-induced differentiation in the explant outgrowth. This resulted in an overall increase in the fraction of differentiated cells from 63.5 {+-} 5.4% to 76.6 {+-} 5.6%. These changes are much greater than that observed for the induction of damage in this model. One interpretation of these results is that in the tissue environment, differentiation is a much more significant response to targeted irradiation and potentially a protective mechanism.

  4. Cartilage Health in Knees Treated with Metal Resurfacing Implants or Untreated Focal Cartilage Lesions: A Preclinical Study in Sheep. (United States)

    Martinez-Carranza, Nicolas; Hultenby, Kjell; Lagerstedt, Anne Sofie; Schupbach, Peter; Berg, Hans E


    Background Full-depth cartilage lesions do not heal and the long-term clinical outcome is uncertain. In the symptomatic middle-aged (35-60 years) patient, treatment with metal implants has been proposed. However, the cartilage health surrounding these implants has not been thoroughly studied. Our objective was to evaluate the health of cartilage opposing and adjacent to metal resurfacing implants. Methods The medial femoral condyle was operated in 9 sheep bilaterally. A metallic resurfacing metallic implant was immediately inserted into an artificially created 7.5 mm defect while on the contralateral knee the defect was left untreated. Euthanasia was performed at 6 months. Six animals, of similar age and study duration, from a previous study were used for comparison in the evaluation of cartilage health adjacent to the implant. Cartilage damage to joint surfaces within the knee, cartilage repair of the defect, and cartilage adjacent to the implant was evaluated macroscopically and microscopically. Results Six animals available for evaluation of cartilage health within the knee showed a varying degree of cartilage damage with no statistical difference between defects treated with implants or left untreated ( P = 0.51; 95% CI -3.7 to 6.5). The cartilage adjacent to the implant (score 0-14; where 14 indicates no damage) remained healthy in these 6 animals showing promising results (averaged 10.5; range 9-11.5, SD 0.95). Cartilage defects did not heal in any case. Conclusion Treatment of a critical size focal lesion with a metal implant is a viable alternative treatment.

  5. A Novel Approach to Stimulate Cartilage Repair: Targeting Collagen Turnover

    NARCIS (Netherlands)

    Y.M. Bastiaansen-Jenniskens (Yvonne)


    textabstractOA is a complex disease of which the ethiopathology is not completely known and therapies to repair cartilage are still under investigation. The increase of collagen type II expression in osteoarthritic cartilage suggests an activated repair mechanism that is however ineffective in

  6. A Dual Flow Bioreactor for Cartilage Tissue Engineering

    NARCIS (Netherlands)

    Spitters, Tim


    Preventing the onset of a degenerative disease like osteoarthritis by restoring tissue function before cartilage degradation occurs will decrease health costs, reduce socio-economic burdens of patients and preserve quality of life. However, producing ex vivo cartilage implants of clinically relevant

  7. Culturing functional cartilage tissue under a novel bionic mechanical condition. (United States)

    Sun, Minglin; Lv, Dan; Zhang, Chunqiu; Zhu, Lei


    Bioreactor, which is used for in vitro construction of tissue-engineered cartilage, has been extensively studied by researchers. The growth and development of articular cartilage tissue are affected by biomechanical and biochemical factors, especially mechanical condition. Kinds of mechanical conditions including compressive and shear force, fluid flow, hydrostatic pressure, and tissue deformation, were developed in the past years. However, most mechanical conditions of improved bioreactor involve only one or two external force, which is merely partial for engineering cartilage tissue. No bioreactor which can simulate a normal articular cartilage in terms of structure and function has been reported. Consequently, simulation of bionic mechanical environment of a normal articular cartilage is considered to be the optimal environment for culturing the functional articular cartilage in vitro. Based upon this purpose, we designed a rolling-compression loading bioreactor. It could provide cultures with multi-mechanical stimulations and sufficiently mimic the complex mechanical environment of a normal articular cartilage. We propose that this comprehensive rolling-compression loading bioreactor can enhance the cultivation of functional cartilage constructs in vitro. Copyright © 2010 Elsevier Ltd. All rights reserved.

  8. Two dimensional spectral camera development for cartilage monitoring (United States)

    Kuehn, A.; Graf, A.; Wenzel, U.; Princz, S.; Miller, R.; Mantz, H.; Hessling, M.


    In the joint project "BioopTiss" between the Ulm University Medical Center and Ulm University of Applied Sciences, a bioreactor is under development to grow facial cartilage by the methods of tissue engineering. In order to ensure a sufficient quality of the cartilage for implantation, the cartilage growth must be monitored continuously. Current monitoring methods destroy the cultured cartilage so that it is no longer suitable for implantation. Alternatively, it is possible to analyze the cartilage using fluorescence spectroscopy with UV light excitation. This allows a non-invasive assessment of cartilage in terms of composition and quality. The cultured cartilage tissue can reach a size of several square centimeters. For recording fluorescence spectra of every point of the cartilage sample, a highly sensitive spectral camera has been developed which allows distinguishing collagen I from collagen II non-invasively by their fluorescence. This spectral camera operates according to the computed tomography imaging spectrometry (CTIS) principle, which allows obtaining many spectra of a small area with only one snapshot.

  9. Cartilage Tissue Engineering: Controversy in the Effect of Oxygen

    NARCIS (Netherlands)

    Malda, J.; Martens, D.E.; Tramper, J.; Blitterswijk, van C.A.; Riesle, J.


    Articular cartilage lacks the ability to repair itself and consequently defects in this tissue do not heal. Tissue engineering approaches, employing a scaffold material and cartilage producing cells (chondrocytes), hold promise for the treatment of such defects. In these strategies the limitation of

  10. Repair of osteochondral defects with allogeneic tissue engineered cartilage implants. (United States)

    Schreiber, R E; Ilten-Kirby, B M; Dunkelman, N S; Symons, K T; Rekettye, L M; Willoughby, J; Ratcliffe, A


    The objective of this study was to evaluate the effect of allogeneic tissue engineered cartilage implants on healing of osteochondral defects. Rabbit chondrocytes were cultured in monolayer, then seeded onto biodegradable, three-dimensional polyglycolic acid meshes. Cartilage constructs were cultured hydrodynamically to yield tissue with relatively more (mature) or less (immature) hyalinelike cartilage, as compared with adult rabbit articular cartilage. Osteochondral defects in the patellar grooves of both stifle joints either were left untreated or implanted with allogeneic tissue engineered cartilage. Histologic samples from in and around the defect sites were examined 3, 6, 9, and 12, and 24 months after surgery. By 9 months after surgery, defects sites treated with cartilage implants contained significantly greater amounts of hyalinelike cartilage with high levels of proteoglycan, and had a smooth, nonfibrillated articular surface as compared to untreated defects. In contrast, the repair tissue formed in untreated defects had fibrillated articular surfaces, significant amounts of fibrocartilage, and negligible proteoglycan. These differences between treated and untreated defects persisted through 24 months after surgery. The results of this study suggest that the treatment of osteochondral lesions with allogenic tissue engineered cartilage implants may lead to superior repair tissue than that found in untreated osteochondral lesions.

  11. Improvement of PHBV scaffolds with bioglass for cartilage tissue engineering.

    Directory of Open Access Journals (Sweden)

    Jun Wu

    Full Text Available Polymer scaffold systems consisting of poly(hydroxybutyrate-co-hydroxyvalerate (PHBV have proven to be possible matrices for the three-dimensional growth of chondrocyte cultures. However, the engineered cartilage grown on these PHBV scaffolds is currently unsatisfactory for clinical applications due to PHBV's poor hydrophilicity, resulting in inadequate thickness and poor biomechanical properties of the engineered cartilage. It has been reported that the incorporation of Bioglass (BG into PHBV can improve the hydrophilicity of the composites. In this study, we compared the effects of PHBV scaffolds and PHBV/BG composite scaffolds on the properties of engineered cartilage in vivo. Rabbit articular chondrocytes were seeded into PHBV scaffolds and PHBV/BG scaffolds. Short-term in vitro culture followed by long-term in vivo transplantation was performed to evaluate the difference in cartilage regeneration between the cartilage layers grown on PHBV and PHBV/BG scaffolds. The results show that the incorporation of BG into PHBV efficiently improved both the hydrophilicity of the composites and the percentage of adhered cells and promoted cell migration into the inner part the constructs. With prolonged incubation time in vivo, the chondrocyte-scaffold constructs in the PHBV/BG group formed thicker cartilage-like tissue with better biomechanical properties and a higher cartilage matrix content than the constructs in the PHBV/BG group. These results indicate that PHBV/BG scaffolds can be used to prepare better engineered cartilage than pure PHBV.

  12. Improvement of PHBV Scaffolds with Bioglass for Cartilage Tissue Engineering (United States)

    Li, Haiyan; Sun, Junying; Liu, Kai


    Polymer scaffold systems consisting of poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) have proven to be possible matrices for the three-dimensional growth of chondrocyte cultures. However, the engineered cartilage grown on these PHBV scaffolds is currently unsatisfactory for clinical applications due to PHBV’s poor hydrophilicity, resulting in inadequate thickness and poor biomechanical properties of the engineered cartilage. It has been reported that the incorporation of Bioglass (BG) into PHBV can improve the hydrophilicity of the composites. In this study, we compared the effects of PHBV scaffolds and PHBV/BG composite scaffolds on the properties of engineered cartilage in vivo. Rabbit articular chondrocytes were seeded into PHBV scaffolds and PHBV/BG scaffolds. Short-term in vitro culture followed by long-term in vivo transplantation was performed to evaluate the difference in cartilage regeneration between the cartilage layers grown on PHBV and PHBV/BG scaffolds. The results show that the incorporation of BG into PHBV efficiently improved both the hydrophilicity of the composites and the percentage of adhered cells and promoted cell migration into the inner part the constructs. With prolonged incubation time in vivo, the chondrocyte-scaffold constructs in the PHBV/BG group formed thicker cartilage-like tissue with better biomechanical properties and a higher cartilage matrix content than the constructs in the PHBV/BG group. These results indicate that PHBV/BG scaffolds can be used to prepare better engineered cartilage than pure PHBV. PMID:23951190

  13. Integrative studies on cartilage tissue engineering and joint homeostasis

    NARCIS (Netherlands)

    Rutgers, M.


    The impact of cartilage injury to the joint is often larger than the initial clinical symptoms suggest. Through an alteration in joint homeostasis and biomechanical loading, cartilage lesions may accelerate osteoarthritis onset. Although good clinical results are achieved in patients treated by the

  14. The histologic relationship of preauricular sinuses to auricular cartilage. (United States)

    Dunham, Brian; Guttenberg, Martha; Morrison, Wynne; Tom, Lawrence


    To determine the histologic relationship and distance between excised preauricular epithelial sinus tract and the adjacent auricular cartilage (sinocartilaginous distance) in a series of patients. The excision of preauricular sinuses is a common surgical procedure. Recurrences are frequent and can be technically challenging. While advocated by several authors, the surgical removal of adjacent auricular cartilage is not universally performed. Retrospective case series. Children's Hospital of Philadelphia. Fifty-two pediatric patients who underwent surgical excision of preauricular sinus tracts and adjacent auricular cartilage. Between September 1, 2005, and July 31, 2007, the preauricular sinus tracts and adjacent auricular cartilage were excised from 52 pediatric patients. A pathologist reviewed a total of 58 specimens to determine the relationship between epithelial tract and cartilage. The sinocartilaginous distance in microns. Patient ages ranged from 8 months to 17 years (mean age, 4 years). In all but 1 case, the tracts were in close proximity to the cartilage. The average sinocartilaginous distance was 472 mum (median distance, 400 mum); the 25th percentile was 250 mum. In over 50% of the specimens, the sinocartilaginous distance was less than 0.5 mm, and in nearly all of the these, the epithelial tract was in continuity with stromal tissue histologically indistinguishable from perichondrium. The observed sinocartilaginous distances suggest that it may be difficult to dissect most sinus tracts from the cartilage. The routine removal of a small portion of auricular cartilage along with the sinus tract may yield a more thorough excision and help to prevent recurrence.

  15. Effect of scopoletin on fascia-wrapped diced cartilage grafts

    African Journals Online (AJOL)

    Surgically wrapped diced cartilages exhibit various degrees of resorption; thus, it has been recommended that fascia be used to wrap diced cartilages. However, few surgeons suggest the use of AlloDerm for wrapping because the harvesting of fascia may cause hematoma and alopecia [17]. Additionally, block grafts have a.

  16. Combined role of type IX collagen and cartilage oligomeric matrix protein in cartilage matrix assembly: Cartilage oligomeric matrix protein counteracts type IX collagen-induced limitation of cartilage collagen fibril growth in mouse chondrocyte cultures

    NARCIS (Netherlands)

    Blumbach, K.; Bastiaansen-Jenniskens, Y.M.; Groot, J. de; Paulsson, M.; Osch, G.J.V.M. van; Zaucke, F.


    Objective. Defects in the assembly and composition of cartilage extracellular matrix are likely to result in impaired matrix integrity and increased susceptibility to cartilage degeneration. The aim of this study was to determine the functional interaction of the collagen fibril-associated proteins

  17. In Vitro Models For Cell-Based Cartilage Regeneration

    NARCIS (Netherlands)

    M.L. de Vries-Van Melle (Marloes)


    markdownabstract__Abstract__ Covering the ends of the long bones in articular joints, cartilage allows smooth, gliding movement of the joints and distributes load evenly across the joint surface. Lesions in the articular cartilage are a major cause of discomfort and disability, especially in


    NARCIS (Netherlands)


    The usefulness of thionin for staining cartilage sections embedded in glycol methacrylate (GMA) and the effect of decalcification on cartilage sections embedded in paraffin and GMA were assessed. Short decalcification periods using 5% formic acid or 10% EDTA did not influence the staining properties

  19. Recent advances in hydrogels for cartilage tissue engineering. (United States)

    Vega, S L; Kwon, M Y; Burdick, J A


    Articular cartilage is a load-bearing tissue that lines the surface of bones in diarthrodial joints. Unfortunately, this avascular tissue has a limited capacity for intrinsic repair. Treatment options for articular cartilage defects include microfracture and arthroplasty; however, these strategies fail to generate tissue that adequately restores damaged cartilage. Limitations of current treatments for cartilage defects have prompted the field of cartilage tissue engineering, which seeks to integrate engineering and biological principles to promote the growth of new cartilage to replace damaged tissue. To date, a wide range of scaffolds and cell sources have emerged with a focus on recapitulating the microenvironments present during development or in adult tissue, in order to induce the formation of cartilaginous constructs with biochemical and mechanical properties of native tissue. Hydrogels have emerged as a promising scaffold due to the wide range of possible properties and the ability to entrap cells within the material. Towards improving cartilage repair, hydrogel design has advanced in recent years to improve their utility. Some of these advances include the development of improved network crosslinking (e.g. double-networks), new techniques to process hydrogels (e.g. 3D printing) and better incorporation of biological signals (e.g. controlled release). This review summarises these innovative approaches to engineer hydrogels towards cartilage repair, with an eye towards eventual clinical translation.

  20. Engineering superficial zone features in tissue engineered cartilage. (United States)

    Chen, Tony; Hilton, Matthew J; Brown, Edward B; Zuscik, Michael J; Awad, Hani A


    A major challenge in cartilage tissue engineering is the need to recreate the native tissue's anisotropic extracellular matrix structure. This anisotropy has important mechanical and biological consequences and could be crucial for integrative repair. Here, we report that hydrodynamic conditions that mimic the motion-induced flow fields in between the articular surfaces in the synovial joint induce the formation of a distinct superficial layer in tissue engineered cartilage hydrogels, with enhanced production of cartilage matrix proteoglycan and Type II collagen. Moreover, the flow stimulation at the surface induces the production of the surface zone protein Proteoglycan 4 (aka PRG4 or lubricin). Analysis of second harmonic generation signature of collagen in this superficial layer reveals a highly aligned fibrillar matrix that resembles the alignment pattern in native tissue's surface zone, suggesting that mimicking synovial fluid flow at the cartilage surface in hydrodynamic bioreactors could be key to creating engineered cartilage with superficial zone features. Copyright © 2012 Wiley Periodicals, Inc.


    Directory of Open Access Journals (Sweden)

    Yuri M. Iryanov, Nikolay A. Kiryanov, Olga V. Dyuriagina , Tatiana Yu. Karaseva, Evgenii A. Karasev


    Full Text Available Background: The damage or loss of articular cartilage is costly medical problem. The purpose of this work – morphological analysis of reparative chondrogenesis when implanted in the area of the knee joint cartilage of granulated mineralized bone matrix. Material and Methods: The characteristic features of the knee cartilage regeneration studied experimentally in pubertal Wistar rats after modeling a marginal perforated defect and implantation of granulated mineralized bone matrix obtained according to original technology without heat and demineralizing processing into the injury zone. Results: This biomaterial established to have pronounced chondro- and osteoinductive properties, and to provide prolonged activation of reparative process, accelerated organotypical remodeling and restoration of the articular cartilage injured. Conclusion: The data obtained demonstrate the efficacy of МВМ in clinical practice for the treatment of diseases and injuries of the articular cartilage.

  2. Injectable hydrogels for cartilage and bone tissue engineering (United States)

    Liu, Mei; Zeng, Xin; Ma, Chao; Yi, Huan; Ali, Zeeshan; Mou, Xianbo; Li, Song; Deng, Yan; He, Nongyue


    Tissue engineering has become a promising strategy for repairing damaged cartilage and bone tissue. Among the scaffolds for tissue-engineering applications, injectable hydrogels have demonstrated great potential for use as three-dimensional cell culture scaffolds in cartilage and bone tissue engineering, owing to their high water content, similarity to the natural extracellular matrix (ECM), porous framework for cell transplantation and proliferation, minimal invasive properties, and ability to match irregular defects. In this review, we describe the selection of appropriate biomaterials and fabrication methods to prepare novel injectable hydrogels for cartilage and bone tissue engineering. In addition, the biology of cartilage and the bony ECM is also summarized. Finally, future perspectives for injectable hydrogels in cartilage and bone tissue engineering are discussed. PMID:28584674

  3. Cell-mediated contraction of vitreous explants from chicken embryo: Possibility of screening for therapeutic agents against proliferative vitreoretinal diseases (United States)

    Oki, Keitaro; Shimada, Arata; Nagase, Terumasa; Katsura, Yoshiya; Kosano, Hiroshi


    Purpose We aimed to establish a novel screening system for identifying potential therapeutic agents for treating proliferative vitreoretinal diseases (PVDs). In this study, we focused on vitreous explants from chicken embryos and evaluated the usefulness of quantitatively analyzing the effects of potential candidates on cell-mediated vitreous contraction, which leads to blindness in PVDs. Methods Vitreous explants were extracted from 19-day-old embryonic chickens and then incubated with retinal Müller cells or endothelial cells to permit cell adhesion. After cell adhesion occurred, we examined the effect of the attached cells on the wet weight of vitreous explants as an index of vitreous contraction. We also performed hematoxylin and eosin staining to characterize the cell morphology on the vitreous surface. Results Contraction of the vitreous explants was observed after cell adhesion of not only retinal Müller cells but also endothelial cells. We confirmed the adhesion of these cells on vitreous explants and estimated the number of adherent cells with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) analysis. The cells on the vitreous surface presented an elongated fibroblast-like phenotype. Integrin was found to be a receptor involved in cell adhesion on the vitreous surface. Discussion Our results suggest that vitreous explants from chicken embryos may be novel useful tools for screening antiadhesion therapeutic agents in PVDs. This preliminary study must be validated with human vitreous and human retinal pigment epithelial cells. PMID:24319331

  4. A rooting procedure for lentil (Lens culinaris Medik.) and other hypogeous legumes (pea, chickpea and Lathyrus) based on explant polarity. (United States)

    Fratini, R; Ruiz, M L


    The present study assessed the rooting response of lentil nodal segments in relation to explant polarity, hormone, salt and carbohydrate concentrations of the medium. Nodal segments of lentil with an axillary bud cultured in an inverted orientation (apical end in medium) showed higher rooting frequencies than explants cultured in a normal orientation (basal end in medium). The highest rooting percentage (95.35%) and average number of shoots regenerated per explant (2.4) were obtained from explants placed in an inverted orientation on Murashige and Skoog (MS) medium salts with 3% sucrose, supplemented with 5 microM indole acetic acid (IAA) and 1 microM kinetin (KN). Reducing or increasing phytohormone concentration did not alter significantly root regeneration of inverted explants. Sucrose at 3% allowed higher root regeneration frequencies compared to 1.5% sucrose. MS full concentration permitted regeneration of longer shoots with more nodes per regenerated shoot, compared to MS half-strength, which regenerated more shoots of shorter length and with less nodes. Inverted nodal segments of other hypogeous legumes (pea, chickpea and Lathyrus) also exhibited higher rooting frequencies than explants cultured in a normal orientation on MS medium with 3% sucrose and supplemented with 5 microM IAA and 1 microM KN. The most novel application of this study is the culture of nodal segments of hypogeous legumes in an inverted orientation. This procedure is a considerable improvement over other published procedures concerning in vitro rooting of lentil, pea, chickpea and Lathyrus.

  5. Application of an acoustofluidic perfusion bioreactor for cartilage tissue engineering. (United States)

    Li, Siwei; Glynne-Jones, Peter; Andriotis, Orestis G; Ching, Kuan Y; Jonnalagadda, Umesh S; Oreffo, Richard O C; Hill, Martyn; Tare, Rahul S


    Cartilage grafts generated using conventional static tissue engineering strategies are characterised by low cell viability, suboptimal hyaline cartilage formation and, critically, inferior mechanical competency, which limit their application for resurfacing articular cartilage defects. To address the limitations of conventional static cartilage bioengineering strategies and generate robust, scaffold-free neocartilage grafts of human articular chondrocytes, the present study utilised custom-built microfluidic perfusion bioreactors with integrated ultrasound standing wave traps. The system employed sweeping acoustic drive frequencies over the range of 890 to 910 kHz and continuous perfusion of the chondrogenic culture medium at a low-shear flow rate to promote the generation of three-dimensional agglomerates of human articular chondrocytes, and enhance cartilage formation by cells of the agglomerates via improved mechanical stimulation and mass transfer rates. Histological examination and assessment of micromechanical properties using indentation-type atomic force microscopy confirmed that the neocartilage grafts were analogous to native hyaline cartilage. Furthermore, in the ex vivo organ culture partial thickness cartilage defect model, implantation of the neocartilage grafts into defects for 16 weeks resulted in the formation of hyaline cartilage-like repair tissue that adhered to the host cartilage and contributed to significant improvements to the tissue architecture within the defects, compared to the empty defects. The study has demonstrated the first successful application of the acoustofluidic perfusion bioreactors to bioengineer scaffold-free neocartilage grafts of human articular chondrocytes that have the potential for subsequent use in second generation autologous chondrocyte implantation procedures for the repair of partial thickness cartilage defects.

  6. Biochemical effects on long-term frozen human costal cartilage

    Energy Technology Data Exchange (ETDEWEB)

    Santin, Stefany P.; Martinho Junior, Antonio C.; Yoshito, Daniele; Soares, Fernando A.N.; Mathor, Monica B., E-mail: mathor@ipen.b [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)


    Currently, the progresses on treatment of musculoskeletal diseases with the evolving of artificial implants and the success of tissue transplantation between genetically different individuals have conducted to an increase in radiosterilization. Regarding to tissue transplantation, it is essential to have sterile tissue and many tissue banks use radiosterilization as an effective method to sterilize these tissues. However, high doses of ionizing radiation and the preservation method may induce structural modifications in the tissues, as degradation of structural scaffold, decreasing its mechanical properties. Particularly, cartilage have been preserved in high concentrations of glycerol or deep-frozen at -70 degree C for storage after radiosterilization. Therefore, it is important to study the modifications induced in cartilage by preservation methods and by radiosterilization to determine the appropriated parameters for high quality of human allografts. Costal cartilages were obtained from cadaveric donors and were frozen at -20 degree C for 2 years long in order to compare with previous studies for fresh, deep-frozen and glycerolised cartilages. The mechanical tests were carried out in a universal testing machine until sample failure. According our results, there is no significant statistical difference between stress at break of fresh, long-term - 20 degree C frozen cartilages and deep-frozen cartilage. This early result suggests, regarding to tensile property, that long-term - 20 degree C frozen cartilages corresponds to glycerolised costal cartilages irradiated with 25 kGy or deep-frozen cartilages irradiated with 25 and 50 kGy. Thus, this long-term frozen cartilages may be used for tissue banks, but more studies about effects of ionizing radiation are necessary. (author)

  7. High fat diet accelerates cartilage repair in DBA/1 mice. (United States)

    Wei, Wu; Bastiaansen-Jenniskens, Yvonne M; Suijkerbuijk, Mathijs; Kops, Nicole; Bos, Pieter K; Verhaar, Jan A N; Zuurmond, Anne-Marie; Dell'Accio, Francesco; van Osch, Gerjo J V M


    Obesity is a well-known risk factor for osteoarthritis, but it is unknown what it does on cartilage repair. Here we investigated whether a high fat diet (HFD) influences cartilage repair in a mouse model of cartilage repair. We fed DBA/1 mice control or HFD (60% energy from fat). After 2 weeks, a full thickness cartilage defect was made in the trochlear groove. Mice were sacrificed, 1, 8, and 24 weeks after operation. Cartilage repair was evaluated on histology. Serum glucose, insulin and amyloid A were measured 24 h before operation and at endpoints. Immunohistochemical staining was performed on synovium and adipose tissue to evaluate macrophage infiltration and phenotype. One week after operation, mice on HFD had defect filling with fibroblast-like cells and more cartilage repair as indicated by a lower Pineda score. After 8 weeks, mice on a HFD still had a lower Pineda score. After 24 weeks, no mice had complete cartilage repair and we did not detect a significant difference in cartilage repair between diets. Bodyweight was increased by HFD, whereas serum glucose, amyloid A and insulin were not influenced. Macrophage infiltration and phenotype in adipose tissue and synovium were not influenced by HFD. In contrast to common wisdom, HFD accelerated intrinsic cartilage repair in DBA/1 mice on the short term. Resistance to HFD induced inflammatory and metabolic changes could be associated with accelerated cartilage repair. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1258-1264, 2017. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  8. Development of a cartilage composite utilizing porous tantalum, fibrin, and rabbit chondrocytes for treatment of cartilage defect. (United States)

    Jamil, Kamal; Chua, Kien-Hui; Joudi, Samad; Ng, Sook-Luan; Yahaya, Nor Hamdan


    Functional tissue engineering has emerged as a potential means for treatment of cartilage defect. Development of a stable cartilage composite is considered to be a good option. The aim of the study was to observe whether the incorporation of cultured chondrocytes on porous tantalum utilizing fibrin as a cell carrier would promote cartilage tissue formation. Rabbit articular chondrocytes were cultured and seeded onto tantalum with fibrin as temporary matrix in a composite, which was divided into three groups. The first group was kept in vitro while a total of 12 constructs were implanted into the dorsum of mice for the second and third groups. The implanted tissues were harvested after 4 weeks (second group) and after 8 weeks (third group). Specific characteristic of cartilage growth were studied by histological and biochemical assessment, immunohistochemistry, and quantitative PCR analysis. Histological and biochemical evaluation of the formed cartilage using hematoxylin and eosin and Alcian blue staining showed lacunae chondrocytes embedded in the proteoglycan rich matrix. Dimethylmethylene blue assay demonstrated high glycosaminoglycans content in the removed tissue following 8 weeks of implantation. Immunohistochemistry results showed the composites after implantation expressed high collagen type II. Quantitative PCR results confirmed a significant increase in cartilage associated genes expression (collagen type II, AggC, Sox 9) after implantation. Tantalum scaffold with fibrin as cell carrier promotes chondrocyte proliferation and cartilaginous tissue formation. Producing hyaline cartilage within a stable construct of tantalum and fibrin has a potential for treatment of cartilage defect.

  9. Effects of platelet-rich plasma on cartilage regeneration after costal cartilage resection: a stereological and histopathological study. (United States)

    Sengul, Aysen Taslak; Buyukkkarabacak, Yasemin Bilgin; Altunkaynak, Berrin Zuhal; Yetim, Tulin Durgun; Altun, Gamze Yayla; Sengul, Bilal; Basoglu, Ahmet


    In cases of congenital chest wall deformities, it is important to maintain the flexibility of the chest wall after rib cartilage resection. In this study, we aimed to determine the regeneration capability of cartilage and the effects of platelet-rich plasma (PRP) on the regeneration process. A total of 16 four-week-old New Zealand rabbits were used in this study. In the 4th-5th right costal cartilages, the perichondrial sheaths were dissected and costal cartilages were excised. Then, the perichondrial sheaths were closed with absorbable material in the sham group (n = 8), and this was done after replacing PRP in the PRP group (n = 8). The left costal cartilages of the animals were used as controls. The volumes of the costal cartilages and their perichondrial sheaths were estimated using Cavalieri's principle. In addition, the mean numerical densities of the chondroblasts and chondrocytes per square millimetre were estimated using unbiased counting frames. In the PRP and sham groups, the volumes of the cartilages and perichondrial sheaths were higher than those of the control group (p < 0.05). The numerical densities of the chondroblasts and chondrocytes increased more in the PRP group than in the sham group (p < 0.05). Applying PRP after resection may provide better healing and faster regeneration of cartilage.

  10. Special pattern of endochondral ossification in human laryngeal cartilages: X-ray and light-microscopic studies on thyroid cartilage. (United States)

    Claassen, Horst; Schicht, Martin; Sel, Saadettin; Paulsen, Friedrich


    Endochondral ossification is a process that also occurs in the skeleton of the larynx. Differences in the ossification mechanism in comparison to growth plates are not understood until now. To get deeper insights into this process, human thyroid cartilage was investigated by the use of X-rays and a series of light-microscopic stainings. A statistical analysis of mineralization was done by scanning areas of mineralized cartilage and of ossification. We detected a special mode of endochondral ossification which differs from the processes in growth plates. Thyroid cartilage ossifies very slowly and in a gender-specific manner. Compared with age-matched women, bone formation in thyroid cartilage of men is significantly higher in the age group 41-60 years. Endochondral ossification is prepared by internal changes of extracellular matrix leading to areas of asbestoid fibers with ingrowing cartilage canals. In contrast to growth plates, bone is deposited on large areas of mineralized cartilage, which appear at the rims of cartilage canals. Furthermore, primary parallel fibered bone was observed which was deposited on woven bone. The predominant bone type is cancellous bone with trabeculae, whereas compact bone with Haversian systems was seldom found. Trabeculae contain a great number of reversal and arresting lines meaning that the former were often reconstructed and that bone formation was arrested and resumed again with advancing age. It is hypothesized that throughout life trabeculae of ossified thyroid cartilage undergo adaptation to different loads due to the use of voice. Copyright © 2014 Wiley Periodicals, Inc.

  11. [Graft integration in the lumbar spine of bovine cancellous bone compared to autologous iliac crest in a sheep model]. (United States)

    Strohm, P C; Kubosch, D C; Sprecher, C M; Schmal, H; Südkamp, N P; Milz, S


    Spinal injuries are common and a standard procedure for the stabilisation of spinal injuries is ventral spondylodesis with an autograft from the iliac crest. Because of the high incidence of harvesting complications there is a need to search for alternative materials. The aim of our study was to evaluate graft integration in the lumbar spine of bovine cancellous bone compared to autologous iliac crest material. Two groups of eight female adult sheep (median age 3 years, range 2.4-3.8 years) received surgical treatment in the form of anterior monosegmental spondylodesis. The spondylodesis was performed in all animals in the motion segment L3/4 through a lateral approach with the animals lying on their right sides. To produce serial sections, the explanted vertebral segments were implanted in methyl methacrylate. On one side the histological preparation was examined qualitatively and in addition we analysed the quantity of the bone structure with special software. The bone structure in both groups did not differ significantly and demonstrated integration of the grafts in the adjacent vertebral bodies. Fractures and lysis occurred in the region of the intervertebral disc and were more frequent in the group with the bovine graft. The bony integration of the grafts of both groups was not significantly different and showed good results. Almost all of the bovine grafts fractured or presented regions of lysis. In our opinion bovine cancellous bone graft is not a good alternative to autologous iliac crest. © Georg Thieme Verlag KG Stuttgart · New York.

  12. Deformation of Nasal Septal Cartilage During Mastication (United States)

    Dayeh, Ayman A. Al; Rafferty, Katherine L.; Egbert, Mark; Herring, Susan W.


    The cartilaginous nasal septum plays a major role in structural integrity and growth of the face, but its internal location has made physiologic study difficult. By surgically implanting transducers in 10 miniature pigs (Sus scrofa), we recorded in vivo strains generated in the nasal septum during mastication and masseter stimulation. The goals were (1) to determine whether the cartilage should be considered as a vertical strut supporting the nasal cavity and preventing its collapse, or as a damper of stresses generated during mastication and (2) to shed light on the overall pattern of snout deformation during mastication. Strains were recorded simultaneously at the septo-ethmoid junction and nasofrontal suture during mastication. A third location in the anterior part of the cartilage was added during masseter stimulation and manipulation. Contraction of jaw closing muscles during mastication was accompanied by anteroposterior compressive strains (around −1,000 με) in the septo-ethmoid junction. Both the orientation and the magnitude of the strain suggest that the septum does not act as a vertical strut but may act in absorbing loads generated during mastication. The results from masseter stimulation and manipulation further suggest that the masticatory strain pattern arises from a combination of dorsal bending and/or shearing and anteroposterior compression of the snout. J. Morphol. PMID:19434723

  13. Sealing and explant types on the mangaba micropropagation Tipo de vedação e explantes na micropropagação de mangabeira

    Directory of Open Access Journals (Sweden)

    Aline de Jesus Sá


    Full Text Available In micropropagation, especially for mangaba tree botanical variety of Northeastern Brazil, limiting aspects such as ethylene accumulation in the cultivation flask and loss of vigor in subcultures have been observed. This study was aimed at assessing the technical and scientific knowledge of the in vitro propagation of botanical mangaba tree variety and at improving the micropropagation protocol, establishing the in vitro cultivation time, the best type of flask sealing and explant at different micropropagation stages. For the establishment phase and for the first and second subcultures, the MS medium with 3% sucrose and 0.6% agar, supplemented with 1 mg L-1 IAA and 1 mg L-1 BA was used. Evaluations were performed at 30, 50 and 65 days of in vitro cultivation. The best types of flask sealing for the establishment phase were the PVC film and Para-film® and for the first subculture the Para-film® seal. In the second subculture the PVC film and Para-film® seals promoted the best growth. The median and basal nodal segments presented the best performance in the first subculture. No significant effect of explant type was observed in the second subculture. The ideal subculture interval in the establishment phase and the first and second subcultures is 50 days.Na micropropagação, especialmente para mangaba, variedade botânica da árvore do Nordeste do Brasil, aspectos limitantes, como acúmulo de etileno no recipiente de cultivo e perda de vigor em subculturas têm sido observados. Neste estudo, objetivou-se avaliar o conhecimento técnico e científico da propagação in vitro de mangabeira, variedade botânica do Nordeste do Brasil, e melhorar o protocolo de micropropagação, o melhor tipo de vedação frasco e explante em diferentes etapas. Para a fase de estabelecimento e para as subculturas primeiro e segundo, foi utlizado o meio MS com 3% de sacarose e agar 0,6%, suplementado com 1 mg L-1 de AIA e 1 mg L-1 de BAP. As avaliações foram

  14. Human Platelet Lysate as a Replacement for Fetal Bovine Serum in Limbal Stem Cell Therapy. (United States)

    Suri, Kunal; Gong, Hwee K; Yuan, Ching; Kaufman, Stephen C


    To evaluate the use of human platelet lysate (HPL) as an alternative supplement for limbal explant culture. Culture media were prepared using either 10% pooled HPL (PHPL), single donor HPL, or fetal bovine serum (FBS). Limbal tissues, obtained from the Minnesota Lions Eye Bank, were cultured in each medium on plastic plates or on denuded amniotic membrane (AM). Immunofluorescence staining was performed for ABCG2, tumor protein p63α, and cytokeratin 3 (K3). Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was used to evaluate the expression of ABCG2 and p63. Limbal explants grown in each medium were labeled with bromodeoxyuridine (BrdU) to assess the proliferative capacity in each medium. Concentration of growth factors including epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), transforming growth factor-β (TGF-β), and platelet derived growth factor (PDGF) in HPL and PHPL was compared to that in human serum (HS). Immunofluorescence staining on AM showed prominent expression of ABCG2, p63α but sparse expression of K3 in HPL and PHPL supplemented medium. Real time-PCR showed 1.7 fold higher expression of ABCG2 in PHPL supplemented medium (p = 0.03), and similar expression of p63 in HPL and PHPL supplemented medium compared to FBS medium. The proliferation assay showed that LSCs retained their proliferative potential in HPL supplemented medium. Higher concentration of growth factors were found in HPL, compared to HS. Human platelet lysate has higher concentration of grown factors and is effective in maintaining growth and stem cell phenotype of corneal limbal explant cultures.

  15. Bovine respiratory disease model based on dual infections with infection with bovine viral diarrhea virus and bovine corona virus (United States)

    Bovine respiratory disease complex (BRDC) is the leading cause of economic loss in the U.S. cattle industry. BRDC likely results from simultaneous or sequential infections with multiple pathogens including both viruses and bacteria. Bovine viral diarrhea virus (BVDV) and bovine corona virus (BoCV...

  16. Deginerative changes of femoral articular cartilage in the knee : comparative study of specimen sonography and pathology

    Energy Technology Data Exchange (ETDEWEB)

    Park, Ju Youn; Hong, Sung Hwan; Sohn, Jin Hee; Wee, Young Hoon; Chang, Jun Dong; Park, Hong Seok; Lee, Eil Seoung; Kang Ik Won [Hallym Univ. College of Medicine, Seoul (Korea, Republic of)


    To determine the sonographic findings of degenerative change in femoral articular cartilage of the knee by comparative study of specimen sonography and pathology. We obtained 40 specimens of cartilage of the femur (20 medial and 20 lateral condylar) from 20 patients with osteoarthritis of the knee who had undergone total knee replacement. The specimens were placed in a saline-filled container and sonography was performed using a 10-MHz linear transducer. Sonographic abnormalities were evaluated at the cartilage surface, within the cartilage, and at the bone-cartilage interface, and were compared with the corresponding pathologic findings. In addition, cartilage thickness was measured at a representative portion of each femoral cartilage specimen and was compared with the thickness determined by sonography. 'Dot' lesions, irregularity or loss of the hyperechoic line, were demonstrated by sonography at the saline-cartilage interface of 14 cartilages. Pathologic examination showed that these findings corresponded to cleft, detachment, erosion, and degeneration. Irregularities in the hyperechoic line at the bone-cartilage interface were revealed by sonography in eight cartilages and were related to irregularity or loss of tidemark, downward displacement of the cartilage, and subchondral callus formation. Dot lesions, corresponding to cleft and degeneration, were noted within one cartilage. Cartilage thickness measured on specimen and by sonography showed no significant difference (p=0.446). Specimen sonography suggested that articular cartilage underwent degenerative histopathological change. Cartilage thickness measured by sonography exactly reflected real thickness.

  17. Articular cartilage thickness measured with US is not as easy as it appears

    DEFF Research Database (Denmark)

    Torp-Pedersen, S; Bartels, E M; Wilhjelm, Jens E.


    Theoretically, the high spatial resolution of US makes it well suited to monitor the decrease in articular cartilage thickness in osteoarthritis. A requirement is, however, that the borders of the cartilage are correctly identified and that the cartilage is measured under orthogonal insonation....... If US measurements are compared to measurements with other techniques, they should be corrected for the higher sound speed in cartilage....

  18. Fine-tuning Cartilage Tissue Engineering by Applying Principles from Embryonic Development

    NARCIS (Netherlands)

    C.A. Hellingman (Catharine)


    textabstractCartilage has a very poor capacity for regeneration in vivo. In head and neck surgery cartilage defects are usually reconstructed with autologous cartilage from for instance the external ear or the ribs. Cartilage tissue engineering may be a promising alternative to supply tissue for

  19. Studies on Callus Induction and Regeneration of Medicinal Plant Chicory (Cichorium intybus L. from Leaf and Petiole Explants

    Directory of Open Access Journals (Sweden)

    H. Hadizadeh


    Full Text Available Introduction: Chicory (Cichorium intybus L. belongs to Asteraceae family is commonly known as witloof chicory. The leaves and the roots of this medicinal plant are edible and commonly used as salad. Some varieties are also cultivated as coffee substitute after roasting the roots. All parts of the plant contain these volatile oils, with the majority of the toxic components concentrated in the plant's root. In folk medicine, the plant is used for the treatment of diarrhea, spleen enlargement, fever, and vomiting. Antihepatotoxic activity on damaged rat’s liver sections and anti-bacterial activity of this crop has been recently reported. In vitro regeneration from leaf explants with various hormonal combinations has been reported previously. Moreover, in vitro regeneration of Chicory from cotyledon explants using different combinations of plant growth regulators has been studied. Also, a protocol for the regeneration of plantlets from leaf and petiole explants of witloof chicory has been developed. The aim of the present investigation was optimization of callus induction and shoot regeneration from leaf and petiole tissues of Chicory (Esfahan genotype. Materials and Methods: In this investigation, Esfahan genotype was used for callus induction and direct shoot regeneration. Seeds were first washed with running tap water for 30 min then seeds were surface sterilized by dipping in 70% ethanol for 90 s and rinsed with sterile distilled water, followed by immersing in 5% sodium hypochlorite solution for 25 min and thereafter rinsed for 30 min with sterile distilled water. The basal medium used in this investigation was MS. For shoot regeneration, leaf and petiole explants (5 mm segments were excised from 4-week-old sterile seedlings and cultured on MS medium containing different combinations of NAA / BA and KIN / BA in two separate experiments. Experiments were performed factorial based on completely randomized design. Cultures were incubated at 25

  20. Leptin plays a catabolic role on articular cartilage. (United States)

    Bao, Jia-peng; Chen, Wei-ping; Feng, Jie; Hu, Peng-fei; Shi, Zhong-li; Wu, Li-dong


    Leptin has been shown to play a crucial role in the regulation of body weight. There is also evidence that this adipokine plays a key role in the process of osteoarthritis. However, the precise role of leptin on articular cartilage metabolism is not clear. We investigate the role of leptin on articular cartilage in vivo in this study. Recombinant rat leptin (100 μg) was injected into the knee joints of rats, 48 h later, messenger RNA (mRNA) expression and protein levels of basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), matrix metalloproteinases 2 and 9 (MMP-2, MMP-9), cathepsin D, and collagen II from articular cartilage were analyzed by real-time quantitative polymerase chain reaction (PCR) and western blot. Two important aggrecanases ADAMTS-4 and -5 (a disintegrin and metalloproteinase with thrombospondin motifs 4 and 5) were also analyzed by real-time quantitative PCR. Besides, articular cartilage was also assessed for proteoglycan/GAG content by Safranin O staining. Leptin significantly increased both gene and protein levels of MMP-2, MMP-9, cathepsin D, and collagen II, while decreased bFGF markedly in cartilage. Moreover, the gene expression of ADAMTS-4 and -5 were markedly increased, and histologically assessed depletion of proteoglycan in articular cartilage was observed after treatment with leptin. These results strongly suggest that leptin plays a catabolic role on cartilage metabolism and may be a disadvantage factor involve in the pathological process of OA.

  1. Comparison of equine articular cartilage thickness in various joints. (United States)

    Lee, Hyeon; Kirkland, W Grant; Whitmore, Ryan N; Theis, Kelcie M; Young, Hannah E; Richardson, Ashton J; Jackson, Robert L; Hanson, R Reid


    Thicknesses of fresh equine articular cartilage surfaces from the fetlock, carpal and stifle joints were measured employing a needle probe test. Eighty-seven samples used in measurement were cultivated from fetlock, carpal and stifle joints of 12 deceased within 4 h of death. After approximately three minutes of exposure to air during dissection, all cartilage samples were preserved in a saline solution to keep the articular cartilage hydrated for testing. The thickness was measured on five different spots on the same sample. The thicknesses of the fetlock, carpus and stifle were compared. The articular cartilage of the stifle was thicker than the fetlock and carpus, while the fetlock and the carpus had similar thickness values. The average thickness of the fetlock, carpal and stifle joint are 0.86, 0.87 and 2.1 mm, respectively. They were statistically compared using the Student t-test. The differences on the articular cartilage thicknesses between the fetlock and stifle, and carpus and stifle were "very highly significant" (p fetlock and carpus. Four different surfaces in the fetlock and four in the carpal joint were also compared. Significant differences between each set of the four surfaces were not observed. In the carpus, the difference in thickness between the distal radius and proximal third carpal bone articular cartilage surfaces as well as the proximal radial carpal bone and distal radial carpal bone articular cartilage surfaces were statistically significant.

  2. The Influence of Articular Cartilage Thickness Reduction on Meniscus Biomechanics. (United States)

    Łuczkiewicz, Piotr; Daszkiewicz, Karol; Chróścielewski, Jacek; Witkowski, Wojciech; Winklewski, Pawel J


    Evaluation of the biomechanical interaction between meniscus and cartilage in medial compartment knee osteoarthritis. The finite element method was used to simulate knee joint contact mechanics. Three knee models were created on the basis of knee geometry from the Open Knee project. We reduced the thickness of medial cartilages in the intact knee model by approximately 50% to obtain a medial knee osteoarthritis (OA) model. Two variants of medial knee OA model with congruent and incongruent contact surfaces were analysed to investigate the influence of congruency. A nonlinear static analysis for one compressive load case was performed. The focus of the study was the influence of cartilage degeneration on meniscal extrusion and the values of the contact forces and contact areas. In the model with incongruent contact surfaces, we observed maximal compressive stress on the tibial plateau. In this model, the value of medial meniscus external shift was 95.3% greater, while the contact area between the tibial cartilage and medial meniscus was 50% lower than in the congruent contact surfaces model. After the non-uniform reduction of cartilage thickness, the medial meniscus carried only 48.4% of load in the medial compartment in comparison to 71.2% in the healthy knee model. We have shown that the change in articular cartilage geometry may significantly reduce the role of meniscus in load transmission and the contact area between the meniscus and cartilage. Additionally, medial knee OA may increase the risk of meniscal extrusion in the medial compartment of the knee joint.

  3. Shark cartilage, cancer and the growing threat of pseudoscience. (United States)

    Ostrander, Gary K; Cheng, Keith C; Wolf, Jeffrey C; Wolfe, Marilyn J


    The promotion of crude shark cartilage extracts as a cure for cancer has contributed to at least two significant negative outcomes: a dramatic decline in shark populations and a diversion of patients from effective cancer treatments. An alleged lack of cancer in sharks constitutes a key justification for its use. Herein, both malignant and benign neoplasms of sharks and their relatives are described, including previously unreported cases from the Registry of Tumors in Lower Animals, and two sharks with two cancers each. Additional justifications for using shark cartilage are illogical extensions of the finding of antiangiogenic and anti-invasive substances in cartilage. Scientific evidence to date supports neither the efficacy of crude cartilage extracts nor the ability of effective components to reach and eradicate cancer cells. The fact that people think shark cartilage consumption can cure cancer illustrates the serious potential impacts of pseudoscience. Although components of shark cartilage may work as a cancer retardant, crude extracts are ineffective. Efficiencies of technology (e.g., fish harvesting), the power of mass media to reach the lay public, and the susceptibility of the public to pseudoscience amplifies the negative impacts of shark cartilage use. To facilitate the use of reason as the basis of public and private decision-making, the evidence-based mechanisms of evaluation used daily by the scientific community should be added to the training of media and governmental professionals. Increased use of logical, collaborative discussion will be necessary to ensure a sustainable future for man and the biosphere.

  4. Multimodal nonlinear optical imaging of cartilage development in mouse model (United States)

    He, Sicong; Xue, Wenqian; Sun, Qiqi; Li, Xuesong; Huang, Jiandong; Qu, Jianan Y.


    Kinesin-1 is a kind of motor protein responsible for intracellular transportation and has been studied in a variety of tissues. However, its roles in cartilage development are not clear. In this study, a kinesin-1 heavy chain (Kif5b) knockout mouse model is used to study the functions of kinesin-1 in the cartilage development. We developed a multimodal nonlinear optical (NLO) microscope system integrating stimulated Raman scattering (SRS), second harmonic generation (SHG) and two-photon excited fluorescence (TPEF) to investigate the morphological and biomedical characteristics of fresh tibial cartilage from normal and mutant mice at different developmental stages. The combined forward and backward SHG imaging resolved the fine structure of collagen fibrils in the extracellular matrix of cartilage. Meanwhile, the chondrocyte morphology in different zones of cartilage was visualized by label-free SRS and TPEF images. The results show that the fibrillar collagen in the superficial zone of cartilage in postnatal day 10 and 15 (P10 and P15) knockout mice was significantly less than that of control mice. Moreover, we observed distorted morphology and disorganization of columnar arrangement of chondrocytes in the growth plate cartilage of mutant mice. This study reveals the significant roles of kinesin-1 in collagen formation and chondrocyte morphogenesis.

  5. New Technology in Imaging Cartilage of the Ankle. (United States)

    Schreiner, Markus M; Mlynarik, Vladimir; Zbýň, Štefan; Szomolanyi, Pavol; Apprich, Sebastian; Windhager, Reinhard; Trattnig, Siegfried


    The incidence of osteochondral lesions, as well as osteoarthritis of the ankle joint following osteochondritis dissecans and trauma, has been reappraised in recent years. Consequently, an increasing number of surgical interventions using different cartilage repair techniques is performed in the ankle joint, which has resulted in a growing demand for repetitive and objective assessment of cartilage tissue and its repair. While morphological imaging does enable monitoring of macroscopic changes with increasing precision, it fails to provide information about the ultrastructural composition of cartilage. The significance of molecular changes in cartilage matrix composition, however, is increasingly recognized, as it is assumed that macroscopic cartilage degeneration is preceded by a loss in glycosaminoglycans and a disorganization of the collagen network. Recent advances in biochemical magnetic resonance imaging (MRI) have yielded sequences sensitive to these changes, thus providing invaluable insight into both early cartilage degeneration and maturation of repair tissue, on a molecular level. The aim of this review was to provide a comprehensive overview of these techniques, including water and collagen-sensitive T2/T2* mapping, as well as glycosaminoglycan-sensitive sequences such as delayed gadolinium-enhanced MRI of cartilage dGEMRIC, and sodium imaging, and describe their applications for the ankle joint.

  6. Functional analysis of CTRP3/cartducin in Meckel's cartilage and developing condylar cartilage in the fetal mouse mandible (United States)

    Yokohama-Tamaki, Tamaki; Maeda, Takashi; Tanaka, Tetsuya S; Shibata, Shunichi


    CTRP3/cartducin, a novel C1q family protein, is expressed in proliferating chondrocytes in the growth plate and has an important role in regulating the growth of both chondrogenic precursors and chondrocytes in vitro. We examined the expression of CTRP3/cartducin mRNA in Meckel's cartilage and in condylar cartilage of the fetal mouse mandible. Based on in situ hybridization studies, CTRP3/cartducin mRNA was not expressed in the anlagen of Meckel's cartilage at embryonic day (E)11.5, but it was strongly expressed in Meckel's cartilage at E14.0, and then reduced in the hypertrophic chondrocytes at E16.0. CTRP3/cartducin mRNA was not expressed in the condylar anlagen at E14.0, but was expressed in the upper part of newly formed condylar cartilage at E15.0. At E16.0, CTRP3/cartducin mRNA was expressed from the polymorphic cell zone to the upper part of the hypertrophic cell zone, but was reduced in the lower part of the hypertrophic cell zone. CTRP3/cartducin-antisense oligodeoxynucleotide (AS-ODN) treatment of Meckel's cartilage and condylar anlagen from E14.0 using an organ culture system indicated that, after 4-day culture, CTRP3/cartducin abrogation induced curvature deformation of Meckel's cartilage with loss of the perichondrium and new cartilage formation. Aggrecan, type I collagen, and tenascin-C were simultaneously immunostained in this newly formed cartilage, indicating possible transformation from the perichondrium into cartilage. Further, addition of recombinant mouse CTRP3/cartducin protein to the organ culture medium with AS-ODN tended to reverse the deformation. These results suggest a novel function for CTRP3/cartducin in maintaining the perichondrium. Moreover, AS-ODN induced a deformation of the shape, loss of the perichondrium/fibrous cell zone, and disorder of the distinct architecture of zones in the mandibular condylar cartilage. Additionally, AS-ODN-treated condylar cartilage showed reduced levels of mRNA expression of aggrecan, collagen types I

  7. Induced collagen cross-links enhance cartilage integration.

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    Aristos A Athens

    Full Text Available Articular cartilage does not integrate due primarily to a scarcity of cross-links and viable cells at the interface. The objective of this study was to test the hypothesis that lysyl-oxidase, a metalloenzyme that forms collagen cross-links, would be effective in improving integration between native-to-native, as well as tissue engineered-to-native cartilage surfaces. To examine these hypotheses, engineered cartilage constructs, synthesized via the self-assembling process, as well as native cartilage, were implanted into native cartilage rings and treated with lysyl-oxidase for varying amounts of time. For both groups, lysyl-oxidase application resulted in greater apparent stiffness across the cartilage interface 2-2.2 times greater than control. The construct-to-native lysyl-oxidase group also exhibited a statistically significant increase in the apparent strength, here defined as the highest observed peak stress during tensile testing. Histology indicated a narrowing gap at the cartilage interface in lysyl-oxidase treated groups, though this alone is not sufficient to indicate annealing. However, when the morphological and mechanical data are taken together, the longer the duration of lysyl-oxidase treatment, the more integrated the interface appeared. Though further data are needed to confirm the mechanism of action, the enhancement of integration may be due to lysyl-oxidase-induced pyridinoline cross-links. This study demonstrates that lysyl-oxidase is a potent agent for enhancing integration between both native-to-native and native-to-engineered cartilages. The fact that interfacial strength increased manifold suggests that cross-linking agents should play a significant role in solving the difficult problem of cartilage integration. Future studies must examine dose, dosing regimen, and cellular responses to lysyl-oxidase to optimize its application.

  8. Effects of thidiazuron and paclobutrazol on regeneration potential of tulip flower stalk explants in vitro and subsequent shoot multiplication

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    Małgorzata Podwyszyńska


    Full Text Available The effects of TDZ and paclobutrazol on the primary regeneration on tulip flower stalk explants of six cultivars and subsequent shoot multiplication were examined. Explants, flower stalk slices, were excised from cooled and subsequently forced bulbs. The explants were incubated for two months in darkness on medium containing NAA and cytokinins, 2iP and BAP, as control, or TDZ (0.5-4 mg l-1 and paclobutrazol (0.05-0.4 mg l-1. Then, the regenerating explants were subcultured on medium with TDZ and NAA applied at low concentrations. Different regeneration capabilities were found depending on cultivar and growth regulators. The percentage of explants forming leaf-like structures ranged, on the control medium, from 80% in 'Blue Parrot' and 'Prominence' to below 30% in 'Apeldoorn' and 'Mirjoran'. TDZ, applied at optimum for each cultivar concentration, greatly increased the regeneration potential up to 70-100%. Paclobutrazol, added to the TDZ-containing medium, significantly enhanced the response of explants, resulting in high numbers of leaf-like structures formed per explant (13.7-22.8. The structures developed gradually into characteristic forms: the growing up cotyledonary leaf, the probable root primordium formed at its base, the growing downwards stolon and the shoot meristem developed finely on its tip. It is suggested that such primary regeneration may have a nature of somatic embryogenesis. Then, the adventitious shoots developed and formed clusters, which were divided into 2-3 smaller ones every two months. The growth regulators, used at initial stage, markedly influenced subsequent shoot multiplication. Thus, the most intensive shoot formation was noted with TDZ at concentrations of 0.5-2 mg l-1 and paclobutrazol of 0.05-0.1 mg l-1.

  9. Simulação da soja geneticamente modificada tolerante ao glyphosate por meio do cultivo de explantes Simulation of the transgenic soybean tolerant to glyphosate through explant cultivation

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    Sérgio C. Siqueira


    Full Text Available O objetivo do experimento consistiu na simulação in vitro da soja transgênica tolerante ao glyphosate, através do cultivo de explantes em meios de cultura contendo aminoácidos aromáticos. As avaliações basearam-se nos efeitos do glyphosate sobre sementes oriundas de explantes de soja (Glycine max (L. Merr. cv. UFV-16. Para tanto, explantes de soja foram cultivados em meios de cultura líquidos com pH em torno de 5,0. Cada explante constou de um legume completamente expandido contendo duas sementes de aproximadamente 100 mg, conectada a um segmento de caule de 45 mm de comprimento. Os tratamentos testados foram: A = glutamina (Gln; B = Gln + fenilalanina (Phe + tirosina (Tyr + triptofano (Trp; C = Gln + glyphosate; D = Gln + Phe + Tyr + Trp + glyphosate. O experimento foi conduzido sob irradiância de 80 mmol-2s-1 a 25oC por 204 horas. Nos tratamentos que receberam aminoácidos aromáticos e glutamina, o herbicida não afetou as massas fresca e seca das sementes, como também, não afetou seus constituintes bioquímicos (proteínas, óleo, ácidos graxos, carboidratos e clorofilas. Portanto, a suplementação exógena de aminoácidos aromáticos suprime os efeitos fitotóxicos do glyphosate sobre explantes de soja, permitindo estudos sobre o seu modo de ação e metabolismo nas sementes, uma vez que os explantes se comportaram analogamente à soja transgênica não suscetível ao herbicida.The objective of this experiment consisted in simulation in vitro of the transgenic soybean tolerant to glyphosate through explant cultivation in culture medium containing aromatic amino acids. The effects of glyphosate on soybean (Glycine max (L. Merrill were evaluated in seeds harvested from explants of cv UFV-16. The soybean explants were cultivated in liquid medium culture with pH about 5.0. Each explant consisted of one fruit completely expanded, containing two seeds of 100 mg approximately, and connected to a stem segment of 45 mm length. The

  10. Influência do cloreto de cálcio no crescimento de explantes de Gypsophila paniculata L. (Caryophyllaceae, cultivados in vitro Influence of calcium chloride on the growth of Gypsophila paniculata L. (Caryophyllaceae explants, cultivated in vitro

    Directory of Open Access Journals (Sweden)

    R. Jun Takane


    Full Text Available Este trabalho teve por finalidade verificar a influência do Cloreto de Cálcio (CaCl12 no crescimento de explantes de Gypsophila paniculata L., cultivados em meio de cultura a fim de fornecer subsídios para a micropropagação desta cultura. Foram utilizados como explantes somente as gemas apicais das plantas em fase de crescimento vegetativo. Os explantes foram inoculados em meio de cultura MS (Murashigue & Skoog modificado com diferentes concentrações de CaCl2: a 440; b 880; c 1.760; d 2.640 e e 3.520 mg/1. Os explantes foram deixados em câmara de crescimento sob uma temperatura constante de 25°C ± 2°C sob fotoperíodo de 16 horas. Os explantes do meio de cultura MS com 1.760 mg/1 de CaCl2 foram os de melhor resultado, com crescimento vigoroso e presença de coloração verde intensa nas folhas. Os explantes do meio de cultura MS com 440 mg/1 de CaCl2 não apresentaram um crescimento satisfatório, com estiolamento e coloração verde pouco intensa nas folhas; os explantes do meio de cultura MS com 3.520 mg/1 de CaCl2, também não apresentaram um crescimento satisfatório, pois com 10 dias de inoculação já se percebia uma menor indução no crescimento, podendo tal efeito ser conseqüência de dois fatores: excesso de cálcio no meio de cultura ou a toxidez ocasionada pelo cloro.Shoot tips from plants still in the vegetative growth phase were used as explants. They were sterilised with tetraciclin 1% for four minutes, sodium hypochloridre (commercial sanitary water with 5% of active cloride 20% (v/v for 15 minutes and alcohol 70 GL for 2 minutes. The explants were inoculated in a growth medium MS (Murashigue & Skoog, with different concentrations of CaCl2: a 440; b 880; c 1,760; d 2,640 and e 3,520 mg/1. Explants were kept in a growth chamber at constant temperature (25°C ± 2°C with a photo period of 16 hours. The explants in the medium modified with 1,760 mg/1 of CaCl2, presented the best results, with a vigorous growth and

  11. Adventitious Shoot Regeneration from Leaf Explant of Dwarf Hygro (Hygrophila polysperma (Roxb. T. Anderson

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    Mehmet Karataş


    Full Text Available Dwarf hygro (Hygrophila polysperma is an ornamental aquatic plant that changes its leaf colours to pinkish in high light. It is listed as a medicinal plant in medicinal plant lists of Indian states of West Bengal and Karnataka. It is also used as a screening tool for toxicities and a bioindicator to detect and control algae. The study reported in vitro adventitious shoot regeneration from leaf explants cultured on MS medium containing 0.10–1.60 mg/L Kin/TDZ with or without 0.10 mg/L IBA and 500 mg/L Amoklavin to eradicate endogenic bacterial contamination. Direct adventitious shoot regeneration started within one week from both culture mediums followed by late callus induction which was more prominent on TDZ containing media compared to Kin containing media. Addition of 0.10 mg/L IBA with both Kin and TDZ increased shoot regeneration frequency, mean number of shoots per explant, and mean shoot length. Maximum number of 16.33 and 20.55 shoots per explant was obtained on MS medium containing 0.80+0.10 mg/L Kin-IBA and 0.10+0.10 mg/L TDZ-IBA, respectively. Regenerated shoots were rooted on MS medium containing 0.20–1.00 mg/L IBA followed by successfull acclimatization in aquariums. Regenerated plantlets were also tested in jars containing distilled water that showed the pH 6–9 for the best plant growth and development.

  12. Glucocorticoids affect 24 h clock genes expression in human adipose tissue explant cultures.

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    Purificación Gómez-Abellán

    Full Text Available to examine firstly whether CLOCK exhibits a circadian expression in human visceral (V and subcutaneous (S adipose tissue (AT in vitro as compared with BMAL1 and PER2, and secondly to investigate the possible effect of the glucocorticoid analogue dexamethasone (DEX on positive and negative clock genes expression.VAT and SAT biopsies were obtained from morbid obese women (body mass index ≥ 40 kg/m(2 (n = 6. In order to investigate rhythmic expression pattern of clock genes and the effect of DEX on CLOCK, PER2 and BMAL1 expression, control AT (without DEX and AT explants treated with DEX (2 hours were cultured during 24 h and gene expression was analyzed at the following times: 10:00 h, 14:00 h, 18:00 h, 22:00 h, 02:00 h and 06:00 h, using qRT-PCR.CLOCK, BMAL1 and PER2 expression exhibited circadian patterns in both VAT and SAT explants that were adjusted to a typical 24 h sinusoidal curve. PER2 expression (negative element was in antiphase with respect to CLOCK and in phase with BMAL1 expression (both positive elements in the SAT (situation not present in VAT. A marked effect of DEX exposure on both positive and negative clock genes expression patterns was observed. Indeed, DEX treatment modified the rhythmicity pattern towards altered patterns with a period lower than 24 hours in all genes and in both tissues.24 h patterns in CLOCK and BMAL1 (positive clock elements and PER2 (negative element mRNA levels were observed in human adipose explants. These patterns were altered by dexamethasone exposure.

  13. Propagation of goldenrod (Solidago canadensis L. from leaf and nodal explants

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    Jun Li


    Full Text Available Goldenrod (Solidago canadensis L. is an invasive plant species in many countries except North America but a cut-flower species worldwide. There is a need to generate and propagate goldenrod clones efficiently for research and commercial purposes. A callus induction and plantlet regeneration system was developed by studying the influence of explant type and different concentrations of plant growth regulators. The highest callus production from leaf segments was obtained on Murashige and Skoog’s medium (MS medium supplemented with 1.0 mg/L naphthalene acetic acid (NAA and 1.0 mg/L 6-benzylaminopurine (BA. Adventitious shoots could be regenerated directly from leaf explants without an intermediate callus phase with the highest shoot induction percentage of 87.2%. The largest number of adventitious shoots per leaf explant (3.2 was obtained on MS medium supplemented with 0.4 mg/L NAA and 2.0 mg/L BA. MS medium supplemented with 0.1 mg/L NAA and 1.0 mg/L BA was the best medium for axillary shoot regeneration from nodal segments. The highest root number and longest roots occurred on half-strength MS without the addition of any growth regulator. Rooted plantlets were then transferred to a soil-based growth medium, placed in a greenhouse, and acclimatized with 100% success. All surviving plants grew normally without showing any morphological varia­tion when compared to those grow from seed. This regeneration protocol may be used to produce certain biotypes of goldenrod suitable for genetic transformation rapid propagation of goldenrod for commercial purposes or for screening fungi and toxins as potential biocontrol agents against this weed.

  14. Agrobacterium-mediated transformation of oat (Avena sativa L.) cultivars via immature embryo and leaf explants. (United States)

    Gasparis, Sebastian; Bregier, Cezary; Orczyk, Waclaw; Nadolska-Orczyk, Anna


    This paper reports on the successful Agrobacterium-mediated transformation of oat, and on some factors influencing this process. In the first step of the experiments, three cultivars, two types of explant, and three combinations of strain/vectors, which were successfully used for transformation of other cereals were tested. Transgenic plants were obtained from the immature embryos of cvs. Bajka, Slawko and Akt and from leaf base explants of cv. Bajka after transformation with A. thumefaciens strain LBA4404(pTOK233). The highest transformation rate (12.3%) was obtained for immature embryos of cv. Bajka. About 79% of the selected plants proved to be transgenic; however, only 14.3% of the T(0) plants and 27.5% of the T(1) showed GUS expression. Cell competence of both types of explant differed in terms of their transformation ability and transgene expression. The next step of the study was to test the suitability for oat transformation of the pGreen binary vector combined with different selection cassettes: nptII or bar under the nos or 35S promoter. Transgenic plants were selected in combinations transformed with nos::nptII, 35S::nptII and nos::bar. The highest transformation efficiency (5.3%) was obtained for cv. Akt transformed with nos::nptII. A detailed analysis of the T(0) plants selected from a given callus line and their progeny revealed that they were the mixture of transgenic, chimeric-transgenic and non-transgenic individuals. Southern blot analysis of T(0) and T(1) showed simple integration pattern with the low copy number of the introduced transgenes.

  15. Prospective Clinical Trial for Septic Arthritis: Cartilage Degradation and Inflammation Are Associated with Upregulation of Cartilage Metabolites

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    Hagen Schmal


    Full Text Available Background. Intra-articular infections can rapidly lead to osteoarthritic degradation. The aim of this clinical biomarker analysis was to investigate the influence of inflammation on cartilage destruction and metabolism. Methods. Patients with acute joint infections were enrolled in a prospective clinical trial and the cytokine composition of effusions (n=76 was analyzed. Characteristics of epidemiology and disease severity were correlated with levels of cytokines with known roles in cartilage turnover and degradation. Results. Higher synovial IL-1β concentrations were associated with clinical parameters indicating a higher disease severity (p<0.03 excluding the incidence of sepsis. Additionally, intra-articular IL-1β levels correlated with inflammatory serum parameters as leucocyte counts (LC and C-reactive protein concentrations (p<0.05 but not with age or comorbidity. Both higher LC and synovial IL-1β levels were associated with increased intra-articular collagen type II cleavage products (C2C indicating cartilage degradation. Joints with preinfectious lesions had higher C2C levels. Intra-articular inflammation led to increased concentrations of typical cartilage metabolites as bFGF, BMP-2, and BMP-7. Infections with Staphylococcus species induced higher IL-1β expression but less cartilage destruction than other bacteria. Conclusion. Articular infections have bacteria-specific implications on cartilage metabolism. Collagen type II cleavage products reliably mark destruction, which is associated with upregulation of typical cartilage turnover cytokines. This trial is registered with DRKS00003536, MISSinG.

  16. Prospective Clinical Trial for Septic Arthritis: Cartilage Degradation and Inflammation Are Associated with Upregulation of Cartilage Metabolites. (United States)

    Schmal, Hagen; Bernstein, Anke; Feucht, Matthias J; Erdle, Benjamin; Pestka, Jan M; Pham, That Minh; Kubosch, Eva Johanna

    Background. Intra-articular infections can rapidly lead to osteoarthritic degradation. The aim of this clinical biomarker analysis was to investigate the influence of inflammation on cartilage destruction and metabolism. Methods. Patients with acute joint infections were enrolled in a prospective clinical trial and the cytokine composition of effusions (n = 76) was analyzed. Characteristics of epidemiology and disease severity were correlated with levels of cytokines with known roles in cartilage turnover and degradation. Results. Higher synovial IL-1β concentrations were associated with clinical parameters indicating a higher disease severity (p < 0.03) excluding the incidence of sepsis. Additionally, intra-articular IL-1β levels correlated with inflammatory serum parameters as leucocyte counts (LC) and C-reactive protein concentrations (p < 0.05) but not with age or comorbidity. Both higher LC and synovial IL-1β levels were associated with increased intra-articular collagen type II cleavage products (C2C) indicating cartilage degradation. Joints with preinfectious lesions had higher C2C levels. Intra-articular inflammation led to increased concentrations of typical cartilage metabolites as bFGF, BMP-2, and BMP-7. Infections with Staphylococcus species induced higher IL-1β expression but less cartilage destruction than other bacteria. Conclusion. Articular infections have bacteria-specific implications on cartilage metabolism. Collagen type II cleavage products reliably mark destruction, which is associated with upregulation of typical cartilage turnover cytokines. This trial is registered with DRKS00003536, MISSinG.

  17. Stable genetic transformation of Jatropha curcas via Agrobacterium tumefaciens-mediated gene transfer using leaf explants

    KAUST Repository

    Kumar, Nitish


    Jatropha curcas is an oil bearing species with multiple uses and considerable economic potential as a biofuel crop. A simple and reproducible protocol was developed for Agrobacterium tumefaciens-mediated stable genetic transformation of J. curcas using leaf explains. Agrobacterium strain LBA 4404 harbouring the binary vector pCAMBIA 1304 having sense-dehydration responsive element binding (S-DREB2A), beta-glucuronidase (gus), and hygromycin-phosphotransferase (hpt) genes were used for gene transfer. A number of parameters such as preculture of explains, wounding of leaf explants, Agrobacterium growth phase (OD), infection duration, co-cultivation period, co-cultivation medium pH, and acetosyringone, were studied to optimized transformation efficiency. The highest transformation efficiency was achieved using 4-day precultured, non-wounded leaf explants infected with Agrobacterium culture corresponding to OD(600)=0.6 for 20 min, followed by co-cultivation for 4 days in a co-cultivation medium containing 100 mu M acetosyringone, pH 5.7. Co-cultivated leaf explants were initially cultured on Murashige and Skoog (MS) medium supplemented with 2.27 mu M thidiazuron (TDZ) for regeneration of shoot buds, followed by selection on same medium with 5 mu g ml(-1) hygromycin. Selected shoot buds were transferred to MS medium containing 10 mu M kinetin (Kn), 4.5 mu M 6-benzyl aminopurine (BA), and 5.5 mu M alpha-naphthaleneacetic acid (NAA) for proliferation. The proliferated shoots were elongated on MS medium supplemented with 2.25 mu M BA and 8.5 mu M indole-3-acetic acid (IAA). The elongated shoots were rooted on half strength MS medium supplemented with 15 mu M indole-3-butyric acid (IBA), 5.7 mu M IAA, 5.5 mu M NAA, and 0.25 mg l(-1) activated charcoal. GUS histochemical analysis of the transgenic tissues further confirmed the transformation event. PCR and DNA gel blot hybridization were performed to confirm the presence of transgene. A transformation efficiency of 29% was

  18. Cultivar-Dependent Direct Organogenesis of Date Palm from Shoot Tip Explants. (United States)

    Abahmane, Larbi


    A number of public and private laboratories are working on date palm micropropagation to meet the increasing worldwide demand for date palm planting material. A standardized direct organogenesis protocol exists for the production of date palm plantlets to maintain the genetic fidelity of regenerated plants. Organogenesis has the advantage of using low concentrations of plant growth regulators and avoiding the callus phase. In addition, direct regeneration of vegetative buds minimizes the risk of somaclonal variation among plant regenerants. However, in vitro multiplication cycles should be limited in duration by frequent renewal of plant material. This chapter describes a simple and routine organogenesis protocol for date palm multiplication using shoot tip explants.

  19. Mercury-induced ethylene formation and abscission in Citrus and Coleus explants

    Energy Technology Data Exchange (ETDEWEB)

    Goren, R.; Siegel, S.M.


    Mercury vapor induces ethylene formation and abscission in Citrus and Coleus explants. Both responses are markedly greater in the absence of CO/sub 2/. The stimulation of these metabolically complex processes indicates that the action of mercury vapor is not consistent with the more popular conception of mercury toxicity. This was manifested in its complete failure to disturb respiratory gas exchange, and in the total absence of any necrosis. Accordingly, the effect of mercury appears to be highly specific. The overall significance of these findings is discussed with respect to physiological, environmental, and methodological aspects.

  20. Influence de la taille des vitroplants et du type d'explant sur la ...

    African Journals Online (AJOL)

    L'aptitude à la callogenèse chez le cotonnier a été estimée par le taux d'induction et le poids sec des cals en fonction de la source de l'explant et de la taille des plantules. Le taux d'induction des cals augmente avec la taille des plantules alors que le poids sec des cals ; résultat de la croissance des cals, évolue en sens ...

  1. Hyperosmolarity normalises serum-induced changes to chondrocyte properties in a model of cartilage injury. (United States)

    Karim, A; Hall, A C


    Partial-thickness cartilage injuries do not heal effectively, potentially leading to degeneration as occurs in post-traumatic osteoarthritis (PTOA). The role of chondrocytes could be crucial in determining the nature of the repair; however, their response to this injury is poorly understood. We have utilised an in vitro bovine osteochondral partial-thickness scalpel injury model and determined chondrocyte properties at and distant from the injury in the presence/absence of (a) serum-free DMEM (340 mOsm), (b) synovial fluid DMEM (SF-DMEM), (c) foetal calf serum DMEM (FCS-DMEM), (d) hyperosmolar serum-free DMEM (600 mOsm), or (e) hyperosmolar FCS-DMEM for up to two weeks. Chondrocytes were fluorescently-labelled with 5-chloromethylfluorescein-diacetate (CMFDA)/propidium iodide (PI) for live/dead cells and imaged using confocal microscopy. Quantitative data were obtained on chondrocyte properties (cell volume, clusters, morphology) at and distant from the injury. In serum-free DMEM, chondrocyte morphology at the injury remained unaffected throughout culture. However, with SF-DMEM or FCS-DMEM the chondrocytes displayed an increase in volume (p serum-free DMEM. Cluster formation and shape changes during FCS-DMEM culture were more pronounced than with SF-DMEM. SF-DMEM or FCS-DMEM stimulated these changes to chondrocytes at the injury with only small effects on distant cells. Hyperosmolarity inhibited the morphological and volume changes to chondrocytes induced by FCS-DMEM (p serum-free DMEM. Raised osmolarity may therefore have benefit in preserving the morphological phenotype of chondrocytes at the site of injury, and thus promote more effective integrative repair in partial-thickness cartilage injury.

  2. Discovery of highly potent and selective small molecule ADAMTS-5 inhibitors that inhibit human cartilage degradation via encoded library technology (ELT). (United States)

    Deng, Hongfeng; O'Keefe, Heather; Davie, Christopher P; Lind, Kenneth E; Acharya, Raksha A; Franklin, G Joseph; Larkin, Jonathan; Matico, Rosalie; Neeb, Michael; Thompson, Monique M; Lohr, Thomas; Gross, Jeffrey W; Centrella, Paolo A; O'Donovan, Gary K; Bedard, Katie L Sargent; van Vloten, Kurt; Mataruse, Sibongile; Skinner, Steven R; Belyanskaya, Svetlana L; Carpenter, Tiffany Y; Shearer, Todd W; Clark, Matthew A; Cuozzo, John W; Arico-Muendel, Christopher C; Morgan, Barry A


    The metalloprotease ADAMTS-5 is considered a potential target for the treatment of osteoarthritis. To identify selective inhibitors of ADAMTS-5, we employed encoded library technology (ELT), which enables affinity selection of small molecule binders from complex mixtures by DNA tagging. Selection of ADAMTS-5 against a four-billion member ELT library led to a novel inhibitor scaffold not containing a classical zinc-binding functionality. One exemplar, (R)-N-((1-(4-(but-3-en-1-ylamino)-6-(((2-(thiophen-2-yl)thiazol-4-yl)methyl)amino)-1,3,5-triazin-2-yl)pyrrolidin-2-yl)methyl)-4-propylbenzenesulfonamide (8), inhibited ADAMTS-5 with IC(50) = 30 nM, showing >50-fold selectivity against ADAMTS-4 and >1000-fold selectivity against ADAMTS-1, ADAMTS-13, MMP-13, and TACE. Extensive SAR studies showed that potency and physicochemical properties of the scaffold could be further improved. Furthermore, in a human osteoarthritis cartilage explant study, compounds 8 and 15f inhibited aggrecanase-mediated (374)ARGS neoepitope release from aggrecan and glycosaminoglycan in response to IL-1β/OSM stimulation. This study provides the first small molecule evidence for the critical role of ADAMTS-5 in human cartilage degradation.

  3. Chondrosarcoma of the Laryngeal Thyroid Cartilage. (United States)

    Righi, Stefano; Boffano, Paolo; Pateras, Dimitrios; Chiodo, Domenico; Zanardi, Fabio; Patetta, Roberta


    Chondrosarcoma of the larynx is a rare tumor, accounting for 0.07% to 2% of all laryngeal cancers. Nevertheless, it represents the most frequent nonepithelial neoplasm of the laryngeal region.Laryngeal chondrosarcomas are usually characterized by slow growth and low metastatic potential.The exact etiopathogenesis of chondrosarcoma is still debated. Diagnosis can be difficult given the slow rate of growth and nonspecific patient symptoms at presentation.Because of its rarity, literature about laryngeal chondrosarcoma is mainly made up of case reports and small case series.Therefore, it is important to add any information regarding this pathology.The aim of this article was to present and discuss a new case of chondrosarcoma located in the thyroid cartilage in a 63-year-old woman.

  4. Organotypic Culture of Breast Tumor Explants as a Multicellular System for the Screening of Natural Compounds with Antineoplastic Potential

    Directory of Open Access Journals (Sweden)

    Irma Edith Carranza-Torres


    Full Text Available Breast cancer is the leading cause of death in women worldwide. The search for novel compounds with antitumor activity, with less adverse effects and higher efficacy, and the development of methods to evaluate their toxicity is an area of ​​intense research. In this study we implemented the preparation and culture of breast tumor explants, which were obtained from precision-cut breast tumor slices. In order to validate the model we are proposing to screen antineoplastic effect of natural compounds, we selected caffeic acid, ursolic acid, and rosmarinic acid. Using the Krumdieck tissue slicer, precision-cut tissue slices were prepared from breast cancer samples; from these slices, 4 mm explants were obtained and incubated with the selected compounds. Viability was assessed by Alamar Blue assay, LDH release, and histopathological criteria. Results showed that the viability of the explants cultured in the presence of paclitaxel (positive control decreased significantly (P<0.05; however, tumor samples responded differently to each compound. When the explants were coincubated with paclitaxel and compounds, a synergic effect was observed. This study shows that ex vivo culture of breast cancer explants offers a suitable alternative model for evaluating natural or synthetic compounds with antitumor properties within the complex microenvironment of the tumor.

  5. Diprosopia em bovino Bovine diprosopus

    Directory of Open Access Journals (Sweden)

    I.T. Rotta


    Full Text Available This work describes a malformation in one newborn female bovine, with two faces and two skull fused, showing one single head. Duplications of the nasal and oral structures, tetraofthalmy, two brains, one single cerebellum, and pons were observed. The right thyroid was hypertrophic and the other organs had normal morphology. Every change observed in this case was compatibles with diprosopus.

  6. Bovine spongiform encephalopathy in sheep?

    NARCIS (Netherlands)

    Schreuder, B.E.C.; Somerville, R.A.


    Bovine spongiform encephalopathy (BSE) in sheep has not been identified under natural conditions at the time of writing and remains a hypothetical issue. However, rumours about the possible finding of a BSE-like isolate in sheep have led to great unrest within the sheep industry, among the general

  7. Infectious bovine rhinotracheitis in Scotland. (United States)


    A cattle dashboard has recently been developed to share surveillance information gathered from submissions to the Great Britain veterinary diagnostic network. Data relating to Scotland come from the SAC C VS. This article, by Tim Geraghty, relates to cases of infectious bovine rhinotracheitis in Scotland, as summarised on the APHA Cattle Dashboard. British Veterinary Association.

  8. New Techniques for Cartilage Magnetic Resonance Imaging Relaxation Time Analysis: Texture Analysis of Flattened Cartilage and Localized Intra- and Inter-subject Comparisons


    Carballido-Gamio, Julio; Link, Thomas M.; Majumdar, Sharmila


    MR relaxation time measurements of knee cartilage have shown potential to characterize knee osteoarthritis (OA). In this work, techniques that allow localized intra- and inter-subject comparisons of cartilage relaxation times, as well as cartilage flattening for texture analysis parallel and perpendicular to the natural cartilage layers, are presented. The localized comparisons are based on the registration of bone structures and the assignment of relaxation time feature vectors to each point...

  9. Identification of lactoferrin in bovine tears. (United States)

    Brown, M H; Brightman, A H; Fenwick, B W; Rider, M A


    To determine whether bovine tear film contains the iron-binding glycoprotein, lactoferrin. 40 Adult Hereford, Angus, and Simmental cattle. Protein analysis: pooled bovine tears were used for protein analysis (size exclusion high-performance liquid chromatography [HPLC] fractionation). HPLC was used for tear analysis. A diode array detector was used (215 and 280 microns) for chromatogram analysis and comparisons. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE): protein electrophoresis was performed, using 7.5% running gels with 4% stacking gels. Molecular weight of proteins in the unknown samples was determined as recommended by the manufacturer of the standards. Protein sequencing: amino acid sequencing, using automated Edman degradation of HPLC purified protein, was performed. The sequence obtained was compared with the known protein sequence of bovine lactoferrin. HPLC analysis of whole bovine tears resulted in a consistent chromatogram. Peak collection was performed to recover a protein from the bovine tear film with chromatogram characteristics nearly identical to purified bovine lactoferrin. Silver-stained SDS-PAGE of this peak revealed a band with molecular mass consistent with bovine lactoferrin (estimated mass of 78 kd). The first 13 amino acid residues of this protein were identical to the amino acid sequence of bovine lactoferrin. Analysis of whole bovine tears, using size exclusion HPLC, SDS-PAGE, and amino acid sequencing, provided evidence that bovine tears contain lactoferrin. Lactoferrin probably exerts a bacteriostatic effect in bovine tear film. Locally produced lactoferrin may bathe the ocular surface and sequester iron from potential pathogens.

  10. Age-associated glycopeptide pigment in human costal cartilage. (United States)

    van der Korst, J K; Willekens, F L; Lansink, A G; Henrichs, A M


    Age-associated pigmentation of human costal cartilage is caused by the accumulation of a brown water-soluble substance which can be only be extracted after proteolytic disruption of the cartilage. After isolation by gel filtration and ion exchange chromatography, the compound was identified as an acid glycopeptide. In contrast to ochronotic pigment and an artificial pigment derived by oxidation of homogentistic acid in alkaline solution, the age-associated cartilage pigment was strongly fluorescent and did not form insoluble complexes with cetylpyridinium chloride. Moreover, age-associated cartilage pigment is alkali resistant, in contrast to the ochronotic pigment. The pigment differs from lipofuscin in being strongly hydrophilic and having no affinity for fat stains. The unidentified chromophore could not be separated from the glycopeptide molecule.

  11. Costal cartilage fractures and disruptions in a rugby football player. (United States)

    Lopez, Victor; Ma, Richard; Li, Xinning; Steele, John; Allen, Answorth A


    Costal cartilage fracture of the rib cage, or costochondral, is a rare sporting injury. For contact athletes, the instability of the rib cage may lead to potential serious complications, similar to rib fractures or thorax disruption. Most authors recommend initial conservative treatment with surgery reserved for only recalcitrant cases. We report a case of an amateur American male rugby football player who sustained a costal cartilage fracture and disruption involving the anterior left fifth and sixth rib costal cartilages. The case highlights the difficulty in establishing the diagnosis based on clinical examination and standard radiographs alone. Computed tomography was used to assist in diagnosing this destabilizing injury to the rib cage. Costal cartilage fractures and disruptions in athletes are rarely reported in the literature and can have serious implications for the athlete's ability to return to play if the rib cage is destabilized.

  12. Does vitamin D affect femoral cartilage thickness? An ultrasonographic study. (United States)

    Malas, Fevziye Unsal; Kara, Murat; Aktekin, Lale; Ersöz, Murat; Ozçakar, Levent


    This study aims to investigate the association between vitamin D levels and distal femoral cartilage thickness in healthy subjects. Eighty patients who were admitted to our outpatient clinic between May and July 2013 were classified into three subgroups according to their 25-OH vitamin D levels of vitamin D deficiency (vitamin D levels and US measurements in the severe vitamin D deficiency group at RLC (r = 444, p = 0.020), LMC (r = 357, p = 0.067), and LLC (r = 568, p = 0.002). Low levels of vitamin D seem to affect the femoral cartilage thickness, adversely. Further studies are necessary to ascertain the clinical relevance of this change in cartilage thickness and whether vitamin D supplementation can reverse the cartilage thinning process or the allied clinical symptoms in the course of knee osteoarthritis.

  13. Starch-modified magnetite nanoparticles for impregnation into cartilage (United States)

    Soshnikova, Yulia M.; Roman, Svetlana G.; Chebotareva, Natalia A.; Baum, Olga I.; Obrezkova, Mariya V.; Gillis, Richard B.; Harding, Stephen E.; Sobol, Emil N.; Lunin, Valeriy V.


    The paper presents preparation and characterization of starch-modified Fe3O4 nanoparticles (NPs) in aqueous dispersion after impregnation into healthy and damaged types of cartilage. We show that starch-modified dispersion has a narrower size distribution than a non-stabilized one. The average hydrodynamic radius of magnetite NPs in a dispersion used for impregnation into cartilage is (48 ± 1) nm with the width of the distribution from 5 to 200 nm. We investigate stability of aqueous magnetite NPs dispersions during storage and with increase in temperature (up to 70 °C). We find that polydisperse magnetite NPs can penetrate into cartilage and the size and concentration of impregnated particles depend on the organization of the tissue structure. The results confirm the possibility of application of magnetite NPs in diagnostics and laser treatment of degenerative cartilage deceases.

  14. Evaluation of early changes of cartilage biomarkers following ...

    African Journals Online (AJOL)

    Evaluation of early changes of cartilage biomarkers following arthroscopic meniscectomy in young Egyptian adults. HK Koryem, MA El Qawy Wanas, MMA Rizk, HT Kotb, AH Naguib, MMA Hamid El Shafei, HMA El Naby ...

  15. Tailored PVA/ECM Scaffolds for Cartilage Regeneration

    Directory of Open Access Journals (Sweden)

    Elena Stocco


    Full Text Available Articular cartilage lesions are a particular challenge for regenerative medicine due to cartilage low self-ability repair in case of damage. Hence, a significant goal of musculoskeletal tissue engineering is the development of suitable structures in virtue of their matrix composition and biomechanical properties. The objective of our study was to design in vitro a supporting structure for autologous chondrocyte growth. We realized a biohybrid composite scaffold combining a novel and nonspecific extracellular matrix (ECM, which is decellularized Wharton’s jelly ECM, with the biomechanical properties of the synthetic hydrogel polyvinyl alcohol (PVA. Wharton’s jelly ECM was tested for its ability in promoting scaffold colonization by chondrocytes and compared with polyvinyl alcohol itself and the more specific decellularized cartilage matrix. Our preliminary evidences highlighted the chance of using Wharton’s jelly ECM in combination with PVA hydrogels as an innovative and easily available scaffold for cartilage restoration.

  16. Namaste (counterbalancing technique: Overcoming warping in costal cartilage

    Directory of Open Access Journals (Sweden)

    Kapil S Agrawal


    Full Text Available Background: Indian noses are broader and lack projection as compared to other populations, hence very often need augmentation, that too by large volume. Costal cartilage remains the material of choice in large volume augmentations and repair of complex primary and secondary nasal deformities. One major disadvantage of costal cartilage grafts (CCG which offsets all other advantages is the tendency to warp and become distorted over a period of time. We propose a simple technique to overcome this menace of warping. Materials and Methods: We present the data of 51 patients of rhinoplasty done using CCG with counterbalancing technique over a period of 4 years. Results: No evidence of warping was found in any patient up to a maximum follow-up period of 4 years. Conclusion: Counterbalancing is a useful technique to overcome the problem of warping. It gives liberty to utilize even unbalanced cartilage safely to provide desired shape and use the cartilage without any wastage.

  17. Cartilage oligomeric matrix protein in patients with juvenile idiopathic arthritis

    DEFF Research Database (Denmark)

    Bjørnhart, Birgitte; Juul, Anders; Nielsen, Susan


    Cartilage oligomeric matrix protein (COMP) has been identified as a prognostic marker of progressive joint destruction in rheumatoid arthritis. In this population based study we evaluated associations between plasma concentrations of COMP, disease activity, and growth velocity in patients...

  18. Articular cartilage repair and the evolving role of regenerative medicine

    Directory of Open Access Journals (Sweden)

    Pieter K Bos


    Full Text Available Pieter K Bos1, Marloes L van Melle1, Gerjo JVM van Osch1,21Department of Orthopaedic Surgery, Erasmus MC, Rotterdam, the Netherlands; 2Department of Otorhinolaryngology, Erasmus MC, Rotterdam, the NetherlandsAbstract: Among the growing applications of regenerative medicine, clinical articular cartilage repair has now been used for 2 decades and forms a successful example of translational medicine. Cartilage is characterized by a limited intrinsic repair capacity following injury. Articular cartilage defects cause symptoms, are not spontaneously repaired, and are generally believed to result in early osteoarthritis. Marrow stimulation techniques, osteochondral transplantation, and cell-based therapies, such as autologous chondrocyte implantation (ACI and use of mesenchymal stem cells (MSCs, are used for tissue regeneration, symptom relief, and prevention of further joint degeneration. The exact incidence of cartilage defects and the natural outcome of joints with these lesions are unclear. Currently available cartilage repair techniques are designed for defect treatment in otherwise healthy joints and limbs, mostly in young adults. The natural history studies presented in this review estimated that the prevalence of cartilage lesions in this patient group ranges from 5% to 11%. The background and results from currently available randomized clinical trials of the three mostly used cartilage repair techniques are outlined in this review. Osteochondral transplantation, marrow stimulation, and ACI show improvement of symptoms with an advantage for cell-based techniques, but only a suggestion that risk for joint degeneration can be reduced. MSCs, characterized by their good proliferative capacity and the potential to differentiate into different mesenchymal lineages, form an attractive alternative cell source for cartilage regeneration. Moreover, MSCs provide a regenerative microenvironment by the secretion of bioactive factors. This trophic activity

  19. Post-traumatic glenohumeral cartilage lesions: a systematic review

    Directory of Open Access Journals (Sweden)

    Stussi Edgar


    Full Text Available Abstract Background Any cartilage damage to the glenohumeral joint should be avoided, as these damages may result in osteoarthritis of the shoulder. To understand the pathomechanism leading to shoulder cartilage damage, we conducted a systematic review on the subject of articular cartilage lesions caused by traumas where non impression fracture of the subchondral bone is present. Methods PubMed (MEDLINE, ScienceDirect (EMBASE, BIOBASE, BIOSIS Previews and the COCHRANE database of systematic reviews were systematically scanned using a defined search strategy to identify relevant articles in this field of research. First selection was done based on abstracts according to specific criteria, where the methodological quality in selected full text articles was assessed by two reviewers. Agreement between raters was investigated using percentage agreement and Cohen's Kappa statistic. The traumatic events were divided into two categories: 1 acute trauma which refers to any single impact situation which directly damages the articular cartilage, and 2 chronic trauma which means cartilage lesions due to overuse or disuse of the shoulder joint. Results The agreement on data quality between the two reviewers was 93% with a Kappa value of 0.79 indicating an agreement considered to be 'substantial'. It was found that acute trauma on the shoulder causes humeral articular cartilage to disrupt from the underlying bone. The pathomechanism is said to be due to compression or shearing, which can be caused by a sudden subluxation or dislocation. However, such impact lesions are rarely reported. In the case of chronic trauma glenohumeral cartilage degeneration is a result of overuse and is associated to other shoulder joint pathologies. In these latter cases it is the rotator cuff which is injured first. This can result in instability and consequent impingement which may progress to glenohumeral cartilage damage. Conclusion The great majority of glenohumeral cartilage

  20. Tissue Engineering Based Therapy for Articular Cartilage Defects - A New Approach

    Directory of Open Access Journals (Sweden)

    Abraham S


    Full Text Available Background: Articular cartilage, the load-bearing tissue in diarthrodial joints, when damaged due to trauma could lead to osteoarthritis. At present Autologous Cartilage Implantation is an established method in which patients own chondrocytes are isolated and then implanted after in vitro expansion over the affected area with bovine or porcine collagen matrix. This procedure results in more of Collagen Type I during in vitro expansion, which eventually becomes fibrocartilage. Also it requires growth factors. We have in this study tried growing human Chondrocytes without growth factors using synthetic scaffolds to grow more Collagen Type II Materials and Methods: Human cartilage specimens were harvested through arthroscopy from the non-weight bearing area of the knee joint from 13 patients who underwent surgical procedures of the knee joint after getting their informed consent. The tissues were transported in saline taking 1 hour to laboratory and subjected to digestion with Collagenase type II for 16~18 Hrs. The chondrocyte cells obtained after dissociation were divided into two groups for culture. Gr. I were embedded in a Thermogelation polymer (TGP and Gr. II in basal culture media (DMEM + Ascorbic Acid without using any growth factors. The Group II cells were viable only for 4 weeks and then started degenerating. The TGP-Chondrocytes scaffolds were grown for 16 weeks and the specimens were harvested at 4, 8, 12 and 16-week intervals and their morphology and molecular characteristics were studied by H&E staining, S-100 protein analysis and RT-PCR.Results: Human chondrocytes could be cultured in both TGP (group I and Basal culture media (group II. The Gr. I cells were viable upto the 16th week while the Group II chondrocytes started degenerating after the 4 week. Both the groups were proven positive for S-100 protein, a Chondrocyte specific marker protein; Gr. II specimens after 4 weeks, and Gr. I specimens after 4, 8, 12 and 16 weeks. RT