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Sample records for cartilage explants cultures

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

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    E.K Moo

    2011-01-01

    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.

  2. Andrographolide Exerts Chondroprotective Activity in Equine Cartilage Explant and Suppresses Interleukin-1β-Induced MMP-2 Expression in Equine Chondrocyte Culture

    OpenAIRE

    Tangyuenyong, Siriwan; Viriyakhasem, Nawarat; Peansukmanee, Siriporn; Kongtawelert, Prachya; Ongchai, Siriwan

    2014-01-01

    Cartilage erosion in degenerative joint diseases leads to lameness in affected horses. It has been reported that andrographolide from Andrographis paniculata inhibited cartilage matrix-degrading enzymes. This study aimed to explore whether this compound protects equine cartilage degradation in the explant culture model and to determine its effect on matrix metalloproteinase-2 (MMP-2) expression, a matrix-degrading enzyme, in equine chondrocyte culture. Equine articular cartilage explant cultu...

  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; Karsdal, Morten A; Madsen, Suzi H; Sondergaard, Bodil-Cecilie; Zheng, Qinlong; Qvist, Per

    2009-01-01

    . 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......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...... cultured in the presence or absence of the catabolic cytokines oncostatin M (OSM) and tumor necrosis factor alpha (TNFalpha). 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...

  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

    2009-12-01

    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. Explant cultures of human colon

    DEFF Research Database (Denmark)

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

    1978-01-01

    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...

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

    International Nuclear Information System (INIS)

    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

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

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    Chenais, Nathalie; Lareyre, Jean-Jacques; Le Bail, Pierre-Yves; Labbe, Catherine, E-mail: catherine.labbe@rennes.inra.fr

    2015-07-01

    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.

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

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    Abeynaike Latasha

    2010-08-01

    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. Explant exenisation for tissue culture in marine macroalgae

    Science.gov (United States)

    Liu, Xuewu; Kloareg, Bernard

    1992-09-01

    Unialgal explants from Laminaria digitata, and from a variety of red algae, were obtained by hand removing the visible epiphytes, and stirring the tissue in the presence of glass beads. Two antibiotic mixtures were found to be efficient in removing the contaminating fungi and bacteria from the algae. The procedure proved suitable as a primary step in the tissue culture of the investigated species.

  10. In vitro Flowering of Shoots Regenerated from Cultured Nodal Explants

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    Kantamaht KANCHANAPOOM

    2011-05-01

    Full Text Available A protocol for the regeneration of Gypsophila paniculata L. using nodal explants from 2-month-old field grown plants was established. The induction of multiple shoots was best obtained on Murashige and Skoog (MS medium supplemented with 13.3 μM BA. Callus growth was observed on MS medium containing 44.3 μM BA. Calluses were transferred to MS medium supplemented with 2, 4-D (4.5, 13.5, 22.6 μM, NAA (5.3, 16.1, 26.8 μM or BA (4.4, 13.3, 22.1 μM for 2 months to induce shoot formation. After 6 weeks of initial culture, multiple shoots were regenerated from calluses cultured on MS medium supplemented with 13.3 μM BA. All regenerated shoots produced roots on 16.1 μM NAA containing MS medium within 4 weeks. Rooted plantlets were hardened and established in pots at 100% survival. For induction of in vitro flowering, regenerated shoots could be induced to flower efficiently when cultured on MS medium containing 13.3 μM BA and 50 g/l sucrose.

  11. The Slice Culture Method for Following Development of Tooth Germs In Explant Culture

    OpenAIRE

    Alfaqeeh, Sarah A; Tucker, Abigail S.

    2013-01-01

    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.

  12. Molecular Profiling of Multiplexed Gene Markers to Assess Viability of Ex Vivo Human Colon Explant Cultures

    OpenAIRE

    Drew, Janice E.; Andrew J Farquharson; Vase, Hollie; Carey, Frank A.; Steele, Robert J C; Ross, Ruth A; Bunton, David C.

    2015-01-01

    Abstract Human colon tissue explant culture provides a physiologically relevant model system to study human gut biology. However, the small (20–30 mg) and complex tissue samples used present challenges for monitoring tissue stability, viability, and provision of sufficient tissue for analyses. Combining molecular profiling with explant culture has potential to overcome such limitations, permitting interrogation of complex gene regulation required to maintain gut mucosa in culture, monitor res...

  13. 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.;

    1978-01-01

    An explant culture system has been developed for the long-term maintenance of colonic tissue from the rat. Explants of 1 cm2 in size were placed in tissue-culture dishes to which was added 2 ml of CMRL-1066 medium supplemented with glucose, hydrocortisone, beta-retinyl acetate, and either 2......,12-dimethylbenz[alpha]anthracene, aflatoxin B1, dimethylnitrosamine, 1,2-dimethylhydrazine, and methylazoxymethanol acetate into chemical species that bind to cellular DNA and protein....

  14. Effect of short-term enzymatic treatment on cell migration and cartilage regeneration: in vitro organ culture of bovine articular cartilage.

    Science.gov (United States)

    Seol, Dongrim; Yu, Yin; Choe, Hyeonghun; Jang, Keewoong; Brouillette, Marc J; Zheng, Hongjun; Lim, Tae-Hong; Buckwalter, Joseph A; Martin, James A

    2014-07-01

    Depending on the damage extent and adjacent tissue condition in traumatic cartilage injury, it is possible to heal the tissue by resident cells. Unlike autologous chondrocyte implantation, short-term enzymatic treatment is an effective single-step procedure without extra cell expansion. Moreover, this method has been shown to significantly increase cellularity in lesion edges, resulting in enhanced integration and interfacial strength. We hypothesize that the locally digested extracellular matrix by treatment allows effortless cell migration from the adjacent tissue. Full-thickness cartilage discs and osteochondral explants were prepared from mature bovine stifle joints. These specimens were treated with collagenase in a culture medium. Two concentrations, 0.25 and 0.5 mg/mL, were used with various treating time of 10, 30, and 180 min. The cartilages were subsequently washed and cultured with fibrin hydrogel. The effect of enzymatic treatment on cell migration was apparent in both experiments of the cartilage disc and full-thickness cartilage defect model. In the disc culture, the treatment resulted in an approximately three to four times higher number of migrated cells than nontreated control. In short-term collagenase-treated groups, the proteoglycan (PG) loss was localized in the edge of tissue with minimal cell death. The treatment also accelerated cell migration in the full-thickness cartilage defects and some cells differentiated into chondrocytes with the deposit of PG. Gene expression results could support the characteristics of migrated cells, which had migratory ability and chondrogenic differentiation potential with overexpression of collagen type I and II, respectively. Based on these results, short-term enzymatic treatment, which can accelerate cell migration into traumatically injured cartilage, has great potential for clinical application. PMID:24428547

  15. A novel method for coral explant culture and micropropagation.

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    Vizel, Maya; Loya, Yossi; Downs, Craig A; Kramarsky-Winter, Esti

    2011-06-01

    We describe here a method for the micropropagation of coral that creates progeny from tissue explants derived from a single polyp or colonial corals. Coral tissue explants of various sizes (0.5-2.5 mm in diameter) were manually microdissected from the solitary coral Fungia granulosa. Explants could be maintained in an undeveloped state or induced to develop into polyps by manipulating environmental parameters such as light and temperature regimes, as well as substrate type. Fully developed polyps were able to be maintained for a long-term in a closed sea water system. Further, we demonstrate that mature explants are also amenable to this technique with the micropropagation of second-generation explants and their development into mature polyps. We thereby experimentally have established coral clonal lines that maintain their ability to differentiate without the need for chemical induction or genetic manipulation. The versatility of this method is also demonstrated through its application to two other coral species, the colonial corals Oculina patigonica and Favia favus. PMID:20700752

  16. Endometrial explant culture to study the response of equine endometrium to insemination.

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    Nash, D M; Sheldon, I M; Herath, S; Lane, E A

    2010-08-01

    Mating-induced endometritis (MIE) is ubiquitous in the horse after natural mating and artificial insemination with frozen/thawed semen causing the most aggressive response. The majority of mares eliminate MIE 24-48 h after insemination. An endometrial explant culture was tested as a potential in vitro exemplar for sperm-induced MIE. Endometrial prostaglandin F(2alpha) (PGF(2alpha)) secretion and expression of interleukin-8 (IL-8) were used as markers of inflammation. Endometrial explants were cultured from uteri collected from follicular phase mares. Explants were challenged with 1 or 10 x 10(6) sperm/ml frozen/thawed semen, chilled semen, washed sperm or seminal plasma. Medium was collected 24 and 72 h after challenge and assayed for PGF(2alpha) by radioimmunoassay. Treatment of endometrial explants with frozen/thawed, chilled semen or washed sperm did not change the secretion of PGF(2alpha) compared with untreated controls. However, 24 h after challenge cultured explants expressed IL-8. The in vitro endometrial explant system did not represent the in vivo response to semen when PGF(2alpha) was used as a marker of inflammation, yet the use of gene expression as an inflammatory marker warrants further investigation. PMID:19144039

  17. Growing Three-Dimensional Cartilage-Cell Cultures

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    Spaulding, Glenn F.; Prewett, Tacey L.; Goodwin, Thomas J.

    1995-01-01

    Process for growing three-dimensional cultures of mammalian cartilage from normal mammalian cells devised. Effected using horizontal rotating bioreactor described in companion article, "Simplified Bioreactor for Growing Mammalian Cells" (MSC-22060). Bioreactor provides quiescent environment with generous supplies of nutrient and oxygen. Initiated with noncartilage cells. Artificially grown tissue resembles that in mammalian cartilage. Potential use in developing therapies for damage to cartilage by joint and back injuries and by such inflammatory diseases as arthritis and temporal-mandibular joint disease. Also used to test nonsteroid anti-inflammation medicines.

  18. Production of immunoglobulins in gingival tissue explant cultures from juvenile periodontitis patients

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    Hall, E.R.; Falkler, W.A. Jr.; Suzuki, J.B. (Univ. of Maryland Dental School, Baltimore (USA))

    1990-10-01

    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.

  19. Production of immunoglobulins in gingival tissue explant cultures from juvenile periodontitis patients

    International Nuclear Information System (INIS)

    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

  20. Histological changes during the adventitious shoot formation in seedling explants of pepper (Capsicum annuum L. cultured in vitro

    Directory of Open Access Journals (Sweden)

    Andrzej Gatz

    2014-02-01

    Full Text Available Adventitious shoots differentiated directly from explant tissue without intermediate callus on all types of examined explants (shoot tip, cotyledonary node, cotyledon and hypocotyl of Capsicum annuum L. cv. Bryza. First cell divisions took place as early as after 3 days of explant culture within epidermal and subepidermal layers of explants, and in the case of cotyledon also within mesophyll cells located near epidermis. Mitotic activity in these layers led to the formation of meristemoids (meristematic centres. In all types of studied explants, meristematic centres appeared approximately at the same time (after about 7 days of culture. In the second week bud primordia began to differentiate from meristematic centres. Subsequently some of shoot primordia developed into leaves and leaf-like structures (mainly on cotyledon explants, and also into adventitious buds with well developed apical meristem and leaf primordia.

  1. MICROPROPAGATION OF BLACK TURMERIC (CURCUMA CAESIA ROXB.) THROUGH IN VITRO CULTURE OF RHIZOME BUD EXPLANTS

    OpenAIRE

    Muhammad Shahinozzaman; Molla Ferdous; Muhammad Faruq; Mustafa Azad; Muhammad Amin

    2013-01-01

    In the present study, preliminary in vitro shoots propagation of Curcuma caesia Roxb. was investigated. Rhizome buds were used as explants and were cultured on Murashige and Skoog MS medium containing 6 Benzyl adenine BA alone or in combination with a Naphthalene acetic acid NAA. The results showed that the optimum shoot proliferation was obtained from MS medium containing 3.0 micro M BA + 0.5 micro M NAA. In this growth regulator combination, maximum 99.97 per cent explants produced 10.38 sh...

  2. In vitro maintenance of spermatogenesis in Xenopus laevis testis explants cultured in serum-free media

    International Nuclear Information System (INIS)

    Spermatogenesis has been maintained for extended periods in Xenopus laevis testis explants cultured in serum-free media supplemented with bovine serum albumin, insulin, transferrin, follicle-stimulating hormone, dihydrotestosterone, testosterone, retinol, ascorbate, and tocopherol. The organization of the testis fragments was maintained for 28 days, and all stages of development were present throughout the culture period. 3H-Thymidine-labeled secondary (Type B) spermatogonia developed in 28 days into spermatids at the acrosomal vesicle stage whereas labeled zygotene spermatocytes became mature spermatids in 28 days. Spermatogonial proliferation also continued in vitro for 28 days. Germ cell differentiation was not dependent upon exogenous testosterone, ascorbate, or tocopherol since 3H-labeled spermatogonia became mature spermatids in testes cultured 35 days in media lacking these supplements. Autoradiography demonstrated that 55% of the luminal sperm present in explants cultured 10 days had differentiated in vitro. Sperm from testes cultured 10-35 days were similar to sperm from freshly dissected testes with regard to motility and fecundity, and eggs fertilized with sperm from explant cultures developed normally into swimming tadpoles. The results demonstrate the feasibility of maintaining vertebrate spermatogenesis in culture and suggest that in vitro analysis of Xenopus spermatogenesis using defined media may provide important insights into the evolution of regulatory mechanisms in spermatogenesis

  3. Depot-dependent effects of adipose tissue explants on co-cultured hepatocytes

    DEFF Research Database (Denmark)

    Du, Zhen-Yu; Ma, Tao; Lock, Erik-Jan;

    2011-01-01

    We have developed an in vitro hepatocyte-adipose tissue explant (ATE) co-culture model enabling examination of the effect of visceral and subcutaneous adipose tissues on primary rat hepatocytes. Initial analyses of inflammatory marker genes were performed in fractionated epididymal or inguinal ad......), particularly macrophages, in inguinal adipose tissue resulting in stronger responses in terms of hepatotoxicity and insulin-resistance....... elicited a stronger cytotoxic response and higher level of insulin resistance in the co-cultured hepatocytes. In conclusion, our results reveal depot-dependent effects of ATEs on co-cultured primary hepatocytes, which in part may be related to a more pronounced infiltration of stromal vascular cells (SVCs......We have developed an in vitro hepatocyte-adipose tissue explant (ATE) co-culture model enabling examination of the effect of visceral and subcutaneous adipose tissues on primary rat hepatocytes. Initial analyses of inflammatory marker genes were performed in fractionated epididymal or inguinal...

  4. Electrical and synaptic properties of embryonic luteinizing hormone-releasing hormone neurons in explant cultures.

    OpenAIRE

    Kusano, K.; Fueshko, S; Gainer, H; Wray, S

    1995-01-01

    Voltage- and ligand-activated channels in embryonic neurons containing luteinizing hormone-releasing hormone (LHRH) were studied by patch-pipette, whole-cell current and voltage clamp techniques. LHRH neurons were maintained in explant cultures derived from olfactory pit regions of embryonic mice. Cells were marked intracellularly with Lucifer yellow following recording. Sixty-two cells were unequivocally identified as LHRH neurons by Lucifer yellow and LHRH immunocytochemistry. The cultured ...

  5. Type II and VI collagen in nasal and articular cartilage and the effect of IL-1 alpha on the distribution of these collagens

    NARCIS (Netherlands)

    I.D.C. Jansen; A.P. Hollander; D.J. Buttle; V. Everts

    2010-01-01

    The distribution of type II and VI collagen was immunocytochemically investigated in bovine articular and nasal cartilage. Cartilage explants were used either fresh or cultured for up to 4 weeks with or without interleukin 1α (IL-1α). Sections of the explants were incubated with antibodies for both

  6. Reactive oxygen species induce expression of vascular endothelial growth factor in chondrocytes and human articular cartilage explants

    OpenAIRE

    Fay, Jakob; Varoga, Deike; Wruck, Christoph J.; Kurz, Bodo; Goldring, Mary B.; Pufe, Thomas

    2006-01-01

    Vascular endothelial growth factor (VEGF) promotes cartilage-degrading pathways, and there is evidence for the involvement of reactive oxygen species (ROS) in cartilage degeneration. However, a relationship between ROS and VEGF has not been reported. Here, we investigate whether the expression of VEGF is modulated by ROS. Aspirates of synovial fluid from patients with osteoarthritis (OA) were examined for intra-articular VEGF using ELISA. Immortalized C28/I2 chondrocytes and human knee cartil...

  7. Prolonged viability of human organotypic skin explant in culture method (hOSEC)*

    OpenAIRE

    Frade, Marco Andrey Cipriani; de Andrade, Thiago Antônio Moretti; Aguiar, Andréia Fernanda Carvalho Leone; Guedes, Flávia Araújo; Leite, Marcel Nani; Passos, Williane Rodrigues; Coelho, Eduardo Barbosa; Das, Pranab Kummar

    2015-01-01

    BACKGROUND: Currently, the cosmetic industry is overwhelmed in keeping up with the safety assessment of the increasing number of new products entering the market. To meet such demand, research centers have explored alternative methods to animal testing and also the large number of volunteers necessary for preclinical and clinical tests. OBJECTIVES: This work describes the human skin ex-vivo model (hOSEC: Human Organotypic Skin Explant Culture) as an alternative to test the effectiveness of co...

  8. In situ fiber-optical monitoring of cytosolic calcium in tissue explant cultures

    CERN Document Server

    Ryser, Manuel; Geiser, Marianne; Frenz, Martin; Rička, Jaro

    2014-01-01

    We present a fluorescence-lifetime based method for monitoring cell and tissue activity in situ, during cell culturing and in the presence of a strong autofluorescence background. The miniature fiber-optic probes are easily incorporated in the tight space of a cell culture chamber or in an endoscope. As a first application we monitored the cytosolic calcium levels in porcine tracheal explant cultures using the Calcium Green-5N (CG5N) indicator. Despite the simplicity of the optical setup we are able to detect changes of calcium concentration as small as 2.5 nM, with a monitoring time resolution of less than 1 s.

  9. Selection of valine-resistance in callus culture of Arabidopsis thaliana (L.) Heynh. derived from leaf explants

    OpenAIRE

    Małgorzata D. Gaj; Grzegorz Czaja; Małgorzata Nawrot

    2014-01-01

    The selection of valine-resistant mutants was carried out in leaf explant cultures of three Arabidopsis thaliana (L.) Heynh. ecotypes: C-24, RLD and Columbia. The valine concentration used for in vitro selection, lethal for seed-growing plants, has not affected callus formation and growth. However, strong inhibition of shoot regeneration ability of calli growing under selection pressure was noticed. In total, 1043 explants were cultured on valine medium and 18 shoots were regenerated with an ...

  10. Prolonged viability of human organotypic skin explant in culture method (hOSEC)*

    Science.gov (United States)

    Frade, Marco Andrey Cipriani; de Andrade, Thiago Antônio Moretti; Aguiar, Andréia Fernanda Carvalho Leone; Guedes, Flávia Araújo; Leite, Marcel Nani; Passos, Williane Rodrigues; Coelho, Eduardo Barbosa; Das, Pranab Kummar

    2015-01-01

    BACKGROUND: Currently, the cosmetic industry is overwhelmed in keeping up with the safety assessment of the increasing number of new products entering the market. To meet such demand, research centers have explored alternative methods to animal testing and also the large number of volunteers necessary for preclinical and clinical tests. OBJECTIVES: This work describes the human skin ex-vivo model (hOSEC: Human Organotypic Skin Explant Culture) as an alternative to test the effectiveness of cosmetics and demonstrate its viability through cutaneous keratinocytes' proliferative capacity up to 75 days in culture. METHODS: The skin explants obtained from surgeries were cultured in CO2-humid incubator. After 1, 7, 30 and 75 days in culture, skin fragments were harvested for analysis with histomorphological exam (HE staining) on all days of follow-up and immunohistochemistry for Ck5/6, Ck10 and Ki-67 only on the 75th day. RESULTS: On the 7th day, the epidermis was perfect in the dermoepidermal junction, showing the viability of the model. On the 30th day, the epidermis was thicker, with fewer layers on the stratum corneum, although the cutaneous structure was unaltered. On the 75th day, the skin became thinner but the dermoepidermal junctions were preserved and epidermal proliferation was maintained. After the 75th day on culture, the skin was similar to normal skin, expressing keratinocytes with Ck5/6 on supra-basal layers; Ck10 on differentiated layers; and viability could be assessed by the positivity of basal cells by Ki-67. CONCLUSION: The hOSEC model seems a good alternative to animal testing; it can be used as a preclinical test analogous to clinical human skin test with similar effectiveness and viability proven by immunohistological analyses. PMID:26131864

  11. Glucocorticoids affect 24 h clock genes expression in human adipose tissue explant cultures.

    Directory of Open Access Journals (Sweden)

    Purificación Gómez-Abellán

    Full Text Available AIMS: 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. SUBJECTS AND METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

  12. Observations on the migratory behaviour of Schwann cells from adult peripheral nerve explant cultures.

    Science.gov (United States)

    Crang, A J; Blakemore, W F

    1987-06-01

    The migration of Schwann cells from adult sciatic nerve explant cultures has been examined by time-lapse photomicrography. Analysis of Schwann cell migratory behaviour indicates that the initial outwandering by individual Schwann cells was random. Although chance cell-cell contacts resulted in temporary immobilization of pairs of cells, stable multicellular structures did not form during this initial phase. As local cell densities increased, Schwann cells assembled networks within which Schwann cell movement continued to be observed. A second form of Schwann cell outgrowth was observed from degenerating fibres in which arrays of highly oriented Schwann cells migrated away from their basal lamina tubes onto the culture dish. These observations of Schwann cell random migration, network self-assembly and coordinated extratubal migration are considered to highlight aspects of Schwann cell behaviour, independent of axonal influences, which may have relevance to their role in peripheral nerve repair following nerve section. PMID:3612187

  13. Development of a pig jejunal explant culture for studying the gastrointestinal toxicity of the mycotoxin deoxynivalenol: histopathological analysis

    OpenAIRE

    Kolf-Clauw, Martine; Castellote, Jessie; Joly, Benjamin; Bourges-Abella, Nathalie; Raymond-Letron, Isabelle; Pinton, Philippe; Oswald, Isabelle P.

    2009-01-01

    The digestive tract is a target for the mycotoxin deoxynivalenol (DON), a major cereals grain contaminant of public health concern in Europe and North America. Pig, the most sensitive species to DON toxicity, can be regarded as the most relevant animal model for studying the intestinal effects of DON. A pig jejunal explants culture was developed to assess short-term effects of DON. In a first step, jejunal explants from 9-13 week-old and from 4-5 week-old pigs were cultured in vitro for up...

  14. Culturated rat cerebral cortex explants and their application in the study of SPECT scan radiopharaceuticals

    International Nuclear Information System (INIS)

    In this thesis mechanics that result in the distinct localization of radiopharmaceuticals within the brain have been investigated. In order to 'get more insight' in uptake and binding of radiopharmaceuticals bu brain tissue, use has been made of the tissue culture technique. Tissue culture privides the opportunity of doing experiments with brain tissue under stable conditions, in the absence of a blood-brain barrier, and without interference by cerebral blood flow. The present thesis is presented in two sections. The first part focusses on longterm culture of 'organotypic' cerebral neocortex tissue, obtained from neonatal rat brain and explanted into a chemically defined medium. Procedures were developed which enabled culturing of this tissue without the occurence of central necrosis and with the preservation of a characteristic histiotypic organization. Morphological characteristics of the cultures were described and measured at various ages in vitro. In the second part, the cultures were used to study mechanisms that might contribute to the tissue uptake of radiopharmaceuticals which are in clinical use for SPECT brain imaging. (author). 369 refs.; 50 figs.; 13 tabs

  15. Blockade of Toll-like receptor 2 prevents spontaneous cytokine release from rheumatoid arthritis ex vivo synovial explant cultures

    LENUS (Irish Health Repository)

    Nic An Ultaigh, Sinead

    2011-02-23

    Abstract Introduction The aim of this study was to examine the effect of blocking Toll-like receptor 2 (TLR2) in rheumatoid arthritis (RA) synovial cells. Methods RA synovial tissue biopsies, obtained under direct visualization at arthroscopy, were established as synovial explant cultures ex vivo or snap frozen for immunohistology. Mononuclear cell cultures were isolated from peripheral blood and synovial fluid of RA patients. Cultures were incubated with the TLR1\\/2 ligand, Pam3CSK4 (200 ng, 1 and 10 μg\\/ml), an anti-TLR2 antibody (OPN301, 1 μg\\/ml) or an immunoglobulin G (IgG) (1 μg\\/ml) matched control. The comparative effect of OPN301 and adalimumab (anti-tumour necrosis factor alpha) on spontaneous release of proinflammatory cytokines from RA synovial explants was determined using quantitative cytokine MSD multiplex assays or ELISA. OPN301 penetration into RA synovial tissue explants cultures was assessed by immunohistology. Results Pam3CSK4 significantly upregulated interleukin (IL)-6 and IL-8 in RA peripheral blood mononuclear cells (PBMCs), RA synovial fluid mononuclear cells (SFMCs) and RA synovial explant cultures (P < 0.05). OPN301 significantly decreased Pam3CSK4-induced cytokine production of tumour necrosis factor alpha (TNF-α), IL-1β, IL-6, interferon (IFN)-γ and IL-8 compared to IgG control in RA PBMCs and SFMCs cultures (all P < 0.05). OPN301 penetration of RA synovial tissue cultures was detected in the lining layer and perivascular regions. OPN301 significantly decreased spontaneous cytokine production of TNF-α, IL-1β, IFN-γ and IL-8 from RA synovial tissue explant cultures (all P < 0.05). Importantly, the inhibitory effect of OPN on spontaneous cytokine secretion was comparable to inhibition by anti-TNFα monoclonal antibody adalimumab. Conclusions These findings further support targeting TLR2 as a potential therapeutic agent for the treatment of RA.

  16. Differential response of normal and tumour oesophageal explant cultures to radiation

    Energy Technology Data Exchange (ETDEWEB)

    Mothersill, C.; Cusack, A.; MacDonnell, M.; Hennessy, T.P.; Seymour, C.B.

    1988-01-01

    An in vitro method is described which allows radiation dose response data to be obtained for samples of oesophageal mucosa obtained from patients undergoing resection for adeno or squamous cell carcinoma. Data are obtained using a growth endpoint from explant cultures and may be expressed in terms of absolute growth inhibition or reduced rate of growth. Radiation dose response curves suggest that cell survival is in the range expected for mammalian cells but that, as is found clinically, tumour cells are far more resistant to radiation than normal cells. The technique provides a means of assessing differential radiation response in normal and tumour tissues from the same patient, as it is unusual for both to be amenable to clonogenic assay.

  17. Micropropagation from cultured nodal explants of rose (Rosa hybrida L. cv. ‘Perfume Delight’

    Directory of Open Access Journals (Sweden)

    Kamnoon Kanchanapoom

    2010-01-01

    Full Text Available A method for the micropropagation of rose (Rosa hybrida L. cv. ‘Perfume Delight’ was developed. First to fifth nodal explants from young healthy shoots were excised and cultured on basal medium of Murashige and Skoog (1962, MS containing several concentrations of BA and NAA. Multiple shoot formation of up to 3 shoots was obtained on MS medium supplemented with 3 mg/l BA and 0.003 mg/l NAA. Shoot readily rooted on ¼MS medium devoid of growth regulators.Rooted plantlets were hardened and established in pots at 100% survival. In vitro flowering was observed on rose plantscultured on MS medium containing 3 mg/l BA and 0.003 mg/l NAA.

  18. Histology of Somatic Embryogenesis in Cultured Leaf Explants of Pistachio (Pistacia veraL)

    OpenAIRE

    ONAY, Ahmet

    2000-01-01

    The histology of somatic embryo initiation and development in pistachio ( Pistacia were L.) embryogenic masses (EMS) derived from leaf explants was examined using light microscopy. Explants with somatic embryos at different developmental stages were fixed for histological examination, cut into 10 µm thick sections, stained with hematoxylin-eosin and observed microscopically. The histological examination showed that the two types of cell clusters induced originated from explants and were mo...

  19. The effect of gamma radiation on in vitro cultured explants of yam (Dioscorea alata L.) cv. Kinampay

    International Nuclear Information System (INIS)

    Various explants of yam were irradiated with gamma rays at doses ranging from 5-50 Gy. Induction of callus was obtained in tuberous root and other vegetative explants: petiole, node, internode and shoot apex. Callus induction was observed in the irradiated and unirradiated fleshy or tuberous root explants grown in Murashige and Skoog's (MS) medium with benzyl adenine (BA) in combination with naphthalene acetic acid (NAA) ranging from 2-6 ppm. Stimulation of callus growth was obtained from tuberous root explants irradiated with 5 and 10 Gy and grown in MS medium supplemented with 2 ppm BA in combination with 6ppm NAA. Similar callus growth (as indicated by their weight) was observed at higher doses of 30 and 40 GY, provided that higher levels of 4 ppm BA and 6 ppm NAA were incorporated into the MS medium. Similarly, induction of callus was enhanced in tuberous root sections of putative mutant lines (for dwarfness and earliness) that were cultured in higher levels of BA and NAA in MS medium. Regeneration of plantlets was obtained from callus-derived shoot apex irradiated with 40 Gy and from calli-derived unirradiated nodal and tuberous root explants. (author). 40 refs.; 2 figs., 6 tabs

  20. Genome-wide transcriptional analysis of super-embryogenic Medicago truncatula explant cultures

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    Rolfe Barry G

    2008-10-01

    Full Text Available Abstract Background The Medicago truncatula (M. truncatula line 2HA has a 500-fold greater capacity to regenerate plants in culture by somatic embryogenesis than its wild type progenitor Jemalong. To understand the molecular basis for the regeneration capacity of this super-embryogenic line 2HA, using Affymetrix GeneChip®, we have compared transcriptomes of explant leaf cultures of these two lines that were grown on media containing the auxin NAA (1-naphthaleneacetic acid and the cytokinin BAP (6-benzylaminopurine for two weeks, an early time point for tissue culture proliferation. Results Using Affymetrix GeneChip®, GCRMA normalisation and statistical analysis, we have shown that more than 196 and 49 probe sets were significantly (p RESPONSE REGULATOR 10, AGAMOUS-LIKE 20, flower promoting factor 1, nodulin 3, fasciclin and lipoxygenase, and a down-regulated gene ETHYLENE INSENSITIVE 3, all of which positively correlated with the microarray data. Conclusion We have described the differences in transcriptomes between the M. truncatula super-embryogenic line 2HA and its non-embryogenic progenitor Jemalong at an early time point. This data will facilitate the mapping of regulatory and metabolic networks involved in the gaining totipotency and regeneration capacity in M. truncatula and provides candidate genes for functional analysis.

  1. Effect of environmental and cultural conditions on medium pH and explant growth performance of Douglas-fir ( Pseudotsuga menziesii) shoot cultures

    Science.gov (United States)

    Chen, Chien-Chih; Bates, Rick; Carlson, John

    2015-01-01

    The medium pH level of plant tissue cultures has been shown to be essential to many aspects of explant development and growth. Sensitivity or tolerance of medium pH change in vitro varies according to specific requirements of individual species. The objectives of this study are to 1) determine medium pH change over time in storage conditions and with presence of explants, 2) evaluate the effects of medium pH change on explant growth performance and 3) assess the effects of adding a pH stabilizer, 2-(N-morpholino)ethanesulfonic acid (MES) that is commonly used in Douglas-fir micropropagation medium. Vegetative buds were collected in the spring before breaking dormancy from juvenile and mature donor trees for conducting these evaluations. Medium, with or without MES, was pre-adjusted to five pH levels before adding MES, agar and autoclaving. Medium pH changes and explant growth parameters were measured at eight different incubation times. Overall, MES provided a more stable medium pH, relative to starting pH values, under both light and dark storage conditions as well as with presence of explants. A general trend of decreasing medium pH over time was found comparing explants from juvenile and mature donor genotypes. Explant height and weight growth increased over time, but differ among explants from juvenile and mature donor genotypes. Our findings suggest that a 21-day subculture practice may best sustain medium freshness, medium pH level and desirable explant growth. PMID:26535110

  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;

    2015-01-01

    Oncostatin M (OSM) were cultured for 21 days with or without a number of inhibitors targeting different types of proteases. Monoclonal antibodies were raised against the active sites of ADAMTS-4, -5, MMP-9 and -13, and 4 ELISAs were developed and technically validated. In addition, the established AGNx...

  3. 3D Hydrogel Scaffolds for Articular Chondrocyte Culture and Cartilage Generation.

    Science.gov (United States)

    Smeriglio, Piera; Lai, Janice H; Yang, Fan; Bhutani, Nidhi

    2015-01-01

    Human articular cartilage is highly susceptible to damage and has limited self-repair and regeneration potential. Cell-based strategies to engineer cartilage tissue offer a promising solution to repair articular cartilage. To select the optimal cell source for tissue repair, it is important to develop an appropriate culture platform to systematically examine the biological and biomechanical differences in the tissue-engineered cartilage by different cell sources. Here we applied a three-dimensional (3D) biomimetic hydrogel culture platform to systematically examine cartilage regeneration potential of juvenile, adult, and osteoarthritic (OA) chondrocytes. The 3D biomimetic hydrogel consisted of synthetic component poly(ethylene glycol) and bioactive component chondroitin sulfate, which provides a physiologically relevant microenvironment for in vitro culture of chondrocytes. In addition, the scaffold may be potentially used for cell delivery for cartilage repair in vivo. Cartilage tissue engineered in the scaffold can be evaluated using quantitative gene expression, immunofluorescence staining, biochemical assays, and mechanical testing. Utilizing these outcomes, we were able to characterize the differential regenerative potential of chondrocytes of varying age, both at the gene expression level and in the biochemical and biomechanical properties of the engineered cartilage tissue. The 3D culture model could be applied to investigate the molecular and functional differences among chondrocytes and progenitor cells from different stages of normal or aberrant development. PMID:26484414

  4. Lubricin is expressed in chondrocytes derived from osteoarthritic cartilage encapsulated in poly(ethylene glycol diacrylate scaffold

    Directory of Open Access Journals (Sweden)

    G. Musumeci

    2011-09-01

    Full Text Available Osteoarthritis (OA is characterized by degenerative changes within joints that involved quantitative and/or qualitative alterations of cartilage and synovial fluid lubricin, a mucinous glycoprotein secreted by synovial fibroblasts and chondrocytes. Modern therapeutic methods, including tissue-engineering techniques, have been used to treat mechanical damage of the articular cartilage but to date there is no specific and effective treatment. This study aimed at investigating lubricin immunohistochemical expression in cartilage explant from normal and OA patients and in cartilage constructions formed by Poly (ethylene glycol (PEG based hydrogels (PEG-DA encapsulated OA chondrocytes. The expression levels of lubricin were studied by immunohistochemistry: i in tissue explanted from OA and normal human cartilage; ii in chondrocytes encapsulated in hydrogel PEGDA from OA and normal human cartilage. Moreover, immunocytochemical and western blot analysis were performed in monolayer cells from OA and normal cartilage. The results showed an increased expression of lubricin in explanted tissue and in monolayer cells from normal cartilage, and a decreased expression of lubricin in OA cartilage. The chondrocytes from OA cartilage after 5 weeks of culture in hydrogels (PEGDA showed an increased expression of lubricin compared with the control cartilage. The present study demonstrated that OA chondrocytes encapsulated in PEGDA, grown in the scaffold and were able to restore lubricin biosynthesis. Thus our results suggest the possibility of applying autologous cell transplantation in conjunction with scaffold materials for repairing cartilage lesions in patients with OA to reduce at least the progression of the disease.

  5. In vitro plant regeneration of two cucumber (Cucumis sativum L. genotypes: Effects of explant types and culture medium

    Directory of Open Access Journals (Sweden)

    Grozeva Stanislava

    2014-01-01

    Full Text Available The effect of different phytohormone concentrations on callusogenesis and organogenesis in two cucumber genotypes were studied. It was established that the rate of plant regeneration depends on genotype, explant type and culture medium. Hypocotyls were found to be more responsive than cotyledons in morphogenesis. In vitro planlet-regenerants have been obtained in hypocotyls explants on culture medium with 1.0 and 2.0 mgL-1 BA for cultivar Gergana and in 1.0 and 3.0 mgL-1K-line 15B. Induction of regeneration in cotyledons were established only in cultivar Gergana on culture medium supplemented with 3.0 mgL-1 BA and in combination of 0.5 mgL-1IAA.

  6. Sonication cultures of explanted components as an add-on test to routinely conducted microbiological diagnostics improve pathogen detection.

    Science.gov (United States)

    Holinka, Johannes; Bauer, Leonhard; Hirschl, Alexander M; Graninger, Wolfgang; Windhager, Reinhard; Presterl, Elisabeth

    2011-04-01

    The purpose of this study was to improve the pathogen detection in prosthetic joint infections, particularly to evaluate the feasibility of the sonication culture method in the clinical routine. Explanted components of all patients with presumptive prosthetic or implant infection were sonicated separately in sterile containers to dislodge the adherent bacteria from the surfaces and cultured. The results of sonication culture were compared to the conventional tissue culture. We investigated 60 consecutive patients with loosening of the prostheses or implants Forty patients had septic and 20 aseptic loosening (24 knee prostheses, 21 hip prostheses, 6 mega-prostheses, 2 shoulder prostheses, 6 osteosynthesis, 1 spinal instrumentation). The sensitivity of sonication fluid culture was 83.3%, of single positive tissue culture was 72.2% and 61.1% when two or more cultures yielded the same microorganism. In patients receiving antibiotic therapy the sensitivity was 65.9%, 57.5%, and 42.5%, respectively. Pathogens detected in a single tissue culture as well as in sonication culture yielded a significantly higher rate of prosthetic infection than conventional tissue culture alone (p = 0.008), even in patients receiving continuous antibiotic therapy before explantation (p = 0.016). The sonication method represents an essential add-on in pathogen detection compared to conventional tissue culture. PMID:21337398

  7. Optimal culture conditions for the initial development of Ilex paraguariensis A.St.-Hil. explants

    Directory of Open Access Journals (Sweden)

    Ana Gláucia Griebeler

    2014-12-01

    Full Text Available In the state of Rio Grande do Sul, Brazil, yerba mate (Ilex paraguariensis A. St.-Hil. is an economically and culturally important species. Because of the difficulty in obtaining plantlets from seeds, micropropagation has emerged as an alternative that could increase production. This work aimed to evaluate the effect that treatment with 6-benzyladenine (BA, at concentrations of 2.2, 8.9, and 20 µM and associated with naphthaleneacetic acid (NAA, 0.5 µM, has on the in vitro growth and leaf emission of I. paraguariensis nodal segments cultivated in 0.5 X Murashige and Skoog (MS medium or 0.5 X Woody Plant Medium (WPM. The most efficient treatment was MS medium with 8.9 µM BA+NAA, in which I. paraguariensis presented greater shoot elongation and higher leaf counts than in the other treatments. Therefore, use of the MS medium with 8.9 µM BA+NAA is recommended in order to increase efficiency in the initial development of I. paraguariensis explants.

  8. Type II and VI collagen in nasal and articular cartilage and the effect of IL-1 alpha on the distribution of these collagens

    OpenAIRE

    Jansen, I.D.C.; Hollander, A P; Buttle, D. J.; Everts, V.

    2010-01-01

    The distribution of type II and VI collagen was immunocytochemically investigated in bovine articular and nasal cartilage. Cartilage explants were used either fresh or cultured for up to 4 weeks with or without interleukin 1α (IL-1α). Sections of the explants were incubated with antibodies for both types of collagen. Microscopic analyses revealed that type II collagen was preferentially localized in the interchondron matrix whereas type VI collagen was primarily found in the direct vicinity o...

  9. The effect of radiation and cytotoxic platinum compounds on the growth of normal and tumour bladder explant cultures

    Energy Technology Data Exchange (ETDEWEB)

    Mothersill, C.; Seymour, C.B.; Cusack, A.; O' Brien, A.; Butler, M. (Saint Luke' s Hospital, Dublin (Ireland) Meath Hospital, Dublin (Ireland). Urology Dept.)

    1990-01-01

    Using an explant tissue culture model developed by this group for use with human surgical and biopsy specimens, data are presented showing the response of normal and tumor bladder urothelium to radiation in combination with cis- and carboplatin. Cellular response is measured after two weeks in culture as a reduction in the extent of outgrowth from the explant. The outgrowth has been shown to be growing and to be epithelial. Results showed that when either drug or radiation is used singly, the tumour is resistant to treatment while the normal cells are severely affected. However, appropriate combinations of either drug with radiation reverse the unfavourable therapeutic ratio and result in higher tumour cell kill. The model may be useful for investigating mechanisms of radiation/chemotherapy action at the cellular level and, if integrated into appropriate clinical trials, may serve as an easy-to-use in vitro test for optimising single agent or combination therapy regimens. (orig.).

  10. Investigating Molecular Mechanisms of Embryonic Mammary Gland Development by Bead-Implantation in Embryonic Flank Explant Cultures – A Protocol

    OpenAIRE

    Veltmaat, Jacqueline M.

    2013-01-01

    The involvement of molecular mechanisms in a particular process such as embryonic mammary gland development, can be revealed by modulation of one or several factors that purportedly act in that process. If those factors or their inhibitors are soluble, their function can be tested by loading them onto small inert beads, which are then implanted in cultured explants of the tissue of interest, in this case embryonic flanks. We here describe a protocol for such experiments.

  11. Effects of donor plant age and explants on in vitro culture of Cedrela montana Moritz ex Turcz

    Directory of Open Access Journals (Sweden)

    Sofía Basto

    2012-12-01

    Full Text Available To evaluate the organogenic potential of Cedrela montana Moritz ex Turcz. Explants from mature (10-20 year-old and juvenile (7-18 month-old trees were collected. The first grouping included buds, leaves, and nodes derived from juvenile basal offshoots and rejuvenated shoots from cuttings. The second, included leaves, petioles, nodes, internodes and nodes of in vitro elongated shoots. The highest organogenic potential was observed in nodes from juvenile trees: 45.8% of explants presented axillary bud elongation, while 56.2% presented rooting in a growth regulator free culture medium. Fifty-one percent of elongated shoots produced adventitious shoots with 0.5 μM NAA and 0.5 μM BA; 30% with 0.5 μM NAA and 1 μM BA; and 30% with 1 μM BA. Twenty percent presented roots with 0.5 μM NAA. Root formation was stimulated in a medium supplemented with activated charcoal (5 gL-1. The acclimatization of eighty percent of plantlets regenerated from nodes, and of 72.5% in vitro generated shoots was successful. On the contrary, mature trees material presented low organogenic response. Axillary bud elongation was recorded just in 10.7% of explants from juvenile shoots and in 6.7% of explants from rejuvenated shoots. The age of donor plant and type of explant affect the organogenic potential of C. montana. This study contributes to the understanding of this species’ response under in vitro conditions.

  12. THE PHENOLS ACCUMULATION IN TRANSFORMED ROOT CULTURES OF DIFFERENT EXPLANTS SOURCES OF COMMON BUCKWHEAT (Fagopyrum esculentum Moench

    Directory of Open Access Journals (Sweden)

    O. V. Sytar

    2013-06-01

    Full Text Available The growth parameters of transformed root cultures, total phenolic content and phenolic acids composition has been studied in root cultures, which were obtained from various explants of buckwheat by Agrobacterium rhizogenes strains A4. The methods of obtaining of the transformed root cultures, total phenol estimation, gas-liquid chromatography and polymerase chain reaction has been used. Elevated levels of total phenols in transformed roots of buckwheat from different sources of explants have been found. The high content of chlorogenic, p-hydroxybenzoic, p-anisic and caffeic acids has been discovered in the root cultures, which can be used for their industrial production. Maximal root growth was equal 21.2 g/l of dry weight in the roots as source for root culture, 17.7 g/l with leaves and 14.6 g/l with stems at 3 week after placement. Molecular analysis by polymerase chain reaction amplification was confirmed that the rol B gene (652 bp which transferred info hairy roots from Ri-plasmid in Agrobacterium rhizogenes is responsible for induction of root from plant species.

  13. Direct regeneration of Periwinkle (Catharanthus roseus via node explants culture and different combinations of plant growth regulators

    Directory of Open Access Journals (Sweden)

    M. Talebi

    2012-09-01

    Full Text Available Periwinkle (Catharanthus roseus L., Apocynaceae contains more than 130 different terpenoid indole alkaloids (TIAs, of which two dimeric alkaloids, Vinblastine and Vincristine, have antineoplastic activity and are useful in treatment of various cancers. Specific production of some alkaloids in differentiated tissues such as leaf and stem led to use direct regeneration of explants in order to increase the production of these important alkaloids in the plant. In this research, 30 combinations of plant growth regulators and activated charcoal were used in MS media for direct regeneration of node explants. Application of BAP in media containing 1 g/l activated charcoal showed the best direct regeneration of node explants and shoot proliferation. Although application of activated charcoal is necessary for periwinkle growth in media due to many phenolic compounds, but it has negative effects on adsorption of plant growth regulators and consequently reduce shoot proliferation. Therefore, it seems that 1 g/l activated charcoal is an appropriate concentration for preparing shoot proliferation media. In addition, transporting regenerated shoots to culture media containing NAA resulted in increasing shoot length. Proliferated shoots rooted in media without PGR and with 2 g/l activated charcoal and acclimated with environmental conditions after transferring to the soil.

  14. Fumonisin B₁ (FB₁) Induces Lamellar Separation and Alters Sphingolipid Metabolism of In Vitro Cultured Hoof Explants.

    Science.gov (United States)

    Reisinger, Nicole; Dohnal, Ilse; Nagl, Veronika; Schaumberger, Simone; Schatzmayr, Gerd; Mayer, Elisabeth

    2016-01-01

    One of the most important hoof diseases is laminitis. Yet, the pathology of laminitis is not fully understood. Different bacterial toxins, e.g. endotoxins or exotoxins, seem to play an important role. Additionally, ingestion of mycotoxins, toxic secondary metabolites of fungi, might contribute to the onset of laminitis. In this respect, fumonsins are of special interest since horses are regarded as species most susceptible to this group of mycotoxins. The aim of our study was to investigate the influence of fumonisin B₁ (FB₁) on primary isolated epidermal and dermal hoof cells, as well as on the lamellar tissue integrity and sphingolipid metabolism of hoof explants in vitro. There was no effect of FB₁ at any concentration on dermal or epidermal cells. However, FB₁ significantly reduced the separation force of explants after 24 h of incubation. The Sa/So ratio was significantly increased in supernatants of explants incubated with FB₁ (2.5-10 µg/mL) after 24 h. Observed effects on Sa/So ratio were linked to significantly increased sphinganine concentrations. Our study showed that FB₁ impairs the sphingolipid metabolism of explants and reduces lamellar integrity at non-cytotoxic concentrations. FB₁ might, therefore, affect hoof health. Further in vitro and in vivo studies are necessary to elucidate the effects of FB₁ on the equine hoof in more detail. PMID:27023602

  15. Effect of Pre-culture Irradiation and Explant Types on Efficiency of Brassica napus Genetic Transformation

    International Nuclear Information System (INIS)

    The irradiated seeds of canola cv. Drakkar ( Brassica napus l. ) were germinated under aspect conditions, cotyledonary petioles and hypocotyl of 6 days old seedlings were used for Agrobacterium-mediated transformation. Agrobacterium tumefaciens has construct with the selectable marker gene (NPT II) and the desirable gene (HPPD). Direct and indirect shoot organogenesis were obtained from the both explants. Cotyledonary petioles was higher responded than hypocotyl with respective 26% and 14% of the explants producing NPT II-positive shoots after the selection on 50mg/l kanamycin. Calli might develop on and not in the agar medium were un transformation. This explains the higher number of escapes detected in hypocotyl explants than in experiments with cotyledons. The frequency of transformation plants as a function of indirect organogenesis was more than direct shoot regeneration from explants. The pre- irradiation with 75 Gy of gamma rays enhanced the genetic transformation frequencies by about 10 % as compared to that of the un-irradiated material. The obtained shoots were rooted and regenerated mature plants

  16. Neuregulin-1β Regulates the migration of Different Neurochemical Phenotypic Neurons from Organotypically Cultured Dorsal Root Ganglion Explants.

    Science.gov (United States)

    Li, Yunfeng; Liu, Guixiang; Li, Hao; Bi, Yanwen

    2016-01-01

    Neuregulin-1β (NRG-1β) has multiple roles in the development and function in the nervous system and exhibits potent neuroprotective properties. In the present study, organotypically cultured dorsal root ganglion (DRG) explants were used to evaluate the effects of NRG-1β on migration of two major phenotypic classes of DRG neurons. The signaling pathways involved in these effects were also determined. Organotypically cultured DRG explants were exposed to NRG-1β (20 nmol/L), the phosphatidylinositol 3-kinase inhibitor LY294002 (10 μmol/L) plus NRG-1β (20 nmol/L), the extracellular signal-regulated protein kinase (ERK1/2) inhibitor PD98059 (10 μmol/L) plus NRG-1β (20 nmol/L), and LY294002 (10 μmol/L) plus PD98059 (10 μmol/L) plus NRG-1β (20 nmol/L), respectively, for 3 days. The DRG explants were continuously exposed to culture media as a control. After that, all above cultures were processed for detecting the mRNA levels of calcitonin gene-related peptide (CGRP) and neurofilament-200 (NF-200) by real-time PCR analysis. CGRP and NF-200 expression in situ was determined by fluorescent labeling technique. The results showed that NRG-1β elevated the mRNA and protein levels of CGRP and NF-200. NRG-1β also increased the number and the percentage of CGRP-immunoreactive (IR) migrating neurons and NF-200-IR migrating neurons. Inhibitors (LY294002, PD98059) either alone or in combination blocked the effects of NRG-1β. The contribution of NRG-1β on modulating distinct neurochemical phenotypic plasticity of DRG neurons suggested that NRG-1β signaling system might play an important role on the biological effects of primary sensory neurons. PMID:26093851

  17. Studies of mineralization in tissue culture: optimal conditions for cartilage calcification

    Science.gov (United States)

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

    1992-01-01

    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.

  18. The use of dynamic culture devices in articular cartilage tissue engineering.

    OpenAIRE

    Akmal, M.

    2006-01-01

    Tissue engineered repair of articular cartilage has now become a clinical reality with techniques for cell culture having advanced from laboratory experimentation to clinical application. Despite the advances in the use of this technology in clinical applications, the basic cell culture techniques for autologous chondrocytes are still based on primitive in-vitro monolayer culture methods. Articular chondrocytes are known to undergo fibroblastic change in monolayer culture as this is not their...

  19. In vitro culture of Cucumis sativus L. VI. Histological analysis of leaf explants cultured on media with 2, 4-D or 2, 4, 5-T

    Directory of Open Access Journals (Sweden)

    Anna Nadolska-Orczyk

    2014-02-01

    Full Text Available The developmental sequence of callus initiation and somatic embryogenesis in leaf explants of Cucumis sativus cv. Borszczagowski was analysed and compared on media containing two different auxin phenoxy-derivatives (2,4-D and 2,4,5-T and cytokinin (BAP or 2iP. During the first 20 days of culture on media with 2,4,5-T proliferation of parenchymatic tissue occurred mainly and only small meristematic centers were observed. There was an intensive detachment of parenchymatic cells and dissociation of their cell walls near vessels and in the lower part of the explant adjacent to the medium. These cells were strongly plasmolysed. On the 2,4-D containing medium mostly meristematic tissue developed, proliferating around vascular bundles and forming meristematic centers or promeristem-like structures. After 35-50 days of culture, secondary callus was formed by separation of meristematic cells from the meristem surface in explants cultured on the 2,4-D containing medium. On medium supplemented with 2, 4, 5-T the detachment of parenchymatic and meristematic cells occurred, along with formation of a gel-like substance. The gel-like callus contained multi-cellular aggregates, proembryoids and embryoids. This type of callus tissue was initiated more intensively on medium with 2, 4, 5-T, but the frequency of somatic embryogenesis was much lower. The periferial cells of aggregates, proembryoids and embryoids showed the tendency to separate from the surface of the tissue. Many embryoids formed adventitious embryos.

  20. Radiation-induced outgrowth inhibition in explant cultures from surgical specimens of five human organs

    Energy Technology Data Exchange (ETDEWEB)

    Mothersill, Carmel; Cusack, Anne; Seymour, C.B.

    1988-03-01

    An explant outgrowth technique to determine the radiation response of five different human organs (bladder, oesophagus, colon, breast and thyroid) is described. In each case except thyroid, where malignancies are rare, data are presented for normal and malignant tissue. Results show that variations in response, consistent with those observed in vivo, can be measured. Tumours were in all cases highly resistant to radiation relative to their corresponding normal tissue. Possible reasons for this are discussed. The method may prove useful in the prediction of the radiobiological response for tumour and surrounding normal tissue where post-operative therapy is planned.

  1. Cartilage-derived extracellular matrix extract promotes chondrocytic phenotype in three-dimensional tissue culture.

    Science.gov (United States)

    Youngstrom, Daniel W; Cakstina, Inese; Jakobsons, Eriks

    2016-05-01

    Cell transplantation is a promising regenerative therapy for cartilage degeneration. However, obtaining sufficient numbers of cells for this purpose is a challenge, due a lack of autologous donor tissue and the difficulty of culturing chondrocytes in vitro. Tissue engineering strategies that induce or maintain chondrocytic phenotype may solve these problems by (1) broadening the range of available donor tissue, and (2) facilitating the expansion of these cells while controlling phenotypic drift. In this study, bone marrow-derived mesenchymal stem cells (MSCs) and cartilage-derived cells (CDCs) were cultured on composite hydrogels containing agarose and homogenized cartilage extracellular matrix (ECM). MSCs cultured on agarose-ECM scaffolds did not show significant signs of chondrogenic differentiation in the absence of additional cues. However, CDCs cultured on agarose-ECM scaffolds proliferated more rapidly than their ECM-free counterparts and MSCs, while retaining chondrocytic morphology. These results were corroborated via expression of cartilage marker genes: in autologous constructs, SOX 9 expression was upregulated by 12.6 ± 5.3-fold, and COL II was upregulated by 2.0 ± 0.3-fold. Agarose-ECM composite hydrogels are therefore useful for expanding partially differentiated CDCs for applications in regenerative medicine. PMID:25707441

  2. Chondroprotective Effect of Kartogenin on CD44-Mediated Functions in Articular Cartilage and Chondrocytes

    OpenAIRE

    Ono, Yohei; Ishizuka, Shinya; Knudson, Cheryl B.; Knudson, Warren

    2014-01-01

    Objective: A recent report identified the small molecule kartogenin as a chondrogenic and chondroprotective agent. Since changes in hyaluronan metabolism occur during cartilage degeneration in osteoarthritis, we began studies to determine whether there was a connection between extracellular hyaluronan, CD44–hyaluronan interactions and the effects of kartogenin on articular chondrocytes. Methods: Chondrocytes cultured in monolayers, bioengineered neocartilages, or cartilage explants were treat...

  3. Modeling IL-1 induced degradation of articular cartilage.

    Science.gov (United States)

    Kar, Saptarshi; Smith, David W; Gardiner, Bruce S; Li, Yang; Wang, Yang; Grodzinsky, Alan J

    2016-03-15

    In this study, we develop a computational model to simulate the in vitro biochemical degradation of articular cartilage explants sourced from the femoropatellar grooves of bovine calves. Cartilage explants were incubated in culture medium with and without the inflammatory cytokine IL-1α. The spatio-temporal evolution of the cartilage explant's extracellular matrix components is modelled. Key variables in the model include chondrocytes, aggrecan, collagen, aggrecanase, collagenase and IL-1α. The model is first calibrated for aggrecan homeostasis of cartilage in vivo, then for data on (explant) controls, and finally for data on the IL-1α driven proteolysis of aggrecan and collagen over a 4-week period. The model was found to fit the experimental data best when: (i) chondrocytes continue to synthesize aggrecan during the cytokine challenge, (ii) a one to two day delay is introduced between the addition of IL-1α to the culture medium and subsequent aggrecanolysis, (iii) collagen degradation does not commence until the total concentration of aggrecan (i.e. both intact and degraded aggrecan) at any specific location within the explant becomes ≤1.5 mg/ml and (iv) degraded aggrecan formed due to the IL-1α induced proteolysis of intact aggrecan protects the collagen network while collagen degrades in a two-step process which, together, significantly modulate the collagen network degradation. Under simulated in vivo conditions, the model predicts increased aggrecan turnover rates in the presence of synovial IL-1α, consistent with experimental observations. Such models may help to infer the course of events in vivo following traumatic joint injury, and may also prove useful in quantitatively evaluating the efficiency of various therapeutic molecules that could be employed to avoid or modify the course of cartilage disease states. PMID:26874194

  4. In vitro cartilage culture: flow, transport and reaction in fibrous porous media

    OpenAIRE

    AHMADI-SENICHAULT, Azita; Lasseux, Didier; LETELLIER, Samuel

    2007-01-01

    Flow and transport in fibrous media are encountered in a wide variety of domains ranging from biotechnology to filtration in chemical engineering. The context of this work is the in vitro cartilage cell culture on a fibrous biodegradable polymer scaffold placed in a bioreactor. A seeding process using a liquid containing cells (chondrocytes) initiates the culture and an imposed continuous flow through the scaffold allows both the transport of nutrients necessary for cell-growth and of metabol...

  5. Changes in the metabolic footprint of placental explant-conditioned medium cultured in different oxygen tensions from placentas of small for gestational age and normal pregnancies.

    LENUS (Irish Health Repository)

    Horgan, R P

    2012-01-31

    Being born small for gestational age (SGA) confers significantly increased risks of perinatal morbidity and mortality. Accumulating evidence suggests that an SGA fetus results from a poorly perfused and abnormally developed placenta. Some of the placental features seen in SGA, such as abnormal cell turnover and impaired nutrient transport, can be reproduced by culture of placental explants in hypoxic conditions. Metabolic footprinting offers a hypothesis-generating strategy to investigate factors absorbed by and released from this tissue in vitro. Previously, metabolic footprinting of the conditioned culture media has identified differences in placental explants cultured under normoxic and hypoxic conditions and between normal pregnancies and those complicated by pre-eclampsia. In this study we aimed to examine the differences in the metabolic footprint of placental villous explants cultured at different oxygen (O(2)) tensions between women who deliver an SGA baby (n = 9) and those from normal controls (n = 8). Placental villous explants from cases and controls were cultured for 96 h in 1% (hypoxic), 6% (normoxic) and 20% (hyperoxic) O(2). Metabolic footprints were analysed by Ultra Performance Liquid Chromatography coupled to an electrospray hybrid LTQ-Orbitrap Mass Spectrometry (UPLC-MS). 574 metabolite features showed significant difference between SGA and normal at one or more of the oxygen tensions. SGA explant media cultured under hypoxic conditions was observed, on a univariate level, to exhibit the same metabolic signature as controls cultured under normoxic conditions in 49% of the metabolites of interest, suggesting that SGA tissue is acclimatised to hypoxic conditions in vivo. No such behaviour was observed under hyperoxic culture conditions. Glycerophospholipid and tryptophan metabolism were highlighted as areas of particular interest.

  6. Plant regeneration of Rhabdadenia Ragonesei (Apocynaceae by in vitro culturing of leaf explants REGENERACIÓN DE PLANTAS DE RHABDADENIA RAGONESEI (APOCYNACEAE POR CULTIVO IN VITRO DE EXPLANTES FOLIARES

    Directory of Open Access Journals (Sweden)

    Eduardo Flachsland

    2010-08-01

    Full Text Available Plants of Rhabdadenia Ragonesei Woodson (Apocynaceae were regenerated in vitro from leaves explants. The procedure employed includes: 1 Surface sterilization of leaves by immersion in 70% ethanol (10 s followed by 1,1%NaOCl (15 min and three wash with sterile distilled water. 2 Callus and buds induction by culture on Murashige and Skoog medium (MS + 3 mg/L benzyladenine (BAP. 3 Subculture of callus and buds on MS + 1 mg/L BAP, and 4 Rooting on MS + 0.5 mg/L naftalenacetic acid Se regeneraron plantas de Rhabdadenia Ragonesei Woodson (Apocynaceae mediante el cultivo in vitro de explantes foliares en condiciones ambientales controladas. El procedimiento consistió en: 1 Desinfección de las hojas por inmersión en etanol al 70% (10 s seguida de Inmersión en NaOCl al 1,1% (15 min y lavado tres veces con agua destilada estéril. 2 Inducción de callos y yemas mediante el cultivo de explantes foliares en el medio de Murashige y Skoog (MS + 3 mg/L de benciladenina (BAP. 3 Subcultivo de callos y yemas en MS + 1 mg/L de BAP y 4 Enraizamiento de los vastagos obtenidos en MS + 0,5 mg/L de ácido naftalenacético

  7. Long-term culture of sponge explants: conditions enhancing survival and growth, and assessment of bioactivity

    NARCIS (Netherlands)

    Caralt, de S.; Agell, G.; Uriz, M.J.

    2003-01-01

    Sponges are an important source of secondary metabolites with pharmaceutical interest. This is the main reason for the increasing interest of sponge culture recent years. The optimal culture system depends on the species to be cultured: while some species easily produce sponge aggregates after disso

  8. The structure of Human Mesenchymal Stem Cells Differentiated into Cartilage in Micromass Culture System

    Directory of Open Access Journals (Sweden)

    Mohamadreza Baghaban Eslaminejad

    2006-01-01

    Full Text Available Introduction: The aim of this study was to differentiate humanmesenchymal stem cells (hMSCs into cartilage in a micromass culturesystem and study of their structure by light and electron microscopy.Material and Methods: Human bone marrow cells obtained from volunteerpatients were plated in 75-cm2 flasks and their MSCs were expandedthrough several sub-cultures. The passage 4 cells were used to establishmicromass culture system for chondrogenic differentiation. For this purpose,200,000 fibroblastic cells were placed in centrifuge tubes and pelleted at 250g for 5 minutes. About 0.5 ml chondrogenic induction medium was thenadded to the pellet and the culture incubated in 5% CO2 at 37°C for 21 days.Then, some pellets were utilized to evaluate chondrogenic differentiation byeither RT-PCR analysis of some cartilage marker molecules or specificstaining for detecting cartilage matrix, and other pellets were used for lightand electron microscopic study of differentiated tissue.Results: Primary culture of the bone marrow cells were initially composed ofthe spindle- and round shaped cells, from which the spindle cells remainedand expanded through several passages. At the end of differentiation period,RT-PCR analysis showed high production of collagen II and X and aggrecanmRNA inside the differentiated cells, and toluidine blue staining indicatedintermediate accumulation of the metachromatic matrix among the inducedcells. In general, light micrograph indicated a rather cellular state of thedifferentiated tissue in which the peripheral part had more metachromaticmatrix than central zone. More detailed study of the sections revealed thatinduced aggregates of the cells were composed externally of very thin layerof elongated cells reminiscent of perichondrium and internally a mass of ovalcells comprising the main part of the pellet. Ultra-thin sections showed thatthe cells in perichondrium-like layer were very similar to fibroblastic cells andthose located

  9. (14C)-glucose metabolism during shoot bud development in cultured cotyledon explants of Pinus radiata

    International Nuclear Information System (INIS)

    Excised cotyledons of Pinus radiata D. Don cultured under shoot-forming (plus benzyladenine) and non shoot-forming (minus benzyladenine) conditions for 10 and 21 days were fed U-[14C]-glucose for 3 h in the light followed by a 3 h chase period. The labelling of individual metabolites as well as 14C incorporation into protein was assessed. It was found that the general metabolic patterns were qualitatively the same in shoot-forming and non shoot-forming conditions, however, metabolism leading to respiration as well as to the synthesis of some amino acids and protein synthesis was enhanced in the shoot-forming cultures. (author)

  10. The effects of low-level ionising radiation on primary explant cultures of rainbow trout Pronephros

    International Nuclear Information System (INIS)

    It has long been known that the haematopoietic tissue of mammals is one of the most radiosensitive tissues. In vitro studies on prawn have also shown that low doses of radiation has an extremely deleterious effect on cells cultured from this animal's blood forming tissues. This raises the question on the relative effects of radiation between animals from different species. One of the most important aquatic animals, from both an economic and ecologic point of view, is the fish. With this in mind, primary cultures of the blood forming tissues of rainbow trout were exposed to radiation followed by a morphological comparison between control and irradiated cultures. The cultured cells were characterised as macrophages following incubation with non-specific fluorescent beads and human apoptotic PMN. The cells demonstrated both specific and non-specific phagocytosis, by consuming the non-indigenous bodies, and were classified as phagocytic leucocytes. These cells were found in two morphological forms, stretched and rounded. It was shown that there was a commensurate increase in the number of stretched cells following application of radiation. Radiation was also shown to cause a dose-dependent increase in the amounts of apoptosis and necrosis in cells over time. The phagocytic efficacy of the irradiated leucocytes compared to controls was also investigated. (author)

  11. Effect of explant treatment with gamma-rays, EMS and SA on somatic tissue culture in rice

    International Nuclear Information System (INIS)

    Dry seeds of two rice cultivars were treated with gamma-rays (0-40 kR), EMS (0-0.4M) and SA (0-4mM) and then inoculated in N6 medium containing 0.5mg/l 2,4-D, 4mg/l NAA and 2mg/l 6-BA to induce callus.Growth and differentiation' of the callus treated rice seeds were observed in subsequent subcultures.Decreased callus induction percentages were observed in treatments with gamma-rays, EMS and SA, but the mutagenic treatments did not reduce the mean weight of primary calli except that the seeds were irradiated with gamma-rays.Moreover, up to 3-fold increase in callus growth rate was shown after treatments of the three mu-tagenic agents.Higher proportion of calli with green spots or small shoots was also seen in one cultivar treated with the three mutagens.Signif icant increase in plant regeneration capacity was found in 60-day-old calli derived from gamma-rays or EMS-treated rice seeds in one cultivar.The results suggested that the following dose or concentration can be used for explant treatment in indica rice before culture, gamma-rays.5-10kR, EMS.<0.2M and SA:2-4mM

  12. A culture system for the live analysis of successive developmental processes and the morphological control of mammalian vertebral cartilage

    OpenAIRE

    Aono, Yuichi; Hirai, Yohei

    2011-01-01

    The mesoderm-derived segmental somite differentiates into dermomyotome and sclerotome, the latter of which undergoes vertebrogenesis to spinal cartilage and ultimately to vertebral bones. However, analysis and manipulation of the developing mammalian vertebrae in the same embryo has been infeasible because of their placental-dependent embryogenesis. Here, we report a novel culture system of the mouse embryonic tailbud, by which the developmental processes of mammalian vertebral cartilage are ...

  13. The Influence of Explant Types and Orientation on in Vitro Culture

    Directory of Open Access Journals (Sweden)

    Kamnoon KANCHANAPOOM

    2011-08-01

    Full Text Available Inflorescence, apical and lateral buds of Musa balbisiana ‘Kluai Hin’ (BBB group were used to culture on MS medium supplemented with 22 μM BA and 15% (v/v coconut water. Comparison of bud orientation showed that the best response of in vitro shoot tip proliferation was obtained with abaxial surface of buds lying down i.e. one side touching the medium (tilt. Mass propagation of shoot tips was obtained when cultured buds on MS medium containing 44 μM BA. Rooting was achieved by transferring the isolated shoots to MS basal medium without growth regulators. Rooted plantlets were acclimatized and successfully established in soil.

  14. AUDITORY HAIR CELL EXPLANT CO-CULTURES PROMOTE THE DIFFERENTIATION OF STEM CELLS INTO BIPOLAR NEURONS

    OpenAIRE

    Coleman, B.; Fallon, J. B.; Gillespie, L.N.; Silva, M.G.; Shepherd, R.K.

    2006-01-01

    Auditory neurons, the target neurons of the cochlear implant, degenerate following a sensorineural hearing loss. The goal of this research is to direct the differentiation of embryonic stem cells (SCs) into bipolar auditory neurons that can be used to replace degenerating neurons in the deafened mammalian cochlea. Successful replacement of auditory neurons is likely to result in improved clinical outcomes for cochlear implant recipients. We examined two post-natal auditory co-culture models w...

  15. Effect of IAA on growth, organogenesis and RNA metabolism during the development of Cichorium intybus root explants cultured „in vitro"

    OpenAIRE

    E. Gwóźdź

    2015-01-01

    The effects of 3-indolylacetic acid (IAA) on growth, organogenesis, RNA content, RNase activity and MAK elution profiles during the development of chicory root explants cultured in vitro were investigated. It was found that the intensive callus growth in the presence of IAA was accompanied by an increase in the RNA content, with simultaneous decrease of RNase activity. Fractionation of RNA by MAK column chromatography showed that the high content of RNA in the callus under the influence of IA...

  16. Tumorigenic risk of human induced pluripotent stem cell explants cultured on mouse SNL76/7 feeder cells

    Energy Technology Data Exchange (ETDEWEB)

    Kamada, Mizuna; Mitsui, Youji, E-mail: y-mitsui8310@hb.tp1.jp; Kumazaki, Tsutomu; Kawahara, Yuta; Matsuo, Taira; Takahashi, Tomoko, E-mail: t-takahashi@kph.bunri-u.ac.jp

    2014-10-24

    Highlights: • hiPS cell explants formed malignant tumors when SNL76/7 feeder cells were used. • Multi type tumors developed by interaction of SNL76/7 feeder cells with hiPS cells. • Tumorigenic risk occurs by co-culture of hiPS cells with SNL76/7 feeder cells. - Abstract: The potential for tumor formation from transplanted human induced pluripotent stem cell (hiPSC) derivatives represents a high risk in their application to regenerative medicine. We examined the genetic origin and characteristics of tumors, that were formed when 13 hiPSC lines, established by ourselves, and 201B7 hiPSC from Kyoto University were transplanted into severe combined immune-deficient (SCID) mice. Though teratomas formed in 58% of mice, five angiosarcomas, one malignant solitary fibrous tumor and one undifferentiated pleomorphic sarcoma formed in the remaining mice. Three malignant cell lines were established from the tumors, which were derived from mitomycin C (MMC)-treated SNL76/7 (MMC-SNL) feeder cells, as tumor development from fusion cells between MMC-SNL and hiPSCs was negative by genetic analysis. While parent SNL76/7 cells produced malignant tumors, neither MMC-SNL nor MMC-treated mouse embryo fibroblast (MEF) produced malignant tumors. When MMC-SNL feeder cells were co-cultured with hiPSCs, growing cell lines were generated, that expressed genes similar to the parent SNL76/7 cells. Thus, hiPSCs grown on MMC-SNL feeder cells have a high risk of generating feeder-derived malignant tumors. The possible mechanism(s) of growth restoration and the formation of multiple tumor types are discussed with respect of the interactions between MMC-SNL and hiPSC.

  17. Tumorigenic risk of human induced pluripotent stem cell explants cultured on mouse SNL76/7 feeder cells

    International Nuclear Information System (INIS)

    Highlights: • hiPS cell explants formed malignant tumors when SNL76/7 feeder cells were used. • Multi type tumors developed by interaction of SNL76/7 feeder cells with hiPS cells. • Tumorigenic risk occurs by co-culture of hiPS cells with SNL76/7 feeder cells. - Abstract: The potential for tumor formation from transplanted human induced pluripotent stem cell (hiPSC) derivatives represents a high risk in their application to regenerative medicine. We examined the genetic origin and characteristics of tumors, that were formed when 13 hiPSC lines, established by ourselves, and 201B7 hiPSC from Kyoto University were transplanted into severe combined immune-deficient (SCID) mice. Though teratomas formed in 58% of mice, five angiosarcomas, one malignant solitary fibrous tumor and one undifferentiated pleomorphic sarcoma formed in the remaining mice. Three malignant cell lines were established from the tumors, which were derived from mitomycin C (MMC)-treated SNL76/7 (MMC-SNL) feeder cells, as tumor development from fusion cells between MMC-SNL and hiPSCs was negative by genetic analysis. While parent SNL76/7 cells produced malignant tumors, neither MMC-SNL nor MMC-treated mouse embryo fibroblast (MEF) produced malignant tumors. When MMC-SNL feeder cells were co-cultured with hiPSCs, growing cell lines were generated, that expressed genes similar to the parent SNL76/7 cells. Thus, hiPSCs grown on MMC-SNL feeder cells have a high risk of generating feeder-derived malignant tumors. The possible mechanism(s) of growth restoration and the formation of multiple tumor types are discussed with respect of the interactions between MMC-SNL and hiPSC

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

    DEFF Research Database (Denmark)

    Stoner, G.D.; Harris, C.C.; Autrup, Herman; Trump, B.F.; Kingsbury, E.W.; Myers, G.A.

    1978-01-01

    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...... types in the peripheral lung can metabolize BP. The major ethylacetate extractable metabolites of BP formed by peripheral lung were tetrols and trans-7,8-diol. The primary water-soluble metabolite released with arylsulfatase and beta-glucuronidase was 3-hydroxybenzo[alpha]pyrene....

  19. Toxic Effects of Lipid-Mediated Gene Transfer in Ventral Mesencephalic Explant Cultures

    DEFF Research Database (Denmark)

    Bauer, Matthias; Kristensen, Bjarne Winther; Meyer, Morten; Gasser, Thomas; Widmer, Hans Ruedi; Rasmussen, Jens Zimmer; Ueffing, Marius

    2006-01-01

    Adverse effects of cDNA and oligonucleotide delivery methods have not yet been systematically analyzed. We introduce a protocol to monitor toxic effects of two non-viral lipid-based gene delivery protocols using CNS primary tissue. Cell membrane damage was monitored by quantifying cellular uptake...... of propidium iodide and release of cytosolic lactate dehydrogenase to the culture medium. Using a liposomal transfection reagent, cell membrane damage was already seen 24 hr after transfection. Nestin-positive target cells, which were used as morphological correlate, were severely diminished in some...

  20. Induction of somatic embryogenesis in explants of shoot cultures established from adult Eucalyptus globulus and E. saligna × E. maidenii trees.

    Science.gov (United States)

    Corredoira, E; Ballester, A; Ibarra, M; Vieitez, A M

    2015-06-01

    A reproducible procedure for induction of somatic embryogenesis (SE) from adult trees of Eucalyptus globulus Labill. and the hybrid E. saligna Smith × E. maidenii has been developed for the first time. Somatic embryos were obtained from both shoot apex and leaf explants of all three genotypes evaluated, although embryogenic frequencies were significantly influenced by the species/genotype, auxin and explant type. Picloram was more efficient for somatic embryo induction than naphthaleneacetic acid (NAA), with the highest frequency of induction being obtained in Murashige and Skoog medium containing 40 µM picloram and 40 mg l(-1) gum Arabic, in which 64% of the shoot apex explants and 68.8% of the leaf explants yielded somatic embryos. The embryogenic response of the hybrid was higher than that of the E. globulus, especially when NAA was used. The cultures initiated on picloram-containing medium consisted of nodular embryogenic structures surrounded by a mucilaginous coating layer that emerged from a watery callus developed from the initial explants. Cotyledonary somatic embryos were differentiated after subculture of these nodular embryogenic structures on a medium lacking plant growth regulators. Histological analysis confirmed the bipolar organization of the somatic embryos, with shoot and root meristems and closed procambial tissue that bifurcated into small cotyledons. The root pole was more differentiated than the shoot pole, which appeared to be formed by a few meristematic layers. Maintenance of the embryogenic lines by secondary SE was attained by subculturing individual cotyledonary embryos or small clusters of globular and torpedo embryos on medium with 16.11 µM NAA at 4- to 5-week intervals. Somatic embryos converted into plantlets after being transferred to liquid germination medium although plant regeneration remained poor. PMID:25877768

  1. Targeting Bone Alleviates Osteoarthritis in Osteopenic Mice and Modulates Cartilage Catabolism

    Science.gov (United States)

    Funck-Brentano, Thomas; Lin, Hilène; Hay, Eric; Ah Kioon, Marie-Dominique; Schiltz, Corinne; Hannouche, Didier; Nizard, Rémy; Lioté, Frédéric; Orcel, Philippe; de Vernejoul, Marie-Christine; Cohen-Solal, Martine Esther

    2012-01-01

    Objective Subchondral bone modifications occur early in the development of osteoarthritis (OA). The level of bone resorption might impact cartilage remodeling. We therefore assessed the in vivo and in vitro effects of targeting bone resorption in OA and cartilage metabolism. Methods OA was induced by meniscectomy (MNX) in ovariectomized osteopenic mice (OP) treated with estradiol (E2), pamidronate (PAM), or phosphate buffered saline (PBS) for 6 weeks. We assessed the subchondral bone and cartilage structure and the expression of cartilage matrix proteases. To assess the involvement of bone soluble factors in cartilage metabolism, supernatant of human bone explants pre-treated with E2 or PAM were transferred to cartilage explants to assess proteoglycan release and aggrecan cleavage. OPG/RANKL mRNA expression was assessed in bone explants by real-time quantitative PCR. The role of osteoprotegerin (OPG) in the bone-cartilage crosstalk was tested using an OPG neutralizing antibody. Results Bone mineral density of OP mice and osteoclast number were restored by E2 and PAM (p<0.05). In OP mice, E2 and PAM decreased ADAMTS-4 and -5 expression, while only PAM markedly reduced OA compared to PBS (2.0±0.63 vs 5.2±0.95; p<0.05). OPG/RANKL mRNA was increased in human bone explants treated with both drugs (2.2–3.7-fold). Moreover, supernatants from bone explants cultured with E2 or PAM reduced aggrecan cleavage and cartilage proteoglycan release (73±8.0% and 80±22% of control, respectively, p<0.05). This effect was reversed with osteoprotegerin blockade. Conclusion The inhibition of bone resorption by pamidronate in osteopenic mice alleviates the histological OA score with a reduction in the expression of aggrecanases. Bone soluble factors, such as osteoprotegerin, impact the cartilage response to catabolic factors. This study further highlights the importance of subchondral bone in the regulation of joint cartilage damage in OA. PMID:22432033

  2. 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.

    2009-01-01

    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

  3. Experimental study on co-culture of ventral spinal cord explants and major pelvic ganglion explants%大鼠脊髓前角与盆神经节之间植块联合培养的实验研究

    Institute of Scientific and Technical Information of China (English)

    程时刚; 耿红琼; 张伊凡; 杨星海; 钟伟

    2013-01-01

    Objective;To co-culture the ventral spinal cord explants and major pelvic ganglion explants from rats and investigate the interaction between co-cultured explants. Methods:We had developed a system for long-term co-culturing of ventral spinal cord explants and major pelvic ganglia explants to observe the variance of neuron growth. Results ;Explanted tissues of both types survived well in co-culture. Spinal motor neurons (SMNs) emitted numerous outgrowing processes, some of which associated with neurons in explanted major pelvic ganglia. Conclusion: Some contacts of specific motor neuron-major pelvic ganglia neuron were observed apparently,suggesting that a functional interaction may develop between the spinal motor neurons and major pelvic ganglion neurons in vitro.%目的:联合培养大鼠脊髓前角与盆神经节(major pelvic ganglia,MPG)植块并初步探讨两者相互作用的可能性.方法:建立大鼠脊髓前角与盆神经节之间植块的联合培养体系,观察神经元的生长变化.结果:神经组织在体外联合培养时均生长良好,脊髓前角植块生长出的运动神经元(spinal motor neurons,SMNs)发出突起与盆神经节的神经元形成明显形态上的接触.结论:联合培养的不同系统的神经元能彼此形成明显接触,初步提示运动神经元与盆神经节神经元之间可能形成功能性的相互作用.

  4. Effects of anti-arthritic drugs on proteoglycan synthesis by equine cartilage.

    Science.gov (United States)

    Frean, S P; Cambridge, H; Lees, P

    2002-08-01

    The concentration-effect relationships of phenylbutazone, indomethacin, betamethasone, pentosan polysulphate (PPS) and polysulphated glycosaminoglycan (PSGAG), on proteoglycan synthesis by equine cultured chondrocytes grown in monolayers, and articular cartilage explants were measured. The effect of PSGAG on interleukin-1beta induced suppression of proteogycan synthesis was also investigated. Proteoglycan synthesis was measured by scintillation assay of radiolabelled sulphate (35SO4) incorporation. Polysulphated glycosaminoglycan and PPS stimulated proteoglycan synthesis in chondrocyte monolayers in a concentration-related manner with maximal effects being achieved at a concentration of 10 microg/mL. Polysulphated glycosaminoglycan reversed the concentration-related suppression of proteoglycan synthesis induced by interleukin-1beta. Neither PSGAG nor PPS exerted significant effects on radiolabel incorporation in cartilage explants. Betamethasone suppressed proteoglycan synthesis by both chondrocytes and explants at high concentrations (0.1-100 microg/mL), but the effect was not concentration-related. At low concentrations (0.001-0.05 microg/mL) betamethasone neither increased nor decreased proteoglycan synthesis. Phenylbutazone and indomethacin increased radiolabel incorporation in chondrocyte cultures but not in cartilage explants at low (0.1, 1 and 10 microg/mL), but not at high (20 and 100 microg/mL) concentrations. These findings may be relevant to the clinical use of these drugs in the treatment of equine disease. PMID:12213118

  5. Fumonisin B1 (FB1) Induces Lamellar Separation and Alters Sphingolipid Metabolism of In Vitro Cultured Hoof Explants

    Science.gov (United States)

    Reisinger, Nicole; Dohnal, Ilse; Nagl, Veronika; Schaumberger, Simone; Schatzmayr, Gerd; Mayer, Elisabeth

    2016-01-01

    One of the most important hoof diseases is laminitis. Yet, the pathology of laminitis is not fully understood. Different bacterial toxins, e.g. endotoxins or exotoxins, seem to play an important role. Additionally, ingestion of mycotoxins, toxic secondary metabolites of fungi, might contribute to the onset of laminitis. In this respect, fumonsins are of special interest since horses are regarded as species most susceptible to this group of mycotoxins. The aim of our study was to investigate the influence of fumonisin B1 (FB1) on primary isolated epidermal and dermal hoof cells, as well as on the lamellar tissue integrity and sphingolipid metabolism of hoof explants in vitro. There was no effect of FB1 at any concentration on dermal or epidermal cells. However, FB1 significantly reduced the separation force of explants after 24 h of incubation. The Sa/So ratio was significantly increased in supernatants of explants incubated with FB1 (2.5–10 µg/mL) after 24 h. Observed effects on Sa/So ratio were linked to significantly increased sphinganine concentrations. Our study showed that FB1 impairs the sphingolipid metabolism of explants and reduces lamellar integrity at non-cytotoxic concentrations. FB1 might, therefore, affect hoof health. Further in vitro and in vivo studies are necessary to elucidate the effects of FB1 on the equine hoof in more detail. PMID:27023602

  6. Cartilage Tissue Engineering: the effect of different biomaterials, cell types and culture methods

    NARCIS (Netherlands)

    W.J.C.M. Marijnissen (Willem)

    2006-01-01

    textabstractChapter 1 outlines the normal structure and composition of articular cartilage and the inefficient spontaneous healing response after focal damage. Current surgical treatment options are briefly discussed and tissue engineering techniques for the repair of articular cartilage defects

  7. Curcumin reduces prostaglandin E2, matrix metalloproteinase-3 and proteoglycan release in the secretome of interleukin 1β-treated articular cartilage [v1; ref status: indexed, http://f1000r.es/1cl

    Directory of Open Access Journals (Sweden)

    Abigail L Clutterbuck

    2013-07-01

    Full Text Available Objective: Curcumin (diferuloylmethane is a phytochemical with potent anti-inflammatory and anti-oxidant properties, and has therapeutic potential for the treatment of a range of inflammatory diseases, including osteoarthritis (OA. The aim of this study was to determine whether non-toxic concentrations of curcumin can reduce interleukin-1beta (IL-1β-stimulated inflammation and catabolism in an explant model of cartilage inflammation. Methods: Articular cartilage explants and primary chondrocytes were obtained from equine metacarpophalangeal joints. Curcumin was added to monolayer cultured primary chondrocytes and cartilage explants in concentrations ranging from 3μM-100μM. Prostaglandin E2 (PGE2 and matrix metalloproteinase (MMP-3 release into the secretome of IL-1β-stimulated explants was measured using a competitive ELISA and western blotting respectively. Proteoglycan (PG release in the secretome was measured using the 1,9-dimethylmethylene blue (DMMB assay. Cytotoxicity was assessed with a live/dead assay in monolayer cultures after 24 hours, 48 hours and five days, and in explants after five days. Results: Curcumin induced chondrocyte death in primary cultures (50μM p<0.001 and 100μM p<0.001 after 24 hours. After 48 hours and five days, curcumin (≥25μM significantly increased cell death (p<0.001 both time points. In explants, curcumin toxicity was not observed at concentrations up to and including 25μM after five days. Curcumin (≥3μM significantly reduced IL-1β-stimulated PG (p<0.05 and PGE2 release (p<0.001 from explants, whilst curcumin (≥12μM significantly reduced MMP-3 release (p<0.01. Conclusion: Non-cytotoxic concentrations of curcumin exert anti-catabolic and anti-inflammatory effects in cartilage explants.

  8. Curcumin reduces prostaglandin E2, matrix metalloproteinase-3 and proteoglycan release in the secretome of interleukin 1β-treated articular cartilage [v2; ref status: indexed, http://f1000r.es/1ks

    Directory of Open Access Journals (Sweden)

    Abigail L Clutterbuck

    2013-08-01

    Full Text Available Objective: Curcumin (diferuloylmethane is a phytochemical with potent anti-inflammatory and anti-oxidant properties, and has therapeutic potential for the treatment of a range of inflammatory diseases, including osteoarthritis (OA. The aim of this study was to determine whether non-toxic concentrations of curcumin can reduce interleukin-1beta (IL-1β-stimulated inflammation and catabolism in an explant model of cartilage inflammation. Methods: Articular cartilage explants and primary chondrocytes were obtained from equine metacarpophalangeal joints. Curcumin was added to monolayer cultured primary chondrocytes and cartilage explants in concentrations ranging from 3μM-100μM. Prostaglandin E2 (PGE2 and matrix metalloproteinase (MMP-3 release into the secretome of IL-1β-stimulated explants was measured using a competitive ELISA and western blotting respectively. Proteoglycan (PG release in the secretome was measured using the 1,9-dimethylmethylene blue (DMMB assay. Cytotoxicity was assessed with a live/dead assay in monolayer cultures after 24 hours, 48 hours and five days, and in explants after five days. Results: Curcumin induced chondrocyte death in primary cultures (50μM p<0.001 and 100μM p<0.001 after 24 hours. After 48 hours and five days, curcumin (≥25μM significantly increased cell death (p<0.001 both time points. In explants, curcumin toxicity was not observed at concentrations up to and including 25μM after five days. Curcumin (≥3μM significantly reduced IL-1β-stimulated PG (p<0.05 and PGE2 release (p<0.001 from explants, whilst curcumin (≥12μM significantly reduced MMP-3 release (p<0.01. Conclusion: Non-cytotoxic concentrations of curcumin exert anti-catabolic and anti-inflammatory effects in cartilage explants.

  9. Effect of microcavitary alginate hydrogel with different pore sizes on chondrocyte culture for cartilage tissue engineering

    International Nuclear Information System (INIS)

    In our previous work, a novel microcavitary hydrogel was proven to be effective for proliferation of chondrocytes and maintenance of chondrocytic phenotype. In present work, we further investigated whether the size of microcavity would affect the growth and the function of chondrocytes. By changing the stirring rate, gelatin microspheres in different sizes including small size (80–120 μm), middle size (150–200 μm) and large size (250–300 μm) were prepared. And then porcine chondrocytes were encapsulated into alginate hydrogel with various sizes of gelatin microspheres. Cell Counting Kit-8 (CCK-8), Live/dead staining and real-time PCR were used to analyze the effect of the pore size on cell proliferation and expression of specific chondrocytic genes. According to all the data, cells cultivated in microcavitary hydrogel, especially in small size, had preferable abilities of proliferation and higher expression of cartilaginous markers including type II collagen, aggrecan and cartilage oligomeric matrix protein (COMP). Furthermore, it was shown by western blot assay that the culture of chondrocytes in microcavitary hydrogel could improve the proliferation of cells potentially by inducing the Erk1/2-MAPK pathway. Taken together, this study demonstrated that chondrocytes favored microcavitary alginate hydrogel with pore size within the range of 80–120 μm for better growth and ECM synthesis, in which Erk1/2 pathway was involved. This culture system would be promising for cartilage tissue engineering. - Highlights: • A novel model with microcavitary structure was set up to study the interaction between cells and materials. • Microcavitary alginate hydrogel could enhance the proliferation of chondrocytes and promote the expression of cartilaginous genes as compared with plain alginate hydrogel. • Cells in microcavitary alginate hydrogel with pore size within the range of 80–120 μm were capable of better growth and ECM synthesis

  10. Effect of microcavitary alginate hydrogel with different pore sizes on chondrocyte culture for cartilage tissue engineering

    Energy Technology Data Exchange (ETDEWEB)

    Zeng, Lei; Yao, Yongchang [School of Materials Science and Engineering, South China University of Technology, Guangzhou 510641 (China); National Engineering Research Center for Tissue Restoration and Reconstruction, Guangzhou 510006 (China); Wang, Dong-an, E-mail: DAWang@ntu.edu.sg [National Engineering Research Center for Tissue Restoration and Reconstruction, Guangzhou 510006 (China); Division of Bioengineering, School of Chemical and Biomedical Engineering, Nanyang Technological University, Singapore 637457 (Singapore); Chen, Xiaofeng, E-mail: chenxf@scut.edu.cn [School of Materials Science and Engineering, South China University of Technology, Guangzhou 510641 (China); National Engineering Research Center for Tissue Restoration and Reconstruction, Guangzhou 510006 (China)

    2014-01-01

    In our previous work, a novel microcavitary hydrogel was proven to be effective for proliferation of chondrocytes and maintenance of chondrocytic phenotype. In present work, we further investigated whether the size of microcavity would affect the growth and the function of chondrocytes. By changing the stirring rate, gelatin microspheres in different sizes including small size (80–120 μm), middle size (150–200 μm) and large size (250–300 μm) were prepared. And then porcine chondrocytes were encapsulated into alginate hydrogel with various sizes of gelatin microspheres. Cell Counting Kit-8 (CCK-8), Live/dead staining and real-time PCR were used to analyze the effect of the pore size on cell proliferation and expression of specific chondrocytic genes. According to all the data, cells cultivated in microcavitary hydrogel, especially in small size, had preferable abilities of proliferation and higher expression of cartilaginous markers including type II collagen, aggrecan and cartilage oligomeric matrix protein (COMP). Furthermore, it was shown by western blot assay that the culture of chondrocytes in microcavitary hydrogel could improve the proliferation of cells potentially by inducing the Erk1/2-MAPK pathway. Taken together, this study demonstrated that chondrocytes favored microcavitary alginate hydrogel with pore size within the range of 80–120 μm for better growth and ECM synthesis, in which Erk1/2 pathway was involved. This culture system would be promising for cartilage tissue engineering. - Highlights: • A novel model with microcavitary structure was set up to study the interaction between cells and materials. • Microcavitary alginate hydrogel could enhance the proliferation of chondrocytes and promote the expression of cartilaginous genes as compared with plain alginate hydrogel. • Cells in microcavitary alginate hydrogel with pore size within the range of 80–120 μm were capable of better growth and ECM synthesis.

  11. Direct regeneration of Periwinkle (Catharanthus roseus) via node explants culture and different combinations of plant growth regulators

    OpenAIRE

    M. Talebi; F Etesam; B.E. Sayed-Tabatabaei; Gh. Khaksar

    2012-01-01

    Periwinkle (Catharanthus roseus L., Apocynaceae) contains more than 130 different terpenoid indole alkaloids (TIAs), of which two dimeric alkaloids, Vinblastine and Vincristine, have antineoplastic activity and are useful in treatment of various cancers. Specific production of some alkaloids in differentiated tissues such as leaf and stem led to use direct regeneration of explants in order to increase the production of these important alkaloids in the plant. In this research, 30 combinations ...

  12. Effect of Different Culture Conditions on Browning Rate of Explants in Tissue Culture of Tea Trees%降低茶树组织培养中外植体褐化程度的研究

    Institute of Scientific and Technical Information of China (English)

    黄燕芬; 周国兰; 赵华富

    2009-01-01

    为降低茶树组培过程中外植体褐化程度,从外植体表面消毒时间、切割方式、转瓶间隔时间、培养基类型、激素配比、抗褐化剂和吸附剂的使用方面进行了综合试验.结果表明:表面消毒最适时间为7~8 min;接种外植体带腋芽叶片保留2/5,3 d转瓶1次降低褐化的效果较好; 经此处理的外植体培养在1/2MS +BA 2 mg/L+NAA 0.1 mg/L+Vc 1.0 g/L+AC 1.0 g/L的培养基中,生长良好,褐化率由86 %下降为41 %.%The sterilized time and cutting pattern of explants, interval time for transferring another bottle, medium types, different hormone combination, anti-browning agents and absorption agents were studied to reduce the browning rate of explants in tissue culture of tea tree. The results showed that the browning rate could reduce from 86 % to 41 % when the explants with 2/5 leaf and a axillary bud were sterilized by 0.1 % mercury solution for 7-8 min, the explants were transferred into another bottle every 3d and the treated explants were cultured on 1/2MS +BA 2 mg/L+NAA 0.1 mg/L+Vc 1.0 g/L+AC 1.0 g/L.

  13. Adrenergic Activation of Melatonin Secretion in Ovine Pineal Explants in Short-Term Superfusion Culture Occurs via Protein Synthesis Independent and Dependent Phenomena

    Directory of Open Access Journals (Sweden)

    Bogdan Lewczuk

    2014-01-01

    Full Text Available The ovine pineal is generally considered as an interesting model for the study on adrenergic regulation of melatonin secretion due to some functional similarities with this gland in the human. The present investigations, performed in the superfusion culture of pineal explants, demonstrated that the norepinephrine-induced elevation of melatonin secretion in ovine pinealocytes comprised of two subsequent periods: a rapid increase phase and a slow increase phase. The first one included the quick rise in release of N-acetylserotonin and melatonin, occurring parallel to elevation of NE concentration in the medium surrounding explants. This rapid increase phase was not affected by inhibition of translation. The second, slow increase phase began after NE level had reached the maximum concentration in the culture medium and lasted about two hours. It was completely abolished by the treatment with translation inhibitors. The obtained results showed for the first time that the regulation of N-acetylserotonin synthesis in pinealocytes of some species like the sheep involves the on/off mechanism, which is completely independent of protein synthesis and works very fast. They provided strong evidence pointing to the need of revision of the current opinion that arylalkylamines N-acetyltransferase activity in pinealocytes is controlled exclusively by changes in enzyme abundance.

  14. Co-culture of placental explants with isolated CD4 and CD8 T cells: a functional model to define the consequences of placental inflammation

    Directory of Open Access Journals (Sweden)

    Hayley Derricott

    2015-11-01

    Full Text Available Appropriate placental function is essential for successful pregnancy and placental dysfunction is associated with fetal growth restriction (FGR and stillbirth. Villitis of unknown etiology (VUE and chronic intervillositis of unknown etiology (CIUE are immune-mediated conditions characterised by placental infiltrates of macrophages, CD4 and CD8 T cells. VUE and CIUE occur more frequently in the placentas of pregnancies complicated by FGR. The mechanisms by which this inflammation induces placental dysfunction are yet to be defined. We aimed to develop an in vitro model of placental inflammation to investigate functional consequences of immune cells in the placental environment. Fragments of placental tissue were co-cultured with CD4 and CD8 T cells isolated from whole blood. CellTrackerTM fluorescence was used to identify T cells in cultured explants. Tissue histology, endocrine and nutrient transport function was assessed using established methods. This novel preparation will enable future investigations into immune cell interactions with placenta.

  15. The effect of radiation on the growth of normal and malignant human oesophageal explant cultures pre-treated with bleomycin

    International Nuclear Information System (INIS)

    A method has been developed for testing the response of oesophageal explants from tumour and surrounding normal tissue in the same patient to chemotherapy and γ-radiation, singly and in combination. The test allows treatment combinations, time and order of administration of agents to the tissue to be accurately controlled. Cytotoxicity, determined by measuring the area of outgrowth from an explant 2 weeks after plating, is the most useful short-term end-point, although many other are possible. Results showing differential cytotoxicity of belomycin with and without radiation in squamous and adenocarcinoma of the oesophagus and surrounding normal tissue from the same patient indicate tumour cells are relatively resistant to radiation alone, low levels of belomycin with or without radiation preferentially spare tumour cells and high levels, in combination with any radiation dose tested, but not without radiation, spare normal cells and give a significantly high amount of relative tumour cell kill. Belomycin must be added to the cells just before or just after irradiation to obtain the normal-tissue sparing effect. (author)

  16. Effect of radiation on the growth of normal and malignant human oesophageal explant cultures pre-treated with bleomycin

    Energy Technology Data Exchange (ETDEWEB)

    Seymour, C.B.; Cusack, A.; Mothersill, C.; Hennessy, T.P.

    1988-05-01

    A method has been developed for testing the response of oesophageal explants from tumour and surrounding normal tissue in the same patient to chemotherapy and ..gamma..-radiation, singly and in combination. The test allows treatment combinations, time and order of administration of agents to the tissue to be accurately controlled. Cytotoxicity, determined by measuring the area of outgrowth from an explant 2 weeks after plating, is the most useful short-term end-point, although many other are possible. Results showing differential cytotoxicity of belomycin with and without radiation in squamous and adenocarcinoma of the oesophagus and surrounding normal tissue from the same patient indicate tumour cells are relatively resistant to radiation alone, low levels of belomycin with or without radiation preferentially spare tumour cells and high levels, in combination with any radiation dose tested, but not without radiation, spare normal cells and give a significantly high amount of relative tumour cell kill. Belomycin must be added to the cells just before or just after irradiation to obtain the normal-tissue sparing effect.

  17. Multiplicação in vitro do porta-enxerto de macieira cv. Marubakaido: efeito da orientação do explante no meio de cultura In vitro multiplication of the apple rootstock cv. Marubakaido: effect of the orientation of explant in the medium of culture

    Directory of Open Access Journals (Sweden)

    ALAN CRISTIANO ERIG

    2002-08-01

    Full Text Available Objetivou-se avaliar o efeito da orientação do explante, vertical ou horizontal, no meio de cultura, na multiplicação in vitro, do porta-enxerto de macieira cv. Marubakaido. O meio de cultura utilizado foi o MS com N (nitrogênio reduzido a ¾ da concentração original, 100mg.L-1 de mio-inositol, 40g.L-1 de sacarose e 6g.L-1 de ágar, suplementado com 4,44mM de BAP (6-benzilaminopurina e 0,2ml.L-1 de PPM TM ("Plant Preservative Mixture". Segmentos caulinares com duas gemas e o ápice excisado foram utilizados como explantes. Após a inoculação, os frascos com os explantes foram incubados a 16 horas de fotoperíodo, à temperatura de 25±2ºC, com radiação de 25µmoles.m-2.s-1. O número de brotações, o número de gemas por explante, a taxa de multiplicação e a altura da brotação maior foram avaliados aos quarenta dias de cultivo. O maior número de brotações, o maior número de gemas e a maior taxa de multiplicação foram obtidos com o explante na orientação horizontal no meio de cultura. Não houve diferença significativa quanto à orientação vertical e horizontal do explante no meio de cultura para a altura da brotação maior.The aim of this study was evaluate the effect of the vertical and the horizontal orientation of the explant in the culture medium, in the in vitro multiplication, for the apple rootstock cv. Marubakaido. The culture medium used was the MS with N reduced to ¾ of the original concentration, myo-inositol (100mg.L-1, sucrose (40g.L-1 and agar (6g.L-1, suplemented with BAP (4.44mM and PPM TM (0.2ml.L-1. Stem segments with two buds and the apex excised were used as explants. After the inoculation, the flasks with the explants were incubated at 16 hour of photoperiod, 25±2ºC temperature, with irradiation of 25µmoles.m-2.s-1. The number of shoots and buds, the rate of multiplication and the height of the larger shoot were evaluated after 40 days of cultivation. The highests shoot number, number of buds

  18. Effect of IAA on growth, organogenesis and RNA metabolism during the development of Cichorium intybus root explants cultured „in vitro"

    Directory of Open Access Journals (Sweden)

    E. Gwóźdź

    2015-05-01

    Full Text Available The effects of 3-indolylacetic acid (IAA on growth, organogenesis, RNA content, RNase activity and MAK elution profiles during the development of chicory root explants cultured in vitro were investigated. It was found that the intensive callus growth in the presence of IAA was accompanied by an increase in the RNA content, with simultaneous decrease of RNase activity. Fractionation of RNA by MAK column chromatography showed that the high content of RNA in the callus under the influence of IAA was due to an increased accumulation of the ribosomal fraction of RNA mainly. Experiments with actinomycin D demonstrated that this antibiotic abolished both the auxin-induced callus growth and the inhibitory effect of IAA on bud formation. No significant inhibition of spontaneous bud formation under the influence of actinomycin D was observed. The possible relationship between the IAA-affected morphogenetic processes and RNA metabolism is discussed.

  19. A new in vitro screening bioassay for the ecotoxicological evaluation of the estrogenic responses of environmental chemicals using roach (Rutilus rutilus) liver explant culture.

    Science.gov (United States)

    Gerbron, Marie; Geraudie, Perrine; Rotchell, Jeanette; Minier, Christophe

    2010-10-01

    There is growing evidence that many chemicals released in the environment are able to disturb the normal endocrinology of organisms affecting the structure and function of their reproductive system. This has prompted the scientific community to develop appropriate testing methods to identify active compounds and elucidate mechanisms of action. Of particular interest are in vitro screening methods that can document the effects of these endocrine disrupting compounds on fish. In this study, an in vitro bioassay was developed in the roach (Rutilus rutilus) for evaluating the estrogenicity or antiestrogenicity potency of environmental pollutants by measuring vitellogenin (VTG) induction in cultured liver explants. The cell viability was assessed by the measurement of nonspecific esterase activity using a fluorescein diacetate hydrolysis assay. Results showed that explants could be cultured for 72 h without any significant loss of activity. Dose-dependent responses have been measured with estrogenic model compounds such as 17-β-estradiol (E2) and 17-α-ethynylestradiol (EE2) or antiestrogenic compounds such as tamoxifen. Lowest observable effective concentrations were 1 nM for E2, 1 nM for EE2, and 100 nM for tamoxifen, showing a good sensitivity of the test system. Estrogenicity of butyl 4-hydroxybenzoate, 4-nonylphenol, and bisphenol A was tested. bisphenol A (100 μM) or butylparaben induced a twofold increase in VTG production when compared with 100 nM E2, whereas this production was only 20% with 100 μM 4-nonylphenol. Overall, this study shows that the bioassay could provide valuable information on endocrine disrupting chemicals including metabolites and mixtures of compounds. PMID:20549626

  20. Stimulation by concanavalin A of cartilage-matrix proteoglycan synthesis in chondrocyte cultures

    Energy Technology Data Exchange (ETDEWEB)

    Yan, W.Q.; Nakashima, K.; Iwamoto, M.; Kato, Y. (Osaka Univ. (Japan))

    1990-06-15

    The effect of concanavalin A on proteoglycan synthesis by rabbit costal and articular chondrocytes was examined. Chondrocytes were seeded at low density and grown to confluency in medium supplemented with 10% fetal bovine serum, and then the serum concentration was reduced to 0.3%. At the low serum concentration, chondrocytes adopted a fibroblastic morphology. Addition of concanavalin A to the culture medium induced a morphologic alteration of the fibroblastic cells to spherical chondrocytes and increased by 3- to 4-fold incorporation of (35S)sulfate and (3H)glucosamine into large chondroitin sulfate proteoglycan that was characteristically found in cartilage. The stimulation of incorporation of labeled precursors reflected real increases in proteoglycan synthesis, as chemical analyses showed a 4-fold increase in the accumulation of macromolecules containing hexuronic acid in concanavalin A-maintained cultures. Furthermore, the effect of concanavalin A on (35S)sulfate incorporation into proteoglycans was greater than that of various growth factors or hormones. However, concanavalin A had smaller effects on (35S)sulfate incorporation into small proteoglycans and (3H)glucosamine incorporation into hyaluronic acid and chondroitinase AC-resistant glycosaminoglycans. Since other lectins tested, such as wheat germ agglutinin, lentil lectin, and phytohemagglutinin, had little effect on (35S)sulfate incorporation into proteoglycans, the concanavalin A action on chondrocytes seems specific. Although concanavalin A decreased (3H)thymidine incorporation in chondrocytes, the stimulation of proteoglycan synthesis could be observed in chondrocytes exposed to the inhibitor of DNA synthesis, cytosine arabinoside. These results indicate that concanavalin A is a potent modulator of proteoglycan synthesis by chondrocytes.

  1. 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

    2009-11-01

    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.

  2. Prevention of the disrupted enamel phenotype in Slc4a4-null mice using explant organ culture maintained in a living host kidney capsule.

    Directory of Open Access Journals (Sweden)

    Xin Wen

    Full Text Available Slc4a4-null mice are a model of proximal renal tubular acidosis (pRTA. Slc4a4 encodes the electrogenic sodium base transporter NBCe1 that is involved in transcellular base transport and pH regulation during amelogenesis. Patients with mutations in the SLC4A4 gene and Slc4a4-null mice present with dysplastic enamel, amongst other pathologies. Loss of NBCe1 function leads to local abnormalities in enamel matrix pH regulation. Loss of NBCe1 function also results in systemic acidemic blood pH. Whether local changes in enamel pH and/or a decrease in systemic pH are the cause of the abnormal enamel phenotype is currently unknown. In the present study we addressed this question by explanting fetal wild-type and Slc4a4-null mandibles into healthy host kidney capsules to study enamel formation in the absence of systemic acidemia. Mandibular E11.5 explants from NBCe1-/- mice, maintained in host kidney capsules for 70 days, resulted in teeth with enamel and dentin with morphological and mineralization properties similar to cultured NBCe1+/+ mandibles grown under identical conditions. Ameloblasts express a number of proteins involved in dynamic changes in H+/base transport during amelogenesis. Despite the capacity of ameloblasts to dynamically modulate the local pH of the enamel matrix, at least in the NBCe1-/- mice, the systemic pH also appears to contribute to the enamel phenotype. Extrapolating these data to humans, our findings suggest that in patients with NBCe1 mutations, correction of the systemic metabolic acidosis at a sufficiently early time point may lead to amelioration of enamel abnormalities.

  3. An exploration of the ability of tepoxalin to ameliorate the degradation of articular cartilage in a canine in vitro model

    Directory of Open Access Journals (Sweden)

    Clegg Peter D

    2009-07-01

    Full Text Available Abstract Background To study the ability of tepoxalin, a dual inhibitor of cyclooxygenase (COX and lipoxygenase (LOX and its active metabolite to reduce the catabolic response of cartilage to cytokine stimulation in an in vitro model of canine osteoarthritis (OA. Grossly normal cartilage was collected post-mortem from seven dogs that had no evidence of joint disease. Cartilage explants were cultured in media containing the recombinant canine interleukin-1β (IL-1β at 100 ng/ml and recombinant human oncostatin-M (OSM at 50 ng/ml. The effects of tepoxalin and its metabolite were studied at three concentrations (1 × 10-5, 1 × 10-6 and 1 × 10-7 M. Total glycosaminoglycan (GAG and collagen (hydroxyproline release from cartilage explants were used as outcome measures of proteoglycan and collagen depletion respectively. PGE2 and LTB4 assays were performed to study the effects of the drug on COX and LOX activity. Results Treatment with IL-1β and OSM significantly upregulated both collagen (p = 0.004 and proteoglycan (p = 0.001 release from the explants. Tepoxalin at 10-5 M and 10-6 M caused a decrease in collagen release from the explants (p = 0.047 and p = 0.075. Drug treatment showed no effect on GAG release. PGE2 concentration in culture media at day 7 was significantly increased by IL-1β and OSM and treatment with both tepoxalin and its metabolite showed a trend towards dose-dependent reduction of PGE2 production. LTB4 concentrations were too low to be quantified. Cytotoxicity assays suggested that neither tepoxalin nor its metabolite had a toxic effect on the cartilage chondrocytes at the concentrations and used in this study. Conclusion This study provides evidence that tepoxalin exerts inhibition of COX and can reduce in vitro collagen loss from canine cartilage explants at a concentration of 10-5 M. We can conclude that, in this model, tepoxalin can partially inhibit the development of cartilage degeneration when it is available locally to

  4. Transforming growth factor-beta 1 stimulates synthesis of proteoglycan aggregates in calf articular cartilage organ cultures

    Energy Technology Data Exchange (ETDEWEB)

    Morales, T.I. (Bone Research Branch, National Institute of Dental Research, National Institutes of Health, Bethesda, MD (United States))

    1991-04-01

    Previous work showed that transforming growth factor-beta 1 (TGF-beta 1), added alone to bovine cartilage organ cultures, stimulated (35S)sulfate incorporation into macromolecular material but did not investigate the fidelity of the stimulated system to maintain synthesis of cartilage-type proteoglycans. This paper provides evidence that chondrocytes synthesize the appropriate proteoglycan matrix under TGF-beta 1 stimulation: (1) there is a coordinated increase in hyaluronic acid and proteoglycan monomer synthesis, (2) link-stable proteoglycan aggregates are assembled, (3) the hybrid chondroitin sulfate/keratan sulfate monomeric species is synthesized, and (4) there is an increase in protein core synthesis. Some variation in glycosylation patterns was observed when proteoglycans synthesized under TGF-beta 1 stimulation were compared to those synthesized under basal conditions. Thus comparing TGF-beta 1 to basal samples respectively, the monomers were larger (Kav on Sepharose CL-2B = 0.29 vs 0.41), the chondroitin sulfate chains were longer by approximately 3.5 kDa, the percentage of total glycosaminoglycan in keratan sulfate increased slightly from approximately 4% (basal) to approximately 6%, and the unsulfated disaccharide decreased from 28% (basal) to 12%. All of these variations are in the direction of a more anionic proteoglycan. Since the ability of proteoglycans to confer resiliency to the cartilage matrix is directly related to their anionic nature, these changes would presumably have a beneficial effect on tissue function.

  5. Lubricin reduces cartilage--cartilage integration.

    Science.gov (United States)

    Schaefer, Dirk B; Wendt, David; Moretti, Matteo; Jakob, Marcel; Jay, Gregory D; Heberer, Michael; Martin, Ivan

    2004-01-01

    Cartilage integration in vivo does not occur, such that even cartilage fissures do not heal. This could be due not only to the limited access of chondrocytes to the wound, but also to exogenous factors. In this paper, we tested the hypothesis that lubricin, a lubricating protein physiologically present in the synovial fluid, reduces the integrative cartilage repair capacity. Disk/ring composites of bovine articular cartilage were prepared using concentric circular blades and cultured for 6 weeks with or without treatment with 250 microg/ml lubricin applied three times per week. Following culture, the percentage of contact area between the disks and the rings, as assessed by light microscopy, were equal in both groups. The adhesive strength of the integration interface, as assessed by push-out mechanical tests, was markedly and significantly lower in lubricin-treated specimens (2.5 kPa) than in the controls (28.7 kPa). Histological observation of Safranin-O stained cross-sections confirmed the reduced integration in the lubricin treated composites. Our findings suggest that the synovial milieu, by providing lubrication of cartilage surfaces, impairs cartilage--cartilage integration. PMID:15299281

  6. IN VITRO CULTURE OF PETIOLE LONGITUDINAL THIN CELL LAYER EXPLANTS OF VIETNAMESE GINSENG ( PANAX VIETNAMENSIS HA ET GRUSHV. AND PRELIMINARY ANALYSIS OF SAPONIN CONTENT

    Directory of Open Access Journals (Sweden)

    Duong Tan Nhut

    2012-08-01

    Full Text Available The present work describes a procedure that allows for the easy and rapid induction of somaticembryos, calli, shoots and adventitious roots of Vietnamese ginseng (Panax vietnamensis Ha et Grushv. fromlongitudinalthin cell layers (lTCLs. In order to investigate the morphogenesis of this medicinal plant, the effect ofseparately–supplemented plant growth regulators and combinatorial effect of co–supplemented auxins and cytokininsin dark or under 16-hour photoperiod was examined. After eight weeks of culture, the lTCL explants excised frompetiole of three-month-old in vitro plants and cultured on a semi solid basal Murashige and Skoog (MS mediasupplemented with 1.0 mg/l 2,4-dichlorophenoxyacetic acid(2,4-Dand 0.1 mg/l thidiazuron(TDZ in dark, 2.0 mg/lα-napththaleneacetic acid (NAA in dark and 1.0 mg/l 2,4-Dunder lightgave the highest rate of callogenesis (100%,embryogenesis (53.3% and adventitious root formation (100% with a mean of 16.7 roots, and shoot formation(26.7%, respectively. The metabolite of petiole lTCL-derived calli qualitative and quantitative analyses wereperformed by using high-performance liquid chromatography and thin layer chromatography. The simple procedure,together with similar saponin profiles between the resulted in vitrotissues and plants grown in nature, suggest itspotential use in generating Vietnamese ginseng in large amount for medicinal purpose.

  7. In vitro flowering of shoots regenerated from cultured nodal explants of Spilanthes acmella Murr. - an ornamental cum medicinal herb

    OpenAIRE

    Kuldeep YADAV; Singh, Narender

    2011-01-01

    An efficient protocol for in vitro flowering of Spilanthes acmella Murr., a medicinally valuable plant, has been developed. Multiple shoot formation of up to 4 shoots was obtained on Murashige and Skoog (MS) medium supplemented with BAP (1.0 mg/l). Regenerated shoots were subcultured cultured on MS medium containing various concentrations of BAP alone or in combination with IAA. Presence of BAP in the culture medium was observed to be absolutely essential for induction of flower. Maximum per...

  8. Pre-metatarsal skeletal development in tissue culture at unit- and microgravity

    Science.gov (United States)

    Klement, B. J.; Spooner, B. S.

    1994-01-01

    Explant organ culture was used to demonstrate that isolated embryonic mouse pre-metatarsal mesenchyme is capable of undergoing a series of differentiative and morphogenetic developmental events. Mesenchyme differentiation into chondrocytes, and concurrent morphogenetic patterning of the cartilage tissue, and terminal chondrocyte differentiation with subsequent matrix mineralization show that cultured tissue closely parallels in vivo development. Whole mount alizarin red staining of the cultured tissue demonstrates that the extracellular matrix around the hypertrophied chondrocytes is competent to support mineralization. Intensely stained mineralized bands are similar to those formed in pre-metatarsals developing in vivo. We have adapted the culture strategy for experimentation in a reduced gravity environment on the Space Shuttle. Spaceflight culture of pre-metatarsals, which have already initiated chondrogenesis and morphogenetic patterning, results in an increase in cartilage rod size and maintenance of rod shape, compared to controls. Older pre-metatarsal tissue, already terminally differentiated to hypertrophied cartilage, maintained rod structure and cartilage phenotype during spaceflight culture.

  9. 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; Karsdal, M A

    2009-01-01

    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...... explants were cultured in Dulbecco's modified Eagle's medium (DMEM):F12 in the presence of 0, 0.01, 0.1, 1, 10, or 100 ng/mL of IGF-I. The viability of the chondrocytes was measured by the colorimetric Alamar blue assay. Collagen formation was assessed from the conditioned medium by the PIINP assay....... Proteoglycan levels retained in the explants after 22 days of culture were extracted and measured by the sulfated glycosaminoglycan (sGAG) assay. RESULTS: In the absence of stimulation, PIINP markedly decreased as a function of time (99.4%, p < 0.001). IGF-I dose-dependently stimulated collagen formation and...

  10. In vitro flowering of shoots regenerated from cultured nodal explants of Spilanthes acmella Murr. - an ornamental cum medicinal herb

    Directory of Open Access Journals (Sweden)

    Kuldeep YADAV

    2011-05-01

    Full Text Available An efficient protocol for in vitro flowering of Spilanthes acmella Murr., a medicinally valuable plant, has been developed. Multiple shoot formation of up to 4 shoots was obtained on Murashige and Skoog (MS medium supplemented with BAP (1.0 mg/l. Regenerated shoots were subcultured cultured on MS medium containing various concentrations of BAP alone or in combination with IAA. Presence of BAP in the culture medium was observed to be absolutely essential for induction of flower. Maximum percentage (50 % of flower induction occurred when regenerated shoots were cultured on MS medium supplemented with BAP (2.0 mg/l + IAA (0.5 mg/l under photoperiod of 16/8 h (light/dark cycle.The 3-week intervals for three consecutive subcultures on this medium were efficient for flower induction. The regenerated shoots rooted best on 1/2 MS medium containing IBA (1.0 mg/l. Rooted plantlets were hardened and established in pots with 70% survival rate.

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

    International Nuclear Information System (INIS)

    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

  12. 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: DRHaudenschild@ucdavis.edu

    2015-05-08

    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.

  13. Meio de cultura e tipo de explante no estabelecimento in vitro de espécies de maracujazeiro Culture medium and type of explant in the in vitro establishment of passion fruit species

    OpenAIRE

    Gláucia Amorim Faria; Maria Angélica Pereira de Carvalho Costa; Carlos Alberto da Silva Ledo; Tatiana Góes Junghans; Antônio da Silva Souza; Mario Augusto Pinto da Cunha

    2007-01-01

    O Brasil é um dos principais centros de dispersão da variabilidade genética do gênero Passiflora. Sua auto-incompatibilidade aliada à incidência de doenças do sistema radicular e da parte aérea, desmatamentos e monocultivos promovem perda de material genético. Tendo-se em vista o risco de erosão genética, torna-se necessário a conservação da variabilidade em bancos de germoplasma, de grande interesse no melhoramento de plantas. Estudos em relação ao tipo de explante e concentração dos meios d...

  14. Engineering Cartilage

    Science.gov (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 ...

  15. Multiplicação e enraizamento in vitro de amoreira-preta 'Xavante': efeito da concentração de sais, do tipo de explante e de carvão ativado no meio de cultura Blackberry 'Xavante' in vitro multiplication and rooting: salt concentrations effect, explant type, and activated coal on culture medium

    Directory of Open Access Journals (Sweden)

    Luciane Nolasco Leitzke

    2009-01-01

    concentrations in culture medium for. The first experiment constituted of nodal segments with 1 cm, originated from of the in vitro cultivation, inoculated at four different MS medium concentrations, supplemented with 7.5 µM BAP, 30 g L-1 sucrose, 100 mg L-1 myo-inositol and 6 g L-1 agar, and with two types of explants, which totaled 8 treatments. The second experiment constituted of apex microcutting, having about 1 to 1.5 cm length, and two leaves were inoculated at three different concentrations of MS medium, added of 30 g L-1 sucrose, 100 mg L-1 myo-inositol and 6 g L-1 agar and supplemented with three different activated coal concentrations, totaling nine treatments. It can be inferred that T1, in other words, explants with leaves, were more efficient at 125% MS salt concentration on culture medium, thereby inducing larger number of leaves, buds, shoots and shoot length, and that the use of 75% salt concentration without activated coal is the most suitable one for the blackberry 'Xavante' in vitro rooting.

  16. PATTERN OF PLANT REGENERATION FROM SHOOT TIP EXPLANTS OF PIGEONPEA (CAJANUS CAJAN L MILLLSP) VAR LRG-41

    OpenAIRE

    T. Raghavendra; P. Sudhakar

    2014-01-01

    An efficient direct shoot bud differentiation and multiple shoot induction from shoot tip explants of pigeon pea (Cajanus cajan L.) has been achieved. The frequency of shoot bud regeneration was influenced by the type of explants, genotype and concentrations of cytokinin. Explants viz. shoot tip isolated from 10 day old seedlings showed better explants response Explants were cultured on Murashige and skoog (MS) medium augmented with different concentrations of BAP and NAA. Among the various c...

  17. Thidiazuron-induced high-frequency plant regeneration from leaf explants of Paulownia tomentosa mature trees

    OpenAIRE

    CORREDOIRA, E.; Ballester, A.; Viéitez Martín, Ana María

    2008-01-01

    Attempts were made to study the effect of thidiazuron (TDZ) on adventitious shoot induction and plant development in Paulownia tomentosa explants derived from mature trees. Media with different concentrations of TDZ in combination with an auxin were used to induce adventitious shoot-buds in two explant types: basal leaf halves with the petiole attached (leaf explant) and intact petioles. Optimal shoot regeneration was obtained in leaf explants cultured on induction medium containing TDZ (22.7...

  18. Prefabrication of 3D cartilage contructs: towards a tissue engineered auricle--a model tested in rabbits.

    Directory of Open Access Journals (Sweden)

    Achim von Bomhard

    Full Text Available The reconstruction of an auricle for congenital deformity or following trauma remains one of the greatest challenges in reconstructive surgery. Tissue-engineered (TE three-dimensional (3D cartilage constructs have proven to be a promising option, but problems remain with regard to cell vitality in large cell constructs. The supply of nutrients and oxygen is limited because cultured cartilage is not vascular integrated due to missing perichondrium. The consequence is necrosis and thus a loss of form stability. The micro-surgical implantation of an arteriovenous loop represents a reliable technology for neovascularization, and thus vascular integration, of three-dimensional (3D cultivated cell constructs. Auricular cartilage biopsies were obtained from 15 rabbits and seeded in 3D scaffolds made from polycaprolactone-based polyurethane in the shape and size of a human auricle. These cartilage cell constructs were implanted subcutaneously into a skin flap (15 × 8 cm and neovascularized by means of vascular loops implanted micro-surgically. They were then totally enhanced as 3D tissue and freely re-implanted in-situ through microsurgery. Neovascularization in the prefabricated flap and cultured cartilage construct was analyzed by microangiography. After explantation, the specimens were examined by histological and immunohistochemical methods. Cultivated 3D cartilage cell constructs with implanted vascular pedicle promoted the formation of engineered cartilaginous tissue within the scaffold in vivo. The auricles contained cartilage-specific extracellular matrix (ECM components, such as GAGs and collagen even in the center oft the constructs. In contrast, in cultivated 3D cartilage cell constructs without vascular pedicle, ECM distribution was only detectable on the surface compared to constructs with vascular pedicle. We demonstrated, that the 3D flaps could be freely transplanted. On a microangiographic level it was evident that all the skin flaps

  19. Xenopus laevis Keller Explants

    OpenAIRE

    sprotocols

    2014-01-01

    Authors: Hazel L. Sive, Robert M. Grainger and Richard M. Harland This protocol was adapted from “Microdissection,” Chapter 10, in [*Early Development of* *Xenopus laevis*](http://www.cshlpress.com/link/xenopus.htm) by Hazel L. Sive, Robert M. Grainger, and Richard M. Harland. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2000. ### INTRODUCTION The basic Keller explant is a rectangle of dorsal mesendoderm and ectoderm from an early-gastrula-stage *Xenopus laevi...

  20. Pancreas Development Ex Vivo: Culturing Embryonic Pancreas Explants on Permeable Culture Inserts, with Fibronectin-Coated Glass Microwells, or Embedded in Three-Dimensional Matrigel™

    OpenAIRE

    Shih, Hung Ping; Sander, Maike

    2014-01-01

    Pancreas development is a complex and dynamic process orchestrated by cellular and molecular events, including morphogenesis and cell differentiation. As a result of recent explorations into possible cell-therapy-based treatments for diabetes, researchers have made significant progress in deciphering the developmental program of pancreas formation. In vitro pancreas organ culture systems provide a valuable tool for exploring the mechanisms of gene regulation, cellular behaviors, and cell diff...

  1. 改良组织块酶消化法培养人龋损牙髓干细胞的实验研究%Culture of human caries dental pulp stem cells with combined explants method and enzymatic separation method

    Institute of Scientific and Technical Information of China (English)

    麻丹丹; 高杰; 吴补领

    2011-01-01

    AIM; To compare the successfulness and the growth of human caries dental pulp stem cells(hCDPSCs) cultured with three different methods. METHODS: Twenty-five normal and caries human third molars were collected, the dental pulp tissues were cultured by the tissue explant method, tissue-explan collagenase digestion method and the combination of explant method and enzymatic separation method, respectively. The adherence of the explants, the morphology and quantity of cells were observed under a phase-contrast microscope. Culture duration was recorded. The clones were identified by expression of Stro-1 and CD90 and the growth curve of normal DPSCs and CDPSCs was drawn. RESULTS; Human normal DPSCs and CDPSCs could be cultured by all the three methods. A large number of human normal DPSCs and CDPSCs were cultured by the com hined explant method and enzymatic separation method in a shorter time, and these cells exhibited more vitality and more different morphologies. The growth rate of CDPSCs was higher than that of normal DPSCs . CONCLISION: The improved combination of explant method and enzymatic separation method is an ideal method for the primary culture of hCDPSCs in vitro, it may provide a methodological foundation for studying the mechanism of the formation of the tertiary dentine when the tooth was damaged.%目的:比较3种方法培养人龋损牙髓干细胞的成功率和细胞生长状态,以探求人龋损牙髓干细胞的最佳培养方法.方法:取18~22岁成人新鲜正常和龋损离体第三磨牙各25个,采用组织块法、酶消化法、改良组织块酶消化法培养牙髓干细胞.通过倒置显微镜观察组织块的贴壁以及细胞的形态和数量,并记录培养所需时间;有限稀释法纯化牙髓干细胞,流式细胞仪检测正常和龋损牙髓干细胞表面标记物Stro-1、CD 90的表达情况,绘制正常和龋损牙髓干细胞生长曲线.结果:组织块法、酶消化法和改良组织块酶消化法均可以培养

  2. The in vitro and in vivo capacity of culture-expanded human cells from several sources encapsulated in alginate to form cartilage

    Directory of Open Access Journals (Sweden)

    MM Pleumeekers

    2014-04-01

    Full Text Available Cartilage has limited self-regenerative capacity. Tissue engineering can offer promising solutions for reconstruction of missing or damaged cartilage. A major challenge herein is to define an appropriate cell source that is capable of generating a stable and functional matrix. This study evaluated the performance of culture-expanded human chondrocytes from ear (EC, nose (NC and articular joint (AC, as well as bone-marrow-derived and adipose-tissue-derived mesenchymal stem cells both in vitro and in vivo. All cells (≥ 3 donors per source were culture-expanded, encapsulated in alginate and cultured for 5 weeks. Subsequently, constructs were implanted subcutaneously for 8 additional weeks. Before and after implantation, glycosaminoglycan (GAG and collagen content were measured using biochemical assays. Mechanical properties were determined using stress-strain-indentation tests. Hypertrophic differentiation was evaluated with qRT-PCR and subsequent endochondral ossification with histology. ACs had higher chondrogenic potential in vitro than the other cell sources, as assessed by gene expression and GAG content (p < 0.001. However, after implantation, ACs did not further increase their matrix. In contrast, ECs and NCs continued producing matrix in vivo leading to higher GAG content (p < 0.001 and elastic modulus. For NC-constructs, matrix-deposition was associated with the elastic modulus (R2 = 0.477, p = 0.039. Although all cells – except ACs – expressed markers for hypertrophic differentiation in vitro, there was no bone formed in vivo. Our work shows that cartilage formation and functionality depends on the cell source used. ACs possess the highest chondrogenic capacity in vitro, while ECs and NCs are most potent in vivo, making them attractive cell sources for cartilage repair.

  3. Effect of ionizing radiation on the growth, morphogenic and metabolic activity of explant cultures of Arctostaphylos uva-ursi (L.) Spreng., Coronilla varia (L.) and Datura meteloides DC. ex Dunal

    International Nuclear Information System (INIS)

    The effect of ionizing radiation on the growth, morphogenic and metabolic activity of explant cultures of Arctostaphylos uva-ursi (L.) Spreng. and Coronilla varia (L.) was studied with doses of 0.5, 1.0, 2.0, 180 and 200 Gy. In the culture of Datura meteloides DC. ex Dunal the doses of radiation used were 0.5, 2.0, 2.5, 5, 10, 20, 40, 80, 120, 160, 180 and 200 Gy. In the first two cultures the results agree with thouse published elsewhere. Doses of 0.5-2.0 Gy slightly inhibited the growth of callus cultures of the bearberry and crown vetch; the development of embryogenic cultures was not influenced. Doses of 180 and 200 Gy strongly inhibited the growth, but no lethal effect was found. The morphogenic activity and the spectrum of metabolites were not influenced. In cultures of stramonium, a statistically significant stimulating effect on the growth after irradiation with a dose of 0.5 Gy was demonstrated. A dose of 200 Gy was not lethal, either. None of the doses used produced a formation of organs or somatic embryos or biosynthesis of tropane alkaloids. (author) 4 figs., 19 refs

  4. Effects of tenoxicam and aspirin on the metabolism of proteoglycans and hyaluronan in normal and osteoarthritic human articular cartilage.

    OpenAIRE

    Manicourt, Daniel; Druetz-Van Egeren, A; Haazen, L.; Nagant de Deuxchaisnes, C

    1994-01-01

    1. As nonsteroidal anti-inflammatory drugs may impair the ability of the chondrocyte to repair its damaged extracellular matrix, we explored the changes in the metabolism of newly synthesized proteoglycan (PG) and hyaluronan (HA) molecules produced by tenoxicam and aspirin in human normal cartilage explants and in osteoarthritic (OA) cartilage from age-matched donors. 2. Explants were sampled from the medial femoral condyle and were classified by use of Mankin's histological-histochemical gra...

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

    Science.gov (United States)

    Dua, Rupak; Comella, Kristin; Butler, Ryan; Castellanos, Glenda; Brazille, Bryn; Claude, Andrew; Agarwal, Arvind; Liao, Jun; Ramaswamy, Sharan

    2016-01-01

    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 engineered

  6. 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

  7. Sour orange bud regeneration and in vitro plant development related to culture medium composition and explant type Regeneração de gemas de laranja-azeda e desenvolvimento in vitro de plantas em função da composição do meio de cultura e tipo de explante

    Directory of Open Access Journals (Sweden)

    Rosely Pereira da Silva

    2010-03-01

    Full Text Available In order to evaluate the formation of adventitious buds and in vitro regeneration of sour orange plants (Citrus aurantium L. two organogenesis-inducing experiments were conducted. In the first experiment, the induction and in vitro regeneration of adventitious buds were tested on epicotyl and internodal segments under the influence of BAP or KIN associated with NAA. The second experiment evaluated the in vitro regeneration of sour orange plants related to different explant types (epicotyls segments, internodal segments of in vitro germinated plantlets and internodal segments of greenhouse cultivated plants. Data collected on both experiments included the percentage of responsive explants (explants that formed buds, and the number of buds per explant. The addition of BAP showed the best organogenic response. In vitro germinated epicotyl segments and internodal segments are recommended as explants for sour orange in vitro organogenesis. Rooting of regenerated shoots was achieved without the need of auxin in the medium.Com o objetivo de avaliar a formação de gemas adventícias e regeneração in vitro de plantas de laranja-azeda (Citrus aurantium L., foram realizados dois experimentos de indução à organogênese. No primeiro experimento, a indução e a regeneração in vitro de gemas adventícias foram investigadas a partir de segmentos internodais e segmentos de epicótilo sob o efeito de BAP ou CIN associados com ANA. O segundo experimento avaliou a regeneração in vitro de plantas de laranja-azeda em função do tipo de explante (segmentos de epicótilo, segmentos internodais de plantas germinadas in vitro e segmentos internodais de plantas cultivadas em casa de vegetação. Os dados coletados em ambos os experimentos incluíram a porcentagem de explantes responsivos (explantes que formaram gemas e número de gemas por explante. A adição de BAP revelou a melhor resposta organogenética. Segmentos de epicótilo e segmentos internodais s

  8. The influence of season collection of explants on micropropagation of peach rootstock GF-677

    Directory of Open Access Journals (Sweden)

    Elektra Spahiu

    2013-02-01

    Full Text Available The influence of season on the rate of multiplication on in vitro culture of peach rootstock GF- 677 was investigated on Murashige and Skoog (MS media, supplemented with GA3 0.1 mg/L and IAA 0.1mg/l. Benzyladenine (BAP at concentrations 1mg/l was used in the multiplication stage and 1mg/l IBA in the stage of rooting. Shoot-tip and nodal segment explants were collected from 5 years old rootstock GF-677 (Prunus persica x Prunus amygdalus in February 24th (from dormant shoots that have been sprouted in climatic room, March 22th, April 20th, May 18th and September 15th during the 2009 growing season and have been sterilized by sodium hypochlorite (NaOCl 10% for 20 min. The data on the effect of the season collection of the explants on number of shoots per explants, the mean shoot length and the percentage of rooted shoots were recorded six weeks after culture. In vitro performance of explants indicated a positive correlation between shoot proliferation and season collection The highest number of shoots per explants (3,5 was obtained on explants collected in March 22th (3,5, which was on a par with explants collected in February 24th (from shoots that have been sprouted in climatic room. Moreover, the highest shoot length was observed on explants collected on February and March (1,53cm and 1,505cm respectively. The percentage of rooted shoots from explants sampled on February was not markedly greater than those sampled on March. The number of shoots per explants, the shoot length and the percentage of rooted shoots on explants sampled in April, May and September were significantly lower than those sampled in February and March. The amount of chlorophyll a + b of the shoots coming from explants collected in March was markedly greater than those collected in February, April, May and September.

  9. The impact of dietary long-chain polyunsaturated fatty acids on bone and cartilage in gilts and sows.

    Science.gov (United States)

    O'Connor-Robison, C I; Spencer, J D; Orth, M W

    2014-10-01

    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 < 0.05) and decreased cortical width of the distal radius (P < 0.05). Sows fed PFO had increased DHA (P < 0.01) and a decrease in the omega-6 to omega-3 ratio (P < 0.05) in cartilage. Gilts fed PFO had increased DHA (P < 0.01), C22:1 (P < 0.01), and docosapentaenoic acid (P < 0.01) and a tendency for increased EPA (P = 0.093) concentrations in cartilage

  10. Plant Regeneration of Sweet Potato via Somatic Embryogenesis from Different Explants

    Institute of Scientific and Technical Information of China (English)

    Ling ZHANG; Hongxuan XU; Baifu QIN; Zhihua LIA0; Min CHEN; Chunxian YANG; Yufan FU; Qitang ZHANG

    2012-01-01

    [Objective] This study aimed to regenerate plants of sweet potato (Ipomoea batatas) cultivar Xushu22 via somatic embryogenesis, using leaf and shoot apex as explants. [Method] The leaf and shoot apex of Xushu 22 were separately cultured on MSB medium and MSD medium. The induced embryogenic calluses were then cultured on MS medium. The regeneration frequency of leaf and shoot apex ex- plants were respectively calculated. [Result] The average frequency of leaf explants developing somatic callus was 95.69% compared to 30.56% in case of shoot apex explants. There were different types of morphogenic structures in the process of so- matic embryo development. Leaf explants gave a high regeneration frequency to 60.61%, while the regeneration frequency of shoot apices was 22%. In addition, no morphological variations were observed in the regeneration plants. [Conclusion] Leaf explant was better than shoot apices in plant regeneration of Xushu22 via somatic embryogenesis.

  11. Chlorophyll Fluorescence as a Tool to Assess the Regeneration Potential of African Violet Leaf Explants

    Directory of Open Access Journals (Sweden)

    Norbert KEUTGEN

    2016-06-01

    Full Text Available Micropropagation of many ornamentals has enabled their efficient commercialisation and many problems have been solved by the elaboration of adequate culture protocols. Nevertheless, a non-destructive technique for monitoring the developmental progress of explants would be desirable. The present study focussed on the applicability of chlorophyll fluorescence in leaf explants of African violet (a Saintpaulia ionantha × confusa – hybrid explanted onto Murashige and Skoog basic medium. The explants that survived on the medium without additional phytohormones had the capacity to develop further into two different kinds of explants: light green explants, characterized by a non-regular size growth and stiffer appearance, and dark green explants capable of organogenesis. Compared to the source leaves of African violet plants, explants were characterized by reduced chlorophyll (Chl and carotenoid (Car contents as well as a tendency towards a higher Car/Chl ratio. The Chl a/b ratio decreased significantly in the light green explants. A reduction of maximum quantum efficiency of photosystem II (Fv/Fm accompanied by a high percentage (> 50% of thermal energy dissipation as a consequence of an elevated light intensity (800 µmol m-2 s-1 quanta indicated photoinhibition in the light green explants, whereas in the dark green explants the largest percentage (> 50% of the light energy was dissipated into the fraction of photon energy trapped by ‘closed’ photosystem II reaction centres. These results suggest that the capacity of organogenesis of leaf explants of African violet can be monitored using chlorophyll fluorescence, because it is related to modifications of the photosynthetic system.

  12. 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

    2009-01-01

    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.

  13. Production of Gymnemic Acid Depends on Medium, Explants, PGRs, Color Lights, Temperature, Photoperiod, and Sucrose Sources in Batch Culture of Gymnema sylvestre

    Directory of Open Access Journals (Sweden)

    A. Bakrudeen Ali Ahmed

    2012-01-01

    Full Text Available Gymnema sylvestre (R.Br. is an important diabetic medicinal plant which yields pharmaceutically active compounds called gymnemic acid (GA. The present study describes callus induction and the subsequent batch culture optimization and GA quantification determined by linearity, precision, accuracy, and recovery. Best callus induction of GA was noticed in MS medium combined with 2,4-D (1.5 mg/L and KN (0.5 mg/L. Evaluation and isolation of GA from the calluses derived from different plant parts, namely, leaf, stem and petioles have been done in the present case for the first time. Factors such as light, temperature, sucrose, and photoperiod were studied to observe their effect on GA production. Temperature conditions completely inhibited GA production. Out of the different sucrose concentrations tested, the highest yield (35.4 mg/g d.w was found at 5% sucrose followed by 12 h photoperiod (26.86 mg/g d.w. Maximum GA production (58.28 mg/g d.w was observed in blue light. The results showed that physical and chemical factors greatly influence the production of GA in callus cultures of G. sylvestre. The factors optimized for in vitro production of GA during the present study can successfully be employed for their large-scale production in bioreactors.

  14. Polyphenol Oxidase (PPO) in Early Stage of Browning of Phalaenopsis Leaf Explants

    OpenAIRE

    Zhiwei Ru; Yanyan Lai; Chuangjun Xu; Ling Li

    2013-01-01

    Explant browning is one of the most common problems during the early stage of tissue culture. The aim of this study was to explore the roles of polyphenol oxidase (PPO, EC 1.10.3.2) in the browning process of leaf explants of Phalaenopsis (Doritaenopsis Queen Bee “Red Sky”) under a routine culture condition. Along with the observation of the time course of browning and PPO distribution, PPO gene expression in leaf explants were investigated. In addition, the expression and activity of PPO wer...

  15. A stable explant culture of HER2/neu invasive carcinoma supported by alpha-Smooth Muscle Actin expressing stromal cells to evaluate therapeutic agents

    International Nuclear Information System (INIS)

    To gain a better understanding of the effects of therapeutic agents on the tumor microenvironment in invasive cancers, we developed a co-culture model from an invasive lobular carcinoma. Tumor cells expressing HER2/neu organize in nests surrounded by alpha-Smooth Muscle Actin (α-SMA) expressing tumor stroma to resemble the morphology of an invading tumor. This co-culture, Mammary Adenocarcinoma Model (MAM-1) maintains a 1:1 ratio of HER2/neu positive tumor cells to α-SMA-reactive stromal cells and renews this configuration for over 20 passages in vitro. We characterized the cellular elements of the MAM-1 model by microarray analysis, and immunocytochemistry. We developed flow cytometric assays to evaluate the relative responses of the tumor and stroma to the tyrosine kinase inhibitor, Iressa. The MAM-1 gene expression profile contains clusters that represent the ErbB-2 breast cancer signature and stroma-specific clusters associated with invasive breast cancers. The stability of this model and the ability to antigenically label the tumor and stromal fractions allowed us to determine the specificity of Iressa, a receptor tyrosine kinase inhibitor, for targeting the tumor cell population. Treatment resulted in a selective dose-dependent reduction in phospho-pMEK1/2 and pp44/42MAPK in tumor cells. Within 24 h the tumor cell fraction was reduced 1.9-fold while the stromal cell fraction increased >3-fold, consistent with specific reductions in phospho-pp44/42 MAPK, MEK1/2 and PCNA in tumor cells and reciprocal increases in the stromal cells. Erosion of the tumor cell nests and augmented growth of the stromal cells resembled a fibrotic response. This model demonstrates the specificity of Iressa for HER2/neu expressing tumor cells versus the tumor associated myofibroblasts and is appropriate for delineating effects of therapy on signal transduction in the breast tumor microenvironment and improving strategies that can dually or differentially target the tumor and stromal

  16. TGF beta suppresses casein synthesis in mouse mammary explants and may play a role in controlling milk levels during pregnancy

    OpenAIRE

    1993-01-01

    Mammary explants from 14-15-d-pregnant mice synthesize and secrete milk proteins in culture in response to insulin, hydrocortisone, and prolactin. Here we demonstrate that transforming growth factor beta (TGF beta) treatment suppresses, in a dose dependent and reversible manner, the ability of explants to synthesize and secrete milk caseins. TGF beta does not affect the level of casein mRNA within explants but inhibits casein synthesis posttranscriptionally. We also show increased expression ...

  17. Collagen metabolism of human osteoarthritic articular cartilage as modulated by bovine collagen hydrolysates

    OpenAIRE

    Saskia Schadow; Hans-Christian Siebert; Günter Lochnit; Jens Kordelle; Markus Rickert; Jürgen Steinmeyer

    2013-01-01

    Destruction of articular cartilage is a characteristic feature of osteoarthritis (OA). Collagen hydrolysates are mixtures of collagen peptides and have gained huge public attention as nutriceuticals used for prophylaxis of OA. Here, we evaluated for the first time whether different bovine collagen hydrolysate preparations indeed modulate the metabolism of collagen and proteoglycans from human OA cartilage explants and determined the chemical composition of oligopeptides representing collagen ...

  18. Imaging of articular cartilage

    Directory of Open Access Journals (Sweden)

    Bhawan K Paunipagar

    2014-01-01

    Full Text Available We tried to review the role of magnetic resonance imaging (MRI in understanding microscopic and morphologic structure of the articular cartilage. The optimal protocols and available spin-echo sequences in present day practice are reviewed in context of common pathologies of articular cartilage. The future trends of articular cartilage imaging have been discussed with their appropriateness. In diarthrodial joints of the body, articular cartilage is functionally very important. It is frequently exposed to trauma, degeneration, and repetitive wear and tear. MRI has played a vital role in evaluation of articular cartilage. With the availability of advanced repair surgeries for cartilage lesions, there has been an increased demand for improved cartilage imaging techniques. Recent advances in imaging strategies for native and postoperative articular cartilage open up an entirely new approach in management of cartilage-related pathologies.

  19. Cartilage (Bovine and Shark) (PDQ)

    Science.gov (United States)

    ... Ask about Your Treatment Research Cartilage (Bovine and Shark) (PDQ®)–Patient Version Overview Go to Health Professional ... 8 ). Questions and Answers About Cartilage (Bovine and Shark) What is cartilage? Cartilage is a type of ...

  20. Effect of vasoactive intestinal peptide on pulmonary surfactants phospholipid synthesis in lung explants

    Institute of Scientific and Technical Information of China (English)

    Lian LI; Zi-qiang LUO; Fu-wen ZHOU; Dan-dan FENG; Cha-xiang GUAN; Chang-qing ZHANG; Xiu-hong SUN

    2004-01-01

    AIM: To investigate the effect of vasoactive intestinal peptide (VIP) on pulmonary surfactants (PS) phospholipid synthesis in cultured lung explants. METHODS: Lung explants were cultured with serum-free medium, [methyl-3H]choline incorporation, total phospholipid, phosphatidylcholine, activity of choline-phosphate cytidylyltransferase (CCT) and CCTα mRNA level in lung explants were determined. RESULTS: (1) VIP (10-10-10-7 mol/L) for 16 h promoted [methyl-3H]choline incorporation in dose dependence and VIP (10-8 mol/L) for 2 h-16 h promoted [methylz3H]choline incorporation in time dependence. (2) VIP (10-8 mol/L) enhanced the contents of total phospholipidsand phosphatidylcholine in lung explants. (3) VIP (10-10-10-7 mol/L) elevated microsomal CCT activity of lung explants in dose dependence. (4) VIP (10-8 mol/L) increased expression of CCTα mRNA in lung explants and alveolar type Ⅱ cells (ATII). (5) [D-P-Cl-Phe(6)-Leu(17)]-VIP (10-6 mol/L), a VIP receptors antagonist, abolished the increase of [3H]choline incorporation, microsomal CCT activity and CCTα mRNA level induced by VIP (10-8 mol/L) in lung explants. CONCLUSION: VIP could enhance synthesis of phosphatidylcholine, the major component of pulmonary surfactants by enhancing microsomal CCT activity and CCTα mRNA level via VIP receptormediated pathway.

  1. Tooth explant cultures in dental apoptosis research

    Czech Academy of Sciences Publication Activity Database

    Matalová, Eva; Šetková, Jana; Míšek, Ivan

    2005-01-01

    Roč. 3, Suppl 1 (2005), S33-S34. ISSN 1214-021X. [Cells VI - Biological Days /18./. 24.10.2005-26.10.2005, České Budějovice] R&D Projects: GA MŠk OC B23.001 Institutional research plan: CEZ:AV0Z50450515 Keywords : dental apoptosis research Subject RIV: EB - Genetics ; Molecular Biology

  2. Life Cycle of Heterodera zeae Koshy, Swarup, and Sethi on Zea mays L. Axenic Root Explants

    OpenAIRE

    Lauritis, J. A.; Rebois, R. V.; Graney, L. S.

    1983-01-01

    Monoxenic cultures of Heterodera zeae, the corn cyst nematode (CCN), were established on root explants of corn Zea mays L., cv. Kenworthy. The life cycle of H. zeae was determined from light anti scanning electron microscopic observations of the root explants grown in the dark at 29.5 ± .5 C under gnotobiotic conditions. The life cycle, from the time the explants were inoculated with second-stage larvae (L2) to the first appearance of newly hatched second-generation L2, required 22 days. The ...

  3. Autofluorescence lifetime metrology for label-free detection of cartilage matrix degradation

    Science.gov (United States)

    Nickdel, Mohammad B.; Lagarto, João. L.; Kelly, Douglas J.; Manning, Hugh B.; Yamamoto, Kazuhiro; Talbot, Clifford B.; Dunsby, Christopher; French, Paul; Itoh, Yoshifumi

    2014-03-01

    Degradation of articular cartilage extracellular matrix (ECM) by proteolytic enzyme is the hallmark of arthritis that leads to joint destruction. Detection of early biochemical changes in cartilage before irreversible structural damages become apparent is highly desirable. Here we report that the autofluorescence decay profile of cartilage is significantly affected by proteolytic degradation of cartilage ECM and can be characterised by measurements of the autofluorescence lifetime (AFL). A multidimensional fluorometer utilizing ultraviolet excitation at 355 nm or 375 nm coupled to a fibreoptic probe was developed for single point time-resolved AFL measurements of porcine articular cartilage explants treated with different proteinases. Degradation of cartilage matrix components by treating with bacterial collagenase, matrix metalloproteinase 1, or trypsin resulted in significant reduction of AFL of the cartilage in both a dose and time dependent manner. Differences in cartilage AFL were also confirmed by fluorescence lifetime imaging microscopy (FLIM). Our data suggest that AFL of cartilage tissue is a potential non-invasive readout to monitor cartilage matrix integrity that may be utilized for diagnosis of arthritis as well as monitoring the efficacy of anti-arthritic therapeutic agents.

  4. 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

    1999-07-01

    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

  5. 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

    2010-05-01

    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

  6. Organogênese direta de explantes cotiledonares e regeneração de plantas de mogango Direct organogenesis of cotyledon explants and plant regeneration of squash

    Directory of Open Access Journals (Sweden)

    André Luís Lopes da Silva

    2006-06-01

    Full Text Available Os objetivos foram induzir a organogênese direta de explantes cotiledonares de mogango e estudar a regeneração de plântulas completas a partir das brotações adventícias. Foram utilizados cotilédones como explantes, originados das plântulas de mogango com 20 dias após a semeadura. O meio basal utilizado foi o MS (MURASHIGE & SKOOG, 1962 suplementado com 30g L-1 de sacarose e 7g L-1 de agar. Foram testadas as concentrações de 6-benzilaminopurina (BAP de 0; 0,5; 1,0 e 2,0mg L-1. Explantes de ápices caulinares e segmentos nodais de brotações adventícias foram então cultivados em meio MS suplementado com 30g L-1 de sacarose e 7g L-1 de agar. Maiores concentrações de BAP no meio MS promoveram um aumento da percentagem de explantes cotiledonares com brotações adventícias e uma redução da percentagem de enraizamento. Explantes de segmentos nodais e ápices caulinares oriundos de brotações adventícias cresceram e enraizaram em meio MS sem reguladores de crescimento. Altas percentagens de enraizamento dependem do tamanho dos explantes utilizados.The objectives were to induce direct organogenesis of squash cotyledons and to study the regeneration of complete plantlets from adventitious shoot. Cotyledon explants of 20-day seedlings were cultured in MS (MURASHIGE & SKOOG, 1962 medium supplemented with 30g L-1 of sucrose and 7g L-1 of agar. The 6-benzilaminopurina (BAP concentrations of 0, 0.5, 1.0 and 2.0mg L-1 were tested. Apical and nodal explants from adventitious shoots were transferred to MS medium supplemented with 30g L-1 of sucrose and 7g L-1 of agar. Increasing BAP concentrations in the MS medium enhance the percentage of adventitious shoot and reduce the percentage of root organogenesis of squash cotyledon explants. Apical and nodal explants from adventitious shoot regenerated plantlets with roots in MS medium without growth regulators. High percentage of plantlet rooting depends upon the size of the explants.

  7. 微载体培养技术在骨与软骨组织工程研究中的应用%Application of microcarrier culture techniques in bone and cartilage tissue engineering

    Institute of Scientific and Technical Information of China (English)

    宁斌; 田周斌; 贾堂宏

    2012-01-01

    BACKGROUND: Proliferation and specific phenotype maintaining of seed cells are the difficulties in bone and cartilage tissueengineering. The microcarrier bioreactor culture system provides a good method to solve this problem.OBJECTIVE: To analyze the domestic and international development of microcarrier culture of bone and cartilage cells, and toprovide the theoretical basis for microcarrier culture of bone and cartilage cells and researches in tissue engineering.METHODS: A computer-based online search of articles published from 1967 to 2011 was performed at November 2010 inPubMed database (http://www.ncbi.nlm.gov/PubMed) and Wanfang database (http://www.wanfangdata.com.cn) using the keywords of "microcarrier, cartilage, tissue engineering" in English and in Chinese, respectively. Articles irrelative to this paper,antiquated or repeated literatures were excluded. Totally 32 articles were chosen for further analysis.RESULTS AND CONCLUSION: Culture conditions of bone and cartilage cells in microcarrier bioreactor culture system can bewell regulated to achieve a large number of proliferation and phenotype maintaining of the cells, or even achieve an enhancementof phenotype. This technology has a good potential application in bone and cartilage tissue engineering and clinical work.%背景 骨与软骨组织工程学中增殖种子细胞和保持细胞特定表型是其难点,微载体生物反应器培养系统提供了很好的条件来解决这个问题.目的 分析近年来国内外骨、软骨细胞微载体培养的研究进展,为骨与软骨细胞微载体培养技术和组织工程研究提供理论基础.方法 由第一作者在2010-11 进行检索.检索数据库:PubMed 数据库(网址http://www.ncbi.nlm.gov/PubMed);万方数据库(网址http://www.wanfangdata.com.cn),资料的检索时间范围为1967/2011.英文检索词为"microcarrier,cartilage,tissue engineering",中文检索词为"微载体,软骨,组织工程学".排除与本文无关及陈

  8. Cartilage Tissue Engineering: What Have We Learned in Practice?

    Science.gov (United States)

    Doran, Pauline M

    2015-01-01

    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. PMID:26445827

  9. Effect of Hormones on Direct Shoot Regeneration in Hypocotyl Explants of Tomato

    Directory of Open Access Journals (Sweden)

    Rizwan RASHID

    2010-03-01

    Full Text Available This study was conducted for developing a high frequency regeneration system in two genotypes of tomato (Lycopersicon esculentum Mill., �Punjab Upma� and �IPA-3� for direct shoot regeneration from hypocotyl explants. The explants were excised from in vitro tomato seedlings and cultured on MS medium supplemented with different concentrations and combinations of hormones. Direct regeneration was significantly influenced by the genotype hormones combination and concentrations. The MS medium supplemented with (Kinetin 0.5 mg/l and (BAP 0.5 mg/l was found optimum for inducing direct shoot regeneration and number of shoots per explant from hypocotyl explants on this medium. Shoot regeneration per cent in �Punjab Upma� and �IPA-3� per cent was recorded to be highest i.e (86.02 and (82.57 respectively. Besides this, average number shoots per explant was also highest i.e (3.16 in case of �Punjab Upma� and (2.93 in case of �IPA-3�. A significant decline was observed in percent shoot regeneration and average number of shoots per explant with increase in the hormonal concentration. Shoots were obtained and transferred to the elongation medium (MS + BAP 0.3 mg/l. Hundred per cent rooting was induced in separated shoots upon culturing on MS and � MS basal media. Hardening on moist cotton showed maximum plantlet survival rate in case of both genotypes. After hardening, plants were transferred to soil. Thus, a tissue culture base line was established in tomato for obtaining direct regeneration using hypocotyl as explants.

  10. Preparation and characterization of a decellularized cartilage scaffold for ear cartilage reconstruction

    International Nuclear Information System (INIS)

    Scaffolds are widely used to reconstruct cartilage. Yet, the fabrication of a scaffold with a highly organized microenvironment that closely resembles native cartilage remains a major challenge. Scaffolds derived from acellular extracellular matrices are able to provide such a microenvironment. Currently, no report specifically on decellularization of full thickness ear cartilage has been published. In this study, decellularized ear cartilage scaffolds were prepared and extensively characterized. Cartilage decellularization was optimized to remove cells and cell remnants from elastic cartilage. Following removal of nuclear material, the obtained scaffolds retained their native collagen and elastin contents as well as their architecture and shape. High magnification scanning electron microscopy showed no obvious difference in matrix density after decellularization. However, glycosaminoglycan content was significantly reduced, resulting in a loss of viscoelastic properties. Additionally, in contact with the scaffolds, human bone-marrow-derived mesenchymal stem cells remained viable and are able to differentiate toward the chondrogenic lineage when cultured in vitro. These results, including the ability to decellularize whole human ears, highlight the clinical potential of decellularization as an improved cartilage reconstruction strategy. (paper)

  11. Responsiveness of mouse calvaria to parathyroid hormone after explant cryopreservation: 45Ca release in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Wezeman, F.H.; Dungan, D.D.

    1986-08-01

    Newborn mouse calvaria prelabeled with /sup 45/Ca and cryopreserved at -196 degrees C in serum-free medium containing dimethylsulfoxide were compared to unpreserved explants for response to parathyroid hormone during subsequent culture. After short-term cryopreservation followed by rapid thawing, the viable explants continued to release /sup 45/Ca to the culture medium but additions of parathyroid hormone to the medium did not cause increased bone resorption. The data suggest that cryopreservation and thawing impairs mechanisms responsible for parathyroid hormone action on bone cells.

  12. Cartilage conduction hearing.

    Science.gov (United States)

    Shimokura, Ryota; Hosoi, Hiroshi; Nishimura, Tadashi; Yamanaka, Toshiaki; Levitt, Harry

    2014-04-01

    Sound information is known to travel to the cochlea via either air or bone conduction. However, a vibration signal, delivered to the aural cartilage via a transducer, can also produce a clearly audible sound. This type of conduction has been termed "cartilage conduction." The aural cartilage forms the outer ear and is distributed around the exterior half of the external auditory canal. In cartilage conduction, the cartilage and transducer play the roles of a diaphragm and voice coil of a loudspeaker, respectively. There is a large gap between the impedances of cartilage and skull bone, such that cartilage vibrations are not easily transmitted through bone. Thus, these methods of conduction are distinct. In this study, force was used to apply a transducer to aural cartilage, and it was found that the sound in the auditory canal was amplified, especially for frequencies below 2 kHz. This effect was most pronounced at an application force of 1 N, which is low enough to ensure comfort in the design of hearing aids. The possibility of using force adjustments to vary amplification may also have applications for cell phone design. PMID:25234994

  13. Mucus glycoprotein secretion by tracheal explants: effects of pollutants

    International Nuclear Information System (INIS)

    Tracheal slices incubated with radioactive precursors in tissue culture medium secrete labeled mucus glycoproteins into the culture medium. We have used an in vivtro approach, a combined method utilizing exposure to pneumotoxins in vivo coupled with quantitation of mucus secretion rates in vitro, to study the effects of inhaled pollutants on mucus biosynthesis by rat airways. In addition, we have purified the mucus glycoproteins secreted by rat tracheal explants in order to determine putative structural changes that might by the basis for the observed augmented secretion rates after exposure of rats to H2SO4 aerosols in combination with high ambient levels of ozone. After digestion with papain, mucus glycoproteins secreted by tracheal explants may be separated into five fractions by ion-exchange chromatography, with recovery in high yield, on columns of DEAE-cellulose. Each of these five fractions, one neutral and four acidic, migrates as a single unique spot upon cellulose acetate electrophoresis at pH values of 8.6 and 1.2. The neutral fraction, which is labeled with [3H] glucosamine, does not contain radioactivity when Na2 35SO4 is used as the precursor. Acidic fractions I to IV are all labeled with either 3H-glucosamine or Na2 35SO4 as precursor. Acidic fraction II contains sialic acid as the terminal sugar on its oligosaccharide side chains, based upon its chromatographic behavior on columns of wheat-germ agglutinin-Agarose. Treatment of this fraction with neuraminidase shifts its elution position in the gradient to a lower salt concentration, coincident with acidic fraction I. After removal of terminal sialic acid residues with either neuraminidase or low pH treatment, the resultant terminal sugar on the oligosaccharide side chains is fucose. These results are identical with those observed with mucus glycoproteins secreted by cultured human tracheal explants and purified by these same techniques

  14. Cartilage Engineering and Microgravity

    Science.gov (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.

    2005-06-01

    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.

  15. Influence of Cartilage Extracellular Matrix Molecules on Cell Phenotype and Neocartilage Formation

    OpenAIRE

    Grogan, Shawn P.; Chen, Xian; Sovani, Sujata; Taniguchi, Noboru; Colwell, Clifford W.; Lotz, Martin K; D'Lima, Darryl D

    2013-01-01

    Interaction between chondrocytes and the cartilage extracellular matrix (ECM) is essential for maintaining the cartilage's role as a low-friction and load-bearing tissue. In this study, we examined the influence of cartilage zone-specific ECM on human articular chondrocytes (HAC) in two-dimensional and three-dimensional (3D) environments. Two culture systems were used. SYSTEM 1: HAC were cultured on cell-culture plates that had been precoated with the following ECM molecules for 7 days: decor...

  16. Proliferation of Female Inflorescences explants of Date Palm

    International Nuclear Information System (INIS)

    This study was conducted to determine the effect of Abscisic acid (ABA) and Ancymidol on proliferation of female inflorescences explants of date palm. In the first experiment two lengths of spath at (5-7 cm) or at (7-10 cm) were cultured on nutrient media which consists of half macro and full micro salts of MS medium supplemented with gradual decreasing in concentration of Abscisic acid (ABA) and Ancymidol from 4.5, 3.0, 1.5 to 0.5 mg-1. In the second experiment two phases of nutrient medium (solid and liquid) and two source of carbon were investigated. Gradual decreasing of ABA concentrations from 4.5 mg-1 to 1.5 mg-1 in culture medium, stimulated the production of direct somatic embryos and accelerated callus initiation, but at last decrement (0.5 mg-1) of Ancymidol concentration few embryos were produced. Callus initiation from inflorescences explants gave high production and well development of somatic embryos when cultured on liquid medium supplemented with 40 g-1 sucrose. All direct or indirect somatic embryos obtained in these experiments were converted successfully to healthy normal plantlets which could be transferred to acclimatization stage.

  17. A novel fibroblast growth factor receptor 1 inhibitor protects against cartilage degradation in a murine model of osteoarthritis

    Science.gov (United States)

    Xu, Wei; Xie, Yangli; Wang, Quan; Wang, Xiaofeng; Luo, Fengtao; Zhou, Siru; Wang, Zuqiang; Huang, Junlan; Tan, Qiaoyan; Jin, Min; Qi, Huabing; Tang, Junzhou; Chen, Liang; Du, Xiaolan; Zhao, Chengguang; Liang, Guang; Chen, Lin

    2016-01-01

    The attenuated degradation of articular cartilage by cartilage-specific deletion of fibroblast growth factor receptor 1 (FGFR1) in adult mice suggests that FGFR1 is a potential target for treating osteoarthritis (OA). The goal of the current study was to investigate the effect of a novel non-ATP-competitive FGFR1 inhibitor, G141, on the catabolic events in human articular chondrocytes and cartilage explants and on the progression of cartilage degradation in a murine model of OA. G141 was screened and identified via cell-free kinase-inhibition assay. In the in vitro study, G141 decreased the mRNA levels of catabolic markers ADAMTS-5 and MMP-13, the phosphorylation of Erk1/2, JNK and p38 MAPK, and the protein level of MMP-13 in human articular chondrocytes. In the ex vivo study, proteoglycan loss was markedly reduced in G141 treated human cartilage explants. For the in vivo study, intra-articular injection of G141 attenuated the surgical destabilization of the medial meniscus (DMM) induced cartilage destruction and chondrocyte hypertrophy and apoptosis in mice. Our data suggest that pharmacologically antagonize FGFR1 using G141 protects articular cartilage from osteoarthritic changes, and intra-articular injection of G141 is potentially an effective therapy to alleviate OA progression. PMID:27041213

  18. MRI of the cartilage

    International Nuclear Information System (INIS)

    With the introduction of fat-suppressed gradient-echo and fast spin-echo (FSE) sequences in clinical routine MR visualization of the hyaline articular cartilage is routinely possible in the larger joints. While 3D gradient-echo with fat suppression allows exact depiction of the thickness and surface of cartilage, FSE outlines the normal and abnormal internal structures of the hyaline cartilage; therefore, both sequences seem to be necessary in a standard MRI protocol for cartilage visualization. In diagnostically ambiguous cases, in which important therapeutic decisions are required, direct MR arthrography is the established imaging standard as an add-on procedure. Despite the social impact and prevalence, until recent years there was a paucity of knowledge about the pathogenesis of cartilage damage. With the introduction of high-resolution MRI with powerful surface coils and fat-suppression techniques, visualization of the articular cartilage is now routinely possible in many joints. After a short summary of the anatomy and physiology of the hyaline cartilage, the different MR imaging methods are discussed and recommended standards are suggested. (orig.)

  19. In vitro morphogenic response of different explants of Gentiana kurroo Royle from Western Himalayas—an endangered medicinal plant

    OpenAIRE

    Sharma, Anshu; Kaur, Rajinder; Sharma, Neha

    2014-01-01

    Micropropagation offers a great potential to produce millions of clonal individuals through tissue culture via induction of morphogenesis. The aim of this work was to obtain an efficient protocol for callus regeneration for Gentiana kurroo Royle. The morphogenic response of different explants (leaves, petioles, roots) varied and responded differently for regeneration according to combinations of growth regulators. The petiole explants were best responding for callus induction and subsequently...

  20. Effect of Macrophage Migration Inhibitory Factor (MIF) in Human Placental Explants Infected with Toxoplasma gondii Depends on Gestational Age

    OpenAIRE

    de Oliveira Gomes, Angelica; de Oliveira Silva, Deise Aparecida; Silva, Neide Maria; de Freitas Barbosa, Bellisa; Franco, Priscila Silva; Angeloni, Mariana Bodini; Fermino, Marise Lopes; Roque-Barreira, Maria Cristina; Bechi, Nicoletta; Paulesu, Luana Ricci; dos Santos, Maria Célia; Mineo, José Roberto; Ferro, Eloisa Amália Vieira

    2011-01-01

    Because macrophage migration inhibitory factor (MIF) is a key cytokine in pregnancy and has a role in inflammatory response and pathogen defense, the objective of the present study was to investigate the effects of MIF in first- and third-trimester human placental explants infected with Toxoplasma gondii. Explants were treated with recombinant MIF, IL-12, interferon-γ, transforming growth factor-β1, or IL-10, followed by infection with T. gondii RH strain tachyzoites. Supernatants of cultured...

  1. 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine destroys dopamine neurons in explants of rat embryo mesencephalon

    International Nuclear Information System (INIS)

    Explants of embryonic rat mesencephalon were grown in organotypic culture. Addition of 10 microM 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to the culture medium for 4 to 7 days resulted in loss of dopamine cell bodies and fiber outgrowths, as observed by fluorescence histochemistry. At the same time, the cultures showed decreased uptake of tritium-labeled dopamine. However, no signs of generalized toxicity were evident when the explant cultures were viewed by light and phase-contrast microscopy. These results show that MPTP exerts a relatively selective destructive action in dopamine neurons in vitro, similar to the action observed in humans and monkeys in vivo. Pargyline (10 microM), a monoamine oxidase inhibitor, protected the dopamine neurons in the explants. Organotypic cultures provide an experimental model for the study of the properties of MPTP in vitro

  2. The Immunosuppressant FTY720 (Fingolimod enhances Glycosaminoglycan depletion in articular cartilage

    Directory of Open Access Journals (Sweden)

    Stradner Martin H

    2011-12-01

    Full Text Available Abstract Background FTY720 (Fingolimod is a novel immunosuppressive drug investigated in clinical trials for organ transplantation and multiple sclerosis. It acts as a functional sphingosine-1-phosphate (S1P receptor antagonist, thereby inhibiting the egress of lymphocytes from secondary lymphoid organs. As S1P is able to prevent IL-1beta induced cartilage degradation, we examined the direct impact of FTY720 on cytokine induced cartilage destruction. Methods Bovine chondrocytes were treated with the bioactive phosphorylated form of FTY720 (FTY720-P in combination with IL-1beta or TNF-alpha. Expression of MMP-1,-3.-13, iNOS and ADAMTS-4,-5 and COX-2 was evaluated using quantitative real-time PCR and western blot. Glycosaminoglycan depletion from cartilage explants was determined using a 1,9-dimethylene blue assay and safranin O staining. Results FTY720-P significantly reduced IL-1beta and TNF-alpha induced expression of iNOS. In contrast FTY720-P increased MMP-3 and ADAMTS-5 mRNA expression. Furthermore depletion of glycosaminoglycan from cartilage explants by IL-1beta and TNF-alpha was significantly enhanced by FTY720-P in an MMP-3 dependent manner. Conclusions Our results suggest that FTY720 may enhance cartilage degradation in pro-inflammatory environment.

  3. In Vitro Propagation of Desmodium gangeticum (L. DC. from Cotyledonary Nodal Explants

    Directory of Open Access Journals (Sweden)

    U R Vishwakarma

    2009-01-01

    Full Text Available An in vitro procedure for rapid multiplication of medicinally important plant Desmodium gangeticum (L. DC. (Fabaceae, has been developed using cotyledonary nodal explant. An average of 9.2 shoots per explant were obtained by culturing cotyledonary nodal explaint on Murashige and Skoog′s medium containing 8.8 μM BAP and 21.2 μM NAA, in combination, within 28 days. These shoots were rooted on half strength MS medium supplemented with IAA 17.1 μM. Rooted plantlets were hardened using 1:1:1 mixture of soil, river sand and vermiculite under green house conditions.

  4. In Vitro Propagation of Desmodium gangeticum (L.) DC. from Cotyledonary Nodal Explants

    OpenAIRE

    U R Vishwakarma; Gurav, A M; Sharma, P.C

    2009-01-01

    An in vitro procedure for rapid multiplication of medicinally important plant Desmodium gangeticum (L.) DC. (Fabaceae), has been developed using cotyledonary nodal explant. An average of 9.2 shoots per explant were obtained by culturing cotyledonary nodal explaint on Murashige and Skoog′s medium containing 8.8 μM BAP and 21.2 μM NAA, in combination, within 28 days. These shoots were rooted on half strength MS medium supplemented with IAA 17.1 μM. Rooted plantlets were hardened using 1:1:1 mix...

  5. 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

    1996-01-01

    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...... developmental stage of the explants. Shoot apices formed the highest percentage of embryogenic callus. There was a difference in the formation of embryogenic callus between leaf explants from in vitro-propagated shoots and greenhouse-grown plants. The best results were obtained from newly formed leaves of in...... vitro-propagated shoots and older leaves of greenhouse-grown plants. Immature inflorescences smaller than 2.5 cm produced a higher percentage of embryogenic callus than larger more mature inflorescences. Embryogenic callus derived from immature inflorescences had the highest regeneration capacity...

  6. Vascularization of engineered cartilage constructs in a mouse model.

    Science.gov (United States)

    Burghartz, Marc; Gehrke, Thomas; Storck, Katharina; Staudenmaier, Rainer; Mandlik, Veronika; Schurr, Christian; Hoang, Nguyen; Hagen, Rudolf; Kleinsasser, Norbert

    2015-02-01

    Tissue engineering of cartilage tissue offers a promising method for reconstructing ear, nose, larynx and trachea defects. However, a lack of sufficient nutrient supply to cartilage constructs limits this procedure. Only a few animal models exist to vascularize the seeded scaffolds. In this study, polycaprolactone (PCL)-based polyurethane scaffolds are seeded with 1 × 10(6) human cartilage cells and implanted in the right hind leg of a nude mouse using an arteriovenous flow-through vessel loop for angiogenesis for the first 3 weeks. Equally seeded scaffolds but without access to a vessel loop served as controls. After 3 weeks, a transposition of the vascularized scaffolds into the groin of the nude mouse was performed. Constructs (verum and controls) were explanted 1 and 6 weeks after transposition. Constructs with implanted vessels were well vascularized. The amount of cells increased in vascularized constructs compared to the controls but at the same time noticeably less extracellular matrix was produced. This mouse model provides critical answers to important questions concerning the vascularization of engineered tissue, which offers a viable option for repairing defects, especially when the desired amount of autologous cartilage or other tissues is not available and the nutritive situation at the implantation site is poor. PMID:25381568

  7. Involvement of ADAMTS5 and hyaluronidase in aggrecan degradation and release from OSM-stimulated cartilage

    Directory of Open Access Journals (Sweden)

    M Durigova

    2011-01-01

    Full Text Available The relative contribution of a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS4 and ADAMTS5 to aggrecan degradation under oncostatin M (OSM stimulation, the role of the ancillary domains of the aggrecanases on their ability to cleave within the chondroitin sulfate (CS-2 region, the role of hyaluronidases (HYAL in stimulating aggrecan release in the absence of proteolysis, and the identity of the hyaluronidase involved in OSM-mediated cartilage breakdown were investigated. Bovine articular cartilage explants were cultured in the presence of interleukin-1beta (IL-1beta, tumor necrosis factor alpha (TNFalpha and/or OSM, or treated with trypsin and/or hyaluronidase. Aggrecan was digested with various domain-truncated isoforms of ADAMTS4 and ADAMTS5. Aggrecan and link protein degradation and release were analyzed by immunoblotting. Aggrecanase and HYAL gene expression were determined. ADAMTS4 was the most inducible aggrecanase upon cytokine stimulation, whereas ADAMTS5 was the most abundant aggrecanase. ADAMTS5 was the most active aggrecanase and was responsible for the generation of an OSM-specific degradation pattern in the CS-2 region. Its ability to cleave at the OSM-specific site adjacent to the aggrecan G3 region was enhanced by truncation of the C-terminal thrombospondin domain, but reduced by further truncation of both the spacer and cysteine-rich domains of the enzyme. OSM has the ability to mediate proteoglycan release through hyaluronan degradation, under conditions where HYAL-2 is the predominant hyaluronidase being expressed. Compared to other catabolic cytokines, OSM exhibits a unique potential at degrading the proteoglycan aggregate, by promoting early robust aggrecanolysis, primarily through the action of ADAMTS5, and hyaluronan degradation.

  8. Failure analysis of explanted sternal wires.

    Science.gov (United States)

    Shih, Chun-Ming; Su, Yea-Yang; Lin, Shing-Jong; Shih, Chun-Che

    2005-05-01

    To classify and understand the mechanisms of surface damages and fracture mechanisms of sternal wires, explanted stainless steel sternal wires were collected from patients with sternal dehiscence following open-heart surgery. Surface alterations and fractured ends of sternal wires were examined and analyzed. Eighty fractured wires extracted from 25 patients from January 1999 to December 2003, with mean implantation interval of 55+/-149 days (range 5-729 days) after cardiac surgery, were studied by various techniques. The extracted wires were cleaned and the fibrotic tissues were removed. Irregularities and fractured ends were assayed by a scanning electron microscopy. After stereomicroscopy and documentation, the explants were cleaned with 1% sodium hypochlorite to remove the blood and tissues and was followed by cleaned with deionized water and alcohol. The explants were examined by stereomicroscopy, and irregularities on surface and fracture surfaces of sternal wires were assayed by scanning electron microscopy, energy dispersive X-ray analysis (EDAX) and X-ray mapping. The explants with surrounding fibrotic tissue were stained and examined with stereomicroscopy and transmission electronic microscopy. Corrosion pits were found on the surface of explanted sternal wires. EDAX and X-ray mapping examinations revealed diminution of nickel concentration in the severely corroded pits on sternal wires. A feature of transgranular cracking was observed for stress corrosion cracking and striation character for typical corrosion fatigue was also identified. TEM examination of tissue showed the metallic particles in phagolysosomes of macrophages inside the surrounding sternal tissue. The synergic effect of hostile environment and the stress could be the precursors of failures for sternal wires. PMID:15576179

  9. CALLUS INDUCTION AND PLANT REGENERATION IN PUNICA GRANATUM L. ?NANA' FROM LEAF EXPLANTS

    Directory of Open Access Journals (Sweden)

    Alireza Bonyanpour

    2013-09-01

    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.

  10. Somatic embryogenesis and massive shoot regeneration from immature embryo explants of tef.

    Science.gov (United States)

    Gugsa, Likyelesh; Kumlehn, Jochen

    2011-01-01

    Tef (Eragrostis tef) provides a major source of human nutrition in the Horn of Africa, but biotechnology has had little impact on its improvement to date. Here, we report the elaboration of an in vitro regeneration protocol, based on the use of immature zygotic embryos as explant. Explant size was an important determinant of in vitro regeneration efficiency, as was the formulation of the culture medium. Optimal results were obtained by culturing 0.2-0.35 mm embryo explants on a medium containing KBP minerals, 9.2-13.8 μM 2,4-dichlorophenoxyacetic acid, 6 mM glutamine, and 0.5% Phytagel. Although this protocol was effective for both the improved cultivar "DZ-01-196" and the landrace "Fesho", the former produced consistently more embryogenic tissue and a higher number of regenerants. An average of more than 2,800 shoots could be obtained from each "DZ-01-196" explant after 12 weeks of in vitro culture. These shoots readily formed roots, and plantlets transferred to soil were able to develop into morphologically normal, fertile plants. This regeneration and multiplication system should allow for the application of a range of biotechnological methods to tef. PMID:22028975

  11. Characterisation of coral explants: a model organism for cnidarian-dinoflagellate studies

    Science.gov (United States)

    Gardner, S. G.; Nielsen, D. A.; Petrou, K.; Larkum, A. W. D.; Ralph, P. J.

    2015-03-01

    Coral cell cultures made from reef-building scleractinian corals have the potential to aid in the pursuit of understanding of the cnidarian-dinoflagellate symbiosis. Various methods have previously been described for the production of cell cultures in vitro with a range of success and longevity. In this study, viable tissue spheroids containing host tissue and symbionts (coral explants) were grown from the tissues of Fungia granulosa. The cultured explants remained viable for over 2 months and showed morphological similarities in tissue structure and internal microenvironment to reef-building scleractinian corals. The photophysiology of the explants (1 week old) closely matched that of the parent coral F. granulosa. This study provides the first empirical basis for supporting the use of coral explants as laboratory models for studying coral symbioses. In particular, it highlights how these small, self-sustaining, skeleton-free models can be useful for a number of molecular, genetic and physiological analyses necessary for investigating host-symbiont interactions at the microscale.

  12. Bovine explant model of degeneration of the intervertebral disc

    Directory of Open Access Journals (Sweden)

    Sivan Sarit

    2008-02-01

    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.

  13. Glucosamine increases hyaluronic acid production in human osteoarthritic synovium explants

    Directory of Open Access Journals (Sweden)

    Uitterlinden EJ

    2008-09-01

    Full Text Available Abstract Background Glucosamine (GlcN used by patients with osteoarthritis was demonstrated to reduce pain, but the working mechanism is still not clear. Viscosupplementation with hyaluronic acid (HA is also described to reduce pain in osteoarthritis. The synthesis of HA requires GlcN as one of its main building blocks. We therefore hypothesized that addition of GlcN might increase HA production by synovium tissue. Methods Human osteoarthritic synovium explants were obtained at total knee surgery and pre-cultured for 1 day. The experimental conditions consisted of a 2 days continuation of the culture with addition of N-Acetyl-glucosamine (GlcN-Ac; 5 mM, glucosamine-hydrochloride (GlcN-HCl; 0.5 and 5 mM, glucose (Gluc; 0.5 and 5 mM. Hereafter HA production was measured in culture medium supernatant using an enzyme-linked binding protein assay. Real time RT-PCR was performed for hyaluronic acid synthase (HAS 1, 2 and 3 on RNA isolated from the explants. Results 0.5 mM and 5 mM GlcN-HCl significantly increased HA production compared to control (approximately 2 – 4-fold, whereas GlcN-Ac had no significant effect. Addition of 5 mM Gluc also increased HA production (approximately 2-fold, but 0.5 mM Gluc did not. Gene expression of the HA forming enzymes HAS 1, 2 and 3 was not altered by the addition of GlcN or Gluc. Conclusion Our data suggest that exogenous GlcN can increase HA production by synovium tissue and is more effective at lower concentrations than Gluc. This might indicate that GlcN exerts its potential analgesic properties through stimulation of synovial HA production.

  14. [Establishment of high frequency regeneration via leaf explants of 'Red Sun' kiwifruit (Actinidia chinensis)].

    Science.gov (United States)

    Zhao, Xupeng; Luo, Keming; Zhou, Yue; Wu, Xiuhua; Yang, Li; Tang, Shaohu

    2013-11-01

    A high efficient in vitro regeneration protocol was developed from leaf explants of the female 'Red Sun' kiwifruit (Actinidia chinensis) and the multiplication coefficient and rooting rate of adventitious buds were also optimized. This method does not require formation of callus tissues which leads to somaclonal variations. The results show that the adventitious buds developing directly from explants tissue were noticed after 30 d of culture. The maximum regeneration frequency of adventitious buds is 100% and 18.67 shoots was observed in each leaf explants when MS medium was supplemented with 3.0 mg/L BA+1.0 mg/L NAA. The optimal culture medium for bud multiplication is MS+2.0 mg/L BA+1.0 mg/L NAA+0.1 mg/L GA3 and the multiplication coefficient reached 8.63. On the rooting medium with 1/2 MS+0.8 mg/L IBA for 15 d, the adventitious plantlets were transferred into matrix perlite supplied with 1/2 MS liquid medium for 15 d and the rooting rate reached 100%. 95 out of 98 plantlets (96.94%) survived acclimatization, producing healthy plants in the greenhouse. Taken together, a highly efficient regeneration method via leaf explants of 'Red Sun' kiwifruit was successfully established. This protocol may be useful for micropropagation and genetic transformation studies of 'Red Sun' kiwifruit. PMID:24701825

  15. Plant regeneration via direct shoot organogenesis from cotyledon explants of Bambara groundnut, Vigna subterranea (L. Verdc.

    Directory of Open Access Journals (Sweden)

    Koné, M.

    2013-01-01

    Full Text Available Bambara groundnut (Vigna subterranea [L.] Verdc. is mainly grown for human consumption. However, several factors limit a wider adoption of the crop including the presence of antinutritional factors in the seeds that lower product quality and protein availability but also the plant susceptibility to pests and diseases. Tissue culture techniques are very scanty in Bambara groundnut and should be developed before carrying out genetic transformation for the crop improvement. Therefore, here, an efficient system for in vitro shoot induction from cotyledons derived from mature seeds has been established. Different types and concentrations of plant growth regulators were used to induce buds in embryo-free cotyledon explants. Cotyledons were cut transversally or longitudinally into three segments: proximal, middle and distal part. The influence of explant orientation on the medium, the type of segment and landrace has then been studied. Benzylaminopurine (3 mg·l-1 alone or combined with α-naphthaleneacetic acid (0.05 mg·l-1 induced multiple shoot formations. The organogenic potential was restricted to the proximal segment of cotyledons. Frequency of bud induction (30% and average number of buds per explant (12 were higher when the adaxial side of the proximal segment was in contact with the medium. Shoot regeneration from cotyledon explants of ten Bambara landraces revealed that the response is genotype-dependent with varieties Ci6, Ci2, Ci4 and Ci15 exhibiting 20 to 30% shoot regeneration and six to ten buds per explant. Regenerated shoot buds excised from explants were elongated and rooted on MS basal medium devoid of plant growth regulators. All rooted plantlets survived to the transfer on a sand soil mixture, and morphologically normal plants were hardened and transferred to greenhouse for further growth to maturity and seed set.

  16. Improved explant method to isolate umbilical cord-derived mesenchymal stem cells and their immunosuppressive properties.

    Science.gov (United States)

    Mori, Yuka; Ohshimo, Jun; Shimazu, Takahisa; He, Haiping; Takahashi, Atsuko; Yamamoto, Yuki; Tsunoda, Hajime; Tojo, Arinobu; Nagamura-Inoue, Tokiko

    2015-04-01

    The umbilical cord (UC) has become one of the major sources of mesenchymal stem cells (MSCs). The common explant method of isolating UC-derived MSCs (UC-MSCs) involves mincing the UCs into small fragments, which are then attached to a culture dish bottom from which the MSCs migrate. However, the fragments frequently float up from the bottom of the dish, thereby reducing the cell recovery rate. To overcome this problem, we demonstrate an improved explant method for UC-MSC isolation, which involves the use of a stainless steel mesh (Cellamigo(®); Tsubakimoto Chain Co.), to protect the tissue from floating after the minced fragments are aligned at regular intervals in culture dishes. The culture medium was refreshed every 3 days and the adherent cells and tissue fragments were harvested using trypsin. The number of UC-MSCs isolated from 1 g of UC using the explant method with Cellamigo was 2.9 ± 1.4 × 10(6)/g, which was significantly higher than that obtained without Cellamigo (0.66 ± 0.53 × 10(6)/g) (n = 6, p < 0.01) when cells reached 80-90% confluence. In addition, the processing and incubation time required to reach 80-90% confluence was reduced in the improved explant method compared with the conventional method. The UC-MSCs isolated using the improved method were positive for CD105, CD73, CD90, and HLA class I expression and negative for CD45 and HLA class II expression. The isolated UC-MSCs efficiently inhibited the responder T cells induced by allogeneic dendritic cells in a mixed lymphocyte reaction. Conclusively, we demonstrated that the use of Cellamigo improves the explant method for isolating UC-MSCs. PMID:25220032

  17. Methylation of inorganic arsenic by murine fetal tissue explants.

    Science.gov (United States)

    Broka, Derrick; Ditzel, Eric; Quach, Stephanie; Camenisch, Todd D

    2016-07-01

    Although it is generally believed that the developing fetus is principally exposed to inorganic arsenic and the methylated metabolites from the maternal metabolism of arsenic, little is known about whether the developing embryo can autonomously metabolize arsenic. This study investigates inorganic arsenic methylation by murine embryonic organ cultures of the heart, lung, and liver. mRNA for AS3mt, the gene responsible for methylation of arsenic, was detected in all embryonic tissue types studied. In addition, methylated arsenic metabolites were generated by all three tissue types. The fetal liver explants yielded the most methylated arsenic metabolites (∼7% of total arsenic/48 h incubation) while the heart, and lung preparations produced slightly greater than 2% methylated metabolites. With all tissues the methylation proceeded mostly to the dimethylated arsenic species. This has profound implications for understanding arsenic-induced fetal toxicity, particularly if the methylated metabolites are produced autonomously by embryonic tissues. PMID:26446802

  18. Chondroptosis in alkaptonuric cartilage.

    Science.gov (United States)

    Millucci, Lia; Giorgetti, Giovanna; Viti, Cecilia; Ghezzi, Lorenzo; Gambassi, Silvia; Braconi, Daniela; Marzocchi, Barbara; Paffetti, Alessandro; Lupetti, Pietro; Bernardini, Giulia; Orlandini, Maurizio; Santucci, Annalisa

    2015-05-01

    Alkaptonuria (AKU) is a rare genetic disease that affects the entire joint. Current standard of treatment is palliative and little is known about AKU physiopathology. Chondroptosis, a peculiar type of cell death in cartilage, has been so far reported to occur in osteoarthritis, a rheumatic disease that shares some features with AKU. In the present work, we wanted to assess if chondroptosis might also occur in AKU. Electron microscopy was used to detect the morphological changes of chondrocytes in damaged cartilage distinguishing apoptosis from its variant termed chondroptosis. We adopted histological observation together with Scanning Electron Microscopy and Transmission Electron Microscopy to evaluate morphological cell changes in AKU chondrocytes. Lipid peroxidation in AKU cartilage was detected by fluorescence microscopy. Using the above-mentioned techniques, we performed a morphological analysis and assessed that AKU chondrocytes undergo phenotypic changes and lipid oxidation, resulting in a progressive loss of articular cartilage structure and function, showing typical features of chondroptosis. To the best of our knowledge, AKU is the second chronic pathology, following osteoarthritis, where chondroptosis has been documented. Our results indicate that Golgi complex plays an important role in the apoptotic process of AKU chondrocytes and suggest a contribution of chondroptosis in AKU pathogenesis. These findings also confirm a similarity between osteoarthritis and AKU. PMID:25336110

  19. PATTERN OF PLANT REGENERATION FROM SHOOT TIP EXPLANTS OF PIGEONPEA (CAJANUS CAJAN L MILLLSP VAR LRG-41

    Directory of Open Access Journals (Sweden)

    T. Raghavendra

    2014-02-01

    Full Text Available An efficient direct shoot bud differentiation and multiple shoot induction from shoot tip explants of pigeon pea (Cajanus cajan L. has been achieved. The frequency of shoot bud regeneration was influenced by the type of explants, genotype and concentrations of cytokinin. Explants viz. shoot tip isolated from 10 day old seedlings showed better explants response Explants were cultured on Murashige and skoog (MS medium augmented with different concentrations of BAP and NAA. Among the various concentrations tested, 2.0mg/l BAP (Benzyl amino purine and 0.1 mg/l Napthalene acetic acid (NAA were found to be the best for maximum shoot bud differentiation. Percentage, as well as the number of shoots per explant showing differentiation of shoot buds was higher on MS media supplement with BAP and optimal BAP concentration for shoot regeneration was 2mg/l. The elongated shoots were successfully rooted on MS medium containing different concentrations of auxins. Among them indole buteric acid (IBA at 1.0mg/l induced maximum frequency of rooting. Regenerated plants were successfully established in soil where 91% of them have been developed into morphologically normal and fertile plants. This method can thus be advantageously applied in the production of transgenic pigeon pea plants.

  20. PATTERN OF PLANT REGENERATION FROM SHOOT TIP EXPLANTS OF PIGEONPEA (CAJANUS CAJAN L MILLLSP VAR LRG-41

    Directory of Open Access Journals (Sweden)

    T. Raghavendra

    2014-03-01

    Full Text Available An efficient direct shoot bud differentiation and multiple shoot induction from shoot tip explants of pigeon pea (Cajanus cajan L. has been achieved. The frequency of shoot bud regeneration was influenced by the type of explants, genotype and concentrations of cytokinin. Explants viz. shoot tip isolated from 10 day old seedlings showed better explants response Explants were cultured on Murashige and skoog (MS medium augmented with different concentrations of BAP and NAA. Among the various concentrations tested, 2.0mg/l BAP (Benzyl amino purine and 0.1 mg/l Napthalene acetic acid (NAA were found to be the best for maximum shoot bud differentiation. Percentage, as well as the number of shoots per explant showing differentiation of shoot buds was higher on MS media supplement with BAP and optimal BAP concentration for shoot regeneration was 2mg/l. The elongated shoots were successfully rooted on MS medium containing different concentrations of auxins. Among them indole buteric acid (IBA at 1.0mg/l induced maximum frequency of rooting. Regenerated plants were successfully established in soil where 91% of them have been developed into morphologically normal and fertile plants. This method can thus be advantageously applied in the production of transgenic pigeon pea plants.

  1. Synergy between Piezo1 and Piezo2 channels confers high-strain mechanosensitivity to articular cartilage

    Science.gov (United States)

    Lee, Whasil; Leddy, Holly A.; Chen, Yong; Lee, Suk Hee; Zelenski, Nicole A.; McNulty, Amy L.; Wu, Jason; Beicker, Kellie N.; Coles, Jeffrey; Zauscher, Stefan; Grandl, Jörg; Sachs, Frederick; Liedtke, Wolfgang B.

    2014-01-01

    Diarthrodial joints are essential for load bearing and locomotion. Physiologically, articular cartilage sustains millions of cycles of mechanical loading. Chondrocytes, the cells in cartilage, regulate their metabolic activities in response to mechanical loading. Pathological mechanical stress can lead to maladaptive cellular responses and subsequent cartilage degeneration. We sought to deconstruct chondrocyte mechanotransduction by identifying mechanosensitive ion channels functioning at injurious levels of strain. We detected robust expression of the recently identified mechanosensitive channels, PIEZO1 and PIEZO2. Combined directed expression of Piezo1 and -2 sustained potentiated mechanically induced Ca2+ signals and electrical currents compared with single-Piezo expression. In primary articular chondrocytes, mechanically evoked Ca2+ transients produced by atomic force microscopy were inhibited by GsMTx4, a PIEZO-blocking peptide, and by Piezo1- or Piezo2-specific siRNA. We complemented the cellular approach with an explant-cartilage injury model. GsMTx4 reduced chondrocyte death after mechanical injury, suggesting a possible therapy for reducing cartilage injury and posttraumatic osteoarthritis by attenuating Piezo-mediated cartilage mechanotransduction of injurious strains. PMID:25385580

  2. Fracture of articular cartilage.

    Science.gov (United States)

    Chin-Purcell, M V; Lewis, J L

    1996-11-01

    Crack formation and propagation is a significant element of the degeneration process in articular cartilage. In order to understand this process, and separate the relative importance of structural overload and material failure, methods for measuring the fracture toughness of cartilage are needed. In this paper, two such methods are described and used to measure fracture properties of cartilage from the canine patella. A modified single edge notch (MSEN) specimen was used to measure J, and a trouser tear test was used to measure T, both measures of fracture toughness with units of kN/m. A pseudo-elastic modulus was also obtained from the MSEN test. Several potential error sources were examined, and results for the MSEN test compared with another method for measuring the fracture parameter for urethane rubber. Good agreement was found. The two test methods were used to measure properties of cartilage from the patellae of 12 canines: 4-9 specimens from each of 12 patellae, with 5 right-left pairs were tested. Values of J ranged from 0.14-1.2 kN/m. J values correlated with T and were an average of 1.7 times larger than T. A variety of failure responses was seen in the MSEN tests, consequently a grade of 0 to 3 was assigned to each test, where 0 represented a brittle-like crack with minimal opening and 3 represented plastic flow with no crack formation. The initial cracks in 12/82 specimens did not propagate and were assigned to grade 3. The method for reducing data in the MSEN test assumed pseudo-elastic response and could not be used for the grade 3 specimens. Stiffness did not correlate with J. Neither J nor T was statistically different between right-left pairs, but varied between animals. The test methods appear useful for providing a quantitative measure of fracture toughness for cartilage and other soft materials. PMID:8950659

  3. The Effects of Extracellular Matrix on Tissue Engineering Construction of Cartilage in Vitro

    Institute of Scientific and Technical Information of China (English)

    YU Li; LI Fa-tao; TANG Ming-qiao; YAN Wei-qun

    2006-01-01

    The effects of various cartilage extracellular matrix on the construction of rabbit growth plate cartilage tissue in vitro were studied. The results show that collagen, proteoglycan and hyaluronic acid can promote the growth of cultured chondrocytes but the effects of various cartilage extracellular matrix(ECM)on chondrocyte differentiation are different. Collagen can promote the hypertrophy of chondrocytes while proteoglycan and hyaluronic acid inhibit the transition of mature chondrocytes into hypertrophied chondrocytes.

  4. 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

    2005-08-01

    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

  5. In Vitro Engineering of High Modulus Cartilage-Like Constructs.

    Science.gov (United States)

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

    2016-04-01

    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. PMID:26850081

  6. In Vitro Engineering of High Modulus Cartilage-Like Constructs

    Science.gov (United States)

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

    2016-01-01

    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. PMID:26850081

  7. Colonies in engineered articular cartilage express superior differentiation.

    Science.gov (United States)

    Selvaratnam, L; Abd Rahim, S; Kamarul, T; Chan, K Y; Sureshan, S; Penafort, R; Ng, C L L

    2005-07-01

    In view of poor regeneration potential of the articular cartilage, in-vitro engineering of cartilage tissue offers a promising option for progressive joint disease. This study aims to develop a biologically engineered articular cartilage for autologous transplantation. The initial work involved determination of chondrocyte yield and viability, and morphological analysis. Cartilage was harvested from the knee, hip and shoulder joints of adult New Zealand white rabbits and chondrocytes were isolated by enzymatic digestion of the extra-cellular matrix before serial cultivation in DMEM/Ham's F12 media as monolayer cultures. No differences were noted in cell yield. Although chondrocytes viability was optimal (>93%) following harvest from native cartilage, their viability tended to be lowered on passaging. Chondrocytes aggregated in isogenous colonies comprising ovoid cells with intimate intracellular contacts and readily exhibited Safranin-O positive matrix; features typically associated with articular cartilage in-vivo. However, chondrocytes also existed concurrently in scattered bipolar/multipolar forms lacking Safranin-O expression. Therefore, early data demonstrated successful serial culture of adult chondrocytes with differentiated morphology seen in established chondrocyte colonies synthesizing matrix proteoglycans. PMID:16381284

  8. Somatic Embryogenesis in Peach-Palm (Bactris gasipaes) Using Different Explant Sources.

    Science.gov (United States)

    Steinmacher, Douglas A; Heringer, Angelo Schuabb; Jiménez, Víctor M; Quoirin, Marguerite G G; Guerra, Miguel P

    2016-01-01

    Peach palm (Bactris gasipaes Kunth) is a member of the family Arecaceae and is a multipurpose but underutilized species. Nowadays, fruit production for subsistence and local markets, and heart-of-palm production for local, national, and international markets are the most important uses of this plant. Conventional breeding programs in peach palm are long-term efforts due to the prolonged generation time, large plant size, difficulties with controlled pollination and other factors. Although it is a caespitose palm, its propagation is currently based on seeds, as off-shoots are difficult to root. Hence, tissue culture techniques are considered to be the most likely strategy for efficient clonal plantlet regeneration of this species. Among various techniques, somatic embryogenesis offers the advantages of potential automated large-scale production and putative genetic stability of the regenerated plantlets. The induction of somatic embryogenesis in peach palm can be achieved by using different explant sources including zygotic embryos, immature inflorescences and thin cell layers from the young leaves and shoot meristems. The choice of a particular explant depends on whether clonal propagation is desired or not, as well as on the plant conditions and availability of explants. Protocols to induce and express somatic embryogenesis from different peach palm explants, up to acclimatization of plantlets, are described in this chapter. PMID:26619867

  9. Effect of explants, hormonal combination and genotype on micropropagation of pepper

    Directory of Open Access Journals (Sweden)

    Z. Moheb Mohamadi

    2014-07-01

    Full Text Available Pepper (Capsicum annuum is generally propagated from seeds. In Iran, imported hybrid seeds are used for this purpose. Germination of the greenhouse pepper seeds and stabilization of their seedlings takes a long time. But, tissue culture technique could rapidly propagate this plant with a homogeneous genetic structure in a large scale. An attempt was made in this study to evaluate the effects of different genotypes, explants and growth regulators on callus production and regeneration and introduce the appropriate protocol for micropropagation of commercial varieties of pepper in Iran. Analysis of variance indicated that effect of different genotypes, interaction of various explants and combination of hormones and interaction between genotypes, explants and hormones were significant on the evaluated traits at 1% level. Based on the results, for micropropagation of pepper with leaves and cotyledons, combination of 1 mg/l IAA and 5 mg/l BAP was selected. It seems that using leaf explants for micropropagation of pepper in large scale in greenhouses would be suitable.

  10. A RIFAMPICINA NA DESCONTAMINAÇÃO BACTERIANA DE EXPLANTES DE MAMOEIRO PROVENIENTES DO CAMPO

    Directory of Open Access Journals (Sweden)

    GIOVANNI RODRIGUES VIANNA

    1997-01-01

    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.

  11. Cartilage analysis by reflection spectroscopy

    Science.gov (United States)

    Laun, T.; Muenzer, M.; Wenzel, U.; Princz, S.; Hessling, M.

    2015-07-01

    A cartilage bioreactor with analytical functions for cartilage quality monitoring is being developed. For determining cartilage composition, reflection spectroscopy in the visible (VIS) and near infrared (NIR) spectral region is evaluated. Main goal is the determination of the most abundant cartilage compounds water, collagen I and collagen II. Therefore VIS and NIR reflection spectra of different cartilage samples of cow, pig and lamb are recorded. Due to missing analytical instrumentation for identifying the cartilage composition of these samples, typical literature concentration values are used for the development of chemometric models. In spite of these limitations the chemometric models provide good cross correlation results for the prediction of collagen I and II and water concentration based on the visible and the NIR reflection spectra.

  12. Cells that emerge from embryonic explants produce fibers of type IV collagen

    OpenAIRE

    1985-01-01

    Double immunofluorescence staining experiments designed to examine the synthesis and deposition of collagen types I and IV in cultured explants of embryonic mouse lung revealed the presence of connective tissue-like fibers that were immunoreactive with anti-type IV collagen antibodies. This observation is contrary to the widely accepted belief that type IV collagen is found only in sheet-like arrangements beneath epithelia or as a sheath-like layer enveloping bundles of nerve or muscle cells....

  13. Protoplast Isolation in Lupin ( Lupinus mutabilis Sweet): Determination of Optimum Explant Sources and Isolation Conditions

    OpenAIRE

    BABAOĞLU, Mehmet

    2000-01-01

    Effects of cultural factors on the yield, viability and division of protoplasts were investigated in Lupinus mutabilis Sweet containing a high protein content as well as a reasonable oil content which may make this species an alternative crop to soybean in Turkey. Explants from different in vitro seedling parts were evaluated on the suitability of protoplast isolation and viability. Leaf mesophyll was the most suitable tissue as a protoplast source. Pectinases as well as cellulases were es...

  14. Shoot regeneration from leaf explants of Cydonia oblonga cultivars in vitro

    OpenAIRE

    Zalunskaitė, Irena; Kavaliauskaitė, Danguolė; Vinskienė, Jurgita; Revin, Viktor V.; Stanys, Vidmantas

    2007-01-01

    Regeneration experiment was started with aim to develop an efficient plant regeneration system for Cydonia oblonga Mill. cultivars. Variation of organogenic capacity of leaves depending on their position on the microshoot, photoperiod influence on differences in regeneration and interaction of these two factors was investigated in this experiment. Shoots were regenerated from leaf obtained from microshoots culture in vitro. Leaf explants of three genotypes were collected from microshoots and ...

  15. Effect of explants, hormonal combination and genotype on micropropagation of pepper

    OpenAIRE

    Z. Moheb Mohamadi; M. Talebi; B.E. Sayed-Tabatabaei; G. Khaksar

    2014-01-01

    Pepper (Capsicum annuum) is generally propagated from seeds. In Iran, imported hybrid seeds are used for this purpose. Germination of the greenhouse pepper seeds and stabilization of their seedlings takes a long time. But, tissue culture technique could rapidly propagate this plant with a homogeneous genetic structure in a large scale. An attempt was made in this study to evaluate the effects of different genotypes, explants and growth regulators on callus production and regeneration and intr...

  16. Micropropagation of Origanum acutidens (HAND.-MAZZ.) IETSWAART Using Stem Node Explants

    OpenAIRE

    Mehmet Ugur Yildirim

    2013-01-01

    Origanum acutidens (HAND.-MAZZ.) IETSWAART is a promising ornamental plant that can be widely used in landscape management. It is endemic to Eastern Anatolian region of Turkey. Tissue culture has not been used to micropropagate it. The study reports stem node explants from one-week-old seedlings of the plant for successful micropropagation. The stem nodes were cultured on MS medium containing 0.6, 1.2, 1.8, and 2.4 mg/L BAP with 0.2 mg/L NAA. Visible effects of culture media on shoot prolifer...

  17. Articular cartilage stem cell signalling

    OpenAIRE

    Karlsson, Camilla; Lindahl, Anders

    2009-01-01

    The view of articular cartilage as a non-regeneration organ has been challenged in recent years. The articular cartilage consists of distinct zones with different cellular and molecular phenotypes, and the superficial zone has been hypothesized to harbour stem cells. Furthermore, the articular cartilage demonstrates a distinct pattern regarding stem cell markers (that is, Notch-1, Stro-1, and vascular cell adhesion molecule-1). These results, in combination with the positive identification of...

  18. Transcriptomic profiling of cartilage ageing

    OpenAIRE

    Mandy Jayne Peffers; Xuan Liu; Peter David Clegg

    2014-01-01

    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 dono...

  19. Encapsulation of in vitro-derived explants: an innovative tool for nurseries.

    Science.gov (United States)

    Standardi, Alvaro; Micheli, Maurizio

    2013-01-01

    The encapsulation technology consists of the inclusion of some millimeter-long plant portions in a nutritive and protective matrix. This technology represents a further and promising tool for exchange of plant material between private and public plant tissue culture laboratories, for short- and medium-term storage of valuable plant material and for use of in vitro-derived or micropropagated propagules directly in farm or in nurseries. After encapsulation, transport, storage and sowing in aseptic conditions, the enclosed explants (capsules) may evolve in shoots (regrowth) and be employed for subsequent micropropagation or culture in vitro. When the encapsulated explant evolves in plantlet (conversion) in in vitro or in vivo conditions, the product of the encapsulation is defined as synthetic seed or artificial seed or synseed. The different evolution of the encapsulated plant material depends on tissue or plant material, genotype, nutritive and culture conditions, and treatments before or after encapsulation. In order to make economical the application of the encapsulation technology in the commercial nursery, research is looking for efficient automation or mechanization of the procedure and for preparation of the encapsulable explants. PMID:23179716

  20. Type II collagen peptide is able to accelerate embryonic chondrocyte differentiation: an association with articular cartilage matrix resorption in osteoarthrosis

    Directory of Open Access Journals (Sweden)

    Elena Vasil'evna Chetina

    2010-01-01

    Conclusion. The effect of CP on gene expression and collagen decomposition activity depends on the morphotype of embryonic chondrocytes. Lack of effect of CP on collagen decomposition activity in both the embryonic hypertrophic chondrocytes and the cartilage explants from OA patients supports the hypothesis that the hypertrophic morphotype is a dominant morphotype of articular chondrocytes in OA. Moreover, collagen decomposition products can be involved in the resorption of matrix in OA and in the maintenance of chronic nature of the pathology.

  1. Comparative effects of plant growth regulators on leaf and stem explants of Labisia pumila var.alata

    Institute of Scientific and Technical Information of China (English)

    Anna Pick Kiong LING; Kinn Poay TAN; Sobri HUSSEIN

    2013-01-01

    Objective:Labisia pumi/a var.a/ata,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.Methods: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.Results: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.Conclusions: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.

  2. Making post-mortem implantable cardioverter defibrillator explantation safe

    DEFF Research Database (Denmark)

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

    2009-01-01

    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...

  3. A novel coculture model of porcine central neuroretina explants and retinal pigment epithelium cells

    Science.gov (United States)

    Di Lauro, Salvatore; Rodriguez-Crespo, David; Gayoso, Manuel J.; Garcia-Gutierrez, Maria T.; Pastor, J. Carlos; Srivastava, Girish K.

    2016-01-01

    Purpose To develop and standardize a novel organ culture model using porcine central neuroretina explants and RPE cells separated by a cell culture membrane. Methods RPE cells were isolated from porcine eyes, expanded, and seeded on the bottom of cell culture inserts. Neuroretina explants were obtained from the area centralis and cultured alone (controls) on cell culture membranes or supplemented with RPE cells in the same wells but physically separated by the culture membrane. Finally, cellular and tissue specimens were processed for phase contrast, cyto-/histological, and immunochemical evaluation. Neuroretina thickness was also determined. Results Compared to the neuroretinas cultured alone, the neuroretinas cocultured with RPE cells maintained better tissue structure and cellular organization, displayed better preservation of photoreceptors containing rhodopsin, lower levels of glial fibrillary acidic protein immunoexpression, and preservation of cellular retinaldehyde binding protein both markers of reactive gliosis. Neuroretina thickness was significantly greater in the cocultures. Conclusions A coculture model of central porcine neuroretina and RPE cells was successfully developed and standardized. This model mimics a subretinal space and will be useful in studying interactions between the RPE and the neuroretina and to preclinically test potential therapies. PMID:27081295

  4. Strain-dependent oxidant release in articular cartilage originates from mitochondria.

    Science.gov (United States)

    Brouillette, M J; Ramakrishnan, P S; Wagner, V M; Sauter, E E; Journot, B J; McKinley, T O; Martin, J A

    2014-06-01

    Mechanical loading is essential for articular cartilage homeostasis and plays a central role in the cartilage pathology, yet the mechanotransduction processes that underlie these effects remain unclear. Previously, we showed that lethal amounts of reactive oxygen species (ROS) were liberated from the mitochondria in response to mechanical insult and that chondrocyte deformation may be a source of ROS. To this end, we hypothesized that mechanically induced mitochondrial ROS is related to the magnitude of cartilage deformation. To test this, we measured axial tissue strains in cartilage explants subjected to semi-confined compressive stresses of 0, 0.05, 0.1, 0.25, 0.5, or 1.0 MPa. The presence of ROS was then determined by confocal imaging with dihydroethidium, an oxidant sensitive fluorescent probe. Our results indicated that ROS levels increased linearly relative to the magnitude of axial strains (r(2) = 0.87, p 40%. By contrast, hydrostatic stress, which causes minimal tissue strain, had no significant effect. Cell-permeable superoxide dismutase mimetic Mn(III)tetrakis (1-methyl-4-pyridyl) porphyrin pentachloride significantly decreased ROS levels at 0.5 and 0.25 MPa. Electron transport chain inhibitor, rotenone, and cytoskeletal inhibitor, cytochalasin B, significantly decreased ROS levels at 0.25 MPa. Our findings strongly suggest that ROS and mitochondrial oxidants contribute to cartilage mechanobiology. PMID:23896937

  5. Extracorporeal shockwave therapy promotes chondrogenesis in cartilage tissue engineering: A hypothesis based on previous evidence.

    Science.gov (United States)

    Ji, Qiaodan; He, Chengqi

    2016-06-01

    The dearth of intrinsic regenerative capacity of articular cartilage makes it a challenge to deal with the cartilage defects. Among all the recommended clinical options, cartilage tissue engineering (CTE) which is highlighted of dominant features and less drawbacks for functional cartilage restoration, has been emphasized recently. Shock waves, a mode of therapeutic mechanical forces, utilized in extracorporeal shockwave therapy (ESWT), is hypothesized to enhance proliferation, chondrogenic differentiation, and cartilage extracellular matrix production of target cells seeded on bioactive scaffolds. The hypothesis is firstly based on cellular mechanotransduction by which cells convent the shockwave mechanical signals into biochemical responses via integrins, iron channels, cytoskeletal filaments, growth factor receptors and nuclei. Secondly, by modulating gene expression and up-regulating the release of various growth factors which are of vital importance in three-dimensional cartilage culture environment, ESWT holds a promising potential to favor the cell sources (e.g. chondrocytes and stem cells) to mimic the optimal functional cartilage. In all, on the basis of cellular mechanotransduction and previous evidence, the hypothesis is developed to support the beneficial effects of ESWT on chondrogenesis in CTE. If this hypothesis is confirmed, shockwaves may allow a better success in combination with other stimulating factors for cartilage repair. There is a paucity of studies investigating the assistant role of shockwave stimulation in CTE. Further research is required to elucidate the mechanisms, and explore effectiveness and appropriate protocols of this novel stimulative factor in cartilage tissue engineering. PMID:27142133

  6. Use of Adult Stem Cells for Cartilage Tissue Engineering: Current Status and Future Developments

    Directory of Open Access Journals (Sweden)

    Catherine Baugé

    2015-01-01

    Full Text Available Due to their low self-repair ability, cartilage defects that result from joint injury, aging, or osteoarthritis, are the most often irreversible and are a major cause of joint pain and chronic disability. So, in recent years, researchers and surgeons have been working hard to elaborate cartilage repair interventions for patients who suffer from cartilage damage. However, current methods do not perfectly restore hyaline cartilage and may lead to the apparition of fibro- or hypertrophic cartilage. In the next years, the development of new strategies using adult stem cells, in scaffolds, with supplementation of culture medium and/or culture in low oxygen tension should improve the quality of neoformed cartilage. Through these solutions, some of the latest technologies start to bring very promising results in repairing cartilage from traumatic injury or chondropathies. This review discusses the current knowledge about the use of adult stem cells in the context of cartilage tissue engineering and presents clinical trials in progress, as well as in the future, especially in the field of bioprinting stem cells.

  7. Shear loading of costal cartilage

    CERN Document Server

    Subit, Damien

    2014-01-01

    A series of tests were performed on a single post-mortem human subject at various length scales. First, tabletop tests were performed. Next, the ribs and intercostal muscles were tested with the view to characterize the load transfer between the ribs. Finally, the costal cartilage was tested under shear loading, as it plays an important in the transfer of the load between the ribs and the sternum. This paper reports the results of dynamic shear loading tests performed on three samples of costal cartilage harvested from a single post-mortem human subject, as well as the quantification of the effective Young's modulus estimated from the amount of cartilage calcification.

  8. Electrospun Cartilage-Derived Matrix Scaffolds for Cartilage Tissue Engineering

    OpenAIRE

    Garrigues, N. William; Little, Dianne; Sanchez-Adams, Johannah; David S Ruch; Guilak, Farshid

    2014-01-01

    Macroscale scaffolds created from cartilage-derived matrix (CDM) demonstrate chondroinductive properties, but many fabrication methods do not allow for control of nanoscale architecture. In this regard, electrospun scaffolds have shown significant promise for cartilage tissue engineering. However, nanofibrous materials generally exhibit a relatively small pore size and require techniques such as multi-layering or the inclusion of sacrificial fibers to enhance cellular infiltration. The object...

  9. Engineered cartilage covered ear implants for auricular cartilage reconstruction.

    Science.gov (United States)

    Lee, Sang Jin; Broda, Christopher; Atala, Anthony; Yoo, James J

    2011-02-14

    Cartilage tissues are often required for auricular tissue reconstruction. Currently, alloplastic ear-shaped medical implants composed of silicon and polyethylene are being used clinically. However, the use of these implants is often associated with complications, including inflammation, infection, erosion, and dislodgement. To overcome these limitations, we propose a system in which tissue-engineered cartilage serves as a shell that entirely covers the alloplastic implants. This study investigated whether cartilage tissue, engineered with chondrocytes and a fibrin hydrogel, would provide adequate coverage of a commercially used medical implant. To demonstrate the in vivo stability of cell-fibrin constructs, we tested variations of fibrinogen and thrombin concentration as well as cell density. After implantation, the retrieved engineered cartilage tissue was evaluated by histo- and immunohistochemical, biochemical, and mechanical analyses. Histomorphological evaluations consistently showed cartilage formation over the medical implants with the maintenance of dimensional stability. An initial cell density was determined that is critical for the production of matrix components such as glycosaminoglycans (GAG), elastin, type II collagen, and for mechanical strength. This study shows that engineered cartilage tissues are able to serve as a shell that entirely covers the medical implant, which may minimize the morbidity associated with implant dislodgement. PMID:21182236

  10. „IN VITRO” EFFECT OF SOME INDUSTRIAL BY-PRODUCTS ON LAVANDULA ANGUSTIFOLIA MILL. EXPLANT GROWTH

    Directory of Open Access Journals (Sweden)

    Corneliu Tanase

    2013-12-01

    Full Text Available After many studies, it was observed that lavender has many therapeutic effects, such as sedation, activities spasmolytic, antiviral, antibacterial. Thus, given the importance of lavender in different areas of human life, in the present study, we studied the influence of natural products bioregulatoars separated from industrial by-products on some lavender stems explants. These explants were inoculated in vitro on MS nutrient media. In these culture media were added polyphenolic extracts obtained from spruce bark and hemp shives, and evaluated their influence on lavender stems explants. The results obtained were compared with those obtained for the control variant, where MS culture medium was used as standard. It was found that the addition of aqueous extract from spruce bark of concentration of 130 mg GAE / L, in the growth of explants of Lavandula angustifolia Mill, an increase in the elongation of the main stem, number of leaves formed, the amount of photoassimilating pigments synthesized and causes the phenomenon of shoots formation. At a higher concentration of the extract (26 mgGAE/100g values are lower.

  11. A novel corneal explant model system to evaluate antiviral drugs against feline herpesvirus type 1 (FHV-1).

    Science.gov (United States)

    Pennington, Matthew R; Fort, Michael W; Ledbetter, Eric C; Van de Walle, Gerlinde R

    2016-06-01

    Feline herpesvirus type-1 (FHV-1) is the most common viral cause of ocular surface disease in cats. Many antiviral drugs are used to treat FHV-1, but require frequent topical application and most lack well-controlled in vivo studies to justify their clinical use. Therefore, better validation of current and novel treatment options are urgently needed. Here, we report on the development of a feline whole corneal explant model that supports FHV-1 replication and thus can be used as a novel model system to evaluate the efficacy of antiviral drugs. The anti-herpes nucleoside analogues cidofovir and acyclovir, which are used clinically to treat ocular herpesvirus infection in cats and have previously been evaluated in traditional two-dimensional feline cell cultures in vitro, were evaluated in this explant model. Both drugs suppressed FHV-1 replication when given every 12 h, with cidofovir showing greater efficacy. In addition, the potential efficacy of the retroviral integrase inhibitor raltegravir against FHV-1 was evaluated in cell culture as well as in the explant model. Raltegravir was not toxic to feline cells or corneas, and most significantly, inhibited FHV-1 replication at 500 µM in both systems. Importantly, this drug was effective when given only once every 24 h. Taken together, our data indicate that the feline whole corneal explant model is a useful tool for the evaluation of antiviral drugs and, furthermore, that raltegravir appears a promising novel antiviral drug to treat ocular herpesvirus infection in cats. PMID:26959283

  12. Relative contribution of matrix metalloprotease and cysteine protease activities to cytokine-stimulated articular cartilage degradation

    DEFF Research Database (Denmark)

    Sondergaard, B C; Henriksen, K; Wulf, H;

    2006-01-01

    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...... vivo in CK null mice. CONCLUSION: Inhibition of MMP activity reduced both proteoglycan loss and type II collagen degradation. 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...

  13. Spinal cord explants use carbon nanotube interfaces to enhance neurite outgrowth and to fortify synaptic inputs.

    Science.gov (United States)

    Fabbro, Alessandra; Villari, Ambra; Laishram, Jummi; Scaini, Denis; Toma, Francesca M; Turco, Antonio; Prato, Maurizio; Ballerini, Laura

    2012-03-27

    New developments in nanotechnology are increasingly designed to modulate relevant interactions between nanomaterials and neurons, with the aim of exploiting the physical properties of synthetic materials to tune desired and specific biological processes. Carbon nanotubes have been applied in several areas of nerve tissue engineering to study cell behavior or to instruct the growth and organization of neural networks. Recent reports show that nanotubes can sustain and promote electrical activity in networks of cultured neurons. However, such results are usually limited to carbon nanotube/neuron hybrids formed on a monolayer of dissociated brain cells. In the present work, we used organotypic spinal slices to model multilayer tissue complexity, and we interfaced such spinal segments to carbon nanotube scaffolds for weeks. By immunofluorescence, scanning and transmission electronic microscopy, and atomic force microscopy, we investigated nerve fiber growth when neuronal processes exit the spinal explant and develop in direct contact to the substrate. By single-cell electrophysiology, we investigated the synaptic activity of visually identified ventral interneurons, within the ventral area of the explant, thus synaptically connected, but located remotely, to the substrate/network interface. Here we show that spinal cord explants interfaced for weeks to purified carbon nanotube scaffolds expand more neuronal fibers, characterized by different mechanical properties and displaying higher growth cones activity. On the other hand, exploring spontaneous and evoked synaptic activity unmasks an increase in synaptic efficacy in neurons located at as far as 5 cell layers from the cell-substrate interactions. PMID:22339712

  14. Microvesicle-mediated release of soluble LH/hCG receptor (LHCGR from transfected cells and placenta explants

    Directory of Open Access Journals (Sweden)

    Randeva Harpal

    2011-05-01

    Full Text Available Abstract Placental hCG and pitutary LH transduce signals in target tissues through a common receptor (LHCGR. We demonstrate that recombinant LHCGR proteins which include the hormone-binding domain are secreted from transfected cells and that natural LHCGR is also secreted from human placental explants. LHCGR recombinant proteins representing varying lengths of the N-terminal extracellular domain were expressed in Chinese Hamster Ovary cells in suspension culture. Secretion was minimal up to 72h but by 96h 24-37% of the LHCGR had been released into the culture medium. The secreted proteins were folded and sensitive to glycosidases suggesting N-linked glycosylation. Secretion was independent of recombinant size and was mediated via structurally defined membrane vesicles (50-150nm. Similarly cultured human early pregnancy placental explants also released LHCGR via microvesicles. These studies provide the first experimental evidence of the possible mechanistic basis of the secretion of LHCGR.

  15. DIRECT IN VITRO SHOOTS PROLIFERATION OF CHICK PEA (CICER ARIETINUM L. FROM SHOOT TIP EXPLANTS INDUCED BY THIDIAZURON

    Directory of Open Access Journals (Sweden)

    Shaheena Parveen

    2012-01-01

    Full Text Available A rapid, simple and efficient protocol for direct in vitro multiple shoot induction and plantlet regeneration was achieved from shoot tip explants of Cicer arietinum. The shoot tips were cultured on MS medium fortified with Thidiazuron (TDZ (1.0-7.0 mg/L for multiple shoot induction. Multiple shoots proliferation was best observed at 3.0 mg/L TDZ from the shoot tip explants within three weeks of culture. Shoot number per explant ranged between 2 and 10. Individual shoots were aseptically excised and sub cultured in the same media for shoot elongation. The elongated shoots were transferred to Indole Butyric Acid (IBA (1.0mg/L–5.0mg/L for root induction. Rooting was observed within two weeks of culture. Rooted plantlets were successfully hardened under culture conditions and subsequently established in the field conditions. The recorded survival rate of the plants was 86%. Plants looked healthy with no visually detectable phenotypic variations.

  16. In Vitro Shoot Regeneration of NAA-Pulse Treated Plumular Leaf Explants of Cowpea

    Directory of Open Access Journals (Sweden)

    Muhammad AASIM

    2010-06-01

    Full Text Available Cowpea (Vigna unguiculata L. is an economically important grain legume crop and is an important source of dietary protein in many of the developing countries. The present study reports the effect of pulse treatment duration, concentration of NAA and presence of NAA in the culture medium on shoot regeneration from plumular leaf explant of Turkish cowpea cv. �Akkiz� and �Karagoz�. Pulse treatment of mature embryos with 20 mg l-1 NAA for 1 and 3 weeks followed by culturing of plumular leaf explant on MS medium containing 0.25, 0.50 and 1.0 BAP with 1.0, 2.0 and 4.0 mg l-1 NAA promoted somatic embryogenesis in both cultivars. Longer duration of pulse treatment was deleterious resulting in browning and consequently death of the embryos on explants. Pulse treatment with 20 mg l-1 NAA for one week was less deleterious and developed two plantlets after the explants were transferred to MS0 medium after 6 weeks through somatic embryogenesis in cv. �Akkiz�. Pulse treatment with 10 mg l-1 NAA for 1 week showed 33.33-50.00 % and 25.00-50.00% shoot regeneration frequency in cv. �Akkiz� and �Karagoz� respectively on MS medium containing 0.25-1.00 mg l-1 BAP. Maximum number of 2.50 shoots each per explant were recorded in cv. �Akkiz� and �Karagoz� on MS medium containing 1.00 and 0.50 mg l-1 BAP respectively. Contrarily, maximum shoot length of 8.98 cm of cv. �Akkiz� and 9.42 cm of cv. �Karagoz� was recorded on MS medium containing 0.50 mg l-1 BAP and 1.00 mg l-1 BAP respectively. Regenerated shoots were rooted on MS medium containing 0.5 mg l-1 IBA and and acclimatized in growth room at room temprature where they produced viable seeds.

  17. In vitro generation of mechanically functional cartilage grafts based on adult human stem cells and 3D-woven poly(ε-caprolactone) scaffolds

    OpenAIRE

    Valonen, P.K.; Moutos, F.T.; Kusanagi, A.; Moretti, M.; Diekman, B.O.; Welter, J. F.; Caplan, A I; Guilak, F.; Freed, L.E.

    2010-01-01

    Three-dimensionally woven poly(ε-caprolactone)(PCL) scaffolds were combined with adult human mesenchymal stem cells (hMSC) to engineer mechanically functional cartilage constructs in vitro. The specific objectives were to: (i) produce PCL scaffolds with cartilage-like mechanical properties, (ii) demonstrate that hMSCs formed cartilage after 21-days of culture on PCL scaffolds, and (iii) study effects of scaffold structure (loosely vs. tightly woven), culture vessel (static dish vs. oscillatin...

  18. MR imaging of cartilage repair procedures

    International Nuclear Information System (INIS)

    It is becoming increasingly important for the radiologist to evaluate the appearance and outcome of cartilage repair procedures. MR imaging is currently the best method for such evaluation but it is necessary to use cartilage-specific sequences and to modify those sequences when necessary to minimize artifacts from retained metal within the joint. This article reviews the surgical technique of the more commonly performed cartilage repair procedures, currently recommended techniques for the MR imaging evaluation of articular cartilage and cartilage repair procedures, and the MR imaging appearance of cartilage repair procedures and of the most frequently encountered complications following such procedures. (orig.)

  19. Effect of a Herbal-Leucine mix on the IL-1β-induced cartilage degradation and inflammatory gene expression in human chondrocytes

    Directory of Open Access Journals (Sweden)

    Haqqi Tariq M

    2011-08-01

    Full Text Available Abstract Background Conventional treatments for the articular diseases are often effective for symptom relief, but can also cause significant side effects and do not slow the progression of the disease. Several natural substances have been shown to be effective at relieving the symptoms of osteoarthritis (OA, and preliminary evidence suggests that some of these compounds may exert a favorable influence on the course of the disease. The objective of this study was to investigate the anti-inflammatory/chondroprotective potential of a Herbal and amino acid mixture containing extract of the Uncaria tomentosa, Boswellia spp., Lepidium meyenii and L-Leucine on the IL-1β-induced production of nitric oxide (NO, glycosaminoglycan (GAG, matrix metalloproteinases (MMPs, aggrecan (ACAN and type II collagen (COL2A1 in human OA chondrocytes and OA cartilage explants. Methods Primary OA chondrocytes or OA cartilage explants were pretreated with Herbal-Leucine mixture (HLM, 1-10 μg/ml and then stimulated with IL-1β (5 ng/ml. Effect of HLM on IL-1β-induced gene expression of iNOS, MMP-9, MMP-13, ACAN and COL2A1 was verified by real time-PCR. Estimation of NO and GAG release in culture supernatant was done using commercially available kits. Results HLM tested in these in vitro studies was found to be an effective anti-inflammatory agent, as evidenced by strong inhibition of iNOS, MMP-9 and MMP-13 expression and NO production in IL-1β-stimulated OA chondrocytes (p Leucine mixture (HLM up-regulation of ACAN and COL2A1 expression in IL-1β-stimulated OA chondrocytes was also noted (p Conclusion Our data suggests that HLM could be chondroprotective and anti-inflammatory agent in arthritis, switching chondrocyte gene expression from catabolic direction towards anabolic and regenerative, and consequently this approach may be potentially useful as a new adjunct therapeutic/preventive agent for OA or injury recovery.

  20. Progesterone regulates the activity of collagenase and related gelatinases A and B in human endometrial explants.

    OpenAIRE

    Marbaix, Etienne; Donnez, Jacques; Courtoy, Pierre J; Eeckhout, Yves

    1992-01-01

    Explants of human endometrium were cultured to study the release of matrix metalloproteinases (MMPs). Analysis of conditioned media by zymography revealed latent and active forms of collagenase (MMP-1, EC 3.4.24.7), 72-kDa gelatinase A (MMP-2, EC 3.4.24.24), and 92-kDa gelatinase B (MMP-9, EC 3.4.24.35). These proteinases were identified by their M(r), their inhibition by tissue inhibitor of metalloproteinases, and the activation of their zymogens by trypsin or aminophenylmercuric acetate. In...

  1. Effect of ZnO nanoparticles on Brassica nigra seedlings and stem explants: growth dynamics and antioxidative response

    Directory of Open Access Journals (Sweden)

    Hira eZafar

    2016-04-01

    Full Text Available Nanoparticles (NPs have diverse properties in comparison to respective chemicals due to structure, surface area ratio, morphology, and reactivity. Toxicological effects of metallic NPs to organisms including plants have been reported. However, to the authors’ knowledge there is no report on the effect of NPs on in vitro culture of plant explants. In this study, ZnO NPs at 500-1500 mg/L badly affected Brassica nigra seed germination and seedling growth and raised antioxidative activities and antioxidants concentrations. On the other hand, culturing the stem explants of B. nigra on Murashige and Skoog (MS medium in presence of low concentration of ZnO NPs (1-20 mg/L produced white thin roots with thick root hairs. At 10 mg/L ZnO NPs shoots emergence was also observed. The developed calli/roots showed 79% DPPH (2,2-diphenyl-1-picryl hydrazyl radical scavenging activity at 10 mg/L. While total antioxidant and reducing power potential were also significantly different in presence of ZnO NPs. Non enzymatic antioxidative molecules, phenolics (up to 0.15 µg GAE/mg FW and flavonoids (up to 0.22 µg QE/mg FW, also raised and found NPs concentration dependent. We state that ZnO NPs may induce roots from explants cultured on appropriate medium and can be cultured for production of valuable secondary metabolites.

  2. Clonal propagation of guava (Psidium guajava L on nodal explants of mature elite cultivar

    Directory of Open Access Journals (Sweden)

    Guochen Yang

    2011-04-01

    Full Text Available Guava (Psidium guajava L. is a very valuable tropical and subtropical fruit. However, guava micro-propagation are genotypes dependent, there are several problems associated with in vitro cultures of guava including browning or blackening of culture medium due to leaching of phenolics, microbial contamination, and in vitro tissue recalcitrance. A micro-propagation system using Murashige and Skoog (MS medium with 6-benzylaminopurine (BA, kinetin and naphthaleneacetic acid (NAA was developed for guava (Psidium guajava L from mature cultivar. As part of this research various disinfection methods and plant growth regulators were tested in vitro. The most effective method involved treating explants in a 15% bleach solution for 20 mins followed by culturing them in MS medium with 250 mg/L polyvinylpyrrolidone (PVP. This method maximized the percentage of bud breakage (53.3%, while producing the minimum browning rate (18.3% for the explants. The best observed proliferation rate (71.2% occurred on the MS medium supplemented with 4.44 μM BA, 4.65 μM kinetin (KT and 0.54 μM NAA. It produced the highest mean number of shoots (2.2. Shoots were then rooted (65% when dipped in 4.9 mM Indole-3-butyric acid (IBA solution for 1 min and rooted plantlets survived (100% after acclimatization to the greenhouse.

  3. ANATOMICAL STUDIES OF IN VITRO ORGANOGENESIS INDUCED IN LEAF-DERIVED EXPLANTS OF PASSIONFRUIT ESTUDOS ANATÔMICOS DA ORGANOGÊNESE IN VITRO INDUZIDA EM EXPLANTES DE FOLHA DE MARACUJÁ

    OpenAIRE

    BEATRIZ APPEZZATO DA GLORIA; MARIA LUCIA CARNEIRO VIEIRA; MARCELO CARNIER DORNELAS

    1999-01-01

    With the aim of studying the organogenesis in vitro in Passiflora edulis Sims f. flavicarpa Deg., the passionfruit, leaf-derived explants were cultured on media containing NAA or BAP and incubated either in continuous darkness or in light. The histological events leading to de novo organ formation were evaluated. Darkness induces rhizogenesis in the presence of NAA, whereas direct shoot regeneration is stimulated by light and BAP. This latter condition is recommended for passionfruit micropro...

  4. T cells fail to develop in the human skin-cell explants system; an inconvenient truth

    Directory of Open Access Journals (Sweden)

    Vanderlocht Joris

    2011-02-01

    Full Text Available Abstract Background Haplo-identical hematopoietic stem cell (HSC transplantation is very successful in eradicating haematological tumours, but the long post-transplant T-lymphopenic phase is responsible for high morbidity and mortality rates. Clark et al. have described a skin-explant system capable of producing host-tolerant donor-HSC derived T-cells. Because this T-cell production platform has the potential to replenish the T-cell levels following transplantation, we set out to validate the skin-explant system. Results Following the published procedures, while using the same commercial components, it was impossible to reproduce the skin-explant conditions required for HSC differentiation towards mature T-cells. The keratinocyte maturation procedure resulted in fragile cells with minimum expression of delta-like ligand (DLL. In most experiments the generated cells failed to adhere to carriers or were quickly outcompeted by fibroblasts. Consequently it was not possible to reproduce cell-culture conditions required for HSC differentiation into functional T-cells. Using cell-lines over-expressing DLL, we showed that the antibodies used by Clark et al. were unable to detect native DLL, but instead stained 7AAD+ cells. Therefore, it is unlikely that the observed T-lineage commitment from HSC is mediated by DLL expressed on keratinocytes. In addition, we did confirm expression of the Notch-ligand Jagged-1 by keratinocytes. Conclusions Currently, and unfortunately, it remains difficult to explain the development or growth of T-cells described by Clark et al., but for the fate of patients suffering from lymphopenia it is essential to both reproduce and understand how these co-cultures really "work". Fortunately, alternative procedures to speed-up T-cell reconstitution are being established and validated and may become available for patients in the near future.

  5. Effect of gamma radiation on different explants of sweet potato (Ipomoea batatas L. (Lam)) to induce NaCl tolerance

    International Nuclear Information System (INIS)

    Clonal lines tolerant to NaCl were obtained by combining in vitro culture and gamma radiation in two Peruvian varieties 'Amarillo de Quillabamba' and 'Nemanete'. The most suitable explants were pedicel sections and leaf blades. Embryogenic callus was induced on basal MS basal medium containing 0.5 ppm 2,4-D. The embryogenic calli were irradiated with 5 Gy from a 137Cs source. Several putative mutants appeared to be stable. (author). 9 refs, 3 tabs

  6. Biosynthesis of intestinal microvillar proteins. Rapid expression of cytoskeletal components in microvilli of pig small intestinal mucosal explants

    DEFF Research Database (Denmark)

    Cowell, G M; Danielsen, E M

    1984-01-01

    Using alkaline extraction to separate cytoskeletal and membrane proteins of intestinal microvilli, the kinetics of assembly of these two microvillar protein compartments was studied by pulse-chase labelling of pig small intestinal mucosal explants, kept in organ culture. Following a 10 min pulse of...... pulse. These different kinetics of appearance indicate that the two microvillar protein compartments have separate mechanisms of biosynthesis and microvillar expression....

  7. Callus Induction and Adventitious Shoot Regeneration from Different Explants of Rootstocks GF-677 (Prunus amygdalus x P. persica)

    OpenAIRE

    ELEKTRA SPAHIU; PETRIT RAMA; BARI HODAJ

    2015-01-01

    GF-677 is one of the most suitable rootstocks for almond and peach used in calcareous soils to overcome lime induced chlorosis. The objective of this study is the in vitro cultivation of the peach rootstock GF-677, in determining the protocol for inoculation and for micropropagation in the medium, influencing the development and the outgrowth of the rootstock. The present study focused on the effect of different explants of this rootstock on regenerative potential tissue culture techniques. T...

  8. Micropropagation of Origanum acutidens (HAND.-MAZZ.) IETSWAART using stem node explants.

    Science.gov (United States)

    Yildirim, Mehmet Ugur

    2013-01-01

    Origanum acutidens (HAND.-MAZZ.) IETSWAART is a promising ornamental plant that can be widely used in landscape management. It is endemic to Eastern Anatolian region of Turkey. Tissue culture has not been used to micropropagate it. The study reports stem node explants from one-week-old seedlings of the plant for successful micropropagation. The stem nodes were cultured on MS medium containing 0.6, 1.2, 1.8, and 2.4 mg/L BAP with 0.2 mg/L NAA. Visible effects of culture media on shoot proliferation were recorded. Shoot regeneration rate was maximum on MS medium containing 1.80 mg/L BAP-0.2 mg/L NAA. The micropropagated shoots were rooted on MS medium containing 0.2 mg/L NAA. All microrooted plantlets survived during acclimatisation on peat moss. It was concluded that O. acutidens can be successfully micropropagated under in vitro conditions. PMID:23983625

  9. Transcriptomic profiling of cartilage ageing.

    Science.gov (United States)

    Peffers, Mandy Jayne; Liu, Xuan; Clegg, Peter David

    2014-12-01

    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). PMID:26484061

  10. Transcriptomic profiling of cartilage ageing

    Directory of Open Access Journals (Sweden)

    Mandy Jayne Peffers

    2014-12-01

    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.

  11. Long-term organ culture of adult rat colon

    DEFF Research Database (Denmark)

    1978-01-01

    Colon explants from adult rats were maintained in culture for over 3 months in our laboratories with good epithelial preservation and cellular differentiation. The light and transmission electron microscopic features of rat colon mucosa during the culture period are described. In all the explants...

  12. PHOTOCROSSLINKABLE HYDROGELS FOR CARTILAGE TISSUE ENGINEERING

    NARCIS (Netherlands)

    Levett, Peter Andrew

    2015-01-01

    For millions of people, damaged cartilage is a major source of pain and disability. As those people often discover upon seeking medical treatment, once damaged, cartilage is very difficult to repair. Finding better clinical therapies for damaged cartilage has generated a huge amount of research inte

  13. Isolation, identification, and comparison of cartilage stem progenitor/cells from auricular cartilage and perichondrium

    OpenAIRE

    Xue, Ke; Zhang, Xiaodie; Qi, Lin; Zhou, Jia; Liu, Kai

    2016-01-01

    Auricular cartilage loss or defect remains a challenge to plastic surgeons, and cartilage regenerative medicine provides a novel method to solve the problem. However, ideal seeding cells seem to be the key point in the development of cartilage regeneration. Although bone marrow-mesenchymal stem cells were considered as the ideal seeding cells in cartilage regeneration, regenerative cartilage differentiated from bone marrow-mesenchymal stem cells still faces some problems. It is reported that ...

  14. MRI EVALUATION OF KNEE CARTILAGE

    Science.gov (United States)

    Rodrigues, Marcelo Bordalo; Camanho, Gilberto Luís

    2015-01-01

    Through the ability of magnetic resonance imaging (MRI) to characterize soft tissue noninvasively, it has become an excellent method for evaluating cartilage. The development of new and faster methods allowed increased resolution and contrast in evaluating chondral structure, with greater diagnostic accuracy. In addition, physiological techniques for cartilage assessment that can detect early changes before the appearance of cracks and erosion have been developed. In this updating article, the various techniques for chondral assessment using knee MRI will be discussed and demonstrated. PMID:27022562

  15. Macrophage Migration Inhibitory Factor Is Up-Regulated in Human First-Trimester Placenta Stimulated by Soluble Antigen of Toxoplasma gondii, Resulting in Increased Monocyte Adhesion on Villous Explants

    OpenAIRE

    Ferro, Eloisa Amália Vieira; Mineo, José Roberto; Ietta, Francesca; Bechi, Nicoletta; Romagnoli, Roberta; Silva, Deise Aparecida Oliveira; Sorda, Giuseppina; Bevilacqua, Estela; Paulesu, Luana Ricci

    2008-01-01

    Considering the potential role of macrophage migration inhibitory factor (MIF) in the inflammation process in placenta when infected by pathogens, we investigated the production of this cytokine in chorionic villous explants obtained from human first-trimester placentas stimulated with soluble antigen from Toxoplasma gondii (STAg). Parallel cultures were performed with villous explants stimulated with STAg, interferon-γ (IFN-γ), or STAg plus IFN-γ. To assess the role of placental MIF on monoc...

  16. Macroscopical, Histological, and In Vitro Characterization of Nonosteoarthritic Versus Osteoarthritic Hip Joint Cartilage

    Science.gov (United States)

    Badendick, Jessica; Godkin, Owen; Kohl, Benjamin; Meier, Carola; Jagielski, Michal; Huang, Zhao; Arens, Stephan; Schneider, Tobias; Schulze-Tanzil, Gundula

    2016-01-01

    Osteoarthritis (OA) might affect chondrocyte culture characteristics and complement expression. Therefore, this study addressed the interrelation between macroscopical and microscopical structure, complement expression, and chondrocyte culture characteristics in non-OA and OA cartilage. Femoral head cartilage samples harvested from patients with femoral neck fractures (FNFs) and OA were analyzed for macroscopical alterations using an in-house scoring system, graded histologically (Mankin score), and immunolabeled for complement regulatory proteins (CRPs) and receptors. Morphology of monolayer cultured chondrocytes isolated from a subset of samples was assessed. The macroscopical score distinguished the FNF and OA cartilage samples and correlated significantly with the histological results. Chondrocyte phenotype from FNF or OA cartilage differed. Complement receptor C5aR, CRPs CD55 and CD59, and weakly receptor C3AR were detected in the investigated FNF and OA cartilage, except for CD46, which was detected in only two of the five investigated donors. The in-house score also allows inexperienced observers to distinguish non-OA and OA cartilage for experimental purposes. PMID:27158224

  17. Direct axillary shoot regeneration from the mature seed explant of the hairy vetch (Vicia villosa Roth

    Directory of Open Access Journals (Sweden)

    Aasim Muhammad

    2011-01-01

    Full Text Available The hairy vetch (Vicia villosa Roth is a climbing, prostrate or trailing legume grown as forage.It fixes atmospheric nitrogen, reduces soil erosion and provides an instant mulch. Multiple axillary shoot regeneration from a mature seed explant (zygotic embryo with two cotyledons was obtained on MS medium containing 0.05 - 1.6 mg/l TDZ with or without 0.10 mg/l IBA. The frequency (% of shoot regeneration ranged from 45.83-75.00% with a maximum number of 28.6 shoots per explant on MS medium containing 0.20 mg/l TDZ-0.10 mg/l IBA. The mean shoot length decreased proportionately with each increase in TDZ concentration irrespective of the IBA concentration in the culture medium. However, comparing the two types of regeneration media, longer shoots were recorded in the presence of IBA in the culture medium. Regenerated shoots were pulse treated with 50 mg/l IBA for 5, 10 and 20 min for rooting.

  18. Utilization of Aseptic Seedling Explants for In vitro Propagation of Indian Red Wood

    Directory of Open Access Journals (Sweden)

    Kishore Kumar CHIRUVELLA

    2013-12-01

    Full Text Available Micropropagation has been advocated as one of the most viable biotechnological tool for ex situ conservation of rare, endangered endemic medicinal plants germplasm. Rapid clonal micropropagation protocol for large-scale multiplication of an endemic medicinal plant Soymida febrifuga (Meliaceae was established from 15-day aseptic seedling cotyledonary node and shoot tip explants. High frequency of sprouting and shoot differentiation was observed from cotyledonary node explants compared to shoot tip, on Murashige and Skoog (MS medium fortified with BA, KN, 2-iP and CM. Of the cytokinins used, BA (3.0 mgl-1 supported highest average number and maximum multiple shoot differentiation (16.6. In vitro proliferated shoots were multiplied rapidly by culturing nodal segments as microcuttings, further subcultured on the same media for elongation. Elongated shoots upon transfer to MS medium fortified with IBA showed rooting within two weeks of culture. Rooted plantlets were successfully hardened and 75% of rooted shoots successfully survived on establishment to the soil. Plants looked healthy with no visually detectable phenotypic variations. This protocol provides a successful and rapid technique that can be used for ex situ conservation minimizing the pressure on wild populations and contributes to the conservation of this endemic medicinally potent flora.

  19. Flower-bud formation in explants of photoperiodic and day-neutral Nicotiana biotypes and its bearing on the regulation of flower formation.

    Science.gov (United States)

    Rajeevan, M S; Lang, A

    1993-05-15

    The capacity to form flower buds in thin-layer explants was studied in flowering plants of several species, cultivars, and lines of Nicotiana differing in their response to photoperiod. This capacity was found in all biotypes examined and could extend into sepals and corolla. It varied greatly, depending on genotype, source tissue and its developmental stage, and composition of the culture medium, particularly the levels of glucose, auxin, and cytokinin. It was greatest in the two day-neutral plants examined, Samsun tobacco and Nicotiana rustica, where it extended from the inflorescence region down the vegetative stem, in a basipetally decreasing gradient; it was least in the two qualitative photoperiodic plants studied, the long-day plant Nicotiana silvestris and the short-day plant Maryland Mammoth tobacco, the quantitative long-day plant Nicotiana alata and the quantitative short-day plant Nicotiana otophora line 38-G-81, where it was limited to the pedicels (and, in some cases, the sepals). Regardless of the photoperiodic response of the source plants, the response was the same in explants cultured under long and short days. The finding that capacity to form flower buds in explants is present in all Nicotiana biotypes studied supports the idea that it is regulated by the same mechanism(s), regardless of the plant's photoperiodic character. However, the source plants were all in the flowering stage, and no flower-bud formation can be obtained in explants from strictly vegetative Nicotiana plants. Hence, flower formation in the explants is not identical with de novo flower formation in a hitherto vegetative plant: it is rather the expression of a floral state already established in the plant, although it can vary widely in extent and spatial distribution. Culture conditions that permit flower-bud formation in an explant are conditions that maintain the floral state and encourage its expression; conditions under which no flower buds are formed reduce this state

  20. Suppression of glycosaminoglycan synthesis by articular cartilage, but not of hyaluronic acid synthesis by synovium, after exposure to radiation

    International Nuclear Information System (INIS)

    We recently found that injection of 2 mCi of yttrium 90 (90Y; approximately 23,000 rads) into normal canine knees stimulated glycosaminoglycan (GAG) synthesis by femoral condylar cartilage. The present investigation was conducted to determine whether radiation affects cartilage metabolism directly. Rates of GAG synthesis and degradation in normal canine articular cartilage were studied following irradiation. Cultured synovium from the same knees was treated similarly, to determine the effects of irradiation on hyaluronic acid synthesis. Twenty-four hours after exposure to 1,000 rads, 10,000 rads, or 50,000 rads, 35S-GAG synthesis by the cartilage was 93%, 69%, and 37%, respectively, of that in control, nonirradiated cartilage. The effect was not rapidly reversible: 120 hours after exposure to 50,000 rads, GAG synthesis remained at only 28% of the control level. Autoradiography showed marked suppression of 35S uptake by chondrocytes after irradiation. Cartilage GAG degradation was also increased following irradiation: 4 hours and 8 hours after exposure to 50,000 rads, the cartilage GAG concentration was only 66% and 54%, respectively, of that at time 0, while corresponding values for control, nonirradiated cartilage were 90% and 87%. In contrast to its effects on cartilage GAG metabolism, radiation at these levels had no effect on synovial hyaluronic acid synthesis

  1. Suppression of glycosaminoglycan synthesis by articular cartilage, but not of hyaluronic acid synthesis by synovium, after exposure to radiation

    Energy Technology Data Exchange (ETDEWEB)

    Hugenberg, S.T.; Myers, S.L.; Brandt, K.D.

    1989-04-01

    We recently found that injection of 2 mCi of yttrium 90 (90Y; approximately 23,000 rads) into normal canine knees stimulated glycosaminoglycan (GAG) synthesis by femoral condylar cartilage. The present investigation was conducted to determine whether radiation affects cartilage metabolism directly. Rates of GAG synthesis and degradation in normal canine articular cartilage were studied following irradiation. Cultured synovium from the same knees was treated similarly, to determine the effects of irradiation on hyaluronic acid synthesis. Twenty-four hours after exposure to 1,000 rads, 10,000 rads, or 50,000 rads, 35S-GAG synthesis by the cartilage was 93%, 69%, and 37%, respectively, of that in control, nonirradiated cartilage. The effect was not rapidly reversible: 120 hours after exposure to 50,000 rads, GAG synthesis remained at only 28% of the control level. Autoradiography showed marked suppression of 35S uptake by chondrocytes after irradiation. Cartilage GAG degradation was also increased following irradiation: 4 hours and 8 hours after exposure to 50,000 rads, the cartilage GAG concentration was only 66% and 54%, respectively, of that at time 0, while corresponding values for control, nonirradiated cartilage were 90% and 87%. In contrast to its effects on cartilage GAG metabolism, radiation at these levels had no effect on synovial hyaluronic acid synthesis.

  2. Stable genetic transformation of Jatropha curcas via Agrobacterium tumefaciens-mediated gene transfer using leaf explants

    KAUST Repository

    Kumar, Nitish

    2010-07-01

    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

  3. Human Endogenous Retrovirus W Activity in Cartilage of Osteoarthritis Patients

    Directory of Open Access Journals (Sweden)

    Signy Bendiksen

    2014-01-01

    Full Text Available The etiology of viruses in osteoarthritis remains controversial because the prevalence of viral nucleic acid sequences in peripheral blood or synovial fluid from osteoarthritis patients and that in healthy control subjects are similar. Until now the presence of virus has not been analyzed in cartilage. We screened cartilage and chondrocytes from advanced and non-/early osteoarthritis patients for parvovirus B19, herpes simplex virus-1, Epstein Barr virus, cytomegalovirus, human herpes virus-6, hepatitis C virus, and human endogenous retroviruses transcripts. Endogenous retroviruses transcripts, but none of the other viruses, were detected in 15 out the 17 patients. Sequencing identified the virus as HERV-WE1 and E2. HERV-W activity was confirmed by high expression levels of syncytin, dsRNA, virus budding, and the presence of virus-like particles in all advanced osteoarthritis cartilages examined. Low levels of HERV-WE1, but not E2 envelope RNA, were observed in 3 out of 8 non-/early osteoarthritis patients, while only 3 out of 7 chondrocytes cultures displayed low levels of syncytin, and just one was positive for virus-like particles. This study demonstrates for the first time activation of HERV-W in cartilage of osteoarthritis patients; however, a causative role for HERV-W in development or deterioration of the disease remains to be proven.

  4. Postnatal development of articular cartilage

    NARCIS (Netherlands)

    Turnhout, van M.C.

    2010-01-01

    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 dep

  5. Effects of Chondroitinase ABC-Mediated Proteoglycan Digestion on Decellularization and Recellularization of Articular Cartilage.

    Directory of Open Access Journals (Sweden)

    Catherine A Bautista

    Full Text Available Articular cartilage has a limited capacity to heal itself and thus focal defects often result in the development of osteoarthritis. Current cartilage tissue engineering strategies seek to regenerate injured tissue by creating scaffolds that aim to mimic the unique structure and composition of native articular cartilage. Decellularization is a novel strategy that aims to preserve the bioactive factors and 3D biophysical environment of the native extracellular matrix while removing potentially immunogenic factors. The purpose of this study was to develop a procedure that can enable decellularization and recellularization of intact articular cartilage matrix. Full-thickness porcine articular cartilage plugs were decellularized with a series of freeze-thaw cycles and 0.1% (w/v sodium dodecyl sulfate detergent cycles. Chondroitinase ABC (ChABC was applied before the detergent cycles to digest glycosaminoglycans in order to enhance donor chondrocyte removal and seeded cell migration. Porcine synovium-derived mesenchymal stem cells were seeded onto the decellularized cartilage scaffolds and cultured for up to 28 days. The optimized decellularization protocol removed 94% of native DNA per sample wet weight, while collagen content and alignment were preserved. Glycosaminoglycan depletion prior to the detergent cycles increased removal of nuclear material. Seeded cells infiltrated up to 100 μm into the cartilage deep zone after 28 days in culture. ChABC treatment enhances decellularization of the relatively dense, impermeable articular cartilage by reducing glycosaminoglycan content. ChABC treatment did not appear to affect cell migration during recellularization under static, in vitro culture, highlighting the need for more dynamic seeding methods.

  6. Cartilage Derived from Bone Marrow Mesenchymal Stem Cells Expresses Lubricin In Vitro and In Vivo.

    Directory of Open Access Journals (Sweden)

    Yusuke Nakagawa

    Full Text Available Lubricin expression in the superficial cartilage will be a crucial factor in the success of cartilage regeneration. Mesenchymal stem cells (MSCs are an attractive cell source and the use of aggregates of MSCs has some advantages in terms of chondrogenic potential and efficiency of cell adhesion. Lubricin expression in transplanted MSCs has not been fully elucidated so far. Our goals were to determine (1 whether cartilage pellets of human MSCs expressed lubricin in vitro chondrogenesis, (2 whether aggregates of human MSCs promoted lubricin expression, and (3 whether aggregates of MSCs expressed lubricin in the superficial cartilage after transplantation into osteochondral defects in rats.For in vitro analysis, human bone marrow (BM MSCs were differentiated into cartilage by pellet culture, and also aggregated using the hanging drop technique. For an animal study, aggregates of BM MSCs derived from GFP transgenic rats were transplanted to the osteochondral defect in the trochlear groove of wild type rat knee joints. Lubricin expression was mainly evaluated in differentiated and regenerated cartilages.In in vitro analysis, lubricin was detected in the superficial zone of the pellets and conditioned medium. mRNA expression of Proteoglycan4 (Prg4, which encodes lubricin, in pellets was significantly higher than that of undifferentiated MSCs. Aggregates showed different morphological features between the superficial and deep zone, and the Prg4 mRNA expression increased after aggregate formation. Lubricin was also found in the aggregate. In a rat study, articular cartilage regeneration was significantly better in the MSC group than in the control group as shown by macroscopical and histological analysis. The transmission electron microscope showed that morphology of the superficial cartilage in the MSC group was closer to that of the intact cartilage than in the control group. GFP positive cells remained in the repaired tissue and expressed lubricin in

  7. Rapid in vitro multiplication of the ethnomedicinal shrub, Acacia caesia (L.) Willd. (Mimosaceae) from leaf explants

    Institute of Scientific and Technical Information of China (English)

    Thambiraj J; Paulsamy S

    2012-01-01

    Objective: To develop an efficient protocol for in vitro multiplication of the ethnomedicinal shrub Acacia caesia (A. caesia) L. Willd., Methods: Leaf explants were inoculated on MS medium supplemented with TDZ and NAA for callus induction. Subculturing experiments were conducted by using leaf derived calli for shoot proliferation on MS medium fortified with various growth regulators like IBA, TDZ, BAP and GA3. The regenerated shoots were rooted in half strength MS medium supplemented with various concentrations of IBA, IAA and Kn. After roots were developed, the plantlets were transplanted to pots filled with garden soil, sand and vermicompost and kept in growth chamber with 70%-80%humidity under16h photoperiod. After acclimatization, the plantlets were transferred to the garden and survival percentage was calculated. Data were statistically analyzed and means were compared using Duncan’s Multiple Range Test (P<0.05). Results: An in vitro multiplication protocol was developed for the locally demanded medicinal plant species, A. caesia by using leaf explant. The study revealed that the callus formation was effective in MS medium containing TDZ and NAA at 1.5 and 0.3 mg/L respectively. Shoot induction was most successful in MS medium supplemented with combination of the auxin, IBA and cytokinin, TDZ at 2.0 and 0.5 mg/L respectively. A single leaf explant was capable of producing 12 shoots/callus after 30 days of culture. The other supplementation in MS medium with IBA and Kn at 2.0 and 0.4 mg/L respectively produced higher rooting frequency, roots/shoot and root length. The survivability rate of leaf callus derived plantlets was significantly higher (84%) in the hardening medium composed by garden soil, sand and vermicompost (1:1:1) by volume. Conclusions: A significant progress has been made in the in vitro regeneration system of this medicinally important plant species, A.caesia.

  8. Effects of gamma-rays irradiation in seed of mungbean (vigna radiata (L.) wilczek) composition of media on shoot regeneration of explants from node of cotyledon

    International Nuclear Information System (INIS)

    Study the effects of gamma-rays irradiation and composition and media on shoot regeneration of explants from node of cotyledon of mungbean. Wallet variety have been conducted. The explants derived of irradiated seeds of 10-20 Gy of gamma rays were planted in the 0.7% agar solution. One day after planting in the agar media the embryo axis of germinate seed were removed and the node of cotyledon were cultured in the regeneration media as examples. The results shown that shoot regeneration was influenced by media composition and the doses of gamma rays irradiation in seed. In the MURASHIGE and SKOOG medium which contain of BAP or 2-iP or Kinetin with 3 ppm concentrate respectively the explants could produced 100% of shoots. However, the highest. number of produced shoot (3 shoots) was showed in the medium which contained of BAP. The medium with I ppm concentrate od BAP could produced 100% shoot regeneration and the maximum number of shoots (4 shoots) per explant was showed in with 5 ppm. concentrate of BAP. The effectivity off BAP for shoot regeneration by enrichment of 12 ppm Ag2SO4 in the media. Irradiation of 10-20 Gy gamma rays on seeds of mungbean walet variety could improved shoot regeneration of explants from node cotyledon. (author)

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

    Directory of Open Access Journals (Sweden)

    Cohn Zachary A

    2007-06-01

    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.

  10. Immunohistochemical localization of IAA in graft union of explanted internode grafting

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The dynamic changes of IAA in graft union of explanted internode autografting of Cucumber (Cucumis sativus Linn.) have been investigated using the immunohistochemical localization technique.It is shown that the efficiency of IAA fixation using lyophilization-gas fixation is higher than that using liquid chemical fixation.In contrast to few silver particles and no significant changes during the development of graft union cultured in hormone-free medium,more silver particles in graft union and significant changes of IAA related to graft union development have been found when graft union was cultured in medium supplemented with appropriate hormones.The fixation procedure of plant hormones and the roles of IAA in graft union are discussed.

  11. Improved cartilage repair via in vitro pre-maturation of MSC-seeded hyaluronic acid hydrogels

    International Nuclear Information System (INIS)

    Functional repair of focal cartilage defects requires filling the space with neotissue that has compressive properties comparable to native tissue and integration with adjacent host cartilage. While poor integration is a common complication with current clinical treatments, reports of tissue engineering advances in the development of functional compressive properties rarely include analyses of their potential for integration. Our objective was thus to assess both the maturation and integration of mesenchymal stem cell (MSC)-laden hyaluronic acid (HA) hydrogels in an in vitro cartilage defect model. Furthermore, we considered the effects of an initial period of pre-maturation as well as various material formulations to maximize both construct compressive properties and integration strength. MSCs were encapsulated in 1%, 3% and 5% methacrylated HA (MeHA) or 2% agarose (Ag) and gelled directly (in situ) within an in vitro cartilage defect or were formed and then pre-cultured for 4 weeks before implantation. Results showed that the integration strength of pre-cultured repair constructs was equal to (1% MeHA) or greater than (2% Ag) the integration of in situ repaired cartilage. Moreover, MSC chondrogenesis and maturation was restricted by the in situ repair environment with constructs maturing to a much lesser extent than pre-matured constructs. These results indicate that construct pre-maturation may be an essential element of functional cartilage repair. (paper)

  12. Factors affecting proliferation and elongation of shoots of Phak Liang (Gnetum gnemon Linn.) through tissue culture technique

    OpenAIRE

    Te-chato, S.; Noochum, P.

    2003-01-01

    The tissue culture of Phak Liang (Gnetum gnemon Linn.) was investigated for micropropagation. The types of explant, culture media, types and concentrations of plant growth regulators, orientation of explant and section of explant were tested for their efficacy in inducing and proliferating shoot buds. The elongation of shoots and root induction was also studied. Young leaves gave the highest number of shoot buds when they were cultured in Murashige and Skoog (MS) medium supplemented with 0.25...

  13. Repair of articular cartilage defects in minipigs by microfracture surgery and BMSCs transplantation

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objective: To investigate the feasibility of minimal invasive repair of cartilage defect by arthroscope-aided microfracture surgery and autologous transplantation of mesenchymal stem cells. Methods: Bone marrow of minipigs was taken out and the bone marrow derived mesenchymal stem cells (BMSCs) were isolated and cultured to passage 3. Then 6 minipigs were randomly divided into 2 groups with 6 knees in each group. After the articular cartilage defect was induced in each knee. the left defect received microfracture surgery and was injected with 2. 5 ml BMSCs cells at a concentration of 3×107 cells/ml into the articular cavity; while right knee got single microfracture or served as blank control group. The animals were killed at 8 or 16 weeks, and the repair tissue was histologically and immunohistochemically examined for the presence of type Ⅱ collagen and glycosaminoglycans (GAGs) at 8 and 16 weeks. Results:Eight weeks after the surgery, the overlying articular surface of the cartilage defect showed normal color and integrated to adjacent cartilage. And 16 weeks after surgery, hyaline cartilage was observed at the repairing tissues and immunostaining indicated the diffuse presence of this type Ⅱ collagen and GAGs throughout the repair cartilage in the treated defects. Single microfracture group had the repairing of fibro-cartilage, while during the treatment, the defects of blank group were covered with fewer fiber tissues, and no blood capillary growth or any immunological rejection was observed. Conclusion:Microfracture technique and BMSCs transplantation to repair cartilage defect is characterized with minimal invasion and easy operation, and it will greatly promote the regeneration repair of articular cartilage defect.

  14. 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.;

    2015-01-01

    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...

  15. A Novel Model for the Mass Transfer of Articular Cartilage: Rolling Depression Load Device

    Science.gov (United States)

    Fan, Zhenmin; Zhang, Chunqiu; Liu, Haiying; Xu, Baoshan; Li, Jiang; Gao, Lilan

    The mass transfer is one of important aspects to maintain the physiological activity proper of tissue, specially, cartilage cannot run without mechanical environment. The mechanical condition drives nutrition in and waste out in the cartilage tissue, the change of this process plays a key role for biological activity. Researchers used to adopt compression to study the mass transfer in cartilage, here we firstly establish a new rolling depression load (RDL) device, and also put this device into practice. The device divided into rolling control system and the compression adjusting mechanism. The rolling control system makes sure the pure rolling and uniform speed of roller applying towards cultured tissue. The compression adjusting mechanism can realize different compressive magnitudes and uniform compression. Preliminary test showed that rolling depression load indeed enhances the process of mass transfer articular cartilage.

  16. Synergistic effect of ethylene inhibitors and putrescine on shoot regeneration from hypocotyl explants of Chinese radish (Raphanus sativus L. var. longipinnatus Bailey) in vitro.

    Science.gov (United States)

    Pua, E C; Sim, G E; Chi, G L; Kong, L F

    1996-05-01

    The role of ethylene and putrescine on shoot regeneration from hypocotyl explants of Chinese radish (Raphanus sativus L. var. longipinnatus Bailey cv. Red Coat) was investigated. Explants were recalcitrant in culture, but exogenous application of ethylene inhibitor [20-30 μM aminoethoxyvinylglycine (AVG) or AgNO3] enhanced shoot regeneration of explants grown on medium supplemented with 2 mg/l N(6)-benzyladenine and 1 mg/l 1-naphthaleneacetic acid. The best regeneration occurred in the medium containing AgNO3 in combination with AVG. Culture medium solidified with agarose in the presence of AgNO3 but not AVG was also beneficial to shoot regeneration. Exogenous putrescine, 2-chloroethylphosphonic acid and 1-aminocyclopropane-1-carboxylate had no effect on shoot regeneration. However, regeneration was greatly promoted by 10-25 mM putrescine in combination with 30 μM AgNO3 or AVG. Explants with high regenerability grown in the presence of AgNO3 or in combination with putrescine emanated high levels of ethylene throughout the 21-d culture period. By contrast, AVG or putrescine alone resulted in a decrease in ethylene production. For rooting of shoot cuttings, IAA and IBA at 1-5 mg/l were more effective than NAA. PMID:24178611

  17. Human sclera maintains common characteristics with cartilage throughout evolution.

    Directory of Open Access Journals (Sweden)

    Yuko Seko

    Full Text Available BACKGROUND: The sclera maintains and protects the eye ball, which receives visual inputs. Although the sclera does not contribute significantly to visual perception, scleral diseases such as refractory scleritis, scleral perforation and pathological myopia are considered incurable or difficult to cure. The aim of this study is to identify characteristics of the human sclera as one of the connective tissues derived from the neural crest and mesoderm. METHODOLOGY/PRINCIPAL FINDINGS: We have demonstrated microarray data of cultured human infant scleral cells. Hierarchical clustering was performed to group scleral cells and other mesenchymal cells into subcategories. Hierarchical clustering analysis showed similarity between scleral cells and auricular cartilage-derived cells. Cultured micromasses of scleral cells exposed to TGF-betas and BMP2 produced an abundant matrix. The expression of cartilage-associated genes, such as Indian hedge hog, type X collagen, and MMP13, was up-regulated within 3 weeks in vitro. These results suggest that human 'sclera'-derived cells can be considered chondrocytes when cultured ex vivo. CONCLUSIONS/SIGNIFICANCE: Our present study shows a chondrogenic potential of human sclera. Interestingly, the sclera of certain vertebrates, such as birds and fish, is composed of hyaline cartilage. Although the human sclera is not a cartilaginous tissue, the human sclera maintains chondrogenic potential throughout evolution. In addition, our findings directly explain an enigma that the sclera and the joint cartilage are common targets of inflammatory cells in rheumatic arthritis. The present global gene expression database will contribute to the clarification of the pathogenesis of developmental diseases such as high myopia.

  18. Mechanical stress and ATP synthesis are coupled by mitochondrial oxidants in articular cartilage.

    Science.gov (United States)

    Wolff, Katherine J; Ramakrishnan, Prem S; Brouillette, Marc J; Journot, Brice J; McKinley, Todd O; Buckwalter, Joseph A; Martin, James A

    2013-02-01

    Metabolic adaptation of articular cartilage under joint loading is evident and matrix synthesis seems to be critically tied to ATP. Chondrocytes utilize the glycolytic pathway for energy requirements but seem to require mitochondrial reactive oxygen species (ROS) to sustain ATP synthesis. The role of ROS in regulating ATP reserves under a mechanically active environment is not clear. It is believed that physiological strains cause deformation of the mitochondria, potentially releasing ROS for energy production. We hypothesized that mechanical loading stimulates ATP synthesis via mitochondrial release of ROS. Bovine osteochondral explants were dynamically loaded at 0.5 Hz with amplitude of 0.25 MPa for 1 h. Cartilage response to mechanical loading was assessed by imaging with dihydroethidium (ROS indicator) and a Luciferase-based ATP assay. Electron transport inhibitor rotenone and mitochondrial ROS scavenger MitoQ significantly suppressed mechanically induced ROS production and ATP synthesis. Our findings indicate that mitochondrial ROS are produced as a result of physiological mechanical strains. Taken together with our previous findings of ROS involvement in blunt impact injuries, mitochondrial ROS are important contributors to cartilage metabolic adaptation and their precise role in the pathogenesis of osteoarthritis warrants further investigation. PMID:22930474

  19. Development of artificial articular cartilage.

    Science.gov (United States)

    Oka, M; Ushio, K; Kumar, P; Ikeuchi, K; Hyon, S H; Nakamura, T; Fujita, H

    2000-01-01

    Attempts have been made to develop an artificial articular cartilage on the basis of a new viewpoint of joint biomechanics in which the lubrication and load-bearing mechanisms of natural and artificial joints are compared. Polyvinyl alcohol hydrogel (PVA-H), 'a rubber-like gel', was investigated as an artificial articular cartilage and the mechanical properties of this gel were improved through a new synthetic process. In this article the biocompatibility and various mechanical properties of the new improved PVA-H is reported from the perspective of its usefulness as an artificial articular cartilage. As regards lubrication, the changes in thickness and fluid pressure of the gap formed between a glass plate and the specimen under loading were measured and it was found that PVA-H had a thicker fluid film under higher pressures than polyethylene (PE) did. The momentary stress transmitted through the specimen revealed that PVA-H had a lower peak stress and a longer duration of sustained stress than PE, suggesting a better damping effect. The wear factor of PVA-H was approximately five times that of PE. Histological studies of the articular cartilage and synovial membranes around PVA-H implanted for 8-52 weeks showed neither inflammation nor degenerative changes. The artificial articular cartilage made from PVA-H could be attached to the underlying bone using a composite osteochondral device made from titanium fibre mesh. In the second phase of this work, the damage to the tibial articular surface after replacement of the femoral surface in dogs was studied. Pairs of implants made of alumina, titanium or PVA-H on titanium fibre mesh were inserted into the femoral condyles. The two hard materials caused marked pathological changes in the articular cartilage and menisci, but the hydrogel composite replacement caused minimal damage. The composite osteochondral device became rapidly attached to host bone by ingrowth into the supporting mesh. The clinical implications of

  20. A Novel Biodegradable Polyurethane Matrix for Auricular Cartilage Repair: An In Vitro and In Vivo Study.

    Science.gov (United States)

    Iyer, Kartik; Dearman, Bronwyn L; Wagstaff, Marcus J D; Greenwood, John E

    2016-01-01

    Auricular reconstruction poses a challenge for reconstructive and burns surgeons. Techniques involving cartilage tissue engineering have shown potential in recent years. A biodegradable polyurethane matrix developed for dermal reconstruction offers an alternative to autologous, allogeneic, or xenogeneic biologicals for cartilage reconstruction. This study assesses such a polyurethane matrix for this indication in vivo and in vitro. To evaluate intrinsic cartilage repair, three pigs underwent auricular surgery to create excisional cartilage ± perichondrial defects, measuring 2 × 3 cm in each ear, into which acellular polyurethane matrices were implanted. Biopsies were taken at day 28 for histological assessment. Porcine chondrocytes ± perichondrocytes were cultured and seeded in vitro onto 1 × 1 cm polyurethane scaffolds. The total culture period was 42 days; confocal, histological, and immunohistochemical analyses of scaffold cultures were performed on days 14, 28, and 42. In vivo, the polyurethane matrices integrated with granulation tissue filling all biopsy samples. Minimal neocartilage invasion was observed marginally on some samples. Tissue composition was identical between ears whether perichondrium was left intact, or not. In vitro, the polyurethane matrix was biocompatible with chondrocytes ± perichondrocytes and supported production of extracellular matrix and Type II collagen. No difference was observed between chondrocyte culture alone and chondrocyte/perichondrocyte scaffold coculture. The polyurethane matrix successfully integrated into the auricular defect and was a suitable scaffold in vitro for cartilage tissue engineering, demonstrating its potential application in auricular reconstruction. PMID:26284639

  1. Variation in phytate accumulation in common bean (Phaseolus vulgaris L. fruit explants

    Directory of Open Access Journals (Sweden)

    Cileide Maria Medeiros Coelho

    2008-02-01

    Full Text Available The in vitro synthesis of phytate was studied in common bean fruit explants. Different concentrations of sucrose; phosphorus (P; myo-inositol; abscisic acid (ABA; glutamine and methionine, were tested. Fixed concentrations of these compounds were tested at different periods (0, 3, 6 and 9 days. Variation in phytate coincided with different concentrations of sucrose, myo-inositol, P and ABA for the duration tested. These compounds caused an accumulation of phytate and were more effective in the presence of myo-inositol and P. The accumulation of P varied less than phytate for the different treatments tested in vitro. In conclusion, P, sucrose, ABA, and myo-inositol caused an increase in the phytate of bean seed, showing that it could be possible to alter its content by culturing bean fruit explants in vitro.O fósforo é armazenado na forma de fitato nas sementes, o qual forma complexos estáveis e insolúveis com minerais e proteínas, conferindo efeito antinutriente. A síntese de fitato foi estudada em cultivo de explantes de fruto de feijão in vitro sob diferentes concentrações de sacarose, fósforo (P, mio-inositol, ácido abscísico (ABA, glutamina e metionina. Fixada a concentração destes compostos, testou-se os diferentes tempos de cultivo (0, 3, 6 e 9 dias. A variação no acúmulo de fitato ocorreu na presença de sacarose, mio-inositol, P e ABA nas diferentes concentrações e tempos testados. O acúmulo mais efetivo de fitato ocorreu na presença de mio-inositol e P. O acúmulo de P variou menos do que fitato em todos os tratamentos. Em conclusão, P, sacarose, ABA e mio-inositol causaram aumento no fitato acumulado nas sementes, mostrando que foi possível alterar a síntese de fitato em cultivo de explantes de fruto de feijão.

  2. Cartilage regeneration by chondrogenic induced adult stem cells in osteoarthritic sheep model.

    Directory of Open Access Journals (Sweden)

    Chinedu C Ude

    Full Text Available OBJECTIVES: In this study, Adipose stem cells (ADSC and bone marrow stem cells (BMSC, multipotent adult cells with the potentials for cartilage regenerations were induced to chondrogenic lineage and used for cartilage regenerations in surgically induced osteoarthritis in sheep model. METHODS: Osteoarthritis was induced at the right knee of sheep by complete resection of the anterior cruciate ligament and medial meniscus following a 3-weeks exercise regimen. Stem cells from experimental sheep were culture expanded and induced to chondrogenic lineage. Test sheep received a single dose of 2 × 10(7 autologous PKH26-labelled, chondrogenically induced ADSCs or BMSCs as 5 mls injection, while controls received 5 mls culture medium. RESULTS: The proliferation rate of ADSCs 34.4 ± 1.6 hr was significantly higher than that of the BMSCs 48.8 ± 5.3 hr (P = 0.008. Chondrogenic induced BMSCs had significantly higher expressions of chondrogenic specific genes (Collagen II, SOX9 and Aggrecan compared to chondrogenic ADSCs (P = 0.031, 0.010 and 0.013. Grossly, the treated knee joints showed regenerated de novo cartilages within 6 weeks post-treatment. On the International Cartilage Repair Society grade scores, chondrogenically induced ADSCs and BMSCs groups had significantly lower scores than controls (P = 0.0001 and 0.0001. Fluorescence of the tracking dye (PKH26 in the injected cells showed that they had populated the damaged area of cartilage. Histological staining revealed loosely packed matrixes of de novo cartilages and immunostaining demonstrated the presence of cartilage specific proteins, Collagen II and SOX9. CONCLUSION: Autologous chondrogenically induced ADSCs and BMSCs could be promising cell sources for cartilage regeneration in osteoarthritis.

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

    Directory of Open Access Journals (Sweden)

    Xia Liu

    2014-01-01

    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.

  4. In vitro direct shoot regeneration from proximal, middle and distal segment of Coleus forskohlii leaf explants.

    Science.gov (United States)

    Krishna, Gaurav; Sairam Reddy, P; Anoop Nair, N; Ramteke, P W; Bhattacharya, P S

    2010-04-01

    Coleus forskohlii is an endangered multipurpose medicinal plant that has widespread applications. In spite of this, there have been few attempts to propagate its cultivation in India. The present communication presents an in vitro rapid regeneration method using leaf explants of Coleus forskohlii through direct organogenesis. Leaf explants that were excised into three different segments i.e. proximal (P), middle (M) and distal (D) were cultured on Murashige and Skoog (MS) basal medium supplemented with cytokinins. MS Media containing 5.0 mg L(-1) BAP (6-Benzylaminopurine) promoted regeneration of multiple shoots through direct organogenesis from the leaf, which were further elongated on MS media augmented with 0.1 mg L-1 BAP and 0.1 mg L(-1) IAA (Indole-3-acetic acid), cytokinin and auxin combination. Regenerated and elongated shoots, when transferred to ose resulted in profuse rooting plants that were transferred to soil after acclimatization and maintained in a green house. The current protocol offers a direct, mass propagation method bypassing the callus phase of C. forskohlii and is suitable for conservation, large-scale commercial cultivation, and genetic transformation with agronomically desirable traits. PMID:23572969

  5. Preclinical Studies for Cartilage Repair

    OpenAIRE

    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.

    2011-01-01

    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...

  6. Diode laser (980nm) cartilage reshaping

    Science.gov (United States)

    El Kharbotly, A.; El Tayeb, T.; Mostafa, Y.; Hesham, I.

    2011-03-01

    Loss of facial or ear cartilage due to trauma or surgery is a major challenge to the otolaryngologists and plastic surgeons as the complicated geometric contours are difficult to be animated. Diode laser (980 nm) has been proven effective in reshaping and maintaining the new geometric shape achieved by laser. This study focused on determining the optimum laser parameters needed for cartilage reshaping with a controlled water cooling system. Harvested animal cartilages were angulated with different degrees and irradiated with different diode laser powers (980nm, 4x8mm spot size). The cartilage specimens were maintained in a deformation angle for two hours after irradiation then released for another two hours. They were serially measured and photographed. High-power Diode laser irradiation with water cooling is a cheep and effective method for reshaping the cartilage needed for reconstruction of difficult situations in otorhinolaryngologic surgery. Key words: cartilage,diode laser (980nm), reshaping.

  7. Propagation of jarrah (Eucalyptus marginata) by organ and tissue culture

    Energy Technology Data Exchange (ETDEWEB)

    Bennett, M.J.; McComb, J.A.

    1982-01-01

    Micropropagation methods are described for the production of clonal lines from Eucalyptus marginata (jarrah) seedlings. Nodal explants from mature trees can also yield shoot cultures, but a high frequency of contamination occurs among such explants. Uncontaminated callus cultures can be produced from mature trees by culturing stamen filaments and shoots can subsequently be regenerated from this callus. The rooting percentage of shoot cultures from either nodes or stamen callus of mature trees is low compared with that from seedling explants. Considerable variation was observed between trees in the ability of stamen callus to regenerate shoots and in the frequency of rooting. (Refs. 27)

  8. Effects of carbon ion beam irradiation on the shoot regeneration from in vitro axillary bud explants of the Impatiens hawkeri

    Science.gov (United States)

    Zhou, Libin; Zhou, Libin; Li, Wenjian; Li, Ping; Dong, Xicun; Qu, Ying; Ma, Shuang; Li, Qiang

    Accelerated ion beams is an excellent mutagen in plant breeding which can induce higher mutation frequencies and wider mutation spectrum than those of low linear energy transfer (LET) irradiations, such as X-rays (Okamura et al. 2003, Yamaguchi et al. 2003). Mutation breeding operation of two Saintpaulia ionahta cultivars using the method combining plant tissue culture technique and carbon ion beam irradiations were set out at Institute of Modern Physics from 2005 (Zhou et al. 2006). The effects of 960 MeV carbon ion beam and 8 MeV X-ray irradiations on regenerated shoots of Impatiens hawkeri from another kind of explants named in vitro axillary buds explants were studied recently. The biology endpoints in this study included relative number of roots (RNR), relative length of roots (RLR), relative height of shoots (RHS), relative number of nodes (RNN), survival fraction (SF) and morphology changes in the regenerated shoots. The experimental results showed that carbon ion beams inhibited the root and stem developments of axillary bud explants more severely than X-rays did. And the 50% lethal dose (LD50 ) is about 23.3 Gy for the carbon ion beam and 49.1 Gy for the X-rays, respectively. Relative biological effectiveness (RBE) of Impatiens hawkeri with respect to X-rays according to 50% SF was about two. Secondly, the percentage of shoots regenerated with malformed shoots including curliness, carnification, nicks in all Impatiens hawkeri axillary bud explants irradiated with carbon ion beam at 20 Gy accounted for 55.6%, while the highest number for the 40 Gy X-ray irradiation was 40%. Last, many regenerated shoots whose vascular bundle fused together were obtained only from explants irradiated with carbon ion beams. Based on the results above, it can be concluded that the effect of mutation induction by carbon ion beam irradiation on the axillary explants of Impatiens hawkeri is better than that by X-ray irradiation; and the optimal mutagenic dose varies from 20 Gy

  9. Multimodal evaluation of tissue-engineered cartilage

    OpenAIRE

    Mansour, Joseph M.; Welter, Jean F.

    2013-01-01

    Tissue engineering (TE) has promise as a biological solution and a disease modifying treatment for arthritis. Although cartilage can be generated by TE, substantial inter- and intra-donor variability makes it impossible to guarantee optimal, reproducible results. TE cartilage must be able to perform the functions of native tissue, thus mechanical and biological properties approaching those of native cartilage are likely a pre-requisite for successful implantation. A quality-control assessment...

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

    DEFF Research Database (Denmark)

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

    2013-01-01

    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 learni...

  11. Advances in treatment of articular cartilage injuries

    Directory of Open Access Journals (Sweden)

    Yuan-cheng LI

    2013-05-01

    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.

  12. Multimodal evaluation of tissue-engineered cartilage.

    Science.gov (United States)

    Mansour, Joseph M; Welter, Jean F

    2013-02-01

    Tissue engineering (TE) has promise as a biological solution and a disease modifying treatment for arthritis. Although cartilage can be generated by TE, substantial inter- and intra-donor variability makes it impossible to guarantee optimal, reproducible results. TE cartilage must be able to perform the functions of native tissue, thus mechanical and biological properties approaching those of native cartilage are likely a pre-requisite for successful implantation. A quality-control assessment of these properties should be part of the implantation release criteria for TE cartilage. Release criteria should certify that selected tissue properties have reached certain target ranges, and should be predictive of the likelihood of success of an implant in vivo. Unfortunately, it is not currently known which properties are needed to establish release criteria, nor how close one has to be to the properties of native cartilage to achieve success. Achieving properties approaching those of native cartilage requires a clear understanding of the target properties and reproducible assessment methodology. Here, we review several main aspects of quality control as it applies to TE cartilage. This includes a look at known mechanical and biological properties of native cartilage, which should be the target in engineered tissues. We also present an overview of the state of the art of tissue assessment, focusing on native articular and TE cartilage. Finally, we review the arguments for developing and validating non-destructive testing methods for assessing TE products. PMID:23606823

  13. Chondroitin Sulfate- and Decorin-Based Self-Assembling Scaffolds for Cartilage Tissue Engineering

    Science.gov (United States)

    Recha-Sancho, Lourdes; Semino, Carlos E.

    2016-01-01

    Cartilage injury and degenerative tissue progression remain poorly understood by the medical community. Therefore, various tissue engineering strategies aim to recover areas of damaged cartilage by using non-traditional approaches. To this end, the use of biomimetic scaffolds for recreating the complex in vivo cartilage microenvironment has become of increasing interest in the field. In the present study, we report the development of two novel biomaterials for cartilage tissue engineering (CTE) with bioactive motifs, aiming to emulate the native cartilage extracellular matrix (ECM). We employed a simple mixture of the self-assembling peptide RAD16-I with either Chondroitin Sulfate (CS) or Decorin molecules, taking advantage of the versatility of RAD16-I. After evaluating the structural stability of the bi-component scaffolds at a physiological pH, we characterized these materials using two different in vitro assessments: re-differentiation of human articular chondrocytes (AC) and induction of human adipose derived stem cells (ADSC) to a chondrogenic commitment. Interestingly, differences in cellular morphology and viability were observed between cell types and culture conditions (control and chondrogenic). In addition, both cell types underwent a chondrogenic commitment under inductive media conditions, and this did not occur under control conditions. Remarkably, the synthesis of important ECM constituents of mature cartilage, such as type II collagen and proteoglycans, was confirmed by gene and protein expression analyses and toluidine blue staining. Furthermore, the viscoelastic behavior of ADSC constructs after 4 weeks of culture was more similar to that of native articular cartilage than to that of AC constructs. Altogether, this comparative study between two cell types demonstrates the versatility of our novel biomaterials and suggests a potential 3D culture system suitable for promoting chondrogenic differentiation. PMID:27315119

  14. Laboratory analyses of two explanted hydrophobic acrylic intraocular lenses

    Directory of Open Access Journals (Sweden)

    Yunhai Dai

    2014-01-01

    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.

  15. Organ culture-cell culture system for studying multistage carcinogenesis in respiratory epithelium. [Mice

    Energy Technology Data Exchange (ETDEWEB)

    Steele, Vernon E.; Marchok, Ann C.; Nettesheim, Paul

    1977-01-01

    An organ culture-cell culture system was used to demonstrate carcinogen dose-dependent transformation of tracheal epithelial cells in vitro. Tracheal explants were exposed to MNNG (N-methyl-N/sup 1/-nitro-N-nitrosoguanidine) in organ culture. Outgrowths from these explants provided epithelial cell cultures. The numbers of long term epithelial cell cultures and cell lines that were established per explant increased as MNNG exposure concentration increased. At the present time, more cell lines derived from explants exposed to the highest MNNG concentration have produced palpable tumors than cell lines derived from explants exposed to lower MNNG concentrations. No cell lines were established from primaries derived from control explants. TPA (12-0-tetradecanoyl-phorbol-13-acetate), stimulates DNA synthesis in tracheal epithelium in organ culture in a manner simular to that described for mouse skin. Short exposures to TPA not only stimulated DNA synthesis earlier, but the stimulation was greater than that obtained with continuous exposure. At the present time, exposure of tracheal organ cultures to MNNG followed by TPA has resulted in an enhanced production of morphologically altered cells in primary epithelial cell cultures, than exposure to either agent alone.

  16. Glucosamine increases hyaluronic acid production in human osteoarthritic synovium explants

    OpenAIRE

    Uitterlinden EJ; Koevoet JLM; Verkoelen CF; Bierma-Zeinstra SMA; Jahr H; Weinans H; Verhaar JAN; GJVM van Osch

    2008-01-01

    Abstract Background Glucosamine (GlcN) used by patients with osteoarthritis was demonstrated to reduce pain, but the working mechanism is still not clear. Viscosupplementation with hyaluronic acid (HA) is also described to reduce pain in osteoarthritis. The synthesis of HA requires GlcN as one of its main building blocks. We therefore hypothesized that addition of GlcN might increase HA production by synovium tissue. Methods Human osteoarthritic synovium explants were obtained at total knee s...

  17. Bovine explant model of degeneration of the intervertebral disc

    OpenAIRE

    Sivan Sarit; Menage Janis; Roberts Sally; Urban Jill PG

    2008-01-01

    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 de...

  18. The effect of auxin 2,4-D and cytokinin 2-ip on direct somatic embryogenesis formation of Coffea arabica L. leaf explant

    Directory of Open Access Journals (Sweden)

    Rina Arimarsetiowati

    2011-08-01

    Full Text Available One of the propagation technique for coffee plant production is tissue culture. Tissue culture technique for Coffea arabica L. faces some problems, mainly in the planlet formation regenerated from explants. The objective of this experiment was to examine the effect 2,4-D and 2-ip combination on the formation of direct somatic embryogenesis of Coffea arabica L. in leaves explant. Auxin (2,4-D and cytokinin (2-ip concentrations of, respectively, 1; 5 µM and 5; 10; 15; 20 were used as treatments. This research was conducted using completely randomized design with 10 replications. Observation to induce somatic embryos was done by quantitatively on number of callus from explant and number of embryogenic callus. Beside that, observation by qualitative descriptive was also done on deve lopment of embryogenesis. The results showed that Arabica coffee leaves explant of AS 2K clones could be induced in all medium combination except 5µM 2,4-D and 20µM 2-ip combination. Arabica coffee leaves explant of S 795, Sigararutang and AS 1 varieties could be induced in all medium combination. The highest frequency of callus formation was found in AS 2K, Sigararutang and AS 1 varieties on medium containing 1µM 2,4-D in combination with 10µM 2-ip, whereas for the S 795 variety on medium containing 5µM 2,4-D in combination with 10µM 2-ip. The highest frequency of embriogenic callus in all Arabica coffee variety could be reached on medium containing 5µM 2,4-D in combination with 15µM 2-ip. Key words : Coffea arabica L., somatic embryogenesis, 2,4-D, 2-ip, tissue culture, leaves, callus embryogenic.

  19. Molecular weight characterization of virgin and explanted polyester arterial prostheses.

    Science.gov (United States)

    Maarek, J M; Guidoin, R; Aubin, M; Prud'homme, R E

    1984-10-01

    The macromolecular properties of 17 virgin commercial arterial prostheses and a series of explanted prostheses, both manufactured from poly(ethylene terephthalate) (PET) yarns, have been studied by gel permeation chromatography (GPC) and by differential scanning calorimetry (DSC). Only small differences were found between the average molecular weights and the degree of crystallinity of the unused reference grafts. A broadening of the DSC curves was observed for the prostheses containing texturized yarns compared with those made solely from flat, untexturized yarns. This broadening may be due to greater heterogeneity of the crystal sizes caused by the texturizing process and to the use of two or more different yarns with dissimilar thermal histories in the same prosthesis. Average molecular weights of the explant series were significantly lower than those of the corresponding reference grafts but almost time independent. The polydispersity index and the degree of crystallinity of the explants remained constant as a function of time. These results are discussed in regard to others available in the literature. PMID:6242474

  20. Characterization of human primary chondrocytes of osteoarthritic cartilage at varying severity

    Institute of Scientific and Technical Information of China (English)

    YIN Jing; YANG Zheng; CAO Yong-ping; GE Zi-gang

    2011-01-01

    Background There is a difficulty in evaluating the in vivo functionality of individual chondrocytes,and there is much heterogeneity among cartilage affected by osteoarthritis (OA).In this study,in vitro cultured chondrocytes harvested from varying stages of degeneration were studied as a projective model to further understand the pathogenesis of osteoarthritis.Methods Cartilage of varying degeneration of end-stage OA was harvested,while cell yield and matrix glycosaminoglycan (GAG) content were measured.Cell morphology,proliferation,and gene expression of collagen type Ⅰ,Ⅱ,and Ⅹ,aggrecan,matrix metalloproteinase 13 (MMP-13),and ADAMTS5 of the acquired chondrocytes were measured during subsequent in vitro culture.Results Both the number of cells and the GAG content increased with increasing severity of OA.Cell spreading area increased and gradually showed spindle-like morphology during in vitro culture.Gene expression of collagen type Ⅱ,collagen type X as well as GAG decreased with severity of cartilage degeneration,while expression of collagen type Ⅰ increased.Expression of MMP-13 increased with severity of cartilage degeneration,while expression of ADAMTS-5 remained stable.Expression of collagen type Ⅱ,X,GAG,and MMP-13 substantially decreased with in vitro culture.Expression of collagen type Ⅰ increased with in vitro cultures,while expression of ADAMTS 5 remained stable.Conclusions Expression of functional genes such as collagen type Ⅱ and GAG decreased during severe degeneration of OA cartilage and in vitro dedifferentiation.Gene expression of collagen Ⅰ and MMP-13 increased with severity of cartilage degeneration.

  1. ORGANOGÊNESE IN VITRO DE Citrus EM FUNÇÃO DE CONCENTRAÇÕES DE BAP E SECCIONAMENTO DO EXPLANTE CITRUS IN VITRO ORGANOGENESIS RELATED TO BAP CONCENTRATIONS AND EXPLANT SECTION

    Directory of Open Access Journals (Sweden)

    THAÍS LACAVA DE MOURA

    2001-08-01

    'Valencia' sweet orange (Citrus sinensis (L. Osbeck. For Rangpur lime, internodal explants were cultured in MT medium, with 25g.L-1 sucrose and supplemented with different concentrations of BAP (0; 2.5; 5; 7.5; 10 mg.L-1. Epicotyl explants were used for 'Pera' and 'Valencia' sweet oranges. For 'Pera' sweet orange, the explants were cultured in MT medium, with 25g.L-1 sucrose and supplemented with different concentrations of BAP (0; 1; 2; 3; 4 mg.L-1. For 'Valencia' sweet orange, the epicotyl explants were cut in half and cultured on MT medium, with 25 g.L-1 of sucrose and 1 mg.L-1 of BAP. Shoot elongation was accomplished on MT + 25 g.L-1 sucrose + 1 mg.L-1 gibberelic acid (GA3 and rooted on MT + 25 g.L-1 of sucrose + 0,5 g.L-1 of activated charcoal + 1 mg.L-1 NAA. The best results for number of buds was 2.5 mg.L-1 of BAP in Rangpur lime and 1 and 2 mg.L-1 in 'Pera' sweet orange. The cut of the explant in half was favorable to the in vitro organogenesis in 'Valencia' sweet orange, but it was not for rooting.

  2. Acetylcholine suppresses shoot formation and callusing in leaf explants of in vitro raised seedlings of tomato, Lycopersicon esculentum Miller var. Pusa Ruby.

    Science.gov (United States)

    Bamel, Kiran; Gupta, Rajendra; Gupta, Shirish C

    2016-06-01

    We present experimental evidence to show that acetylcholine (ACh) causes decrease in shoot formation in leaf explants of tomato (Lycopersicon esculentum Miller var Pusa Ruby) when cultured on shoot regeneration medium. The optimum response was obtained at 10(-4) M ACh-enriched medium. ACh also causes decrease in percentage of cultures forming callus and reduces the callus mass. Inhibitors of enzymatic hydrolysis of ACh, neostigmine and physostigmine, also suppresses callogenesis and caulogenesis. On the other hand, the breakdown products of Ach, choline and acetate, do not alter the morphogenic response induced on the shoot regeneration medium. Neostigmine showed optimal reduction in shoot formation at 10(-5) M. The explants cultured on neostigmine augmented medium showed decline in the activity of ACh hydrolyzing enzyme acetylcholinesterase. ACh and neostigmine added together showed marked reduction in callus mass. These results strongly support the role of ACh as a natural regulator of morphogenesis in tomato plants. PMID:27348536

  3. Cold Atmospheric Plasma Modified Electrospun Scaffolds with Embedded Microspheres for Improved Cartilage Regeneration.

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    Wei Zhu

    Full Text Available Articular cartilage is prone to degeneration and possesses extremely poor self-healing capacity due to inherent low cell density and the absence of a vasculature network. Tissue engineered cartilage scaffolds show promise for cartilage repair. However, there still remains a lack of ideal biomimetic tissue scaffolds which effectively stimulate cartilage regeneration with appropriate functional properties. Therefore, the objective of this study is to develop a novel biomimetic and bioactive electrospun cartilage substitute by integrating cold atmospheric plasma (CAP treatment with sustained growth factor delivery microspheres. Specifically, CAP was applied to a poly(ε-caprolactone electrospun scaffold with homogeneously distributed bioactive factors (transforming growth factor-β1 and bovine serum albumin loaded poly(lactic-co-glycolic acid microspheres. We have shown that CAP treatment renders electrospun scaffolds more hydrophilic thus facilitating vitronectin adsorption. More importantly, our results demonstrate, for the first time, CAP and microspheres can synergistically enhance stem cell growth as well as improve chondrogenic differentiation of human marrow-derived mesenchymal stem cells (such as increased glycosaminoglycan, type II collagen, and total collagen production. Furthermore, CAP can substantially enhance 3D cell infiltration (over two-fold increase in infiltration depth after 1 day of culture in the scaffolds. By integrating CAP, sustained bioactive factor loaded microspheres, and electrospinning, we have fabricated a promising bioactive scaffold for cartilage regeneration.

  4. Development of cartilage conduction hearing aid

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    H. Hosoi

    2010-04-01

    Full Text Available Purpose: The potential demand for hearing aids is increasing in accordance with aging of populations in many developed countries. Because certain patients cannot use air conduction hearing aids, they usually use bone conduction hearing aids. However, bone does not transmit sound as efficiently as air, and bone conduction hearing aids require surgery (bone anchored hearing aid or great pressure to the skull. The first purpose of this study is to examine the efficacy of a new sound conduction pathway via the cartilage. The second purpose is to develop a hearing aid with a cartilage conduction transducer for patients who cannot use regular air conduction hearing aids.Design/methodology/approach: We examined the hearing ability of a patient with atresia of both external auditory meatuses via three kinds of conduction pathways (air, bone, and cartilage. After the best position for the cartilage conduction transducer was found, audiometric evaluation was performed for his left ear with an insertion earphone (air conduction, a bone conduction transducer, and a cartilage conduction transducer. Then we made a new hearing aid using cartilage conduction and got subjective data from the patients.Findings: The tragal cartilage was the best position for the cartilage conduction transducer. The patient’s mean hearing levels were 58.3 dBHL, 6.7 dBHL, and 3.3 dBHL for air conduction, bone conduction, and cartilage conduction respectively. The hearing ability of the patients obtained from the cartilage conduction hearing aid was comparable to those from the bone conduction hearing aid.Practical implications: Hearing levels using cartilage conduction are very similar to those via bone conduction. Cartilage conduction hearing aids may overcome the practical disadvantages of bone conduction hearing aids such as pain and the need for surgery.Originality/value: We have clarified the efficacy of the cartilage conduction pathway and developed a prototype ‘cartilage

  5. Callus induction and biomass accumulation in vitro in explants from chokeberry (Aronia melanocarpa (Michx. Elliot fruit

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    Tatiana I. Calalb

    2014-09-01

    Full Text Available In this study, the following features were determined: biological (the optimal histogen as explant and the optimal age of donor fruit, biotechnological (type, dosage and combination of growth regulators supplements in culture medium Murashige and Skoog as well as sucrose dosage, and physical (light regime, to induce callusing and biomass accumulation in vitro from the succulent chokeberry (Aronia melanocarpa (Michx. Elliot fruit. It turned out that it was much easier to induce callus from explants composed of the epicarp and hypoderm cut from fruits at 50–60 days after flowering. The role of light regime and varied supplementation of the basic MS medium with different doses of growth regulators was established; they resulted in four pigmented carpomass: violet, cream-pink, cream-white and green. The best combinations for the proliferation of fruit callus were culture media with 0.2–2.5 mg × dm-3 2,4-D+0.5 mg × dm-3 KIN +60 g × dm-3sucrose, while for fruit biomass accumulation enriched with phenolic substances – 2.5–3.5 mg × dm-3 NAA+0.5 mg × dm-3 KIN+60 g × dm-3sucrose. The chemical study of phenolic compounds by HPLC coupled with the mass spectrometry method identified chlorogenic acid, hiperozide, quercetrin, isoquercitrin and rutozide quantitatively and qualitatively in all pigmented carpomass and fruits; an exception is p-coumaric present only qualitatively in green carpomass and absent in fruit and quercetol absent in green carpomass.

  6. Anatomical study of nasal cartilage in buffalo (Bubalus bubulus

    Directory of Open Access Journals (Sweden)

    Mahdi Yeganehzad

    2011-07-01

    Full Text Available This study used ten heads of adult buffalo taken from slaughterhouse. After transferring the samples to the anatomy hall, a split was carefully created on skin of muzzle and the skin was slowly separated from muscles and hypodermal connective tissue. Place of connection of cartilages to bone, cartilages to each other and shape of the cartilages were specified. In buffalo, nose apex has two nostrils fixed by bone and cartilage. After identifying and separating the cartilages, it was found that nasal cartilages in buffalo consisted of: 1 septum nasal located between two nostrils and reinforces it from inside. 2 dorso-lateral nasal cartilage constituting dorsal and lateral parts of the nostril. 3 ventro-lateral nasal cartilage constituting ventral and lateral parts of the nostril. 4 lateral accessory cartilage constituting lateral and ventral parts of the nostril. 5 medial accessory nasal cartilage located at Alar fold and connected to ventro-lateral nasal cartilage.

  7. Comprehensive Profiling of Cartilage Extracellular Matrix Formation and Maturation Using Sequential Extraction and Label-free Quantitative Proteomics*

    OpenAIRE

    Wilson, Richard; Diseberg, Anders F.; Gordon, Lavinia; Zivkovic, Snezana; Tatarczuch, Liliana; Mackie, Eleanor J.; Gorman, Jeffrey J.; Bateman, John F.

    2010-01-01

    Articular cartilage is indispensable for joint function but has limited capacity for self-repair. Engineering of neocartilage in vitro is therefore a major target for autologous cartilage repair in arthritis. Previous analysis of neocartilage has targeted cellular organization and specific molecular components. However, the complexity of extracellular matrix (ECM) development in neocartilage has not been investigated by proteomics. To redress this, we developed a mouse neocartilage culture sy...

  8. Prolactin promotes cartilage survival and attenuates inflammation in inflammatory arthritis

    Science.gov (United States)

    Adán, Norma; Guzmán-Morales, Jessica; Ledesma-Colunga, Maria G.; Perales-Canales, Sonia I.; Quintanar-Stéphano, Andrés; López-Barrera, Fernando; Méndez, Isabel; Moreno-Carranza, Bibiana; Triebel, Jakob; Binart, Nadine; Martínez de la Escalera, Gonzalo; Thebault, Stéphanie; Clapp, Carmen

    2013-01-01

    Chondrocytes are the only cells in cartilage, and their death by apoptosis contributes to cartilage loss in inflammatory joint diseases, such as rheumatoid arthritis (RA). A putative therapeutic intervention for RA is the inhibition of apoptosis-mediated cartilage degradation. The hormone prolactin (PRL) frequently increases in the circulation of patients with RA, but the role of hyperprolactinemia in disease activity is unclear. Here, we demonstrate that PRL inhibits the apoptosis of cultured chondrocytes in response to a mixture of proinflammatory cytokines (TNF-α, IL-1β, and IFN-γ) by preventing the induction of p53 and decreasing the BAX/BCL-2 ratio through a NO-independent, JAK2/STAT3–dependent pathway. Local treatment with PRL or increasing PRL circulating levels also prevented chondrocyte apoptosis evoked by injecting cytokines into the knee joints of rats, whereas the proapoptotic effect of cytokines was enhanced in PRL receptor–null (Prlr–/–) mice. Moreover, eliciting hyperprolactinemia in rats before or after inducing the adjuvant model of inflammatory arthritis reduced chondrocyte apoptosis, proinflammatory cytokine expression, pannus formation, bone erosion, joint swelling, and pain. These results reveal the protective effect of PRL against inflammation-induced chondrocyte apoptosis and the therapeutic potential of hyperprolactinemia to reduce permanent joint damage and inflammation in RA. PMID:23908112

  9. Regeneration of articular cartilage using adipose stem cells.

    Science.gov (United States)

    Im, Gun-Il

    2016-07-01

    Articular cartilage (AC) has limited potential for self-regeneration and damage to AC eventually leads to the development and progression of osteoarthritis (OA). Cell implantation strategies have emerged as a new treatment modality to regenerate AC. Adipose stem cells/adipose-derived stromal cells (ASCs) have gained attention due to their abundance, excellent proliferative potential, and minimal morbidity during harvest. These advantages lower the cost of cell therapy by circumventing time-consuming procedure of culture expansion. ASCs have drawn attention as a potential source for cartilage regeneration since the feasibility of chondrogenesis from ASCs was first reported. After several groups reported inferior chondrogenesis from ASCs, numerous methods were devised to overcome the intrinsic properties. Most in vivo animal studies have reported good results using predifferentiated or undifferentiated, autologous or allogeneic ASCs to regenerate cartilage in osteochondral defects or surgically-induced OA. In this review, we summarize literature on the isolation and in vitro differentiation processes of ASCs, in vivo studies to regenerate AC in osteochondral defects and OA using ASCs, and clinical applications of ASCs. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1830-1844, 2016. PMID:26990234

  10. Effect of ZnO Nanoparticles on Brassica nigra Seedlings and Stem Explants: Growth Dynamics and Antioxidative Response

    Science.gov (United States)

    Zafar, Hira; Ali, Attarad; Ali, Joham S.; Haq, Ihsan U.; Zia, Muhammad

    2016-01-01

    Nanoparticles (NPs) have diverse properties when compared to respective chemicals due to their structure, surface to volume ratio, morphology, and reactivity. Toxicological effects of metallic NPs on organisms including plants have been reported. However, to the best of our knowledge, there is still not any report on the effect of NPs on in vitro culture of plant explants. In this study, ZnO NPs concentration ranging from 500 to 1500 mg/L adversely affects the Brassica nigra seed germination and seedling growth and also lead to an increase in the antioxidative activities and non-enzymatic antioxidants. While, culturing the stem explants of B. nigra on Murashige and Skoog (MS) medium at lower concentration of ZnO NPs (1–20 mg/L) resulted in the production of white thin roots with thick root hairs. At 10 mg/L ZnO NPs, shoots emergence is also observed. The developed calli/roots showed 79% DPPH (2,2-diphenyl-1-picryl hydrazyl) radical scavenging activity at 10 mg/L. The total antioxidant and reducing power potential also significantly affected in presence of ZnO NPs. Moreover, an increase in non-enzymatic antioxidative molecules, phenolics (up to 0.15 μg GAE/mg FW) and flavonoids (up to 0.22 μg QE/mg FW), depending on NPs concentration is also observed. We conclude that ZnO NPs may induce roots from explants cultured on appropriate medium that can be used for production of valuable secondary metabolites. PMID:27148347

  11. Cysts of the semilunar cartilage

    International Nuclear Information System (INIS)

    On the basis of the studies listed in the bibliography, this dissertation reports on the pathology, clinical symptoms and radiology of cysts of the semilunar cartilage. The author analyses 118 cases of his own, with special regard to the results of pneumo-arthrographic investigations carried through according to a special technique by Schaefer. In the course of this work, measurements of the meniscal base are for the first time used as radiological criteria indicating the presence of a cyst of the semilunar cartilage. Furthermore the well-known radiological signs of cysts, such as bone defects according to Albert and Keller, light central spot in the meniscal body, as well as Rauber's sign and horizontal rupture, are investigated as to the frequency of their incidence. For that purpose all the X-ray pictures were subjected to a further dose scrutiny. A list of all the 118 cases with their clinical and radiological data is found in the annex, together with the results of the operations and patho-anatomical investigations. (orig.)

  12. In vitro of quantitative MR imaging of early degenerative changes in human articular cartilage

    International Nuclear Information System (INIS)

    To assess the applicability of quantitative MR microscopy for the detection of glycosaminoglycan (GAG) depletion as an early sign of degeneration in the articular cartilage of humans treated by trypsin. Four cartilage-bone blocks were obtained from the patient who had suffered from osteoarthritis of the knee and underwent a total knee replacement arthroscopy. Each articular cartilage segment was resected as to a round disk shape (8 mm in diameter) with a remnant of subchondral bone 1 mm in thickness. Four different culture solutions were prepared, and these solutions were 0.2 mg/ml of trypsin solution (group 1), 1 mM of Gd (DTPA) 2-mixed trypsin solution (group 2), phosphate buffered saline (PBS) (group 3), and 1 mM of Gd (DTPA) 2-mixed PBS (group 4). The cartilages were cultured and then MR imagings were performed every hour for 5 hrs, and we continued the additional cultures of 24 hrs, 36 hrs and 48 hrs. Three imaging sequences were used: T1-weighted spin echo (TR/TE, 450/22), proton density turbo spin echo with fat suppression (TR/TE, 3000/25), and CPMG (Carr-Purcell-Meiboom-Gill) (TR/TE/TI, 760/21-168, 360). MR imaging data were analyzed with pixel-by-pixel comparisons in all groups. The GAG loss in the articular cartilage was increased proportionately to the culture duration. Mean changes of T1 relaxation time were 1.2% for group 1, -1.9% for group 3, -54.7% for group 2 and -64.2% for group 4 (p< 0.05). When comparing by linear profile on the T1-weighted images, SNR increased and T1 relaxation time decreased for group 2 and 4, as the culture duration increased (p< 0.05). On the correlation analysis, there is significant correlation between GAG loss and Gd (DTPA) 2-enhancement for group 2 (p=0.0431), but there was no significant difference for group 4 (p=0.0918). More enhancement with Gd (DTPA) 2-was noted for group 2 than for group 4. Group 2 showed a diffuse enhancement in all the layers of cartilage, but for group 4, prominent enhancement was noted only in

  13. Nondestructive Assessment of Engineered Cartilage Composition by Near Infrared Spectroscopy.

    Science.gov (United States)

    McGoverin, Cushla M; Hanifi, Arash; Palukuru, Uday P; Yousefi, Farzad; Glenn, Padraig B M; Shockley, Michael; Spencer, Richard G; Pleshko, Nancy

    2016-03-01

    Tissue engineering presents a strategy to overcome the limitations of current tissue healing methods. Scaffolds, cells, external growth factors and mechanical input are combined in an effort to obtain constructs with properties that mimic native tissues. However, engineered constructs developed using similar culture environments can have very different matrix composition and biomechanical properties. Accordingly, a nondestructive technique to assess constructs during development such that appropriate compositional endpoints can be defined is desirable. Near infrared spectroscopy (NIRS) analysis is a modality being investigated to address the challenges associated with current evaluation techniques, which includes nondestructive compositional assessment. In the present study, cartilage tissue constructs were grown using chondrocytes seeded onto polyglycolic acid (PGA) scaffolds in similar environments in three separate tissue culture experiments and monitored using NIRS. Multivariate partial least squares (PLS) analysis models of NIR spectra were calculated and used to predict tissue composition, with biochemical assay information used as the reference data. Results showed that for combined data from all tissue culture experiments, PLS models were able to assess composition with significant correlations to reference values, including engineered cartilage water (at 5200 cm(-1), R = 0.68, p = 0.03), proteoglycan (at 4310 cm(-1), R = 0.82, p = 0.007), and collagen (at 4610 cm(-1), R = 0.84, p = 0.005). In addition, degradation of PGA was monitored using specific NIRS frequencies. These results demonstrate that NIR spectroscopy combined with multivariate analysis provides a nondestructive modality to assess engineered cartilage, which could provide information to determine the optimal time for tissue harvest for clinical applications. PMID:26817457

  14. Imaging diagnosis of the articular cartilage disorders

    International Nuclear Information System (INIS)

    Objective: To evaluate the diagnosis and differential diagnosis among the chronic osteoarthritis, rheumatoid arthritis and other chronic cartilage lesions on the plain films and MR images. Methods: Eighty-nine cases, including 115 joints, underwent plain film and MRI examination, and enhanced MRI scan was performed on 32 of them, including 44 joints. MRI scan sequences consisted of T1WI, T2WI + PDWI, STIR, and 3D FS SPGR. There were 90 knee joints in this group and each of the articular cartilage was divided into four parts: patella, femoral medial condyle, femoral lateral condyle, and tibia facet on MR images. The cartilage disorders were classified according to the outerbridge method. In addition, 61 cases including 75 joints were observed as a control group on the plain films and MR images. Results: 115 cartilage lesions were found on MR images, in which thinness of the cartilage (58 cases, 50.4%), bone changes under the cartilage (22 cases, 19.7%), medullar edema (22 cases, 19.7%), and synovial hyperplasia (52 cases, 45.2%) were seen. The patella cartilage was the most likely affected part (81/90, 90%). So the patellar cartilage lesions were divided as group 1 (grade I-II) and group 2 (grade III-IV) on MR images, which were compared with the plain film signs. The narrowing of the joint space and saccules under the articular surface were statistically significant with each other, and χ2 values were 9.349 and 9.885, respectively (P=0.002). Conclusion: No constant signs could be seen on the plain films with grade I-II cartilage disorders. While the narrowing joint space and saccules under the joint surface could be seen on them with grade III-IV cartilage disorders, which were mainly correlated with the cartilage disorders and bone changes under the articular cartilages. A combination of the plain films and MR images is the best imaging method for examining the joints and joint cartilages. Enhanced MRI scan is very helpful on the diagnosis and differential

  15. Transforming growth factor-β1 regulated phosphorylated AKT and interferon gamma expressions are associated with epithelial cell survival in rhesus macaque colon explants.

    Science.gov (United States)

    Pahar, Bapi; Pan, Diganta; Lala, Wendy; Kenway-Lynch, Carys S; Das, Arpita

    2015-05-01

    Transforming growth factor-β1 (TGF-β1) is an important immunoregulatory cytokine that plays an obligate role in regulating T-cell functions. Here, we demonstrated the role of TGF-β1 in regulating the survival of intestinal epithelial cells (ECs) in rhesus colon explant cultures using either anti-TGF-β1 antibody or recombinant TGF-β1 proteins. Neutralization of endogenous TGF-β1 using anti-TGF-β1 antibodies induced apoptosis of both intestinal ECs and lamina propria (LP) cells. Additionally, endogenous TGF-β1 blocking significantly increased expression of IFNγ, TNFα, CD107a and Perforin in LP cells compared to media and isotype controls. A significant decrease in pAKT expression was detected in anti-TGF-β1 MAbs treated explants compared to isotype and rTGF-β1 protein treated explants. Our results demonstrated TGF-β1 regulated pAKT and IFNγ expressions were associated with epithelial cell survival in rhesus macaque colon explants and suggest a potential role of mucosal TGF-β1 in regulating intestinal homeostasis and EC integrity. PMID:25769244

  16. Establishment of high frequency shoot regeneration system in Himalayan poplar (Populus ciliata Wall. ex Royle) from petiole explants using Thidiazuron cytokinin as plant growth regulator

    Institute of Scientific and Technical Information of China (English)

    G Aggarwal; A Gaur; D K Srivastava

    2015-01-01

    Populus species are important resources for industry and in scientific study on biological and agricul-tural systems. Our objective was to enhance the frequency of plant regeneration in Himalayan poplar (Populus ciliata wall. ex Royle). The effect of TDZ alone and in combi-nation with adenine and NAA was studied on the regen-eration potential of petiole explants. The explants were excised from Himalayan poplar plants grown in glass-houses. After surface sterilization the explants were cul-tured on shoot induction medium. High percentage shoot regeneration (86%) was recorded on MS medium sup-plemented with 0.004 mg L-1 TDZ and 79.7 mg L-1 adenine. The regenerated shoots for elongation and multi-plication were transferred to MS ? 0.5 mg L-1 BAP ? 0.2 mg L-1 IAA ? 0.3 mg L-1 GA3. Root re-generation from shoots developed in vitro was observed on MS medium supplemented with 0.10 mg L-1 IBA. Hi-malayan poplar plantlets could be produced within 2 months after acclimatization in a sterile mixture of sand and soil. We developed a high efficiency plant regeneration protocol from petiole explants of P. ciliata.

  17. TRANSFORMING GROWTH FACTOR-β AND FIBROBLAST GROWTH FACTOR INDUCE LENS EPITHELIAL EXPLANT METAPLASIA: IMPLICATIONS FOR THE FORMATION OF SUBCAPSULAR OPACIFICATION

    Institute of Scientific and Technical Information of China (English)

    刘颉; 叶俊杰

    1998-01-01

    Objective. This study was to investigate the effects of transforming growth factor-β(TGFβ) and fibroblast growth factor (FGF) in the subcapsular opaeification formation of the lens. Methods. Lens epithelial explants from 10-day-old rats were cultured with TGFβ1 or TGFβ2 in the presence of FGF for 5 days, then were examined by light and electron microscopy, and by immunolocalization of α-smooth muscle(α-sm) actin and type Ⅰ collagen. Resets. In TGFβ/FGF-treated explants,extensive proliferation oeeured, with formation of spindle and star-shaped cells. These cells showed ultrastructure and biochemical features of fibroblast or myofibroblast.Prominent Golgi apparatus and rough endoplaie reticulum were observed in scene cells, Intracellular microfilaments with cytoplasmic dense bodies and membrane associated dense bodies, features of smooth muscle cells, were also observed. Some cells showed reactivity to α-sin actin antibody. TGFβ/FGF-treated explants were strongly stained with type I collagen antibody. Conclusion. In the presence of FGF, TGFβ1 and TGFβ2 induced lens epithelial cell(LEC)proliferation and transformation into fibroblast or myofibroblast like ceils, with producing of abundant collagen matrix in the explants. The changes are similar to the metaplasia that oeeurrs in subeapsular opacification of the lens. The findings suggest that TGFβ and FGF plays a role in the pathogenesis of subcapsular opacification of the lens.

  18. Usual interstitial pneumonia end-stage features from explants with radiologic and pathological correlations.

    Science.gov (United States)

    Rabeyrin, Maud; Thivolet, Françoise; Ferretti, Gilbert R; Chalabreysse, Lara; Jankowski, Adrien; Cottin, Vincent; Pison, Christophe; Cordier, Jean-François; Lantuejoul, Sylvie

    2015-08-01

    Idiopathic pulmonary fibrosis (IPF) is the most frequent and severe idiopathic interstitial pneumonia, with typical high-resolution computed tomography (HRCT) features and histologic pattern of usual interstitial pneumonia (UIP); its main differential diagnosis is fibrotic nonspecific interstitial pneumonia (F-NSIP). Usual interstitial pneumonia was mainly described from lung biopsies, and little is known on explants. Twenty-two UIP/IPF explants were analyzed histologically and compared with previous open lung biopsies (OLBs; n = 11) and HRCT (n = 19), when available. Temporospatial heterogeneity and subpleural and paraseptal fibrosis were similarly found in UIP/IPF explants and OLB (91%-95%). Fibroblastic foci were found in 82% of OLBs and 100% of explants, with a higher mean score in explants (P = .023). Honeycombing was present in 64% of OLBs and 95% of explants, with a higher mean score in explants (P = .005). Almost 60% of UIP/IPF explants showed NSIP areas and 41% peribronchiolar fibrosis; inflammation, bronchiolar metaplasia, and vascular changes were more frequent in UIP/IPF explants; and Desquamative Interstitial Pneumonia (DIP)-like areas were not common (18%-27%). Numerous large airspace enlargements with fibrosis were frequent in UIP/IPF explants (59%). On HRCT, honeycombing was observed in 95% of the cases and ground-glass opacities in 53%, correlating with NSIP areas or acute exacerbation at histology. Six patients had combined IPF and emphysema. Lesions were more severe in UIP/IPF explants, reflecting the worsening of the disease. Usual interstitial pneumonia/IPF explants more frequently presented with confounding lesions such as NSIP areas, peribronchiolar fibrosis, and airspace enlargements with fibrosis sometimes associated with emphysema. PMID:26025258

  19. Replication of avian, human and swine influenza viruses in porcine respiratory explants and association with sialic acid distribution

    Directory of Open Access Journals (Sweden)

    Nauwynck Hans J

    2010-02-01

    Full Text Available Abstract Background Throughout the history of human influenza pandemics, pigs have been considered the most likely "mixing vessel" for reassortment between human and avian influenza viruses (AIVs. However, the replication efficiencies of influenza viruses from various hosts, as well as the expression of sialic acid (Sia receptor variants in the entire porcine respiratory tract have never been studied in detail. Therefore, we established porcine nasal, tracheal, bronchial and lung explants, which cover the entire porcine respiratory tract with maximal similarity to the in vivo situation. Subsequently, we assessed virus yields of three porcine, two human and six AIVs in these explants. Since our results on virus replication were in disagreement with the previously reported presence of putative avian virus receptors in the trachea, we additionally studied the distribution of sialic acid receptors by means of lectin histochemistry. Human (Siaα2-6Gal and avian virus receptors (Siaα2-3Gal were identified with Sambucus Nigra and Maackia amurensis lectins respectively. Results Compared to swine and human influenza viruses, replication of the AIVs was limited in all cultures but most strikingly in nasal and tracheal explants. Results of virus titrations were confirmed by quantification of infected cells using immunohistochemistry. By lectin histochemistry we found moderate to abundant expression of the human-like virus receptors in all explant systems but minimal binding of the lectins that identify avian-like receptors, especially in the nasal, tracheal and bronchial epithelium. Conclusions The species barrier that restricts the transmission of influenza viruses from one host to another remains preserved in our porcine respiratory explants. Therefore this system offers a valuable alternative to study virus and/or host properties required for adaptation or reassortment of influenza viruses. Our results indicate that, based on the expression of Sia

  20. The effects of Brazilian propolis on etiological agents of mastitis and the viability of bovine mammary gland explants.

    Science.gov (United States)

    Fiordalisi, Samira A L; Honorato, Luciana A; Loiko, Márcia R; Avancini, César A M; Veleirinho, Maria B R; Machado Filho, Luiz C P; Kuhnen, Shirley

    2016-03-01

    The objective of this study was to evaluate in vitro the antimicrobial activity of Brazilian propolis from Urupema, São Joaquim, and Agua Doce (Santa Catarina State) and green propolis from Minas Gerais State, and the effects of propolis on bovine mammary gland explant viability. The propolis samples differed in flavonoid content and antioxidant activity. Green propolis showed the highest content of flavonoids, followed by the sample from São Joaquim. The propolis from Urupema showed the lowest flavonoid content along with the lowest antioxidant activity. The total phenolics were similar across all studied samples. Despite phytochemical differences, the propolis samples from Minas Gerais, São Joaquim, and Urupema presented the same level of antimicrobial activity against Staphylococcus aureus strains. The reduction in S. aureus growth was, on average, 1.5 and 4 log10 times at 200 and 500 μg/mL, respectively. At concentrations of 1,000 μg/mL, all propolis reduced bacterial growth to zero. On the other hand, when the propolis were tested against strains of Escherichia coli, the samples presented weak antimicrobial activity. Mammary explants were maintained in culture for 96h without a loss in viability, demonstrating the applicability of the model in evaluating the toxicity of propolis. The origin and chemical composition of the propolis had an effect on mammary explant viability. We encountered inhibitory concentrations of 272.4, 171.8, 63.85, and 13.26 μg/mL for the propolis from Água Doce, Urupema, São Joaquim, and Mina Gerais, respectively. A clear association between greater antimicrobial activity and toxicity for mammary explants was observed. Of all propolis tested, the Urupema sample was noteworthy, as it showed antimicrobial activity at less toxic concentrations than the other samples, reducing bacterial growth to an average of 9.3 × 10(2) cfu/mL after 6h of contact using 200 μg/mL of extract. The results demonstrate the potential for Brazilian

  1. Novel Elements of the Chondrocyte Stress Response Identified Using an in Vitro Model of Mouse Cartilage Degradation.

    Science.gov (United States)

    Wilson, Richard; Golub, Suzanne B; Rowley, Lynn; Angelucci, Constanza; Karpievitch, Yuliya V; Bateman, John F; Fosang, Amanda J

    2016-03-01

    The destruction of articular cartilage in osteoarthritis involves chondrocyte dysfunction and imbalanced extracellular matrix (ECM) homeostasis. Pro-inflammatory cytokines such as interleukin-1α (IL-1α) contribute to osteoarthritis pathophysiology, but the effects of IL-1α on chondrocytes within their tissue microenvironment have not been fully evaluated. To redress this we used label-free quantitative proteomics to analyze the chondrocyte response to IL-1α within a native cartilage ECM. Mouse femoral heads were cultured with and without IL-1α, and both the tissue proteome and proteins released into the media were analyzed. New elements of the chondrocyte response to IL-1α related to cellular stress included markers for protein misfolding (Armet, Creld2, and Hyou1), enzymes involved in glutathione biosynthesis and regeneration (Gstp1, Gsto1, and Gsr), and oxidative stress proteins (Prdx2, Txn, Atox1, Hmox1, and Vnn1). Other proteins previously not associated with the IL-1α response in cartilage included ECM components (Smoc2, Kera, and Crispld1) and cysteine proteases (cathepsin Z and legumain), while chondroadherin and cartilage-derived C-type lectin (Clec3a) were identified as novel products of IL-1α-induced cartilage degradation. This first proteome-level view of the cartilage IL-1α response identified candidate biomarkers of cartilage destruction and novel targets for therapeutic intervention in osteoarthritis. PMID:26794603

  2. Priming Adipose-Derived Mesenchymal Stem Cells with Hyaluronan Alters Growth Kinetics and Increases Attachment to Articular Cartilage

    Directory of Open Access Journals (Sweden)

    Peter Succar

    2016-01-01

    Full Text Available Background. Biological therapeutics such as adipose-derived mesenchymal stem cell (MSC therapy are gaining acceptance for knee-osteoarthritis (OA treatment. Reports of OA-patients show reductions in cartilage defects and regeneration of hyaline-like-cartilage with MSC-therapy. Suspending MSCs in hyaluronan commonly occurs in animals and humans, usually without supporting data. Objective. To elucidate the effects of different concentrations of hyaluronan on MSC growth kinetics. Methods. Using a range of hyaluronan concentrations, we measured MSC adherence and proliferation on culture plastic surfaces and a novel cartilage-adhesion assay. We employed time-course and dispersion imaging to assess MSC binding to cartilage. Cytokine profiling was also conducted on the MSC-secretome. Results. Hyaluronan had dose-dependent effects on growth kinetics of MSCs at concentrations of entanglement point (1 mg/mL. At higher concentrations, viscosity effects outweighed benefits of additional hyaluronan. The cartilage-adhesion assay highlighted for the first time that hyaluronan-primed MSCs increased cell attachment to cartilage whilst the presence of hyaluronan did not. Our time-course suggested patients undergoing MSC-therapy for OA could benefit from joint-immobilisation for up to 8 hours. Hyaluronan also greatly affected dispersion of MSCs on cartilage. Conclusion. Our results should be considered in future trials with MSC-therapy using hyaluronan as a vehicle, for the treatment of OA.

  3. Fetal mesenchymal stromal cells differentiating towards chondrocytes acquire a gene expression profile resembling human growth plate cartilage.

    Directory of Open Access Journals (Sweden)

    Sandy A van Gool

    Full Text Available We used human fetal bone marrow-derived mesenchymal stromal cells (hfMSCs differentiating towards chondrocytes as an alternative model for the human growth plate (GP. Our aims were to study gene expression patterns associated with chondrogenic differentiation to assess whether chondrocytes derived from hfMSCs are a suitable model for studying the development and maturation of the GP. hfMSCs efficiently formed hyaline cartilage in a pellet culture in the presence of TGFβ3 and BMP6. Microarray and principal component analysis were applied to study gene expression profiles during chondrogenic differentiation. A set of 232 genes was found to correlate with in vitro cartilage formation. Several identified genes are known to be involved in cartilage formation and validate the robustness of the differentiating hfMSC model. KEGG pathway analysis using the 232 genes revealed 9 significant signaling pathways correlated with cartilage formation. To determine the progression of growth plate cartilage formation, we compared the gene expression profile of differentiating hfMSCs with previously established expression profiles of epiphyseal GP cartilage. As differentiation towards chondrocytes proceeds, hfMSCs gradually obtain a gene expression profile resembling epiphyseal GP cartilage. We visualized the differences in gene expression profiles as protein interaction clusters and identified many protein clusters that are activated during the early chondrogenic differentiation of hfMSCs showing the potential of this system to study GP development.

  4. Regulatory Challenges for Cartilage Repair Technologies.

    Science.gov (United States)

    McGowan, Kevin B; Stiegman, Glenn

    2013-01-01

    In the United States, few Food and Drug Administration (FDA)-approved options exist for the treatment of focal cartilage and osteochondral lesions. Developers of products for cartilage repair face many challenges to obtain marketing approval from the FDA. The objective of this review is to discuss the necessary steps for FDA application and approval for a new cartilage repair product. FDA Guidance Documents, FDA Panel Meetings, scientific organization recommendations, and clinicaltrials.gov were reviewed to demonstrate the current thinking of FDA and the scientific community on the regulatory process for cartilage repair therapies. Cartilage repair therapies can receive market approval from FDA as medical devices, drugs, or biologics, and the specific classification of product can affect the nonclinical, clinical, and regulatory strategy to bring the product to market. Recent FDA guidance gives an outline of the required elements to bring a cartilage repair product to market, although these standards are often very general. As a result, companies have to carefully craft their study patient population, comparator group, and clinical endpoint to best showcase their product's attributes. In addition, regulatory strategy and manufacturing process validation need to be considered early in the clinical study process to allow for timely product approval following the completion of clinical study. Although the path to regulatory approval for a cartilage repair therapy is challenging and time-consuming, proper clinical trial planning and attention to the details can eventually save companies time and money by bringing a product to the market in the most expeditious process possible. PMID:26069647

  5. The bone-cartilage unit in osteoarthritis.

    Science.gov (United States)

    Lories, Rik J; Luyten, Frank P

    2011-01-01

    Osteoarthritis (OA) refers to a group of mechanically-induced joint disorders to which both genetic and acquired factors contribute. Current pathophysiological concepts focus on OA as a disease of the whole joint. Within these models, the functional unit formed by the articular cartilage and the subchondral bone seems to be of particular interest. Cartilage and bone receive and dissipate the stress associated with movement and loading, and are therefore continuously challenged biomechanically. Recent data support the view that cartilage and bone can communicate over the calcified tissue barrier; vessels reach out from bone into the cartilage zone, patches of uncalcified cartilage are in contact with bone, and microcracks and fissures further facilitate transfer of molecules. Several molecular signaling pathways such as bone morphogenetic proteins and Wnts are hypothesized to have a role in OA and can activate cellular and molecular processes in both cartilage and bone cells. In addition, intracellular activation of different kinase cascades seems to be involved in the molecular crosstalk between cartilage and bone cells. Further research is required to integrate these different elements into a comprehensive approach that will increase our understanding of the disease processes in OA, and that could lead to the development of specific therapeutics or treatment strategies. PMID:21135881

  6. Micropropagation of Origanum acutidens (HAND.-MAZZ. IETSWAART Using Stem Node Explants

    Directory of Open Access Journals (Sweden)

    Mehmet Ugur Yildirim

    2013-01-01

    Full Text Available Origanum acutidens (HAND.-MAZZ. IETSWAART is a promising ornamental plant that can be widely used in landscape management. It is endemic to Eastern Anatolian region of Turkey. Tissue culture has not been used to micropropagate it. The study reports stem node explants from one-week-old seedlings of the plant for successful micropropagation. The stem nodes were cultured on MS medium containing 0.6, 1.2, 1.8, and 2.4 mg/L BAP with 0.2 mg/L NAA. Visible effects of culture media on shoot proliferation were recorded. Shoot regeneration rate was maximum on MS medium containing 1.80 mg/L BAP-0.2 mg/L NAA. The micropropagated shoots were rooted on MS medium containing 0.2 mg/L NAA. All microrooted plantlets survived during acclimatisation on peat moss. It was concluded that O. acutidens can be successfully micropropagated under in vitro conditions.

  7. High frequency plant regeneration with histological analysis of organogenic callus from internode explants of Asteracantha longifolia Nees

    Directory of Open Access Journals (Sweden)

    Muthusamy Senthil Kumar

    2015-06-01

    Full Text Available Asteracantha longifolia Nees is an ayurvedic medicinal herb. The internode explants of this plant were used for high frequency plant regeneration on Murashige and Skoog (MS medium supplemented with various plant growth regulators (PGRs in different concentrations. Apical meristem and leaf primordium formations were confirmed through microscopic analysis of histological sections of the organogenic callus tissues. The synergistic effect of α-naphthaleneacetic acid (NAA 0.5 mg/l with N6 benzyladenine (BA 0.25 mg/l increased the percentage of explants response for callus induction while comparing other treatments. Various concentrations of NAA were also found to be best for explants response to callus induction than 2,4-dichlorophenoxyacetic acid (2,4-D. The callus morphology (color and texture was different according to the growth regulators and their concentrations. The highest percentage of response per culture for shoot bud regeneration was noted for the concentration of NAA 0.5 mg/l with BA 2.0 mg/l, the same concentration effectively increased the number of shoots per culture. Different concentrations of indol-3-butyric acid (IBA and NAA were used in half strength MS medium for in vitro rooting of regenerated shoots. The maximum percentage of shoot response for rooting and the highest number of root formations per shoot were observed on the medium containing 0.5 mg/l of IBA. The survival rate (86.7% of the regenerated plants was noted after 20 days of transplantation.

  8. Herpes simplex virus-1 infection of colonic explants as a model of viral-induced activation of Crohn's disease.

    Science.gov (United States)

    Silva, Manuel A; Menezes, José; Dionne, Serge; Levy, Emile; Amre, Devendra K; Seidman, Ernest G

    2012-05-01

    The exogenous triggers responsible for Crohn's disease (CD) relapses are not often identified. Cytomegalovirus and other members of the herpesvirus family have been implicated in precipitating relapses. However, the role of viral infections in the immunopathogenesis of CD remains poorly understood. We describe an ex-vivo model of primary viral infection of CD tissue with Herpes Simplex Virus type I (HSV-1). IL-6 and CD68 served as markers for CD inflammation, type I IFNs for viral infection. Colonic explants obtained from CD resections were infected via the luminal or the submucosal compartments with HSV-1 or mock virus solution, at varying concentrations for up to 20 h. Serial tissue sections were assayed for expression of HSV-1 specific antigens, CD-68, IL-6 and DC-SIGN. Culture supernatants were tested for IL-6 and type I IFN production. Positive immunostaining for HSV-1 specific antigens was consistently detectable using 11×10(6)PFU from 13 h onwards, mainly on cells located in the submucosa, and in the perivascular area. CD68 was up-regulated in lamina propria macrophages from mildly and non-inflamed CD tissue after HSV-1 infection. IL-6+ cells in the infected tissues were mainly submucosal DC-SIGN+ dendritic cells. IL-6 and IFN-β levels were higher in the supernatants from HSV-1-infected explants compared to controls after 20 h of culture (p<0.01). These data show increased expression of inflammatory markers during the initial stages of HSV-1 primary infection using CD colonic explants. This in vitro model appears promising to study the immunoregulatory changes induced by microbial infection in reactivation of CD. PMID:22398063

  9. Flower-bud formation in explants of photoperiodic and day-neutral Nicotiana biotypes and its bearing on the regulation of flower formation.

    OpenAIRE

    Rajeevan, M S; Lang, A.

    1993-01-01

    The capacity to form flower buds in thin-layer explants was studied in flowering plants of several species, cultivars, and lines of Nicotiana differing in their response to photoperiod. This capacity was found in all biotypes examined and could extend into sepals and corolla. It varied greatly, depending on genotype, source tissue and its developmental stage, and composition of the culture medium, particularly the levels of glucose, auxin, and cytokinin. It was greatest in the two day-neutral...

  10. In vitro propagation and genetic fidelity study of plant regenerated from inverted hypocotyl explants of eggplant (Solanum melongena L.) cv. Arka Shirish

    OpenAIRE

    Padma Mallaya, N.; Ravishankar, G.A

    2012-01-01

    Genetic variation due to somaclonal variation in micropropagated plants is a beneficial phenomenon for crop improvement. Genetic integrity of the plants derived through micropropagation becomes crucial if genetic transformation studies have to be carried out. Somaclonal variation in tissue culture is a common phenomenon which makes it mandatory to check for genetic stability of plants. Hypocotyl explants of Solanum melongena L. cv. Arka Shirish inoculated with inverted polarity in MS media su...

  11. Organ of Corti explants direct tonotopically graded morphology of spiral ganglion neurons in vitro.

    Science.gov (United States)

    Smith, Felicia L; Davis, Robin L

    2016-08-01

    The spiral ganglion is a compelling model system to examine how morphological form contributes to sensory function. While the ganglion is composed mainly of a single class of type I neurons that make simple one-to-one connections with inner hair cell sensory receptors, it has an elaborate overall morphological design. Specific features, such as soma size and axon outgrowth, are graded along the spiral contour of the cochlea. To begin to understand the interplay between different regulators of neuronal morphology, we cocultured neuron explants with peripheral target tissues removed from distinct cochlear locations. Interestingly, these "hair cell microisolates" were capable of both increasing and decreasing neuronal somata size, without adversely affecting survival. Moreover, axon characteristics elaborated de novo by the primary afferents in culture were systematically regulated by the sensory endorgan. Apparent peripheral nervous system (PNS)-like and central nervous system (CNS)-like axonal profiles were established in our cocultures allowing an analysis of putative PNS/CNS axon length ratios. As predicted from the in vivo organization, PNS-like axon bundles elaborated by apical cocultures were longer than their basal counterparts and this phenotype was methodically altered when neuron explants were cocultured with microisolates from disparate cochlear regions. Thus, location-dependent signals within the organ of Corti may set the "address" of neurons within the spiral ganglion, allowing them to elaborate the appropriate tonotopically associated morphological features in order to carry out their signaling function. J. Comp. Neurol. 524:2182-2207, 2016. © 2015 Wiley Periodicals, Inc. PMID:26663318

  12. [Surgical therapeutic possibilities of cartilage damage].

    Science.gov (United States)

    Burkart, A C; Schoettle, P B; Imhoff, A B

    2001-09-01

    Therapy of cartilage damage is a frequent problem, especially in the young and active patient. For the treatment of a cartilage damage we have to consider the size of the defect, age and weight of the patient, meniscal tears, ligament instabilities and varus-/valgus-malalignment. Lavage, shaving and debridement are only sufficient for a short time and have no long term effect. Abrasio and drilling could be useful in eldery people. Microfracturing seems to be an effective alternative for small defects. The restoration of the cartilage surface with the use of autologous chondrocyte transplantation, osteochondral autograft transplantation and posterior condyle transfer seems to be an adequate treatment for younger patients. PMID:11572120

  13. Inter-subject comparison of MRI knee cartilage thickness

    OpenAIRE

    Carballido-Gamio, Julio; Jan S. Bauer; Stahl, Robert; Lee, Keh-Yang; Krause, Stefanie; Link, Thomas M.; Majumdar, Sharmila

    2007-01-01

    In this paper, we present the development and application of current image processing techniques to perform MRI inter-subject comparison of knee cartilage thickness based on the registration of bone structures. Each point in the bone surface which is part of the bone–cartilage interface is assigned a cartilage thickness value. Cartilage and corresponding bone structures are segmented and their shapes interpolated to create isotropic voxels. Cartilage thicknesses are computed for each point in...

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

    Energy Technology Data Exchange (ETDEWEB)

    Simon, V., E-mail: viosimon@phys.ubbcluj.ro [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)

    2015-01-15

    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.

  15. Effect of explant density and volume of cultivation medium on in-vitro multiplication of blueberry (Vaccinium corymbosum L. varieties "Brigitta" and "Legacy"

    Directory of Open Access Journals (Sweden)

    Mario Rodríguez Beraud

    2015-03-01

    Full Text Available The objective of the investigation was to evaluate the in-vitro multiplication of two varieties of blueberry (Vaccinium corymbosum L., “Brigitta” and “Legacy” in response to five explants densities (5, 10, 15, 20 and 25 and four flask volumes (10, 20, 30 and 40 mL for cultivation. For both varieties the cultivation medium WPM (Woody Plant Medium was used. The experiment was completely randomized with 20 treatments and 12 repetitions per treatment. After 45 days of cultivation we evaluated the height of shoots, number of shoots/explant, number of nodes/shoot and number of shoots/flask. Variety “Brigitta” had highest shoots at higher densities and flask volumes, while variety “Legacy” had the highest average shoot height with intermediate densities and high volumes. Regarding the number of shoots/explant, the volume of the medium had no influence on “Brigitta”, however, higher plant densities affected this parameter. With variety “Legacy” the maximum number of shoots was achieved with lower plant densities and intermediate culture volumes per flask. In relation to the number of nodes per explant "Brigitta had lower numbers as compared to “Legacy”, but with both varieties the number of nodes decresed with smaller volumes of medium in the flasks. For the number of shoots per flask, “Brigitta” responsed best at higher densities exceeding 40 shoots per flask. In contrast, “Legacy” produced maximum results at density of 25 explants in 30 mL of medium. It is concluded that for the optimum multiplication of both varieties the correct selection of both, the planting density and the volume of multiplication medium are important.

  16. Biochemical and metabolic abnormalities in normal and osteoarthritic human articular cartilage

    Energy Technology Data Exchange (ETDEWEB)

    Ryu, J.; Treadwell, B.V.; Mankin, H.J.

    1984-01-01

    Incorporation of radioactive precursors into macromolecules was studied with human normal and osteoarthritic articular cartilage organ culture. Analysis of the salt extracted matrix components separated by cesium chloride buoyant density gradient centrifugation showed an increase in the specific activities of all gradient fractions prepared from the osteoarthritic cartilage. Further analysis of these fractions showed the osteoarthritic cartilage contained 5 times as much sulfate incorporated into proteoglycans, and an even greater amount of 3H-glucosamine incorporated into material sedimenting to the middle of the gradient. Greater than half of this radioactive middle fraction appears to be hyaluronate, as judged by the position it elutes from a DEAE column and its susceptibility to hyaluronidase digestion. This study supports earlier findings showing increased rates of macromolecular synthesis in osteoarthritis, and in addition, an even greater synthetic rate for hyaluronic acid is demonstrated.

  17. Controlled-Potential Electromechanical Reshaping of Cartilage.

    Science.gov (United States)

    Hunter, Bryan M; Kallick, Jeremy; Kissel, Jessica; Herzig, Maya; Manuel, Cyrus; Protsenko, Dmitri; Wong, Brian J F; Hill, Michael G

    2016-04-25

    An alternative to conventional "cut-and-sew" cartilage surgery, electromechanical reshaping (EMR) is a molecular-based modality in which an array of needle electrodes is inserted into cartilage held under mechanical deformation by a jig. Brief (ca. 2 min) application of an electrochemical potential at the water-oxidation limit results in permanent reshaping of the specimen. Highly sulfated glycosaminoglycans within the cartilage matrix provide structural rigidity to the tissue through extensive ionic-bonding networks; this matrix is highly permselective for cations. Our studies indicate that EMR results from electrochemical generation of localized, low-pH gradients within the tissue: fixed negative charges in the proteoglycan matrix are protonated, resulting in chemically induced stress relaxation of the tissue. Re-equilibration to physiological pH restores the fixed negative charges, and yields remodeled cartilage that retains a new shape approximated by the geometry of the reshaping jig. PMID:27059655

  18. Soursop contamination and sprouting in function of the fungicide in the culture medium

    OpenAIRE

    Renata Aparecida de Andrade; Amanda Garcia Bagatim; Samir Paulo Jasper

    2014-01-01

    The present research was conducted to verify the effectiveness of adding different doses of fungicide to the culture medium, in the contamination of in vitro culture of the explants of soursop. The experiment was conducted using explants collected from seedlings of soursop that were kept on lath house conditions (50% brightness), underwent three treatments, consisting in doses of fungicide carbendazim added to the MS culture medium: 5 ml per liter of culture medium, 10 ml/L, 15 mL /L, more...

  19. A Synthetic Thermosensitive Hydrogel for Cartilage Bioprinting and Its Biofunctionalization with Polysaccharides.

    Science.gov (United States)

    Abbadessa, Anna; Mouser, Vivian H M; Blokzijl, Maarten M; Gawlitta, Debby; Dhert, Wouter J A; Hennink, Wim E; Malda, Jos; Vermonden, Tina

    2016-06-13

    Hydrogels based on triblock copolymers of polyethylene glycol and partially methacrylated poly[N-(2-hydroxypropyl) methacrylamide mono/dilactate] make up an attractive class of biomaterials because of their biodegradability, cytocompatibility, and tunable thermoresponsive and mechanical properties. If these properties are fine-tuned, the hydrogels can be three-dimensionally bioprinted, to generate, for instance, constructs for cartilage repair. This study investigated whether hydrogels based on the polymer mentioned above with a 10% degree of methacrylation (M10P10) support cartilage formation by chondrocytes and whether the incorporation of methacrylated chondroitin sulfate (CSMA) or methacrylated hyaluronic acid (HAMA) can improve the mechanical properties, long-term stability, and printability. Chondrocyte-laden M10P10 hydrogels were cultured for 42 days to evaluate chondrogenesis. M10P10 hydrogels with or without polysaccharides were evaluated for their mechanical properties (before and after UV photo-cross-linking), degradation kinetics, and printability. Extensive cartilage matrix production occurred in M10P10 hydrogels, highlighting their potential for cartilage repair strategies. The incorporation of polysaccharides increased the storage modulus of polymer mixtures and decreased the degradation kinetics in cross-linked hydrogels. Addition of HAMA to M10P10 hydrogels improved printability and resulted in three-dimensional constructs with excellent cell viability. Hence, this novel combination of M10P10 with HAMA forms an interesting class of hydrogels for cartilage bioprinting. PMID:27171342

  20. The structure and function of cartilage proteoglycans

    Directory of Open Access Journals (Sweden)

    P J Roughley

    2006-11-01

    Full Text Available Cartilage contains a variety of proteoglycans that are essential for its normal function. These include aggrecan, decorin, biglycan, fibromodulin and lumican. Each proteoglycan serves several functions that are determined by both its core protein and its glycosaminoglycan chains. This review discusses the structure/function relationships of the cartilage proteoglycans, and the manner in which perturbations in proteoglycan structure or abundance can adversely affect tissue function.

  1. Fibrin for tissue engineering of cartilage

    OpenAIRE

    Eyrich, Daniela

    2006-01-01

    Since the beginning of the 1990s a plethora of research approaches towards cartilage engineering for plastic and reconstructive surgery have been undertaken. However, a general standard method for generation of cartilage tissue equivalent is still lacking. The goal of this thesis is based on the project �Bavarian Research Cooperation for Tissue Engineering and Rapid Prototyping� (ForTEPro) for development of individually customized implants for facial and reconstructive surgery. The main o...

  2. Materials science: Like cartilage, but simpler

    DEFF Research Database (Denmark)

    Skov, Anne Ladegaard

    2015-01-01

    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....

  3. Magnetic resonance imaging of articular cartilage at 3 tesla

    International Nuclear Information System (INIS)

    Smooth motor function can be maintained by articular cartilage. When the cartilage is injured, edema occurs, and as degeneration progresses, the cartilage thins and the cartilage matrix decreases. Magnetic resonance (MR) imaging allows noninvasive evaluation of these changes. Fat suppression proton density- and T2-weighted imaging are useful in the morphologic evaluation of articular cartilage. High resolution, 3-tesla MR imaging provides more detailed evaluation. Biochemical information from T2 mapping, T1ρ mapping, and delayed gadolinium-enhanced MR imaging of cartilage (dGEMRIC) is useful for early diagnosis of cartilage injury and evaluation of cartilage repair. The role of MR imaging in evaluating articular cartilage will increase in the future aging society. (author)

  4. Intra-articular injection of synovium-derived mesenchymal stem cells and hyaluronic acid promote regeneration of massive cartilage defects in rabbits

    Directory of Open Access Journals (Sweden)

    Vyacheslav Ogay

    2014-01-01

    Full Text Available Introduction: The purpose of this study was to investigate whether intra-articular injection of synovium-derived mesenchymal stem cells (SD MSCs with low molecular weight hyaluronic acid (HA could promote regeneration of massive cartilage in rabbits. Material and methods: The SD MSCs were harvested from the knees of 10 Flemish giant rabbits, expanded in culture, and characterized. A reproducible 4-mm cylindrical defect was created in the intercondylar groove area using a kit for the mosaic chondroplasty of femoral condyle COR (De Puy, Mitek. The defect was made within the cartilage layer without destruction of subchondral bone. Two weeks after the cartilage defect, SD MSCs (2 × 106 cell/0.15 ml were suspended in 0.5% low molecular weight HA (0.15 ml and injected into the left knee, and HA solution (0.30 ml alone was placed into the right knee. Cartilage regeneration in the experimental and control groups were evaluated by macroscopically and histologically at 10, 30, and 60 days. Results: On day 10, after intra-articular injection of SD MSCs, we observed an early process of cartilage regeneration in the defect area. Histological studies revealed that cartilage defect was covered by a thin layer of spindle-shaped undifferentiated cells and proliferated chodroblasts. In contrast, an injection of HA did not induce reparation of cartilage in the defect area. At 30 days, macroscopic observation showed that the size of cartilage defect after SD MSC injection was significantly smaller than after HA injection. Histological score was also better in the MSC- treated intercondylar defect. At 60 days after MSC treatment, cartilage defect was nearly nonexistent and looked similar to an intact cartilage. Conclusion: Thus, intra-articular injection of SD MSCs can adhere to the defect in the intercondylar area, and promote cartilage regeneration in rabbits.

  5. In vitro cartilage production using an extracellular matrix-derived scaffold and bone marrow-derived mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    ZHAO Yan-hong; YANG Qiang; XIA Qun; PENG Jiang; LU Shi-bi; GUO Quan-yi; MA Xin-long

    2013-01-01

    Background Cartilage repair is a challenging research area because of the limited healing capacity of adult articular cartilage.We had previously developed a natural,human cartilage extracellular matrix (ECM)-derived scaffold for in vivo cartilage tissue engineering in nude mice.However,before these scaffolds can be used in clinical applications in vivo,the in vitro effects should be further explored.Methods We produced cartilage in vitro using a natural cartilage ECM-derived scaffold.The scaffolds were fabricated by combining a decellularization procedure with a freeze-drying technique and were characterized by scanning electron microscopy (SEM),micro-computed tomography (micro-CT),histological staining,cytotoxicity assay,biochemical and biomechanical analysis.After being chondrogenically induced,the induction results of BMSCs were analyzed by histology and Immunohisto-chemistry.The attachment and viability assessment of the cells on scaffolds were analyzed using SEM and LIVE/DEAD staining.Cell-scaffold constructs cultured in vitro for 1 week and 3 weeks were analyzed using histological and immunohistochemical methods.Results SEM and micro-CT revealed a 3-D interconnected porous structure.The majority of the cartilage ECM was found in the scaffold following the removal of cellular debris,and stained positive for safranin O and collagen Ⅱ.Viability staining indicated no cytotoxic effects of the scaffold.Biochemical analysis showed that collagen content was (708.2±44.7)μg/mg,with GAG (254.7±25.9) μg/mg.Mechanical testing showed the compression moduli (E) were (1.226±0.288) and (0.052±0.007) MPa in dry and wet conditions,respectively.Isolated canine bone marrow-derived stem cells (BMSCs) were induced down a chondrogenic pathway,labeled with PKH26,and seeded onto the scaffold.Immunofluorescent staining of the cell-scaffold constructs indicated that chondrocyte-like cells were derived from seeded BMSCs and excreted ECM.The cell-scaffold constructs contained

  6. 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, T.N.de; Groot, J.de; Polak, A.A.; Lafeber, F.P.J.G.

    2012-01-01

    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 propertie

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

    Directory of Open Access Journals (Sweden)

    Jin Suk Byun, MD, PhD

    2013-09-01

    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.

  8. High-frequency in vitro plantlet regeneration from apical bud as a novel explant of Carum copticum L.

    Institute of Scientific and Technical Information of China (English)

    Mansoureh Salehi; Bahman Hosseini; Zohreh Jabbarzadeh

    2014-01-01

    Objective: To develop an in vitro regeneration system to increase the recovery of Carum copticum L. plantlets as a part of developing a metabolic engineering program.Methods:3-acetic acid and indole butyric acid on direct shoot regeneration and rooting of ajowan from apical bud explants were assessed. All explants were cultured on Murashige and Skoog (MS) medium supplemented with different combinations of 6-benzyl amino purine (BAP) (0, 2.2, 4.4, 8.8µ The efficacy of different concentrations and combinations of 6-benzyladenine, indole-Results: The maximum shoot regeneration frequency (97.5%) and the highest number of shoots produced from apical buds (34 shoots per explant) were obtained on MS medium fortified with BAP (4.4 µmol/L) and IAA (0.5 µmol/L). Low shoot regeneration frequency was observed in BAP free treatments. The effects of different strengths of MS medium and various concentrations of IAA and indole-3- butyric acid on rooting rate, length and average number of roots were also investigated. Application of indole-3- butyric acid (6 µmol/L) in full-strength MS medium, was more effective than IAA and resulted in highest shoot regeneration frequency with the rooting rate of 100% and highest mean number of roots per shoot (41.8). The rooted plantlets were acclimatized successfully in greenhouse conditions with a survival rate of 90%. mol/L) and indole-3-acetic acid (IAA) (0, 0.5, 1.1, 2.2 µmol/L). Conclusion: In this study, a simple and reliable regeneration and acclimatization protocol for Carum copticum has been presented. This protocol can be found very advantageous for a variety of purposes, including mass multiplication of Carum species, medicinal plant breeding studies and transgenic plant production.

  9. IL-1 beta and TGF-beta 1 modulate the sulphation grade of chondro-disaccharides in porcine articular cartilage: a capillary electrophoresis study.

    Science.gov (United States)

    Zanni, M; Tamburro, A; Rotilio, D

    1995-07-01

    This report describes the effect of interleukin-1 beta (IL-1 beta) and transforming growth factor-beta 1 (TGF-beta 1) on proteoglycan release from cartilage explants and modification at the sulphation level. Matrix proteoglycans purified by ion-exchange chromatography were composed of two distinct peaks (1 and 2) each showing a different Kav value when they were subjected to size-exclusion chromatography on a Sepharose CL-2B column. Glycosaminoglycans (GAGs) of conditioned medium and extracellular matrix proteoglycans were digested by chondroitin ABC and AC lyase, suggesting that chondroitin sulphate (CS) is the major GAG present (80-90%). Structural analysis of disaccharides, by capillary zone electrophoresis, revealed a different pattern of sulphated glycosaminoglycans when cartilage was treated with either IL-1 beta or TGF-beta 1. Analysis of GAGs released into the medium from TGF-beta 1 treated cartilage showed a reduction in the level of 4-S-disaccharide (delta Di4S) and an increase in non-sulphated disaccharides (delta Di0S), while no significant changes were found in IL-1 beta treated cartilage. In the extracellular matrix, IL-1 beta and TGF-beta 1 induced a more complex rearrangement of the GAGs. The level of non-sulphated disaccharides was increased whereas that of total sulphated disaccharides was reduced. Taken together, these results suggest that both cytokines modify the structure of GAGs, probably by interfering with the activity or the synthesis of sulphotransferases involved in GAG turnover. PMID:7551687

  10. Corneal recovery in a rabbit limbal stem cell deficiency model by autologous grafts of tertiary outgrowths from cultivated limbal biopsy explants

    Science.gov (United States)

    Durak, Ismet; Gürdal, Mehmet; Baysal, Kemal; Ates, Halil; Ozbek, Zeynep; Wang, Zheng; Wu, Albert; Wolosin, J. Mario

    2016-01-01

    Purpose To determine the corneal regenerative capacity of sequentially generated primary, secondary, and tertiary limbal explant outgrowths in a limbal stem cell deficiency (LSCD) surgical model. Methods Two-millimeter-long limbal shallow biopsies were surgically excised from the upper quadrant of the right eye of rabbits and set on preserved amniotic membrane for explant culture. After the generation of primary outgrowth, the biopsies were sequentially transferred to new amniotic membrane to generate secondary and then tertiary outgrowths. Eighteen rabbits were subjected to a 360° limbal peritomy extending into the scleral zone and combined with superficial keratectomy of the corneal periphery and thorough mechanical debridement of the central cornea in their left eye. Right eye outgrowths, six of each generation, were engrafted on the ocular surface. Clinical outcomes (neovascularization, corneal clarity, and corneal fluorescein staining) were graded after 6 months. Post-mortem corneas were compared with histology, immunochemistry for p63 and Krt3, ABCG2-dependent dye exclusion, and capacity for outgrowths in explant culture. Results Immunohistology and western blot of the outgrowths for p63 and Krt3 indicated no differences in expression between the primary and tertiary outgrowths for these two markers of growth and differentiation. Clinically, all rabbits treated with amniotic membrane alone developed severe LSCD. Most rabbits grafted with cell outgrowths from all three outgrowth generations achieved stable (>6 months) recovery of the ocular surface. There were partial failures of grafts performed with two secondary and tertiary outgrowths. However, Kruskal–Wallis statistical analysis of the clinical scores yielded no significant difference between the three groups (p=0.524). Histology showed full anatomic recovery of grafts made with primary and tertiary outgrowths. Krt3 and p63 expression throughout the whole limbal corneal epithelium with primary or

  11. Nonparenchymal cells cultivated from explants of fibrotic liver resemble endothelial and smooth muscle cells from blood vessel walls

    International Nuclear Information System (INIS)

    Tissue specimens from human fibrotic liver obtained by needle biopsy were cultured. Two cell types emerged from the tissue explants. From their morphology and biosynthetic products they resembled smooth muscle cells and endothelial cells from blood vessel walls. In the endothelial cells, factor VIII-associated protein was demonstrated by indirect immunofluorescence. Synthesis of collagen types I and III, basement membrane collagen types IV and V, and fibronectin by both cell types was observed by immunofluorescence microscopy. Homogeneous cultures of smooth muscle cells were observed in subcultures. After incubation with [14C]glycine, collagen was isolated and characterized by CM cellulose chromatography, and consisted mainly of types I and III. These data suggest involvement of mesenchymal cells in hepatic fibrosis; they presumably originate from blood vessel or sinusoidal walls

  12. Thidiazuron induces shoot organogenesis at low concentrations and somatic embryogenesis at high concentrations on leaf and petiole explants of African violet (Saintpaulia ionantha Wendl).

    Science.gov (United States)

    Mithila, J; Hall, J C; Victor, J M R; Saxena, P K

    2003-01-01

    Regeneration via shoot organogenesis and somatic embryogenesis was observed from thidiazuron (TDZ)-treated leaf and petiole explants of greenhouse- and in vitro-grown African violet plants. The response of cultures to other growth regulators over a range of 0.5 microM to 10 microM was 50% less than that observed with TDZ. A comparative study among several cultivars of African violet indicated that "Benjamin" and "William" had the highest regeneration potential. In "Benjamin", higher frequencies of shoot organogenesis (twofold) and somatic embryogenesis (a 50% increase) were observed from in vitro- and greenhouse-grown plants, respectively. At concentrations lower than 2.5 microM, TDZ induced shoot organogenesis, whereas at higher doses (5-10 microM) somatic embryos were formed. These findings provide the first report of simultaneous shoot organogenesis and somatic embryogenesis of African violet explants in response to TDZ. PMID:12789442

  13. Plant tissue culture

    International Nuclear Information System (INIS)

    Plant tissue culture refers to growing and multiplication of cells, tissues and organs of plants on defined solid or liquid media under aseptic and controlled environment. The commercial technology is primarily based on micropropagation, in which rapid proliferation is achieved from tiny stem cuttings, axillary buds, and to a limited extent from somatic embryos, cell clumps in suspension cultures and bioreactors. The cultured cells and tissue can take several pathways. The pathways that lead to the production of true-to-type plants in large numbers are the preferred ones for commercial multiplication. The process of micropropagation is usually divided into several stages i.e., pre-propagation, initiation of explants, subculture of explants for proliferation, shooting and rooting, and hardening. These stages are universally applicable in large-scale multiplication of plants. The delivery of hardened small micropropagated plants to growers and market also requires extra care. (author)

  14. Promotion of the articular cartilage proteoglycan degradation by T-2 toxin and selenium protective effect

    Institute of Scientific and Technical Information of China (English)

    Si-yuan LI; Jun-ling CAO; Zhong-li SHI; Jing-hong CHEN; Zeng-tie ZHANG; Clare E. HUGHES; Bruce CATERSON

    2008-01-01

    Objective: To identify the relationship between T-2 toxin and Kashin-Beck disease (KBD), the effects of T-2 toxin on aggrecan metabolism in human chondrocytes and cartilage were investigated in vitro. Methods: Chondrocytes were isolated from human articular cartilage and cultured in vitro. Hyaluronic acid (HA), soluble CD44 (sCD44), IL-1β and TNF-α levels in supernatants were measured by enzyme-linked immunosorbent assay (ELISA). CD44 content in chondrocyte membrane was determined by flow cytometry (FCM). CD44, hyaluronic acid synthetase-2 (HAS-2) and aggrecanases mRNA levels in chondrocytes were determined using reverse transcription polymerase chain reaction (RT-PCR). Immunocytochemical method was used to investigate expressions of BC-13, 3-B-3(-) and 2-B-6 epitopes in the cartilage reconstructed in vitro. Results: T-2 toxin inhibited CD44, HAS-2, and aggrecan mRNA expressions, but promoted aggrecanase-2 mRNA expression. Meanwhile, CD44 expression was found to be the lowest in the chondrocytes cultured with T-2 toxin and the highest in control plus selenium group. In addition,ELISA results indicated that there were higher sCD44, IL-1β and TNF-α levels in T-2 toxin group. Similarly, higher HA levels were also observed in T-2 toxin group using radioimmunoprecipitation assay (RIPA). Furthermore, using monoclonal antibodies BC-13, 3-B-3 and 2-B-6, strong positive immunostaining was found in the reconstructed cartilage cultured with T-2 toxin, whereas no positive staining or very weak staining was observed in the cartilage cultured without T-2 toxin. Selenium could partly inhibit the effects of T-2 toxin above. Conclusion: T-2 toxin could inhibit aggrecan synthesis, promote aggrecanases and pro-inflammatory cytokines production, and consequently induce aggrecan degradation in chondrocytes. These will perturb metabolism balance between aggrecan synthesis and degradation in cartilage, inducing aggrecan loss in the end, which may be the initiation of the cartilage

  15. Aggrecan structure in amphibian cartilage

    Directory of Open Access Journals (Sweden)

    Covizi D.Z.

    2000-01-01

    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.

  16. Streamlined bioreactor-based production of human cartilage tissues.

    Science.gov (United States)

    Tonnarelli, B; Santoro, R; Adelaide Asnaghi, M; Wendt, D

    2016-01-01

    Engineered tissue grafts have been manufactured using methods based predominantly on traditional labour-intensive manual benchtop techniques. These methods impart significant regulatory and economic challenges, hindering the successful translation of engineered tissue products to the clinic. Alternatively, bioreactor-based production systems have the potential to overcome such limitations. In this work, we present an innovative manufacturing approach to engineer cartilage tissue within a single bioreactor system, starting from freshly isolated human primary chondrocytes, through the generation of cartilaginous tissue grafts. The limited number of primary chondrocytes that can be isolated from a small clinically-sized cartilage biopsy could be seeded and extensively expanded directly within a 3D scaffold in our perfusion bioreactor (5.4 ± 0.9 doublings in 2 weeks), bypassing conventional 2D expansion in flasks. Chondrocytes expanded in 3D scaffolds better maintained a chondrogenic phenotype than chondrocytes expanded on plastic flasks (collagen type II mRNA, 18-fold; Sox-9, 11-fold). After this "3D expansion" phase, bioreactor culture conditions were changed to subsequently support chondrogenic differentiation for two weeks. Engineered tissues based on 3D-expanded chondrocytes were more cartilaginous than tissues generated from chondrocytes previously expanded in flasks. We then demonstrated that this streamlined bioreactor-based process could be adapted to effectively generate up-scaled cartilage grafts in a size with clinical relevance (50 mm diameter). Streamlined and robust tissue engineering processes, as the one described here, may be key for the future manufacturing of grafts for clinical applications, as they facilitate the establishment of compact and closed bioreactor-based production systems, with minimal automation requirements, lower operating costs, and increased compliance to regulatory guidelines. PMID:27232665

  17. ANATOMICAL STUDIES OF IN VITRO ORGANOGENESIS INDUCED IN LEAF-DERIVED EXPLANTS OF PASSIONFRUIT ESTUDOS ANATÔMICOS DA ORGANOGÊNESE IN VITRO INDUZIDA EM EXPLANTES DE FOLHA DE MARACUJÁ

    Directory of Open Access Journals (Sweden)

    BEATRIZ APPEZZATO DA GLORIA

    1999-11-01

    Full Text Available With the aim of studying the organogenesis in vitro in Passiflora edulis Sims f. flavicarpa Deg., the passionfruit, leaf-derived explants were cultured on media containing NAA or BAP and incubated either in continuous darkness or in light. The histological events leading to de novo organ formation were evaluated. Darkness induces rhizogenesis in the presence of NAA, whereas direct shoot regeneration is stimulated by light and BAP. This latter condition is recommended for passionfruit micropropagation as several adventitious shoot buds were formed from meristemoids of parenchymal origin.Com o objetivo de estudar a organogênese in vitro em Passiflora edulis Sims f. flavicarpa Deg., o maracujá-amarelo, explantes derivados de folha foram cultivados em meio contendo NAA ou BAP, no escuro e na presença de luz. Foram descritos os eventos histológicos que levam à formação de novo de órgãos. Concluiu-se que o escuro induz a rizogênese, na presença de NAA, enquanto a regeneração de brotos é estimulada pela luz e BAP. Esta condição é recomendada para micropropagar o maracujá uma vez que vários brotos adventícios são formados a partir de meristemóides de origem parenquimática.

  18. In vitro precultivation alleviates post-implantation inflammation and enhances development of tissue-engineered tubular cartilage

    Energy Technology Data Exchange (ETDEWEB)

    Luo Xusong; Zhou Guangdong; Liu Wei; Zhang Wenjie; Cui Lei; Cao Yilin [Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai 9th People' s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200011 (China); Cen Lian, E-mail: guangdongzhou@126.co, E-mail: yilincao@yahoo.co [National Tissue Engineering Center of China, Shanghai 200011 (China)

    2009-04-15

    Tissue-engineered tubular cartilage is a promising graft for tracheal reconstruction. But polylactic acid/polyglycolic acid (PLA/PGA) fibers, the frequently used scaffolds for cartilage engineering, often elicit an obvious inflammation response following implantation into immunocompetent animals. We propose that the inflammation could be alleviated by in vitro precultivation. In this study, after in vitro culture for either 2 days (direct implantation group (DI)) or for 2 weeks (precultivation implantation group (PI)), autologous tubular chondrocyte-PLA/PGA constructs were subcutaneously implanted into rabbits. In the PI group, after 2 weeks of precultivation, most of the fibers were found to be completely embedded in an extracellular matrix (ECM) produced by the chondrocytes. Importantly, no obvious inflammatory reaction was observed after in vivo implantation and homogeneous cartilage-like tissue was formed with biomechanical properties close to native tracheal cartilage at 4 weeks post-implantation. In the DI group, however, an obvious inflammatory reaction was observed within and around the cell-scaffold constructs at 1 week implantation and only sporadic cartilage islands separated by fibrous tissue were observed at 4 weeks. These results demonstrated that the post-implantation inflammatory reaction could be alleviated by in vitro precultivation, which contributes to the formation of satisfactory tubular cartilage for tracheal reconstruction.

  19. In vitro precultivation alleviates post-implantation inflammation and enhances development of tissue-engineered tubular cartilage

    International Nuclear Information System (INIS)

    Tissue-engineered tubular cartilage is a promising graft for tracheal reconstruction. But polylactic acid/polyglycolic acid (PLA/PGA) fibers, the frequently used scaffolds for cartilage engineering, often elicit an obvious inflammation response following implantation into immunocompetent animals. We propose that the inflammation could be alleviated by in vitro precultivation. In this study, after in vitro culture for either 2 days (direct implantation group (DI)) or for 2 weeks (precultivation implantation group (PI)), autologous tubular chondrocyte-PLA/PGA constructs were subcutaneously implanted into rabbits. In the PI group, after 2 weeks of precultivation, most of the fibers were found to be completely embedded in an extracellular matrix (ECM) produced by the chondrocytes. Importantly, no obvious inflammatory reaction was observed after in vivo implantation and homogeneous cartilage-like tissue was formed with biomechanical properties close to native tracheal cartilage at 4 weeks post-implantation. In the DI group, however, an obvious inflammatory reaction was observed within and around the cell-scaffold constructs at 1 week implantation and only sporadic cartilage islands separated by fibrous tissue were observed at 4 weeks. These results demonstrated that the post-implantation inflammatory reaction could be alleviated by in vitro precultivation, which contributes to the formation of satisfactory tubular cartilage for tracheal reconstruction.

  20. Articular cartilage: from formation to tissue engineering.

    Science.gov (United States)

    Camarero-Espinosa, Sandra; Rothen-Rutishauser, Barbara; Foster, E Johan; Weder, Christoph

    2016-05-26

    Hyaline cartilage is the nonlinear, inhomogeneous, anisotropic, poro-viscoelastic connective tissue that serves as friction-reducing and load-bearing cushion in synovial joints and is vital for mammalian skeletal movements. Due to its avascular nature, low cell density, low proliferative activity and the tendency of chondrocytes to de-differentiate, cartilage cannot regenerate after injury, wear and tear, or degeneration through common diseases such as osteoarthritis. Therefore severe damage usually requires surgical intervention. Current clinical strategies to generate new tissue include debridement, microfracture, autologous chondrocyte transplantation, and mosaicplasty. While articular cartilage was predicted to be one of the first tissues to be successfully engineered, it proved to be challenging to reproduce the complex architecture and biomechanical properties of the native tissue. Despite significant research efforts, only a limited number of studies have evolved up to the clinical trial stage. This review article summarizes the current state of cartilage tissue engineering in the context of relevant biological aspects, such as the formation and growth of hyaline cartilage, its composition, structure and biomechanical properties. Special attention is given to materials development, scaffold designs, fabrication methods, and template-cell interactions, which are of great importance to the structure and functionality of the engineered tissue. PMID:26923076

  1. Projection Stereolithographic Fabrication of Human Adipose Stem Cell-incorporated Biodegradable Scaffolds for Cartilage Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Aaron X Sun

    2015-08-01

    Full Text Available Poor self-healing ability of cartilage necessitates the development of methods for cartilage regeneration. Scaffold construction with live stem cell incorporation and subsequent differentiation presents a promising route. Projection stereolithography (PSL offers high resolution and processing speed as well as the ability to fabricate scaffolds that precisely fit the anatomy of cartilage defects using medical imaging as the design template. We report here the use of a visible-light based PSL (VL-PSL system to encapsulate human adipose-derived stem cells (hASCs into a biodegradable polymer (poly-D,L-lactic acid/polyethylene glycol/ poly-D,L-lactic acid (PDLLA-PEG/hyaluronic acid (HA matrix to produce live cell constructs with customized architectures. After fabrication, hASCs showed high viability (84% and were uniformly distributed throughout the constructs, which possessed high mechanical property with a compressive modulus of 780 kPa. The hASC-seeded constructs were then cultured in Control or TGF-β3-containing chondrogenic medium for up to 28 days. In chondrogenic medium treated group (TGF-β3 group hASCs maintained 77% viability and expressed chondrogenic genes Sox9, collagen type II, and aggrecan at 11, 232, and 2.29 x 10(5 fold increases, respectively, compared to levels at day 0 in non-chondrogenic medium. The TGF-β3 group also produced a collagen type II and glycosaminoglycan (GAG-rich extracellular matrix, detected by immunohistochemistry, and Alcian blue and Safranin O staining suggesting robust chondrogenesis within the scaffold. Without chondroinductive addition (Control group, cell viability decreased with time (65% at 28 days and showed poor cartilage matrix deposition. After 28 days, mechanical strength of the TGF-β3 group remained high at 240 kPa. Thus, the PSL- and PLLA-PEG/HA based fabrication method using adult stem cells is a promising approach in producing mechanically competent engineered cartilage for joint cartilage

  2. Cartilage regeneration using a porous scaffold, a collagen sponge incorporating a hydroxyapatite/chondroitinsulfate composite

    Energy Technology Data Exchange (ETDEWEB)

    Ohyabu, Yohimi, E-mail: ooyabu.yoshimi@aist.go.jp [Department of Metallurgy and Ceramics Science, Tokyo Institute of Technology, 2-12-1, S7-5 Ookayama, Meguro, Tokyo 152-8550 (Japan); Nanotechnology Research Institute (NRI), National Institute of Advanced Industrial Science and Technology (AIST), Central-4, 1-1-1 Higashi, Tsukuba, Ibaraki, 305-8566 (Japan); Adegawa, Takuro; Yoshioka, Tomohiko [Department of Metallurgy and Ceramics Science, Tokyo Institute of Technology, 2-12-1, S7-5 Ookayama, Meguro, Tokyo 152-8550 (Japan); Ikoma, Toshiyuki [Biomaterials Center, National Institute for Materials Science, 1-1 Sengen, Tsukuba, Ibaraki, 305-0047 (Japan); Uemura, Toshimasa [Nanotechnology Research Institute (NRI), National Institute of Advanced Industrial Science and Technology (AIST), Central-4, 1-1-1 Higashi, Tsukuba, Ibaraki, 305-8566 (Japan); Tanaka, Junzo [Department of Metallurgy and Ceramics Science, Tokyo Institute of Technology, 2-12-1, S7-5 Ookayama, Meguro, Tokyo 152-8550 (Japan)

    2010-10-15

    Because cartilage has limited potential for self-repair, tissue engineering is expected to replace the present therapies for damaged cartilage, such as total knee arthroplasty. However, scaffolds suitable for cartilage tissue engineering have not been established. We synthesized a novel porous scaffold, a collagen sponge incorporating a hydroxyapatite/chondroitinsulfate composite (pCol-HAp/ChS), containing materials which resemble extracellular matrices in bone and cartilage tissues, which needs high compressive strength for clinical use. HAp/ChS had smaller crystals and a larger total surface area than HAp. SEM images showed pCol-HAp/ChS to have the roughest surface compared with pCol and pCol-HAp. The mechanical properties suggest that pCol-HAp/ChS and pCol/HAp are similar, and superior to pCol. Seeding experiments showed a uniform distribution of mesenchymal stem cells (MSCs) in pCol-HAp/ChS and pCol/HAp. Safranin O, Toluidine blue and Alcian blue staining after 2 weeks of culture revealed pCol-HAp/ChS to be the most chondrogenic in each case. In addition, MSCs in pCol-HAp/ChS produced more glycosaminoglycans, a cartilage matrix, than those in pCol-HAp. Further, pCol-HAp/ChS regenerated 15 times more cartilaginous tissue than pCol. From these results, pCol-HAp/ChS is expected to be a candidate for a scaffold for cartilage tissue engineering in place of collagen sponge.

  3. Cartilage regeneration using a porous scaffold, a collagen sponge incorporating a hydroxyapatite/chondroitinsulfate composite

    International Nuclear Information System (INIS)

    Because cartilage has limited potential for self-repair, tissue engineering is expected to replace the present therapies for damaged cartilage, such as total knee arthroplasty. However, scaffolds suitable for cartilage tissue engineering have not been established. We synthesized a novel porous scaffold, a collagen sponge incorporating a hydroxyapatite/chondroitinsulfate composite (pCol-HAp/ChS), containing materials which resemble extracellular matrices in bone and cartilage tissues, which needs high compressive strength for clinical use. HAp/ChS had smaller crystals and a larger total surface area than HAp. SEM images showed pCol-HAp/ChS to have the roughest surface compared with pCol and pCol-HAp. The mechanical properties suggest that pCol-HAp/ChS and pCol/HAp are similar, and superior to pCol. Seeding experiments showed a uniform distribution of mesenchymal stem cells (MSCs) in pCol-HAp/ChS and pCol/HAp. Safranin O, Toluidine blue and Alcian blue staining after 2 weeks of culture revealed pCol-HAp/ChS to be the most chondrogenic in each case. In addition, MSCs in pCol-HAp/ChS produced more glycosaminoglycans, a cartilage matrix, than those in pCol-HAp. Further, pCol-HAp/ChS regenerated 15 times more cartilaginous tissue than pCol. From these results, pCol-HAp/ChS is expected to be a candidate for a scaffold for cartilage tissue engineering in place of collagen sponge.

  4. In vitro morphogenic response of different explants of Gentiana kurroo Royle from Western Himalayas-an endangered medicinal plant.

    Science.gov (United States)

    Sharma, Anshu; Kaur, Rajinder; Sharma, Neha

    2014-04-01

    Micropropagation offers a great potential to produce millions of clonal individuals through tissue culture via induction of morphogenesis. The aim of this work was to obtain an efficient protocol for callus regeneration for Gentiana kurroo Royle. The morphogenic response of different explants (leaves, petioles, roots) varied and responded differently for regeneration according to combinations of growth regulators. The petiole explants were best responding for callus induction and subsequently for indirect and direct regeneration. The callus induction was achieved on MS basal + 1.0 mg/l benzyladenine (BA) and 3.00 mg/l naphthalene acetic acid (NAA). MS medium supplemented with 0.10 mg/l NAA and 1.0 mg/l thidiazuron (TDZ) was recorded as the best medium for indirect regeneration. However, for direct regeneration the maximum number of shoot emergence was observed on MS basal fortified with 0.10 mg/l NAA + 0.75 mg/l TDZ. Half strength MS basal supplemented with indole-3-butyric acid (IBA) 1.00 mg/l gave best response for root induction. Subsequently, the plantlets were transferred and 100 % survival rate was recorded only on autoclaved cocopeat. No morphological variations were recorded in the callus regenerated plantlets. PMID:24757329

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

    Science.gov (United States)

    Akkiraju, Hemanth; Nohe, Anja

    2016-01-01

    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. PMID:27347486

  6. Effect of estrogen and dietary loading on rat condylar cartilage

    OpenAIRE

    Orajärvi, M. (Marko)

    2015-01-01

    Abstract The temporomandibular joint (TMJ) is a synovial joint which attaches the mandible to the skull. The head of the mandibular condyle is covered by condylar cartilage, which functions as both growth and articular cartilage. Masticatory forces are transmitted to the condylar cartilage, and the consistency of a person’s diet partly defines the loading force. Condylar cartilage acts as a load-absorbing structure together with the articular disc. Temporomandibular disorders (TMDs) are...

  7. Engineering articular cartilage using newly developed carrageenan basedhydrogels

    OpenAIRE

    Popa, Elena Geta

    2014-01-01

    Articular cartilage holds specific functionality in the human body creating smooth gliding areas and allowing the joints to move easily without pain. However, due to its avascular nature and to the low metabolic activity of the constituent cells-the chondrocytes, cartilage has a low regenerative potential. The current surgical options to treat damaged cartilage are not long lasting and involve frequent revisions. Tissue engineering may provide an alternative approach for cartilage...

  8. Type III Collagen, a Fibril Network Modifier in Articular Cartilage*

    OpenAIRE

    Wu, Jiann-Jiu; Weis, Mary Ann; Kim, Lammy S.; Eyre, David R.

    2010-01-01

    The collagen framework of hyaline cartilages, including articular cartilage, consists largely of type II collagen that matures from a cross-linked heteropolymeric fibril template of types II, IX, and XI collagens. In the articular cartilages of adult joints, type III collagen makes an appearance in varying amounts superimposed on the original collagen fibril network. In a study to understand better the structural role of type III collagen in cartilage, we find that type III collagen molecules...

  9. Irradiated homologous costal cartilage for augmentation rhinoplasty

    International Nuclear Information System (INIS)

    Although the ideal reconstructive material for augmentation rhinoplasty continues to challenge plastic surgeons, there exists no report in the literature that confines the use of irradiated homologous costal cartilage, first reported by Dingman and Grabb in 1961, to dorsal nasal augmentation. The purpose of this paper is to present a retrospective analysis of the author's experience using irradiated homologous costal cartilage in augmentation rhinoplasty. Twenty-seven dorsal nasal augmentations were performed in 24 patients between 16 and 49 years of age with a follow-up ranging from 1 to 27 months. Good-to-excellent results were achieved in 83.3% (20 of 24). Poor results requiring revision were found in 16.7% (4 of 24). Complication rates included 7.4% infection (2 of 27) and 14.8% warping (4 of 27). The resorption rate was zero. These results compare favorably with other forms of nasal augmentation. Advantages and disadvantages of irradiated homologous costal cartilage are discussed

  10. Development of artificial articular cartilage

    Indian Academy of Sciences (India)

    Biswajit Bera

    2009-10-01

    The present study describes the development of artificial articular cartilage on the basis of mimicking structural gel properties and mechanical gel properties of natural articular cartilage. It is synthesized from PVA/Si nanocomposite containing 20% Tetra ethoxy silane (TEOS) by sol–gel method. Mechanical strength of Poly(vinyl alcohol), PVA is improved up to 35 MPa. Manufacturing method is adopted considering colloidal stability of nano silica particle in PVA sol at specific pH = 1. An adhesive is also prepared from PVA/Si nanocomposite containing 40% TEOS for firm attachment of artificial articular cartilage on underlying bone with high bond strength.

  11. Reproducible simulation of respiratory motion in porcine lung explants

    International Nuclear Information System (INIS)

    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 motion. (orig.)

  12. 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)

    2006-11-15

    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

  13. Semi-automatic knee cartilage segmentation

    Science.gov (United States)

    Dam, Erik B.; Folkesson, Jenny; Pettersen, Paola C.; Christiansen, Claus

    2006-03-01

    Osteo-Arthritis (OA) is a very common age-related cause of pain and reduced range of motion. A central effect of OA is wear-down of the articular cartilage that otherwise ensures smooth joint motion. Quantification of the cartilage breakdown is central in monitoring disease progression and therefore cartilage segmentation is required. Recent advances allow automatic cartilage segmentation with high accuracy in most cases. However, the automatic methods still fail in some problematic cases. For clinical studies, even if a few failing cases will be averaged out in the overall results, this reduces the mean accuracy and precision and thereby necessitates larger/longer studies. Since the severe OA cases are often most problematic for the automatic methods, there is even a risk that the quantification will introduce a bias in the results. Therefore, interactive inspection and correction of these problematic cases is desirable. For diagnosis on individuals, this is even more crucial since the diagnosis will otherwise simply fail. We introduce and evaluate a semi-automatic cartilage segmentation method combining an automatic pre-segmentation with an interactive step that allows inspection and correction. The automatic step consists of voxel classification based on supervised learning. The interactive step combines a watershed transformation of the original scan with the posterior probability map from the classification step at sub-voxel precision. We evaluate the method for the task of segmenting the tibial cartilage sheet from low-field magnetic resonance imaging (MRI) of knees. The evaluation shows that the combined method allows accurate and highly reproducible correction of the segmentation of even the worst cases in approximately ten minutes of interaction.

  14. Effects of natural cartilaginous extracellular matrix on chondrogenic potential for cartilage cell transplantation.

    Science.gov (United States)

    Yang, W; Lee, S; Jo, Y H; Lee, K M; Nemeno, J G; Nam, B M; Kim, B Y; Jang, I J; Kim, H N; Takebe, T; Lee, J I

    2014-05-01

    Autologous chondrocyte transplantation (ACT) has been established to contribute cartilage regeneration over the past years; however, many obstacles need to be overcome. Recently, newer ACT technique involves cotransplantation of chondrocytes and biomaterial. Although various proposed intelligent biomaterials exist, many of them remain insufficient and controversial. In this study, we aimed to examine the effects of natural extracellular matrix (ECM) to the proliferation rate and differentiation on the chondrocytes. We first derived a natural ECM sheet from 10-μm-thick frozen sections of porcine knee cartilages. We then cultured the chondrocytes derived from a rabbit's knee on a dish precoated with the natural ECM. Then we assessed differentiation and chondrogenic potential of the cells compared with those grown in untreated culture dishes. We characterized the gene expression of chondrogenic markers, such as collagen type II, SOX-9, and aggrecan, as well as the level of ECM protein with the use of reverse-transcription polymerase chain reaction analysis. The cells cultured with the ECM sheet showed highest chondrogenic potential and differentiation. Therefore, we can induce good chondrogenesis by with the use of a natural ECM sheet on the culture dish. The readily available and easy-to-handle thin ECM sheets create an environment that promotes efficient cartilage regeneration. Our data suggest that this natural ECM scaffold improved the chondrogenic differentiation of the cells in vitro by providing a favorable microenvironment. PMID:24815172

  15. In Vitro Callus Induction and Embryogenesis of Oil Palm (Elaeis guineensis Jacq. from Leaf Explants

    Directory of Open Access Journals (Sweden)

    DWI HAPSORO

    2011-06-01

    Full Text Available This research was to study in vitro callus induction and somatic embryogenesis in oil palm from leaf explants. Young leaf segments from mature oil palm were cultured on MS medium supplemented with different concentrations of 2,4-D with or without addition of 2 g/l activated charcoal (AC or 2,4-D and picloram. Embryogenesis induction was done using MS medium containing 2,4-D 450 M and benziladenine 4.4 M with 3g/l activated charcoal. The treatment of 2,4-D 15 M resulted in the highest percentage of callus induction. The treatment of 2,4-D and AC showed that 2,4-D 450 M and AC led to higher percentage of callus induction than that of 2,4-D 400 M and 2 g/l AC. Embryogenesis occured in 27 out of 250 clumps of primary callus was occurred after 2-3 times subcultures. Somatic embryo development occurred when the embryogenic callus was transferred on the same basal medium supplemented with casein hydrolysate with 1 M BA or growth regulator free basal medium with 2 g/l activated charcoal.

  16. Effect of prolactin on enzymes of lipid biosynthesis in mammary gland explants

    International Nuclear Information System (INIS)

    Prolactin (PRL) stimulates an increased rate of incorporation of [14C]acetate and [3H]glucose into lipids in cultured mammary gland explants from 10-to 14-day-pregnant mice. This response is biphasic with an early increase occurring from 6 through 12 h, and an additional increase from 16 to 24 h. Enzymes likely to be rate limiting to this process include acetyl CoA carboxylase, fatty acid synthetase, acetyl CoA synthetase, and/or pyruvate dehydrogenase. Of these enzymes only pyruvate dehydrogenase activity was elevated at 6 h, suggesting that this enzymatic activity is important in stimulating early increases in lipogenesis after PRL treatment. In addition, the PRL stimulation of pyruvate dehydrogenase may also indirectly stimulate acetyl CoA carboxylase through the generation of citrate; this may explain the early (6-12 h) effect of PRL on [14C]acetate incorporation. After 16 h of PRL treatment, the activities of all the lipogenic enzymes were enhanced. The second phase of PRLs stimulation of lipogenesis thus likely involves the enhanced activities of more than one of the lipogenic enzymes

  17. Articular chondrocytes and mesenchymal stem cells seeded on biodegradable scaffolds for the repair of cartilage in a rat osteochondral defect model.

    Science.gov (United States)

    Dahlin, Rebecca L; Kinard, Lucas A; Lam, Johnny; Needham, Clark J; Lu, Steven; Kasper, F Kurtis; Mikos, Antonios G

    2014-08-01

    This work investigated the ability of co-cultures of articular chondrocytes and mesenchymal stem cells (MSCs) to repair articular cartilage in osteochondral defects. Bovine articular chondrocytes and rat MSCs were seeded in isolation or in co-culture onto electrospun poly(ɛ-caprolactone) (PCL) scaffolds and implanted into an osteochondral defect in the trochlear groove of 12-week old Lewis rats. Additionally, a blank PCL scaffold and untreated defect were investigated. After 12 weeks, the extent of cartilage repair was analyzed through histological analysis, and the extent of bone healing was assessed by quantifying the total volume of mineralized bone in the defect through microcomputed tomography. Histological analysis revealed that the articular chondrocytes and co-cultures led to repair tissue that consisted of more hyaline-like cartilage tissue that was thicker and possessed more intense Safranin O staining. The MSC, blank PCL scaffold, and empty treatment groups generally led to the formation of fibrocartilage repair tissue. Microcomputed tomography revealed that while there was an equivalent amount of mineralized bone formation in the MSC, blank PCL, and empty treatment groups, the defects treated with chondrocytes or co-cultures had negligible mineralized bone formation. Overall, even with a reduced number of chondrocytes, co-cultures led to an equal level of cartilage repair compared to the chondrocyte samples, thus demonstrating the potential for the use of co-cultures of articular chondrocytes and MSCs for the in vivo repair of cartilage defects. PMID:24927682

  18. Tissue engineering of cartilage in space

    OpenAIRE

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

    1997-01-01

    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...

  19. Directing chondrogenic differentiation of mesenchymal stem cells with a solid-supported chitosan thermogel for cartilage tissue engineering

    International Nuclear Information System (INIS)

    Hydrogels are attractive for cartilage tissue engineering because of their high plasticity and similarity with the native cartilage matrix. However, one critical drawback of hydrogels for osteochondral repair is their inadequate mechanical strength. To address this limitation, we constructed a solid-supported thermogel comprising a chitosan hydrogel system and demineralized bone matrix. Scanning electron microscopy, the equilibrium scanning ratio, the biodegradation rate, biomechanical tests, biochemical assays, metabolic activity tests, immunostaining and cartilage-specific gene expression analysis were used to evaluate the solid-supported thermogel. Compared with pure hydrogel or demineralized matrix, the hybrid biomaterial showed superior porosity, equilibrium swelling and degradation rate. The hybrid scaffolds exhibited an increased mechanical strength: 75% and 30% higher compared with pure hydrogels and demineralized matrix, respectively. After three days culture, bone-derived mesenchymal stem cells (BMSCs) maintained viability above 90% in all three materials; however, the cell retention of the hybrid scaffolds was more efficient and uniform than the other materials. Matrix production and chondrogenic differentiation of BMSCs in the hybrid scaffolds were superior to its precursors, based on glycosaminoglycan quantification and hyaline cartilage marker expression after three weeks in culture. Its easy preparation, favourable biophysical properties and chondrogenic capacity indicated that this solid-supported thermogel could be an attractive biomaterial framework for cartilage tissue engineering. (paper)

  20. RESEARCH ON BEHAVIOUR OF GINKGO BILOBA IN THE INITIATION STAGE OF IN VITRO CULTURE

    Directory of Open Access Journals (Sweden)

    Ana-Maria Radomir

    2012-04-01

    Full Text Available This article presents the realizations of the tehnology of producing biological material with rapidly clonal multiplication with reference at the phase of in vitro initiation. The growth of Ginkgo biloba explants was influenced by the period of explants sampling and by the composition of culture medium. The explants sampled from the herbaceous shoots a year old cropping at the end of the summer have the best behavior. They have registered 80% explants growth on culture medium MS with 20 mg/l benzyladenine. As one goes along the concentration of benzyladenine decreased, has been found a diminution of the number of explants growth until 25% and a progresive growth of the length of the shoots obtained.

  1. Efficient in vitro direct shoot organogenesis and regeneration of fertile plants from embryo explants of Bambara groundnut ( Vigna subterranea L. Verdc.).

    Science.gov (United States)

    Lacroix, B; Assoumou, Y; Sangwan, R S

    2003-08-01

    An efficient protocol has been developed for direct shoot organogenesis from embryo axes derived from mature seeds of two different landraces of Bambara groundnut. Multiple shoots were initiated on several media containing different concentrations and combinations of benzylaminopurine (BAP) or thidiazuron (TDZ). Efficient regeneration occurred when the embryo axes were first plated for 6 days on a medium containing high concentrations of BAP (1 mg/l) and alpha-naphthaleneacetic acid (NAA, 1 mg/l) and then cut transversely and transferred onto a medium containing 1.5 mg/l BAP. Shoot regeneration frequency was 100% and from five to eight shoots per explant were obtained. The importance of using embryo explants and cytokinins in the culture media, with respect to controlling the development of a highly organogenic system, was demonstrated. Histological studies revealed that proliferating buds originated directly from the superficial layers of the explants without an intermediate callus phase. The regenerated shoots were rooted on a medium containing 1 mg/l NAA and then transferred to the greenhouse. Flow cytometric analyses and chloroplast counts of guard cells suggested that the regenerants were diploid. All were morphologically normal and fertile. The short duration, high efficiency and low frequency of somaclonal variation of this system make it well suited for wider biotechnological applications of Bambara groundnut-a neglected and under-utilized crop. PMID:12910367

  2. Effect of Age of Explant on Transgenic Cotton (Gossypium Plant Due to Expression of Mannose-Binding Lectin Gene from Allium sativum

    Directory of Open Access Journals (Sweden)

    Lynelle van Emmenes

    2011-09-01

    Full Text Available Cotton is the most important textile plant in the world and is one of the most important crops for the production of oilseed. Because of its worldwide economic importance, new cultivars are constantly being released in the world. Although great improvements have been achieved through traditional breeding methods, cotton breeders are facing many problems, i.e., narrow genetic base, inability to use alien genes and difficulty in breaking gene linkages. Genetic transformations analyses are main tools used by breeders to overcome these problems. The aim of the study reported in this paper is to determine the effect of age of explant on regeneration response of apical shoot for tissue culture and gene transfer systems of cotton. This enabled us evaluate it effects on cotton transformation. The age of explants was observed to have significant effect on shoot tip elongation. The elongation rates of the three varieties studied were not significantly different from each other (p = 0.1573 and was observed to be affected by the size of isolated tips. It was observed that if the starting size of the apex was less than 1 mm, the tips would not grow at all. Insecticidal lectin gene from Allium sativum was transferred into the transgenic cotton plants via Agrobacterium-mediated transformation using shoot apices as explants. Putative transgenic plants were confirmed by leaf GUS assay, kanamycin leaf test and molecular analysis of plantlet.

  3. Establishment of in vitro culture of grapes

    International Nuclear Information System (INIS)

    The establishment of in vitro culture from shoot tip explants (meristemetic tissue) of grapes was investigated through tissue culture technique. These explants were collected from gene bank of Institute of Agricultural Biotechnology and Genetic Resources (IABGR), National Agricultural Research Centre (NARC), Islamabad, Pakistan. Fifteen accessions of grapes were surface sterilized and tested on 75% MS media for germination and initial growth parameters. Accession No. 020017 (Dakh-1) exhibited highest viability (100%), shoot length (4.12 cm) and nodes plant (3.8). Moreover, it was found that response of cultures to different treatments was dependent both on accession and treatment duration. In conclusion, this protocol proved to be useful in optimizing the dose and duration of the treatment of grape explants with the surface disinfectant. (author)

  4. Somatic embryogenesis in anthurium (Anthurium andraeanum cv. Eidibel as affected by different explants - doi: 10.4025/actasciagron.v36i1.16557

    Directory of Open Access Journals (Sweden)

    Marcos Vinícius Marques Pinheiro

    2014-01-01

    Full Text Available This study establishes a protocol for the induction of somatic embryogenesis in Anthurium andraeanum cv. Eidibel. The experiment was arranged in a completely randomized 5 x 5 x 5 factorial design using five explant types (whole leaves, half leaves; petiole; nodal segments and root segments from in vitro plantlets; five auxins: indole-3-acetic acid (IAA, α-naphthaleneacetic acid (NAA, indole-3-butyric acid (IBA, 2,4-dichlorophenoxyacetic acid (2,4-D, and 4-amino-3,5,6-trichloropicolinic acid (Picloram; at five concentrations (0, 2.5, 5, 7.5 and 10 µM, with five replications using five Petri dishes. The cultures were maintained in a growth room at 25 ± 2ºC in the dark. The explant type was investigated for the induction of somatic embryogenesis in anthurium cv. Eidibel, and nodal segments were shown to be the most suitable explant for this process. After 60 days in culture, the highest number of embryogenic calli was recorded for the nodal segments cultured in NAA (5, 7.5 and 10 µM, 2,4-D (10 µM and Picloram (7.5 and 10 µM. The histological analysis confirmed the presence of embryos with established polarization, procambium, ground meristem and protoderm in the nodal segments cultured in Pierik medium containing 10 µM NAA. After the conversion of the somatic embryos into plantlets, these plantlets were acclimatized transferred to in vivo conditions and grown into normal plants.

  5. Proteomic Profile of Brucella abortus-Infected Bovine Chorioallantoic Membrane Explants

    Science.gov (United States)

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

    2016-01-01

    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

  6. cultural

    Directory of Open Access Journals (Sweden)

    Irene Kreutz

    2006-01-01

    Full Text Available Es un estudio cualitativo que adoptó como referencial teorico-motodológico la antropología y la etnografía. Presenta las experiencias vivenciadas por mujeres de una comunidad en el proceso salud-enfermedad, con el objetivo de comprender los determinantes sócio-culturales e históricos de las prácticas de prevención y tratamiento adoptados por el grupo cultural por medio de la entrevista semi-estructurada. Los temas que emergieron fueron: la relación entre la alimentación y lo proceso salud-enfermedad, las relaciones con el sistema de salud oficial y el proceso salud-enfermedad y lo sobrenatural. Los dados revelaron que los moradores de la comunidad investigada tienen un modo particular de explicar sus procedimientos terapéuticos. Consideramos que es papel de los profesionales de la salud en sus prácticas, la adopción de abordajes o enfoques que consideren al individuo en su dimensión sócio-cultural e histórica, considerando la enorme diversidad cultural en nuestro país.

  7. Flavonoid Compound Icariin Activates Hypoxia Inducible Factor-1α in Chondrocytes and Promotes Articular Cartilage Repair.

    Science.gov (United States)

    Wang, Pengzhen; Zhang, Fengjie; He, Qiling; Wang, Jianqi; Shiu, Hoi Ting; Shu, Yinglan; Tsang, Wing Pui; Liang, Shuang; Zhao, Kai; Wan, Chao

    2016-01-01

    Articular cartilage has poor capability for repair following trauma or degenerative pathology due to avascular property, low cell density and migratory ability. Discovery of novel therapeutic approaches for articular cartilage repair remains a significant clinical need. Hypoxia is a hallmark for cartilage development and pathology. Hypoxia inducible factor-1alpha (HIF-1α) has been identified as a key mediator for chondrocytes to response to fluctuations of oxygen availability during cartilage development or repair. This suggests that HIF-1α may serve as a target for modulating chondrocyte functions. In this study, using phenotypic cellular screen assays, we identify that Icariin, an active flavonoid component from Herba Epimedii, activates HIF-1α expression in chondrocytes. We performed systemic in vitro and in vivo analysis to determine the roles of Icariin in regulation of chondrogenesis. Our results show that Icariin significantly increases hypoxia responsive element luciferase reporter activity, which is accompanied by increased accumulation and nuclear translocation of HIF-1α in murine chondrocytes. The phenotype is associated with inhibiting PHD activity through interaction between Icariin and iron ions. The upregulation of HIF-1α mRNA levels in chondrocytes persists during chondrogenic differentiation for 7 and 14 days. Icariin (10-6 M) increases the proliferation of chondrocytes or chondroprogenitors examined by MTT, BrdU incorporation or colony formation assays. Icariin enhances chondrogenic marker expression in a micromass culture including Sox9, collagen type 2 (Col2α1) and aggrecan as determined by real-time PCR and promotes extracellular matrix (ECM) synthesis indicated by Alcian blue staining. ELISA assays show dramatically increased production of aggrecan and hydroxyproline in Icariin-treated cultures at day 14 of chondrogenic differentiation as compared with the controls. Meanwhile, the expression of chondrocyte catabolic marker genes

  8. Flavonoid Compound Icariin Activates Hypoxia Inducible Factor-1α in Chondrocytes and Promotes Articular Cartilage Repair.

    Directory of Open Access Journals (Sweden)

    Pengzhen Wang

    Full Text Available Articular cartilage has poor capability for repair following trauma or degenerative pathology due to avascular property, low cell density and migratory ability. Discovery of novel therapeutic approaches for articular cartilage repair remains a significant clinical need. Hypoxia is a hallmark for cartilage development and pathology. Hypoxia inducible factor-1alpha (HIF-1α has been identified as a key mediator for chondrocytes to response to fluctuations of oxygen availability during cartilage development or repair. This suggests that HIF-1α may serve as a target for modulating chondrocyte functions. In this study, using phenotypic cellular screen assays, we identify that Icariin, an active flavonoid component from Herba Epimedii, activates HIF-1α expression in chondrocytes. We performed systemic in vitro and in vivo analysis to determine the roles of Icariin in regulation of chondrogenesis. Our results show that Icariin significantly increases hypoxia responsive element luciferase reporter activity, which is accompanied by increased accumulation and nuclear translocation of HIF-1α in murine chondrocytes. The phenotype is associated with inhibiting PHD activity through interaction between Icariin and iron ions. The upregulation of HIF-1α mRNA levels in chondrocytes persists during chondrogenic differentiation for 7 and 14 days. Icariin (10-6 M increases the proliferation of chondrocytes or chondroprogenitors examined by MTT, BrdU incorporation or colony formation assays. Icariin enhances chondrogenic marker expression in a micromass culture including Sox9, collagen type 2 (Col2α1 and aggrecan as determined by real-time PCR and promotes extracellular matrix (ECM synthesis indicated by Alcian blue staining. ELISA assays show dramatically increased production of aggrecan and hydroxyproline in Icariin-treated cultures at day 14 of chondrogenic differentiation as compared with the controls. Meanwhile, the expression of chondrocyte catabolic

  9. Thermosensitive chitosan-Pluronic hydrogel as an injectable cell delivery carrier for cartilage regeneration.

    Science.gov (United States)

    Park, Kyung Min; Lee, Sang Young; Joung, Yoon Ki; Na, Jae Sik; Lee, Myung Chul; Park, Ki Dong

    2009-07-01

    Injectable hydrogels have been studied for potential applications for articular cartilage regeneration. In this study, a thermosensitive chitosan-Pluronic (CP) hydrogel was designed as an injectable cell delivery carrier for cartilage regeneration. The CP conjugate was synthesized by grafting Pluronic onto chitosan using EDC/NHS chemistry. The sol-gel phase transition and mechanical properties of the CP hydrogel were examined by rheological experiments. The CP solution underwent a sol-gel transition around 25 degrees C at which the storage modulus (G') approaches 10(4)Pa, highlighting the potential of this material as an injectable scaffold for cartilage regeneration. The CP hydrogel was formed rapidly by increasing the temperature. The morphology of the dried CP hydrogel was observed by scanning electron microscopy. In vitro cell culture was performed using bovine chondrocytes. The proliferation of bovine chondrocytes and the amount of synthesized glycosaminoglycan increased for 28 days. These results suggested that the CP hydrogel has potential as an injectable cell delivery carrier for cartilage regeneration and could serve as a new biomaterial for tissue engineering. PMID:19261553

  10. Naringin Protects Against Cartilage Destruction in Osteoarthritis Through Repression of NF-κB Signaling Pathway.

    Science.gov (United States)

    Zhao, Yunpeng; Li, Zhong; Wang, Wenhan; Zhang, Hui; Chen, Jianying; Su, Peng; Liu, Long; Li, Weiwei

    2016-02-01

    Naringin was previously reported as a multifunctional agent. Recently, naringin was found to play a protective role in various inflammatory conditions. However, the role of naringin in cartilage degeneration and osteoarthritis (OA) progression is still unknown. TNF-α is reported to play a detrimental role in OA. Herein, primary murine chondrocytes were isolated and cultured with stimulation of TNF-α, in the presence or absence of naringin treatment. As a result, naringin attenuated TNF-α-mediated inflammation and catabolism in chondrocyte. Besides, surgically induced OA mice models were established. Cartilage degradation and OA severity were evaluated using Safranin-O staining, immunohistochemistry, and ELISA. Moreover, levels of inflammatory cytokines and catabolic markers in OA were analyzed. Oral administration of naringin alleviated degradation of cartilage matrix and protected against OA development in the surgically induced OA models. Furthermore, the protective function of naringin in cartilage and chondrocyte was possibly due to suppression of NF-κB signaling pathway. Collectively, this study presents naringin as a potential target for the treatment of joint degenerative diseases, including OA. PMID:26438631

  11. Development of large engineered cartilage constructs from a small population of cells.

    Science.gov (United States)

    Brenner, Jillian M; Kunz, Manuela; Tse, Man Yat; Winterborn, Andrew; Bardana, Davide D; Pang, Stephen C; Waldman, Stephen D

    2013-01-01

    Confronted with articular cartilage's limited capacity for self-repair, joint resurfacing techniques offer an attractive treatment for damaged or diseased tissue. Although tissue engineered cartilage constructs can be created, a substantial number of cells are required to generate sufficient quantities of tissue for the repair of large defects. As routine cell expansion methods tend to elicit negative effects on chondrocyte function, we have developed an approach to generate phenotypically stable, large-sized engineered constructs (≥3 cm(2) ) directly from a small amount of donor tissue or cells (as little as 20,000 cells to generate a 3 cm(2) tissue construct). Using rabbit donor tissue, the bioreactor-cultivated constructs were hyaline-like in appearance and possessed a biochemical composition similar to native articular cartilage. Longer bioreactor cultivation times resulted in increased matrix deposition and improved mechanical properties determined over a 4 week period. Additionally, as the anatomy of the joint will need to be taken in account to effectively resurface large affected areas, we have also explored the possibility of generating constructs matched to the shape and surface geometry of a defect site through the use of rapid-prototyped defect tissue culture molds. Similar hyaline-like tissue constructs were developed that also possessed a high degree of shape correlation to the original defect mold. Future studies will be aimed at determining the effectiveness of this approach to the repair of cartilage defects in an animal model and the creation of large-sized osteochondral constructs. PMID:23197468

  12. Effects of laser irradiation on immature olfactory neuroepithelial explants from the rat

    Energy Technology Data Exchange (ETDEWEB)

    Mester, A.F.; Snow, J.B. Jr.

    1988-07-01

    The photobiological effect of low-output laser irradiation on the maturation and regeneration of immature olfactory bipolar receptor cells of the rat was studied. The maturation and regeneration of the receptor cells of rat fetuses were quantified in neuroepithelial explants with morphometric analysis. The number of explants with outgrowth and the number and length of neuritic outgrowths were determined on a regular basis for 12 days. Explants in the experimental group were irradiated with a helium-neon laser using different incident energy densities (IED). Explants in the fluorescent light control group were exposed to fluorescent light for the same periods of time as those in the experimental group were exposed to laser irradiation. Explants in another control group were not exposed to laser or fluorescent light irradiation. The IED of 0.5 J/cm2 laser irradiation has been found to increase significantly the number of explants with outgrowth and the number and length of the outgrowths. Other laser IEDs or fluorescent light irradiation did not influence maturation or regeneration.

  13. Molecular analysis of chondrocytes cultured in agarose in response to dynamic compression

    Directory of Open Access Journals (Sweden)

    Mallein-Gerin Frédéric

    2008-09-01

    Full Text Available Abstract Background Articular cartilage is exposed to high mechanical loads under normal physiological conditions and articular chondrocytes regulate the composition of cartilaginous matrix, in response to mechanical signals. However, the intracellular pathways involved in mechanotransduction are still being defined. Using the well-characterized chondrocyte/agarose model system and dynamic compression, we report protocols for preparing and characterizing constructs of murine chondrocytes and agarose, and analyzing the effect of compression on steady-state level of mRNA by RT-PCR, gene transcription by gene reporter assay, and phosphorylation state of signalling molecules by Western-blotting. The mouse model is of particular interest because of the availability of a large choice of bio-molecular tools suitable to study it, as well as genetically modified mice. Results Chondrocytes cultured in agarose for one week were surrounded by a newly synthesized pericellular matrix, as revealed by immunohistochemistry prior to compression experiments. This observation indicates that this model system is suitable to study the role of matrix molecules and trans-membrane receptors in cellular responsiveness to mechanical stress. The chondrocyte/agarose constructs were then submitted to dynamic compression with FX-4000C™ Flexercell® Compression Plus™ System (Flexcell. After clearing proteins off agarose, Western-blotting analysis showed transient activation of Mitogen-activated protein kinases (MAPK in response to dynamic compression. After assessment by capillary electrophoresis of the quality of RNA extracted from agarose, steady-state levels of mRNA expression was measured by real time PCR. We observed an up-regulation of cFos and cJun mRNA levels as a response to compression, in accordance with the mechanosensitive character observed for these two genes in other studies using cartilage explants submitted to compression. To explore further the

  14. Interactive segmentation of Hip Joint Cartilage

    Czech Academy of Sciences Publication Activity Database

    Dvořák, Pavel; Juráš, V.; Vogl, W.; Chytil, J.

    Cambridge: The Electromagnetics Academy, 2014, s. 2369-2372. ISBN 978-1-934142-28-8. [PIERS 2014. Progress In Electromagnetics Research Symposium /35./. Guangzhou (CN), 25.08.2014-28.08.2014] R&D Projects: GA ČR GAP102/12/1104 Institutional support: RVO:68081731 Keywords : hip joint * MRI * segmentation of cartilage Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering

  15. Birth injuries to the epiphyseal cartilage

    International Nuclear Information System (INIS)

    A birth injury in the vicinity of a joint might lead to a fracture through the epiphyseal cartilage. The criteria for diagnosing such a fracture at radiography are considered and the continued remodelling of the bone demonstrated. The history of 2 cases with late diagnosis and serious long-term sequelae are described, in order to emphasize the necessity of early radiography. (Auth.)

  16. Oxygen, nitric oxide and articular cartilage

    Directory of Open Access Journals (Sweden)

    B Fermor

    2007-04-01

    Full Text Available Molecular oxygen is required for the production of nitric oxide (NO, a pro-inflammatory mediator that is associated with osteoarthritis and rheumatoid arthritis. To date there has been little consideration of the role of oxygen tension in the regulation of nitric oxide production associated with arthritis. Oxygen tension may be particularly relevant to articular cartilage since it is avascular and therefore exists at a reduced oxygen tension. The superficial zone exists at approximately 6% O2, while the deep zone exists at less than 1% O2. Furthermore, oxygen tension can alter matrix synthesis, and the material properties of articular cartilage in vitro.The increase in nitric oxide associated with arthritis can be caused by pro-inflammatory cytokines and mechanical stress. Oxygen tension significantly alters endogenous NO production in articular cartilage, as well as the stimulation of NO in response to both mechanical loading and pro-inflammatory cytokines. Mechanical loading and pro-inflammatory cytokines also increase the production of prostaglandin E2 (PGE2. There is a complex interaction between NO and PGE2, and oxygen tension can alter this interaction. These findings suggest that the relatively low levels of oxygen within the joint may have significant influences on the metabolic activity, and inflammatory response of cartilage as compared to ambient levels. A better understanding of the role of oxygen in the production of inflammatory mediators in response to mechanical loading, or pro-inflammatory cytokines, may aid in the development of strategies for therapeutic intervention in arthritis.

  17. Spatially resolved elemental distributions in articular cartilage

    International Nuclear Information System (INIS)

    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

  18. Advanced Strategies for Articular Cartilage Defect Repair

    Directory of Open Access Journals (Sweden)

    Fergal J. O'Brien

    2013-02-01

    Full Text Available Articular cartilage is a unique tissue owing to its ability to withstand repetitive compressive stress throughout an individual’s lifetime. However, its major limitation is the inability to heal even the most minor injuries. There still remains an inherent lack of strategies that stimulate hyaline-like articular cartilage growth with appropriate functional properties. Recent scientific advances in tissue engineering have made significant steps towards development of constructs for articular cartilage repair. In particular, research has shown the potential of biomaterial physico-chemical properties significantly influencing the proliferation, differentiation and matrix deposition by progenitor cells. Accordingly, this highlights the potential of using such properties to direct the lineage towards which such cells follow. Moreover, the use of soluble growth factors to enhance the bioactivity and regenerative capacity of biomaterials has recently been adopted by researchers in the field of tissue engineering. In addition, gene therapy is a growing area that has found noteworthy use in tissue engineering partly due to the potential to overcome some drawbacks associated with current growth factor delivery systems. In this context, such advanced strategies in biomaterial science, cell-based and growth factor-based therapies that have been employed in the restoration and repair of damaged articular cartilage will be the focus of this review article.

  19. Zn deposition at the bone cartilage interface in equine articular cartilage

    Science.gov (United States)

    Bradley, D. A.; Moger, C. J.; Winlove, C. P.

    2007-09-01

    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 μm 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.

  20. 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: D.A.Bradley@surrey.ac.uk; Moger, C.J.; Winlove, C.P. [School of Physics, University of Exeter, Exeter, EX4 4QL (United Kingdom)

    2007-09-21

    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.

  1. Zn deposition at the bone-cartilage interface in equine articular cartilage

    International Nuclear Information System (INIS)

    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 μm 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

  2. Effect of Basal Medium, Explants Size and Density on the In Vitro Proliferation and Growth of Date Palm (Phoenix dactylifera L. Cultivar ‘16-bis’

    Directory of Open Access Journals (Sweden)

    Mouaad Amine MAZRI

    2013-08-01

    Full Text Available The effect of basal medium, explant size and density on shoot multiplication, growth, rooting and acclimatization of date palm cv. ‘16-bis’ was evaluated. Bud clusters of different sizes (2, 3, 4 and 5 buds per cluster were cultured at density of 1, 2, 3 and 4 clusters on Murashige and Skoog medium (MS, woody plant medium (WPM and Nitsch medium (NM supplemented with 0.5 mg/L 2-naphthoxyacetic acid and 0.5 mg/L kinetin for three months (multiplication phase. Separated shoots of different sizes (<3 cm; 3 to 4.5 cm and 4.5 to 6 cm were cultured at density of 1, 2, 3 and 4 shoots on hormone free MS medium, WPM or NM for three months (Elongation-rooting phase. The proliferation and development of shoots were affected by the basal medium, explant size and density. The optimal shoot proliferation (18.1 was observed when 4 buds clusters were cultured at the density of 2 clusters per jar in MS medium. Separated shoots of 4.5 to 6 cm length exhibited the optimal in vitro development in terms of leaf length and greening, and root number and length when cultured on MS medium. In addition, these shoots reached the highest acclimation frequency with 80%. Our results would be utilized for an efficient propagation of plantlets of cv. ‘16-bis’, a selected date palm cultivar resistant to the bayoud disease.

  3. Cartilage Regeneration in Human with Adipose Tissue-Derived Stem Cells: Current Status in Clinical Implications

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    Jaewoo Pak

    2016-01-01

    Full Text Available Osteoarthritis (OA is one of the most common debilitating disorders among the elderly population. At present, there is no definite cure for the underlying causes of OA. However, adipose tissue-derived stem cells (ADSCs in the form of stromal vascular fraction (SVF may offer an alternative at this time. ADSCs are one type of mesenchymal stem cells that have been utilized and have demonstrated an ability to regenerate cartilage. ADSCs have been shown to regenerate cartilage in a variety of animal models also. Non-culture-expanded ADSCs, in the form of SVF along with platelet rich plasma (PRP, have recently been used in humans to treat OA and other cartilage abnormalities. These ADSCs have demonstrated effectiveness without any serious side effects. However, due to regulatory issues, only ADSCs in the form of SVF are currently allowed for clinical uses in humans. Culture-expanded ADSCs, although more convenient, require clinical trials for a regulatory approval prior to uses in clinical settings. Here we present a systematic review of currently available clinical studies involving ADSCs in the form of SVF and in the culture-expanded form, with or without PRP, highlighting the clinical effectiveness and safety in treating OA.

  4. Factors affecting proliferation and elongation of shoots of Phak Liang (Gnetum gnemon Linn. through tissue culture technique

    Directory of Open Access Journals (Sweden)

    Te-chato, S.

    2003-09-01

    Full Text Available The tissue culture of Phak Liang (Gnetum gnemon Linn. was investigated for micropropagation. The types of explant, culture media, types and concentrations of plant growth regulators, orientation of explant and section of explant were tested for their efficacy in inducing and proliferating shoot buds. The elongation of shoots and root induction was also studied. Young leaves gave the highest number of shoot buds when they were cultured in Murashige and Skoog (MS medium supplemented with 0.25 mg/l IBA and 1.53 mg/l BA. The medium supplemented with 0.25 mg/l thidiazuron (TDZ alone provided the best result on multiple shoot bud induction both in percentage of explant forming shoots and number of shoot buds per explant. The percentage of explant forming shoot buds and number of shoot buds obtained from leaves were 90% and 26.50 shoot buds, while those from stems were 96.25% and 23.00 shoot buds, respectively. One hundred percent friable callus was induced from stem explant in the same medium supplemented with 1.0 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D after 2 months of culture. Culturing whole leaf in the position of dorsal contact with medium gave the best multiple shoot bud formation of 92% and 23.00 shoot buds/explant. Cutting stem into half and culturing in horizontal position gave the best multiple shoot bud formation of 96% and 23.00 shoot buds/explant after culture for 2 months. The best elongation of shoot buds (2.54 shoots derived from cultured leaves was induced in the liquid medium. While stem-derived shoot buds (3.45 shoots was induced in the solid medium of the same medium components. However, root could not be induced from elongated shoots.

  5. STIMULATION OF RAPID REGENERATION BY A MAGNETIC FIELD IN PAULOWNIA NODE CULTURES

    OpenAIRE

    Özge Çelik; Çimen Atak; Aitekin Rzakulieva

    2008-01-01

    In this study, the aim was to determine the effect of magnetic fields on regeneration of Paulownia node cultures. Paulownia tomentosa node cultures were used to generate explants and these explants were passed through a 2.9- 4.6-mT magnetic flux density 1 and 9 times at 2.2 and 19.8 seconds, respectively. Chlorophyll quantities, total RNA concentrations of shoots and shoot formation rates from control and treated explants were determined. While the shoot formation rate was 61.9% in the contro...

  6. Enhanced mechanical properties of thermosensitive chitosan hydrogel by silk fibers for cartilage tissue engineering

    Energy Technology Data Exchange (ETDEWEB)

    Mirahmadi, Fereshteh [Faculty of Biomedical Engineering, Amirkabir University of Technology, Tehran (Iran, Islamic Republic of); National Cell Bank of Iran, Pasteur Institute of Iran, Tehran (Iran, Islamic Republic of); Tafazzoli-Shadpour, Mohammad, E-mail: Tafazoli@aut.ac.ir [Faculty of Biomedical Engineering, Amirkabir University of Technology, Tehran (Iran, Islamic Republic of); Shokrgozar, Mohammad Ali, E-mail: mashokrgozar@pasteur.ac.ir [National Cell Bank of Iran, Pasteur Institute of Iran, Tehran (Iran, Islamic Republic of); Bonakdar, Shahin [National Cell Bank of Iran, Pasteur Institute of Iran, Tehran (Iran, Islamic Republic of)

    2013-12-01

    Articular cartilage has limited repair capability following traumatic injuries and current methods of treatment remain inefficient. Reconstructing cartilage provides a new way for cartilage repair and natural polymers are often used as scaffold because of their biocompatibility and biofunctionality. In this study, we added degummed chopped silk fibers and electrospun silk fibers to the thermosensitive chitosan/glycerophosphate hydrogels to reinforce two hydrogel constructs which were used as scaffold for hyaline cartilage regeneration. The gelation temperature and gelation time of hydrogel were analyzed by the rheometer and vial tilting method. Mechanical characterization was measured by uniaxial compression, indentation and dynamic mechanical analysis assay. Chondrocytes were then harvested from the knee joint of the New Zealand white rabbits and cultured in constructs. The cell proliferation, viability, production of glycosaminoglycans and collagen type II were assessed. The results showed that mechanical properties of the hydrogel were significantly enhanced when a hybrid with two layers of electrospun silk fibers was made. The results of GAG and collagen type II in cell-seeded scaffolds indicate support of the chondrogenic phenotype for chondrocytes with a significant increase in degummed silk fiber–hydrogel composite for GAG content and in two-layer electrospun fiber–hydrogel composite for Col II. It was concluded that these two modified scaffolds could be employed for cartilage tissue engineering. - Highlights: • Chitosan hydrogel composites fabricated by two forms of silk fiber • Silk fibers provide structural support for the hydrogel matrix. • The mechanical properties of hydrogel significantly improved by associating with silk. • Production of GAG and collagen type II was demonstrated within the scaffolds.

  7. Ventricular Recovery and Pump Explantation in Patients Supported by Left Ventricular Assist Devices: A Systematic Review.

    Science.gov (United States)

    Phan, Kevin; Huo, Ya Ruth; Zhao, Dong Fang; Yan, Tristan D; Tchantchaleishvili, Vakhtang

    2016-01-01

    Several studies have reported that a portion of patients who exhibit cardiac recovery during left ventricular assist device (LVAD) support can have their device explanted with reasonable long-term survival. The aim of this systematic review is to assess the survival and cardiac function in patients with explanted LVADs from the current literature. Electronic search was performed to identify all studies in English literature assessing LVAD explantation. All identified articles were systematically assessed using the inclusion and exclusion criteria. Selected studies were subjected to quantitative assessment. From 5 electronic databases, 11 studies (213 patients) were included. Pooled mean perioperative mortality rate of those explanted was 9.2% (95% CI, 5.0-14.5%; I = 0). Pooled mean late mortality rate was 15% (95% CI, 9.0-22.1%; I = 31%). The pooled 1, 5, and 10 year survival postexplant was 91, 76, and 65.7%, respectively. Pooled postweaning freedom from heart failure (HF) recurrence reached 81.3%. Subset analysis demonstrated that patients explanted from a continuous-flow LVAD versus pulsatile LVAD had a lower rate of HF recurrence (6.6 vs. 28.3%, p = 0.03) and LVAD reimplantation (7.5 vs. 37%, p = 0.001). Before LVAD explantation, overall mean left ventricular ejection fraction (LVEF) was 49%. Weighted pooled early and late postexplantation mean LVEF was 47.3 and 41.2%, respectively. Late postexplantation LVEF was significantly higher in the continuous-flow versus pulsatile LVAD subgroup (41.5 vs. 24%, p = 0.001). This review shows encouraging safety and 10 year survival outcomes after explantation of LVADs in carefully selected patients, with rates better than expected after a heart transplant. Recovery of the native heart is the most desirable clinical outcome in patients supported with LVADs and should be actively sought. PMID:26735559

  8. BACTERIAL MICROORGANISMS ASSOCIATED WITH THE PLANT TISSUE CULTURE: IDENTIFICATION AND POSSIBLE ROLE (review)

    OpenAIRE

    S.E. DUNAEVA; Yu, S.

    2015-01-01

    Effective sterilization of plant explants and antiseptics rules compliance do not exclude the presence of so-called covert (endophytic) bacteria in in vitro cultures. But the role of these bacteria in tissues cultures has been not enough studied whereas it was related to the explants regeneration capacity and the possibility of animal and human cells transformation under in vitro cultivation. Bacterial strains pathogenic to humans can be stably maintained in cultivated tissues and ex vitro pl...

  9. Elimination of browning exudate and in vitro development of shoots in Pistacia vera L. cv. mateur and Pistacia atlantica Desf. Culture

    OpenAIRE

    M. I. Trujillo; S. Mederos-Molina

    2014-01-01

    We report diminution and/or elimination of browning exudate followed by in vitro establishment of in Pistacia vera cv. mateur and Pistacia atlantica explants. Soaking P. vera cv. mateur explants prior to culture in L-cysteine HCl for 15 min (100 µM) inhibits blackening of the modified Murashige and Skoog medium - MS + 400 mg/l NH4NO3 - and of the explants; while shoot formation was increased. The browning in P. vera cv. mateur and P. atlantica explants dissolved when modified MS and Quoirin a...

  10. The effects of exposure dose and dose rate of gamma radiation on in vitro shoot-forming capacity of cotyledon explants in red pepper (Capsicum annuum L. cv. Yatsufusa)

    International Nuclear Information System (INIS)

    The effect of exposure dose and dose rate of gamma radiation on shoot-forming capacity of cotyledon explants of red pepper were investigated. The twelve-day-old seedlings were irradiated by gamma radiation with varying exposure doses of O, 0.5, 0.75, 1.0, 1.25, 1.5, 1.75, 2.0 and 2.5 krad delivered at the dose rates of l, 5 and 10 krad/ hr. After irradiation, cotyledons were excised from the seedlings and cultured on MURASHIGE and SKOOG medium supplemented with benzyladenine at the concentration of 3 mg/l to induce shoot formation. It was found that the percentage of shoot-forming explants and the number of shoots per explant were decreased as the exposure dose increased. Moreover, the increase in exposure dose delayed callus and shoot bud formation and inhibited the development of shoot buds into vigorous shoots. At the same exposure dose, the higher dose rate (10 krad/hr) was more detrimental for shoot formation than the lower ones (1 and 5 krad/hr). The exposure dose which caused 50 percent reduction in the number of shoots per explant (RD50) was around 1.0 krad at the dose rates of 1 and 5 krad/hr whereas RD50 was 0.75 krad at the rate of 10 krad/hr. (author)

  11. Analyzing Biological Performance of 3D-Printed, Cell-Impregnated Hybrid Constructs for Cartilage Tissue Engineering.

    Science.gov (United States)

    Izadifar, Zohreh; Chang, Tuanjie; Kulyk, William; Chen, Xiongbiao; Eames, B Frank

    2016-03-01

    Three-dimensional (3D) bioprinting of hybrid constructs is a promising biofabrication method for cartilage tissue engineering because a synthetic polymer framework and cell-impregnated hydrogel provide structural and biological features of cartilage, respectively. During bioprinting, impregnated cells may be subjected to high temperatures (caused by the adjacent melted polymer) and process-induced mechanical forces, potentially compromising cell function. This study addresses these biofabrication issues, evaluating the heat distribution of printed polycaprolactone (PCL) strands and the rheological property and structural stability of alginate hydrogels at various temperatures and concentrations. The biocompatibility of parameters from these studies was tested by culturing 3D hybrid constructs bioprinted with primary cells from embryonic chick cartilage. During initial two-dimensional culture expansion of these primary cells, two morphologically and molecularly distinct cell populations ("rounded" and "fibroblastic") were isolated. The biological performance of each population was evaluated in 3D hybrid constructs separately. The cell viability, proliferation, and cartilage differentiation were observed at high levels in hybrid constructs of both cell populations, confirming the validity of these 3D bioprinting parameters for effective cartilage tissue engineering. Statistically significant performance variations were observed, however, between the rounded and fibroblastic cell populations. Molecular and morphological data support the notion that such performance differences may be attributed to the relative differentiation state of rounded versus fibroblastic cells (i.e., differentiated chondrocytes vs. chondroprogenitors, respectively), which is a relevant issue for cell-based tissue engineering strategies. Taken together, our study demonstrates that bioprinting 3D hybrid constructs of PCL and cell-impregnated alginate hydrogel is a promising approach for

  12. Productivity of the improvement in secondary metabolites by gamma-irradiation and tissue culture in vitro of Phyllanthus niruri L. (Euphorbiaceae)

    International Nuclear Information System (INIS)

    The in vitro culture of Phyllanthus niruri L. stem explants irradiated with increasing doses of gamma radiation on medium MS supplemented with IBA/BAP and albumen liquid of C. nucifera milk showed that the callogenesis decrease with the dose of irradiation. The alkaloids, tanins catechic, saponins and terpenoids were identified in the extracts of the calli of irradiated explants, contrary to the extracts of the original plant. The production of the secondary metabolites was abundant in the aqueous extracts of the calli of explants irradiated compared to those of the calli of explants not exposed to radiation and the original plant. She was higher on the extracts of the calli of irradiated explants with 5Gy. The sensitivity of Plasmodium falciparum stain was maximum (100%) with the extracts of calli's explants irradiated with 1Gy, with the concentrations of 300 and 3μg/ml. The extract of the original plant gave the smallest percentage of the inhibition (65,25%)

  13. Analysis of friction between articular cartilage and polyvinyl alcohol hydrogel artificial cartilage.

    Science.gov (United States)

    Li, Feng; Wang, Anmin; Wang, Chengtao

    2016-05-01

    Many biomaterials are being used to repair damaged articular cartilage. In particular, poly vinyl alcohol hydrogel has similar mechanical properties to natural cartilage under compressive and shearing loading. Here, three-factor and two-level friction experiments and long-term tests were conducted to better evaluate its tribological properties. The friction coefficient between articular cartilage and the poly vinyl alcohol hydrogel depended primarily on the three factors of load, speed, and lubrication. When the speed increased from 10 to 20 mm/s under a load of 10 N, the friction coefficient increased from 0.12 to 0.147. When the lubricant was changed from Ringer's solution to a hyaluronic acid solution, the friction coefficient decreased to 0.084 with loads as high as 22 N. The poly vinyl alcohol hydrogel was severely damaged and lost its top surface layers, which were transferred to the articular cartilage surface. Wear was observed in the surface morphologies, which indicated the occurrence of surface adhesion of bovine cartilage. Surface fatigue and adhesive wear was the dominant wear mechanism. PMID:26970769

  14. Novel nano-rough polymers for cartilage tissue engineering

    Directory of Open Access Journals (Sweden)

    Balasundaram G

    2014-04-01

    Full Text Available Ganesan Balasundaram,1 Daniel M Storey,1 Thomas J Webster2,31Surfatek, Longmont, CO, USA; 2Department of Chemical Engineering, Northeastern University, Boston, MA, USA; 3Center of Excellence for Advanced Materials Research, King Abdulaziz University, Jeddah, Saudi ArabiaAbstract: This study presents an innovative method for creating a highly porous surface with nanoscale roughness on biologically relevant polymers, specifically polyurethane (PU and polycaprolactone (PCL. Nanoembossed polyurethane (NPU and nanoembossed polycaprolactone (NPCL were produced by the casting of PU and PCL over a plasma-deposited, spiky nanofeatured crystalline titanium (Ti surface. The variables used in the process of making the spiky Ti surface can be altered to change the physical properties of the spiky particles, and thus, the cast polymer substrate surface can be altered. The spiky Ti surface is reusable to produce additional nanopolymer castings. In this study, control plain PU and PCL polymers were produced by casting the polymers over a plain Ti surface (without spikes. All polymer surface morphologies were characterized using both scanning electron microscopy and atomic force microscopy, and their surface energies were measured using liquid contact angle measurements. The results revealed that both NPU and NPCL possessed a higher degree of nanometer surface roughness and higher surface energy compared with their respective unaltered polymers. Further, an in vitro study was carried out to determine chondrocyte (cartilage-producing cells functions on NPU and NPCL compared with on control plain polymers. Results of this study provided evidence of increased chondrocyte numbers on NPU and NPCL compared with their respective plain polymers after periods of up to 7 days. Moreover, the results provide evidence of greater intracellular protein production and collagen secretion by chondrocytes cultured on NPU and NPCL compared with control plain polymers. In summary

  15. Cartilage restoration technique of the hip.

    Science.gov (United States)

    Mardones, Rodrigo; Larrain, Catalina

    2016-04-01

    Hip cartilage lesions represent a diagnostic challenge and can be an elusive source of pain. Treatment may present difficulties due to localization and spherical form of the joint and is most commonly limited to excision, debridement, thermal chondroplasty and microfractures. This chapter will focus in new technologies to enhance the standard techniques. These new technologies are based in stem cells therapies; as intra-articular injections of expanded mesenchymal stem cells, mononuclear concentrate in a platelet-rich plasma matrix and expanded mesenchymal stem cells seeded in a collagen membrane. This review will discuss the bases, techniques and preliminary results obtained with the use of stem cells for the treatment of hip cartilage lesions. PMID:27026816

  16. Bioprinted Scaffolds for Cartilage Tissue Engineering.

    Science.gov (United States)

    Kang, Hyun-Wook; Yoo, James J; Atala, Anthony

    2015-01-01

    Researchers are focusing on bioprinting technology as a viable option to overcome current difficulties in cartilage tissue engineering. Bioprinting enables a three-dimensional (3-D), free-form, computer-designed structure using biomaterials, biomolecules, and/or cells. The inner and outer shape of a scaffold can be controlled by this technology with great precision. Here, we introduce a hybrid bioprinting technology that is a co-printing process of multiple materials including high-strength synthetic polymer and cell-laden hydrogel. The synthetic polymer provides mechanical support for shape maintenance and load bearing, while the hydrogel provides the biological environment for artificial cartilage regeneration. This chapter introduces the procedures for printing of a 3-D scaffold using our hybrid bioprinting technology and includes the source materials for preparation of 3-D printing. PMID:26445837

  17. Cartilage Tissue Engineering with Silk Fibroin Scaffolds Fabricated by Indirect Additive Manufacturing Technology

    Directory of Open Access Journals (Sweden)

    Chih-Hao Chen

    2014-03-01

    Full Text Available Advanced tissue engineering (TE technology based on additive manufacturing (AM can fabricate scaffolds with a three-dimensional (3D environment suitable for cartilage regeneration. Specifically, AM technology may allow the incorporation of complex architectural features. The present study involves the fabrication of 3D TE scaffolds by an indirect AM approach using silk fibroin (SF. From scanning electron microscopic observations, the presence of micro-pores and interconnected channels within the scaffold could be verified, resulting in a TE scaffold with both micro- and macro-structural features. The intrinsic properties, such as the chemical structure and thermal characteristics of SF, were preserved after the indirect AM manufacturing process. In vitro cell culture within the SF scaffold using porcine articular chondrocytes showed a steady increase in cell numbers up to Day 14. The specific production (per cell basis of the cartilage-specific extracellular matrix component (collagen Type II was enhanced with culture time up to 12 weeks, indicating the re-differentiation of chondrocytes within the scaffold. Subcutaneous implantation of the scaffold-chondrocyte constructs in nude mice also confirmed the formation of ectopic cartilage by histological examination and immunostaining.

  18. Time-Dependent Nanomechanics of Cartilage

    OpenAIRE

    Han, Lin; Frank, Eliot H.; Greene, Jacqueline J.; Lee, Hsu-Yi; Hung, Han-Hwa K.; Grodzinsky, Alan J.; Ortiz, Christine

    2011-01-01

    In this study, atomic force microscopy-based dynamic oscillatory and force-relaxation indentation was employed to quantify the time-dependent nanomechanics of native (untreated) and proteoglycan (PG)-depleted cartilage disks, including indentation modulus Eind, force-relaxation time constant τ, magnitude of dynamic complex modulus |E∗|, phase angle δ between force and indentation depth, storage modulus E′, and loss modulus E″. At ∼2 nm dynamic deformation amplitude, |E∗| increased significant...

  19. Cartilage restoration technique of the hip

    OpenAIRE

    Mardones, Rodrigo; Larrain, Catalina

    2015-01-01

    Hip cartilage lesions represent a diagnostic challenge and can be an elusive source of pain. Treatment may present difficulties due to localization and spherical form of the joint and is most commonly limited to excision, debridement, thermal chondroplasty and microfractures. This chapter will focus in new technologies to enhance the standard techniques. These new technologies are based in stem cells therapies; as intra-articular injections of expanded mesenchymal stem cells, mononuclear conc...

  20. Oxygen, nitric oxide and articular cartilage

    OpenAIRE

    Fermor, B.; Christensen, S. E.; I Youn; J M Cernanec; C M Davies; Weinberg, J. B.

    2007-01-01

    Molecular oxygen is required for the production of nitric oxide (NO), a pro-inflammatory mediator that is associated with osteoarthritis and rheumatoid arthritis. To date there has been little consideration of the role of oxygen tension in the regulation of nitric oxide production associated with arthritis. Oxygen tension may be particularly relevant to articular cartilage since it is avascular and therefore exists at a reduced oxygen tension. The superficial zone exists at approximately 6% O...

  1. Articular cartilage collagen: an irreplaceable framework?

    OpenAIRE

    Eyre, D. R.; Weis, M A; J-J Wu

    2006-01-01

    Adult articular cartilage by dry weight is two-thirds collagen. The collagen has a unique molecular phenotype. The nascent type II collagen fibril is a heteropolymer, with collagen IX molecules covalently linked to the surface and collagen XI forming the filamentous template of the fibril as a whole. The functions of collagens IX and XI in the heteropolymer are far from clear but, evidently, they are critically important since mutations in COLIX and COLXI genes can result in chondrodysplasia ...

  2. Processed bovine cartilage: an improved biosynthetic implant for contour defects

    Energy Technology Data Exchange (ETDEWEB)

    Ersek, R.A.; Hart, W.G. Jr.; Greer, D.; Beisang, A.A.; Flynn, P.J.; Denton, D.R.

    1984-05-01

    Irradiated human cartilage has been found to be a superior implant material for correction of contour defects; however, availability problems have prevented this material from gaining wide acceptance. Implantation of processed irradiated bovine cartilage in primates and rabbits, as described here, provides strong evidence that this material performs like irradiated allograft cartilage antigenically and has certain cosmetic advantages over allograft cartilage. Our studies in primates have shown that there is no systemically measurable antibody-antigen reaction, either cellular or noncellular, to irradiated processed bovine cartilage. Neither primary nor second-set provocative implantations produced any measurable rejection. In rabbits, composite grafts of two pieces of irradiated bovine cartilage adjacent to each other were also well tolerated, with no measurable absorption and with capsule formation typical of a foreign body reaction to an inert object.

  3. Respiratory syncytial virus infection in sheep bronchial explants is associated with enhanced ETB receptor-mediate contractile functional and autoradiographic studies

    International Nuclear Information System (INIS)

    Full text: Respiratory syncytial virus (RSV) is an important precipitant of asthma in children. The impact of RSV infection on endothelin (ET) receptor density and function in airways is unknown. In the present study, sheep bronchial rings were maintained as explants in culture for up to 48 h. During this time, both the structural integrity of the epithelium and carbachol responsiveness were preserved. Bronchial rings in culture were exposed to non-infected culture medium or to RSV (1/50 TCID50) for 0, 24 and 48 h which caused marked damage to and loss of the epithelium. RSV infection did not significantly alter responsiveness to ET-1 at either 24 (Control EC40 = 102 nM, 95% confidence limits, 76-138 nM vs RSV EC40 = 66 nM, 95% confidence limits, 48-91 nM, n=5-6, P>0.05) or 48 h (Control EC40 35 nM, 95% confidence limits, 19-66 nM vs RSV EC40 = 55 nM, 95% confidence limits, 32-93 nM, n=8, P>0.05). As seen previously (Goldie et al., 1994), sarafotoxin S6c (StxS6c, ETB-selective) did not cause contraction in non-infected sheep bronchial explants. In contrast, StxS6c (300 nM) increased tone by 8±3% carbachol Emax (n=6-8) in explants exposed to RSV for 24 or 48 h. Light microscopic autoradiography was used to determine the relative distribution of ETA and ETB receptors using [125I]-ET-1, BQ-123 (ETA-selective) and StxS6c. Sheep airway smooth muscle contains a homogeneous population of ETA receptors (Goldie et al., 1994). Since StxS6c caused significant contraction in RSV-infected bronchial explants, it was surprising that autoradiographic techniques failed to detect airway smooth muscle ETB receptors in these preparations. It is likely that ETB receptors fell below the level of detection of autoradiography. The significant StxS6c-induced contraction of sheep bronchi suggests the novel expression of ETB receptors triggered by RSV which might be relevant to RSV-associated asthma. Copyright (2001) Australasian Society of Clinical and Experimental Pharmacologists and

  4. Time-dependent nanomechanics of cartilage.

    Science.gov (United States)

    Han, Lin; Frank, Eliot H; Greene, Jacqueline J; Lee, Hsu-Yi; Hung, Han-Hwa K; Grodzinsky, Alan J; Ortiz, Christine

    2011-04-01

    In this study, atomic force microscopy-based dynamic oscillatory and force-relaxation indentation was employed to quantify the time-dependent nanomechanics of native (untreated) and proteoglycan (PG)-depleted cartilage disks, including indentation modulus E(ind), force-relaxation time constant τ, magnitude of dynamic complex modulus |E(∗)|, phase angle δ between force and indentation depth, storage modulus E', and loss modulus E″. At ∼2 nm dynamic deformation amplitude, |E(∗)| increased significantly with frequency from 0.22 ± 0.02 MPa (1 Hz) to 0.77 ± 0.10 MPa (316 Hz), accompanied by an increase in δ (energy dissipation). At this length scale, the energy dissipation mechanisms were deconvoluted: the dynamic frequency dependence was primarily governed by the fluid-flow-induced poroelasticity, whereas the long-time force relaxation reflected flow-independent viscoelasticity. After PG depletion, the change in the frequency response of |E(∗)| and δ was consistent with an increase in cartilage local hydraulic permeability. Although untreated disks showed only slight dynamic amplitude-dependent behavior, PG-depleted disks showed great amplitude-enhanced energy dissipation, possibly due to additional viscoelastic mechanisms. Hence, in addition to functioning as a primary determinant of cartilage compressive stiffness and hydraulic permeability, the presence of aggrecan minimized the amplitude dependence of |E(∗)| at nanometer-scale deformation. PMID:21463599

  5. Irradiated homologous costal cartilage for augmentation rhinoplasty

    Energy Technology Data Exchange (ETDEWEB)

    Lefkovits, G. (Lenox Hill Hospital, New York, NY (USA))

    1990-10-01

    Although the ideal reconstructive material for augmentation rhinoplasty continues to challenge plastic surgeons, there exists no report in the literature that confines the use of irradiated homologous costal cartilage, first reported by Dingman and Grabb in 1961, to dorsal nasal augmentation. The purpose of this paper is to present a retrospective analysis of the author's experience using irradiated homologous costal cartilage in augmentation rhinoplasty. Twenty-seven dorsal nasal augmentations were performed in 24 patients between 16 and 49 years of age with a follow-up ranging from 1 to 27 months. Good-to-excellent results were achieved in 83.3% (20 of 24). Poor results requiring revision were found in 16.7% (4 of 24). Complication rates included 7.4% infection (2 of 27) and 14.8% warping (4 of 27). The resorption rate was zero. These results compare favorably with other forms of nasal augmentation. Advantages and disadvantages of irradiated homologous costal cartilage are discussed.

  6. Interleukin 17A evoked mucosal damage is attenuated by cannabidiol and anandamide in a human colonic explant model.

    Science.gov (United States)

    Harvey, B S; Sia, T C; Wattchow, D A; Smid, S D

    2014-02-01

    Interleukin 17A (IL-17A) is a cytokine linked to inflammatory bowel disease. We investigated IL-17A expression in human colonic mucosa, whether IL-17A can elicit colonic mucosal damage in a human explant model and modulate gastrointestinal epithelial permeability in cell culture. We also tested if select cannabinoid ligands, shown to be protective in colitis models could attenuate damage caused by IL-17A. In addition, the ability of pro-inflammatory cytokines TNF-α and IL-1β to modulate levels of IL-17A in the explant colitis model was also explored. IL-17A incubation caused significant mucosal epithelial and crypt damage which were attenuated following hydrocortisone treatment, and also reduced following anandamide or cannabidiol incubation. IL-17A-evoked mucosal damage was also associated with an increase in matrix metalloprotease activity. However, IL-17A did not induce any significant changes in epithelial permeability in confluent Caco-2 cell monolayers over a 48h incubation period. IL-17A was located predominantly in human mucosal epithelium together with IL-17C, but both IL-17A and IL-17C were also expressed in the lamina propria and submucosa. Incubation of human colonic mucosal tissue or Caco-2 cells with pro-inflammatory cytokines TNF-α and IL-1β however did not alter IL-17A expression. These results indicate IL-17A has a widespread distribution in the human colon and the capacity to elicit mucosal damage which can be attenuated by cannabinoid ligands. PMID:24238999

  7. Long-term gene therapy with Del1 fragment using nonviral vectors in mice with explanted tumors.

    Science.gov (United States)

    Kitano, Hisataka; Mamiya, Atsushi; Ishikawa, Tomomi; Egoshi, Kayo; Kokubun, Shinichiro; Hidai, Chiaki

    2016-01-01

    Cancer gene therapy using nonviral vectors is useful for long periods of treatment because such vectors are both safe and inexpensive, and thus can be used repeatedly. It has been reported that gene therapy with an E3C1 fragment of Del1 in a mouse explanted tumor model improved prognosis. The present study aimed to analyze the long-term effects of repeated non-viral gene transfer of E3C1. Mice with explanted tumors of SCCKN cells, a human squamous carcinoma, were treated with a plasmid encoding E3C1. Plasmids were injected locally every week using a transfection reagent. Control mice treated with mock DNA started to be euthanized on day 18, because the tumors had grown to over 15% of the body weight, and all of them had died by day 43. On the other hand, the tumors in two of ten mice treated with E3C1 had disappeared. The other eight mice started to be euthanized on day 46 and eight of ten mice had been euthanized by day 197. After 18 days of therapy, the tumor volume of control mice was 2,804±829 mm(3) and that of the E3C1 mice was 197±159 mm(3). Histochemical studies showed enhanced apoptosis in the E3C1-treated tumors, as compared with controls. Changes in cell morphology and decreased polymerized actin induced by E3C1 indicated disturbed cell adhesion to the matrix. In in vitro studies of SCCKN cells, prolonged administration of an E3C1 recombinant protein to cultured cells reduced adhesion-independent growth of cancer cells, as compared with control cells. These data suggest that E3C1 treatment induces anoikis. PMID:26889088

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

    Directory of Open Access Journals (Sweden)

    Elisa Cristina Soares de Carvalho Alves

    2004-05-01

    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.

  9. Comprehensive profiling of cartilage extracellular matrix formation and maturation using sequential extraction and label-free quantitative proteomics.

    Science.gov (United States)

    Wilson, Richard; Diseberg, Anders F; Gordon, Lavinia; Zivkovic, Snezana; Tatarczuch, Liliana; Mackie, Eleanor J; Gorman, Jeffrey J; Bateman, John F

    2010-06-01

    Articular cartilage is indispensable for joint function but has limited capacity for self-repair. Engineering of neocartilage in vitro is therefore a major target for autologous cartilage repair in arthritis. Previous analysis of neocartilage has targeted cellular organization and specific molecular components. However, the complexity of extracellular matrix (ECM) development in neocartilage has not been investigated by proteomics. To redress this, we developed a mouse neocartilage culture system that produces a cartilaginous ECM. Differential analysis of the tissue proteome of 3-week neocartilage and 3-day postnatal mouse cartilage using solubility-based protein fractionation targeted components involved in neocartilage development, including ECM maturation. Initially, SDS-PAGE analysis of sequential extracts revealed the transition in protein solubility from a high proportion of readily soluble (NaCl-extracted) proteins in juvenile cartilage to a high proportion of poorly soluble (guanidine hydrochloride-extracted) proteins in neocartilage. Label-free quantitative mass spectrometry (LTQ-Orbitrap) and statistical analysis were then used to filter three significant protein groups: proteins enriched according to extraction condition, proteins differentially abundant between juvenile cartilage and neocartilage, and proteins with differential solubility properties between the two tissue types. Classification of proteins differentially abundant between NaCl and guanidine hydrochloride extracts (n = 403) using bioinformatics revealed effective partitioning of readily soluble components from subunits of larger protein complexes. Proteins significantly enriched in neocartilage (n = 78) included proteins previously not reported or with unknown function in cartilage (integrin-binding protein DEL1; coiled-coil domain-containing protein 80; emilin-1 and pigment epithelium derived factor). Proteins with differential extractability between juvenile cartilage and neocartilage

  10. Cartilage-Specific Near-Infrared Fluorophores for Biomedical Imaging.

    Science.gov (United States)

    Hyun, Hoon; Owens, Eric A; Wada, Hideyuki; Levitz, Andrew; Park, GwangLi; Park, Min Ho; Frangioni, John V; Henary, Maged; Choi, Hak Soo

    2015-07-20

    A novel class of near-infrared fluorescent contrast agents was developed. These agents target cartilage with high specificity and this property is inherent to the chemical structure of the fluorophore. After a single low-dose intravenous injection and a clearance time of approximately 4 h, these agents bind to all three major types of cartilage (hyaline, elastic, and fibrocartilage) and perform equally well across species. Analysis of the chemical structure similarities revealed a potential pharmacophore for cartilage targeting. Our results lay the foundation for future improvements in tissue engineering, joint surgery, and cartilage-specific drug development. PMID:26095685

  11. Cutaneous Squamous Cell Carcinoma with Invasion through Ear Cartilage

    Directory of Open Access Journals (Sweden)

    Julie Boisen

    2016-01-01

    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.

  12. Secondary cartilage revealed in a non-avian dinosaur embryo.

    Science.gov (United States)

    Bailleul, Alida M; Hall, Brian K; Horner, John R

    2013-01-01

    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. PMID:23418610

  13. 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.

  14. Development and potential of a biomimetic chitosan/type Ⅱ collagen scaffold for cartilage tissue engineering

    Institute of Scientific and Technical Information of China (English)

    SHI De-hai; CAI Dao-zhang; ZHOU Chang-ren; RONG Li-min; WANG Kun; XU Yi-chun

    2005-01-01

    Background Damaged articular cartilage has very limited capacity for spontaneous healing. Tissue engineering provides a new hope for functional cartilage repair. Creation of an appropriate cell carrier is one of the critical steps for successful tissue engineering. With the supposition that a biomimetic construct might promise to generate better effects, we developed a novel composite scaffold and investigated its potential for cartilage tissue engineering. Methods Chitosan of 88% deacetylation was prepared via a modified base reaction procedure. A freeze-drying process was employed to fabricate a three-dimensional composite scaffold consisting of chitosan and type Ⅱcollagen. The scaffold was treated with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide. Ultrastructure and tensile strength of the matrix were carried out to assess its physico-chemical properties. After subcutaneous implantation in rabbits, its in vivo biocompatibility and degradability of the scaffold were determined. Its capacity to sustain chondrocyte growth and biosynthesis was evaluated through cell-scaffold co-culture in vitro. Results The fabricated composite matrix was porous and sponge-like with interconnected pores measuring from 100-250 μm in diameter. After cross-linking, the scaffold displayed enhanced tensile strength. Subcutaneous implantation results indicated the composite matrix was biocompatible and biodegradable. In intro cell-scaffold culture showed the scaffold sustained chondrocyte proliferation and differentiation, and maintained the spheric chondrocytic phenotype. As indicated by immunohistochemical staining, the chondrocytes synthesized type Ⅱ collagen. Conclusions Chitosan and type Ⅱ collagen can be well blended and developed into a porous 3-D biomimetic matrix. Results of physico-chemical and biological tests suggest the composite matrix satisfies the constraints specified for a tissue-engineered construct and may be used as a chondrocyte

  15. A HYBRID SCAFFOLD OF POLY(LACTIDE-CO-GLYCOLIDE) SPONGE FILLED WITH FIBRIN GEL FOR CARTILAGE TISSUE ENGINEERING

    Institute of Scientific and Technical Information of China (English)

    Wei Wang; Dan Li; Mei-cong Wang; Yang-lin Li; Chang-you Gao

    2011-01-01

    The poly(lactide-co-glycolide) (PLGA) sponge fabricated by a gelatin porogen leaching method was filled with fibrin gel to obtain a hybrid scaffold for chondrocytes culture in vitro. The fibrin gel evenly distributed in the hybrid scaffold with visible fibrinogen fibers after drying. In vitro culture it was found that in the hybrid scaffold the chondrocytes distributed more evenly and kept a round morphology as that in the normal cartilage. Although the chondrocytes seeded in the control PLGA sponges showed similar proliferation behavior with that in the hybrid scaffolds, they were remarkably elongated, forming a fibroblast-like morphology. Moreover, a larger amount of glycosaminoglycans was secreted in the hybrid scaffolds than that in the PLGA sponges after in vitro culture of chondrocytes for 4 weeks. The results suggest that the fibrin/PLGA hybrid scaffold may be favorably applied for cartilage tissue engineering.

  16. Direct axillary shoot regeneration from the mature seed explant of the hairy vetch (Vicia villosa Roth)

    OpenAIRE

    Aasim Muhammad; Sahin-Demirbag Nurdan; Khawar Mahmood Khalid; Kendir Hayrettin; Özcan Sebahattin

    2011-01-01

    The hairy vetch (Vicia villosa Roth) is a climbing, prostrate or trailing legume grown as forage.It fixes atmospheric nitrogen, reduces soil erosion and provides an instant mulch. Multiple axillary shoot regeneration from a mature seed explant (zygotic embryo with two cotyledons) was obtained on MS medium containing 0.05 - 1.6 mg/l TDZ with or without 0.10 mg/l IBA. The frequency (%) of shoot regeneration ranged from 45.83-75.00% with a maximum number of 28.6 shoots per explant on MS me...

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

    OpenAIRE

    Amir Ghaffar SHAHRIARI; Bagheri, Abdolreza; Sharifi, Ahmad; Nasrin MOSHTAGHI

    2012-01-01

    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 ...

  18. Early somatic embryogenesis in Heliconia chartacea Lane ex Barreiros cv. Sexy Pink ovary section explants

    Directory of Open Access Journals (Sweden)

    Cláudia Ulisses

    2010-02-01

    Full Text Available The present work evaluated the development of embryogenic callus from transversal ovary sections. The experiments were carried out under two experimental regimes using combinations of IAA (0; 5.71; 8.56; 11.42; 14.27μM and 2,4-D (0; 13.57; 18.10; 22.62μM or combinations of 2,4-D with BA (0; 4.43; 6.65; 8.87; 11.09μM. Assessments were made of anatomical aspects of the callus and for the presence of embryogenic structures using cytochemical and histological analyses and stereomicroscopic and scanning electronic microscopic observations. Treatments with 2,4-D and IAA produced friable calluses demonstrating cellular acquisition of morphogenetic competence as well as the formation of pro-embryogenic sectors. The expression of embryogenic program could be observed, with proembryogenic cell clusters developing into globular embryos. These results offer the possibility of using new types of explants for culturing helicons that avoid the growth of endophytic bacteria.Este trabalho teve como objetivo avaliar a resposta de secções transversais de ovários e o desenvolvimento de calos embriogênicos. O experimento constou de dois ensaios. No primeiro avaliou-se combinações entre AIA (0; 5.71; 8.56; 11.42; 14.27μM e 2,4-D (0; 13.57; 18.10; 22.62μM e no segundo avaliou-se as concentrações de 2,4-D supracitadas, combinadas com concentrações de BA (0; 4.43; 6.65; 8.87; 11.09μM. Os calos formados foram avaliados quanto à presença de estruturas embriogênicas utilizando-se estereomicroscópio, microscópio eletrônico de varredura, além de análises citoquímicas e histológicas. Combinações entre 2,4-D e AIA induziram a formação de calos friáveis com setores pró-embriogênicos, refletindo a aquisição de competência morfogenética. Posteriormente foi observada a expressão do programa embriogênico quando massas pró-embriogências desenvolveram-se formando embriões somáticos. Esses resultados apresentam uma alternativa para a utiliza

  19. Plant regeneration from cotyledonary explants of Eucalyptus camaldulensis dehn and histological study of organogenesis in vitro

    Directory of Open Access Journals (Sweden)

    Roberson Dibax

    2010-04-01

    Full Text Available The present work aimed at regenerating plants of Eucalyptus camaldulensis from the cotyledonary explants and describing the anatomy of the tissues during callogenesis and organogenesis processes, in order to determine the origin of the buds. The cotyledonary leaves of E. camaldulensis were cultured in Murashige and Skoog (MS, WPM and JADS media supplemented with 2.7 µM NAA and 4.44 µM BAP. The best results for bud regeneration were obtained on MS and WPM media (57.5 and 55% of calluses formed buds, respectively. Shoot elongation and rooting (80% were obtained on MS/2 medium (with half-strength salt concentration with 0.2% activated charcoal. Acclimatization was performed in the growth chamber for 48 h and then the plants were transferred to a soil:vermiculite mixture and cultured in a greenhouse. Histological studies revealed that the callogenesis initiated in palisade parenchyma cells and that the adventitious buds were formed from the calluses, indicating indirect organogenesis.Este trabalho teve como objetivo a obtenção de plantas de Eucalyptus camaldulensis a partir de folhas cotiledonares e o estudo da anatomia dos tecidos durante a calogênese e organogênese para determinar a origem das gemas. Folhas cotiledonares foram cultivadas em meios de cultura MS, WPM e JADS suplementados com 2,7 µM de ANA e 4,44 µM de BAP. Os melhores resultados para a regeneração de gemas foram obtidos com os meios MS e WPM. Para o alongamento e enraizamento, o meio de cultura MS/2 contendo 0,2% de carvão ativado apresentou-se eficiente para ambas as etapas. A aclimatização foi realizada mediante a abertura dos frascos na sala de crescimento por 48 horas, seguido da transferência para casa-de-vegetação com nebulização intermitente. Estudos histológicos foram conduzidos e revelaram que a calogênese teve início nas células do parênquima paliçádico e que as gemas adventícias formaram-se a partir dos calos, indicando a organogênese indireta.

  20. FT-IR Microspectroscopy of Rat Ear Cartilage.

    Directory of Open Access Journals (Sweden)

    Benedicto de Campos Vidal

    Full Text Available Rat ear cartilage was studied using Fourier transform-infrared (FT-IR microspectroscopy to expand the current knowledge which has been established for relatively more complex cartilage types. Comparison of the FT-IR spectra of the ear cartilage extracellular matrix (ECM with published data on articular cartilage, collagen II and 4-chondroitin-sulfate standards, as well as of collagen type I-containing dermal collagen bundles (CBs with collagen type II, was performed. Ear cartilage ECM glycosaminoglycans (GAGs were revealed histochemically and as a reduction in ECM FT-IR spectral band heights (1140-820 cm-1 after testicular hyaluronidase digestion. Although ear cartilage is less complex than articular cartilage, it contains ECM components with a macromolecular orientation as revealed using polarization microscopy. Collagen type II and GAGs, which play a structural role in the stereo-arrangement of the ear cartilage, contribute to its FT-IR spectrum. Similar to articular cartilage, ear cartilage showed that proteoglycans add a contribution to the collagen amide I spectral region, a finding that does not recommend this region for collagen type II quantification purposes. In contrast to articular cartilage, the symmetric stretching vibration of -SO3- groups at 1064 cm-1 appeared under-represented in the FT-IR spectral profile of ear cartilage. Because the band corresponding to the asymmetric stretching vibration of -SO3- groups (1236-1225 cm-1 overlapped with that of amide III bands, it is not recommended for evaluation of the -SO3- contribution to the FT-IR spectrum of the ear cartilage ECM. Instead, a peak (or shoulder at 1027-1016 cm-1 could be better considered for this intent. Amide I/amide II ratios as calculated here and data from the literature suggest that protein complexes of the ear cartilage ECM are arranged with a lower helical conformation compared to pure collagen II. The present results could motivate further studies on this tissue

  1. FT-IR Microspectroscopy of Rat Ear Cartilage.

    Science.gov (United States)

    Vidal, Benedicto de Campos; Mello, Maria Luiza S

    2016-01-01

    Rat ear cartilage was studied using Fourier transform-infrared (FT-IR) microspectroscopy to expand the current knowledge which has been established for relatively more complex cartilage types. Comparison of the FT-IR spectra of the ear cartilage extracellular matrix (ECM) with published data on articular cartilage, collagen II and 4-chondroitin-sulfate standards, as well as of collagen type I-containing dermal collagen bundles (CBs) with collagen type II, was performed. Ear cartilage ECM glycosaminoglycans (GAGs) were revealed histochemically and as a reduction in ECM FT-IR spectral band heights (1140-820 cm-1) after testicular hyaluronidase digestion. Although ear cartilage is less complex than articular cartilage, it contains ECM components with a macromolecular orientation as revealed using polarization microscopy. Collagen type II and GAGs, which play a structural role in the stereo-arrangement of the ear cartilage, contribute to its FT-IR spectrum. Similar to articular cartilage, ear cartilage showed that proteoglycans add a contribution to the collagen amide I spectral region, a finding that does not recommend this region for collagen type II quantification purposes. In contrast to articular cartilage, the symmetric stretching vibration of -SO3- groups at 1064 cm-1 appeared under-represented in the FT-IR spectral profile of ear cartilage. Because the band corresponding to the asymmetric stretching vibration of -SO3- groups (1236-1225 cm-1) overlapped with that of amide III bands, it is not recommended for evaluation of the -SO3- contribution to the FT-IR spectrum of the ear cartilage ECM. Instead, a peak (or shoulder) at 1027-1016 cm-1 could be better considered for this intent. Amide I/amide II ratios as calculated here and data from the literature suggest that protein complexes of the ear cartilage ECM are arranged with a lower helical conformation compared to pure collagen II. The present results could motivate further studies on this tissue under

  2. Sugarcane Juice as an Alternative Carbon Source for in vitro Culture of Plantains and Bananas

    OpenAIRE

    G. Addae; J.W. Tachie-Menson; J.N. Buah; Asare, P.

    2011-01-01

    The suitability of sugarcane juice for the in vitro culture of bananas and plantains was investigated in this study. Two concentrations of sugar cane juice 5, 10% and 30 g L-1 laboratory sucrose were used. There was also a control medium without carbon source. Explants were prepared from field grown sword suckers. The preparation of the explants involved, sterilization with 70% ethanol, followed by 1% Sodium hypochlorite+1% tween 20. The explants were cultured at 26°C, under 16 h photoperiod,...

  3. Meristem Culture of Aloe vera L

    International Nuclear Information System (INIS)

    Shoot tips of Aloe vera L were excised and inoculated in basal and modified MS (Murashige and Skoog, 1962) medium. Growth regulators and growth factors were supplemented to the basal medium. Development of explants, initiation of callus and multiplication of shoots in culture were recorded.

  4. Deficient Mechanical Activation of Anabolic Transcripts and Post-Traumatic Cartilage Degeneration in Matrilin-1 Knockout Mice

    Science.gov (United States)

    Chen, Yupeng; Cossman, Jack; Jayasuriya, Chathuraka T.; Li, Xin; Guan, Yingjie; Fonseca, Vera; Yang, Kun; Charbonneau, Cherie; Yu, Hongchuan; Kanbe, Katsuaki; Ma, Peter; Darling, Eric; Chen, Qian

    2016-01-01

    Matrilin-1 (Matn1), a cartilage-specific peri-cellular and extracellular matrix (ECM) protein, has been hypothesized to regulate ECM interactions and transmit mechanical signals in cartilage. Since Matn1 knock-out (Matn1-/-) mice exhibit a normal skeleton, its function in vivo is unclear. In this study, we found that the anabolic Acan and Col2a transcript levels were significantly higher in wildtype (Matn1+/+) mouse cartilage than that of MATN1-/- mice in vivo. However, such difference was not observed between Matn1+/+ and MATN1-/- chondrocytes cultured under stationary conditions in vitro. Cyclic loading significantly stimulated Acan and Col2a transcript levels in Matn1+/+ but not in MATN1-/- chondrocytes. This suggests that, while Matn1+/+ chondrocytes increase their anabolic gene expression in response to mechanical loading, the MATN1-/- chondrocytes fail to do so because of the deficiency in mechanotransduction. We also found that altered elastic modulus of cartilage matrix in Matn1-/- mice, suggesting the mechanotransduction has changed due to the deficiency of Matn1. To understand the impact of such deficiency on joint disease, mechanical loading was altered in vivo by destabilization of medial meniscus. While Matn1+/+ mice exhibited superficial fissures and clefts consistent with mechanical damage to the articular joint, Matn1-/- mice presented more severe cartilage lesions characterized by proteoglycan loss and disorganization of cells and ECM. This suggests that Matn1 deficiency affects pathogenesis of post-traumatic osteoarthritis by failing to up-regulate anabolic gene expression. This is the first demonstration of Matn1 function in vivo, which suggests its protective role in cartilage degeneration under altered mechanical environment. PMID:27270603

  5. Computational model for the analysis of cartilage and cartilage tissue constructs.

    Science.gov (United States)

    Smith, David W; Gardiner, Bruce S; Davidson, John B; Grodzinsky, Alan J

    2016-04-01

    We propose a new non-linear poroelastic model that is suited to the analysis of soft tissues. In this paper the model is tailored to the analysis of cartilage and the engineering design of cartilage constructs. The proposed continuum formulation of the governing equations enables the strain of the individual material components within the extracellular matrix (ECM) to be followed over time, as the individual material components are synthesized, assembled and incorporated within the ECM or lost through passive transport or degradation. The material component analysis developed here naturally captures the effect of time-dependent changes of ECM composition on the deformation and internal stress states of the ECM. For example, it is shown that increased synthesis of aggrecan by chondrocytes embedded within a decellularized cartilage matrix initially devoid of aggrecan results in osmotic expansion of the newly synthesized proteoglycan matrix and tension within the structural collagen network. Specifically, we predict that the collagen network experiences a tensile strain, with a maximum of ~2% at the fixed base of the cartilage. The analysis of an example problem demonstrates the temporal and spatial evolution of the stresses and strains in each component of a self-equilibrating composite tissue construct, and the role played by the flux of water through the tissue. Copyright © 2013 John Wiley & Sons, Ltd. PMID:23784936

  6. Computational model for the analysis of cartilage and cartilage tissue constructs

    Science.gov (United States)

    Smith, David W.; Gardiner, Bruce S.; Davidson, John B.; Grodzinsky, Alan J.

    2013-01-01

    We propose a new non-linear poroelastic model that is suited to the analysis of soft tissues. In this paper the model is tailored to the analysis of cartilage and the engineering design of cartilage constructs. The proposed continuum formulation of the governing equations enables the strain of the individual material components within the extracellular matrix (ECM) to be followed over time, as the individual material components are synthesized, assembled and incorporated within the ECM or lost through passive transport or degradation. The material component analysis developed here naturally captures the effect of time-dependent changes of ECM composition on the deformation and internal stress states of the ECM. For example, it is shown that increased synthesis of aggrecan by chondrocytes embedded within a decellularized cartilage matrix initially devoid of aggrecan results in osmotic expansion of the newly synthesized proteoglycan matrix and tension within the structural collagen network. Specifically, we predict that the collagen network experiences a tensile strain, with a maximum of ~2% at the fixed base of the cartilage. The analysis of an example problem demonstrates the temporal and spatial evolution of the stresses and strains in each component of a self-equilibrating composite tissue construct, and the role played by the flux of water through the tissue. PMID:23784936

  7. 99mTc-labeled chondroitin sulfate-uptake by chondrocytes and cartilage. Potential agent for osteoarthritis imaging?

    International Nuclear Information System (INIS)

    Aim: Chondroitin sulfate (CS) is an endogenous component of cartilage proteoglycan which could monitor osteoarthritic cartilage degradation after radiolabeling. This substance is used in the treatment of human osteoarthritis as a slow acting symptomatic drug (CONDROSULF; Sanova Pharma, Vienna; Ibsa, Switzerland). Material and Methods: Radiolabeling of CS was performed using 99mTcO4-/stannous chloride in 0.50 M sodium acetate buffer at pH 5.0. The quality control of the tracer was performed using ITLC-SG chromatography and 0.2 M saline in 10% ethanol as solvent to detect colloid content. Aluminium oxide IB-F TLC-sheets and ethanol as solvent were used to estimate free pertechnetate. For uptake studies cultured human chondrocytes and age-matched cartilage were used. Uptake of the tracer in chondrocytes was studied in monolayer and in suspension cultures at 370C. Uptake was monitored for a total of 120-180 minutes, samples being drawn every 10 minutes. Because the commercially available drug Condrosulf contains calcium stearate as additive to improve the resorption of the drug, we investigated also the uptake with and without additive. Results: The tracer was stable over 6h period after labeling (95% of the radiochemical purity). In plasma the stability was lower amounting to 75%. Viability of chondrocytes after incubation with either CS-preparation was found by trypan blue exclusion to be above 95 %. Uptake of the tracer performed in monolayer ± additives was low and amounted to 0.5%±0.05%, n=6. The cells were saturated already after an incubation interval of 10 minutes. In suspension cultures a maximal uptake of 1.0%±0.1%, n=6 and 5.9%±0.65%, n=6 was found, without and with additives, respectively, the saturation was achieved after 100 min. Thus, not only the resorption of the drug is enhanced by Ca-stearate, but also uptake increases in presence of this additive. Using human rib cartilage the uptake of the tracer was much higher amounting to 4.9%±2.3%, n=6

  8. An Efficient Protocol for Plantlet Regeneration via Direct Organogenesis by Using Nodal Segments from Embryo-Cultured Seedlings of Cinnamomum camphora L.

    Science.gov (United States)

    Du, Li; Li, Yongpeng; Yao, Yao; Zhang, Liwei

    2015-01-01

    A simple and efficient plantlet regeneration protocol via direct organogenesis was established for camphor tree (Cinnamomum camphora L.). Stem segments with one node (SN explants) from embryo-cultured seedlings (EC seedlings) were used as explants. Murashige and Skoog (MS) medium supplemented with 0.5 mg/L 2, 4-dichlorophenoxyacetic acid and 2.0 mg/L 6-benzyladenine was used to induce cotyledonary embryo germination. This medium was also used for EC seedlings propagation and adventitious bud induction from SN explants. Regenerated plantlets were cultured on hormone-free MS medium for elongation and root induction. The regeneration capability of SN explants was compared by using EC seedling lines established in this research. EC seedling line EL6 exhibited the highest adventitious bud induction frequency (91.7%) and the highest number of buds per responding explant (5.2), which was considered as the most efficient EC seedling line for further gene transformation research. PMID:25962170

  9. Phosphorylation of proteoglycans from human articular cartilage

    International Nuclear Information System (INIS)

    Previous studies have shown that sulfated proteoglycans from human articular and epiphyseal cartilage were phosphorylated. These macromolecules contribute to the stiffness and resiliency of this tissue. We demonstrate here that the phosphate moieties are an integral part of proteoglycan subunits. Specifically, evidence is presented which indicates that proteoglycan monomers contain 3 to 4 phosphate moieties per core protein and that these appear to exist as phosphoserine residues. Furthermore, the data illustrate that human articular cartilage also contains more than 20 different phosphoproteins, some of which are closely associated with proteoglycan aggregates. Proteoglycan subunits were purified from extracts of articular cartilage or from media fractions which had been used to label tissue specimens with 32P-orthophosphate. Chemical and radiographic analyses revealed that the phosphate concentration with respect to sulfate and uronic acid content remained constant when purified proteoglycan monomers were subjected to equilibrium ultracentrifugation and size-exclusion chromatography. That the phosphate moieties were bound to proteoglycan monomers via monoester linkages was indicated by the release of 32P-orthophosphate from proteoglycan subunits incubated under mild alkaline conditions or reacted with acid or alkaline phosphatases. Identification of serine residues in the core protein as the sites of phosphorylation was made by autoradiography of thin layer plates on which hydrolyzed samples of purified 32P-proteoglycan subunits had been subjected to 2-dimensional electrophoresis/chromatography. Quantification of 3 to 4 phosphate moieties per core protein of 200,000 daltons was made by chemical analysis of inorganic phosphate released from proteoglycans by acid hydrolysis

  10. The Application of Polysaccharide Biocomposites to Repair Cartilage Defects

    Directory of Open Access Journals (Sweden)

    Feng Zhao

    2014-01-01

    Full Text Available Owing to own nature of articular cartilage, it almost has no self-healing ability once damaged. Despite lots of restore technologies having been raised in the past decades, no repair technology has smoothly substituted for damaged cartilage using regenerated cartilage tissue. The approach of tissue engineering opens a door to successfully repairing articular cartilage defects. For instance, grafting of isolated chondrocytes has huge clinical potential for restoration of cartilage tissue and cure of chondral injury. In this paper, SD rats are used as subjects in the experiments, and they are classified into three groups: natural repair (group A, hyaluronic acid repair (group B, and polysaccharide biocomposites repair (hyaluronic acid hydrogel containing chondrocytes, group C. Through the observation of effects of repairing articular cartilage defects, we concluded that cartilage repair effect of polysaccharide biocomposites was the best at every time point, and then the second best was hyaluronic acid repair; both of them were better than natural repair. Polysaccharide biocomposites have good biodegradability and high histocompatibility and promote chondrocytes survival, reproduction, and spliting. Moreover, polysaccharide biocomposites could not only provide the porous network structure but also carry chondrocytes. Consequently hyaluronic acid-based polysaccharide biocomposites are considered to be an ideal biological material for repairing articular cartilage.

  11. MORPHOMETRIC STUDY OF THYROID CARTILAGES IN WESTERN INDIA

    Directory of Open Access Journals (Sweden)

    Mohini M.Joshi

    2015-06-01

    Full Text Available Background: Morphometrical evaluation of the larynx has always been interesting for both morphologists and the physicians. A good understanding of the anatomy and the knowledge of variations in the laryngeal cartilages is important Objective: Objective of the present study was to collect exact and reliable morphometric data of thyroid cartilage in adult human larynx of regional population. Methods: The totals of 50 thyroid cartilage specimens were studied. The cartilages were preserved in 5% formalin. The measurements were taken with the help of Digital Vernier Caliper. The cartilages were weighed on Single pan electronic balance. For each of the parameters, the mean, standard deviation (S.D. and range was calculated. Results: Mean depth of superior thyroid notch was 9.7± 3.36 mm. Asymmetry between the length of superior horn of thyroid cartilages in left and right sides can be seen, but difference was not statistically significant (p>0.05. It is observed that inner thyroid angle varies from 55 to 1040 and outer thyroid angle varies from 53 to 990. In present study mean weight of thyroid cartilage was 6.70±1.55 grams. Conclusions: A fair amount of intersubject variability in the dimensions was observed. Bilateral asymmetry, though present in majority of specimens, was insignificant. Various dimensions of thyroid cartilages are smaller as compared to the western population.

  12. Crosstalk between cartilage and bone: when bone cytokines matter.

    Science.gov (United States)

    Funck-Brentano, Thomas; Cohen-Solal, Martine

    2011-04-01

    The cartilage damage which characterizes osteoarthritis is often accompanied by bone lesions. Joint integrity results from the balance in the physiological interactions between bone and cartilage. Several local factors regulate the physiological remodeling of cartilage, the disequilibrium of these leading to a higher cartilage catabolism. Several cytokines secreted by bone cells can induce chondrocyte differentiation, which suggests their role in the dialogue between both cells. Accumulative in vivo evidence shows that increased bone resorption occurs at an early stage in the development of osteoarthritis and that blocking bone-resorbing cytokines prevents cartilage damage, confirming the role of bone factors in the crosstalk of both tissues. Recently, molecules of the Wnt pathway have emerged as key regulators of bone and cartilage. Activation of Wnt/βcatenin induces an imbalance in cartilage homeostasis, and agonists/antagonists of Wnt are potential candidates for this interaction. This review will summarize what is known about the contribution of bone cytokines to the physiological remodeling of cartilage and in the pathophysiology of osteoarthritis. PMID:21596615

  13. Endogenous Cartilage Repair by Recruitment of Stem Cells.

    Science.gov (United States)

    Im, Gun-Il

    2016-04-01

    Articular cartilage has a very limited capacity for repair after injury. The adult body has a pool of stem cells that are mobilized during injury or disease. These cells exist inside niches in bone marrow, muscle, adipose tissue, synovium, and other connective tissues. A method that mobilizes this endogenous pool of stem cells will provide a less costly and less invasive alternative if these cells successfully regenerate defective cartilage. Traditional microfracture procedures employ the concept of bone marrow stimulation to regenerate cartilage. However, the regenerated tissue usually is fibrous cartilage, which has very poor mechanical properties compared to those of normal hyaline cartilage. A method that directs the migration of a large number of autologous mesenchymal stem cells toward injury sites, retains these cells around the defects, and induces chondrogenic differentiation that would enhance success of endogenous cartilage repair. This review briefly summarizes chemokines and growth factors that induce recruitment, proliferation, and differentiation of endogenous progenitor cells, endogenous cell sources for regenerating cartilage, scaffolds for delivery of bioactive factors, and bioadhesive materials that are necessary to bring about endogenous cartilage repair. PMID:26559963

  14. A Novel Approach to Stimulate Cartilage Repair: Targeting Collagen Turnover

    NARCIS (Netherlands)

    Y.M. Bastiaansen-Jenniskens (Yvonne Maria)

    2009-01-01

    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 repai

  15. THIONIN STAINING OF PARAFFIN AND PLASTIC EMBEDDED SECTIONS OF CARTILAGE

    NARCIS (Netherlands)

    BULSTRA, SK; DRUKKER, J; KUIJER, R; BUURMAN, WA; VANDERLINDEN, AJ

    1993-01-01

    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

  16. Derivation of chondrogenically-committed cells from human embryonic cells for cartilage tissue regeneration.

    Directory of Open Access Journals (Sweden)

    Nathaniel S Hwang

    Full Text Available BACKGROUND: Heterogeneous and uncontrolled differentiation of human embryonic stem cells (hESCs in embryoid bodies (EBs limits the potential use of hESCs for cell-based therapies. More efficient strategies are needed for the commitment and differentiation of hESCs to produce a homogeneous population of specific cell types for tissue regeneration applications. METHODOLOGY/PRINCIPAL FINDINGS: We report here that significant chondrocytic commitment of feeder-free cultured human embryonic stem cells (FF-hESCs, as determined by gene expression and immunostaining analysis, was induced by co-culture with primary chondrocytes. Furthermore, a dynamic expression profile of chondrocyte-specific genes was observed during monolayer expansion of the chondrogenically-committed cells. Chondrogenically-committed cells synergistically responded to transforming growth factor-beta1 (TGF-beta1 and beta1-integrin activating antibody by increasing tissue mass in pellet culture. In addition, when encapsulated in hydrogels, these cells formed cartilage tissue both in vitro and in vivo. In contrast, the absence of chondrocyte co-culture did not result in an expandable cell population from FF-hESCs. CONCLUSIONS/SIGNIFICANCE: The direct chondrocytic commitment of FF-hESCs can be induced by morphogenetic factors from chondrocytes without EB formation and homogenous cartilage tissue can be formed in vitro and in vivo.

  17. Glucocorticoids affect 24 h clock genes expression in human adipose tissue explant cultures

    Science.gov (United States)

    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 ...

  18. Poroelasticity of Cartilage at the Nanoscale

    OpenAIRE

    Nia, Hadi Tavakoli; Han, Lin; Li, Yang; Ortiz, Christine; Grodzinsky, Alan

    2011-01-01

    Atomic-force-microscopy-based oscillatory loading was used in conjunction with finite element modeling to quantify and predict the frequency-dependent mechanical properties of the superficial zone of young bovine articular cartilage at deformation amplitudes, δ, of ∼15 nm; i.e., at macromolecular length scales. Using a spherical probe tip (R ∼ 12.5 μm), the magnitude of the dynamic complex indentation modulus, |E∗|, and phase angle, ϕ, between the force and tip displacement sinusoids, were me...

  19. Cartilage Aggrecan Can Undergo Self-Adhesion

    OpenAIRE

    Han, Lin; Dean, Delphine; Daher, Laura A.; Grodzinsky, Alan J.; Ortiz, Christine

    2008-01-01

    Here it is reported that aggrecan, the highly negatively charged macromolecule in the cartilage extracellular matrix, undergoes Ca2+-mediated self-adhesion after static compression even in the presence of strong electrostatic repulsion in physiological-like solution conditions. Aggrecan was chemically end-attached onto gold-coated planar silicon substrates and gold-coated microspherical atomic force microscope probe tips (end radius R ≈ 2.5 μm) at a density (∼40 mg/mL) that simulates physiolo...

  20. Tissue engineering of cartilages using biomatrices

    DEFF Research Database (Denmark)

    Melrose, J.; Chuang, C.; Whitelock, J.

    2008-01-01

    cartilage engineering approaches and many of these are discussed and their in vitro and in vivo applications covered in this review. Tissue engineering is entering an exciting era; significant advances have been made; however, many technical challenges remain to be solved before this technology becomes......Tissue engineering is an exciting new cross-disciplinary methodology which applies the principles of engineering and structure-function relationships between normal and pathological tissues to develop biological substitute to restore, maintain or improve tissue function. Tissue engineering...

  1. New developments in osteoarthritis and cartilage biology.

    Science.gov (United States)

    Poulet, Blandine; Staines, Katherine A

    2016-06-01

    Osteoarthritis (OA) is a degenerative joint disease and the most common form of arthritis. Characterised by articular cartilage loss, subchondral bone thickening and osteophyte formation, the OA joint afflicts much pain and disability. Whilst OA has been associated with many contributing factors, its underpinning molecular mechanisms are, nevertheless, not fully understood. Clinical management of OA is largely palliative and there is an ever growing need for an effective disease modifying treatment. This review discusses some of the recent progress in OA therapies in the different joint tissues affected by OA pathology. PMID:26921602

  2. Mutation induction in mangosteen : Response of explants to mutagens

    OpenAIRE

    Phrommee, V.; Te-chato, S.

    1999-01-01

    Young purplish red leaves of mangosteen raised in two-phase medium and calli were collected and treated with two mutagens, gamma rays and ethylethane sulfonate (EMS). After treating with both mutagens, leaves and calli were cultured onto callus induction medium to evaluate the effect of mutagens on percent leaf forming callus and survival percentage of callus. The results showed that an increament of EMS caused decreased in survival rate of calli. The concentration that inhibited 50%growth of...

  3. Kartogenin induces cartilage-like tissue formation in tendon–bone junction

    Science.gov (United States)

    Zhang, Jianying; Wang, James H-C

    2014-01-01

    Tendon–bone junctions (TBJs) are frequently injured, especially in athletic settings. Healing of TBJ injuries is slow and is often repaired with scar tissue formation that compromises normal function. This study explored the feasibility of using kartogenin (KGN), a biocompound, to enhance the healing of injured TBJs. We first determined the effects of KGN on the proliferation and chondrogenic differentiation of rabbit bone marrow stromal cells (BMSCs) and patellar tendon stem/progenitor cells (PTSCs) in vitro. KGN enhanced cell proliferation in both cell types in a concentration-dependent manner and induced chondrogenic differentiation of stem cells, as demonstrated by high expression levels of chondrogenic markers aggrecan, collagen II and Sox-9. Besides, KGN induced the formation of cartilage-like tissues in cell cultures, as observed through the staining of abundant proteoglycans, collagen II and osteocalcin. When injected into intact rat patellar tendons in vivo, KGN induced cartilage-like tissue formation in the injected area. Similarly, when KGN was injected into experimentally injured rat Achilles TBJs, wound healing in the TBJs was enhanced, as evidenced by the formation of extensive cartilage-like tissues. These results suggest that KGN may be used as an effective cell-free clinical therapy to enhance the healing of injured TBJs. PMID:25419468

  4. Kartogenin induces cartilage-like tissue formation in tendon-bone junction

    Institute of Scientific and Technical Information of China (English)

    Jianying Zhang; James H-C Wang

    2014-01-01

    Tendon-bone junctions (TBJs) are frequently injured, especially in athletic settings. Healing of TBJ injuries is slow and is often repaired with scar tissue formation that compromises normal function. This study explored the feasibility of using kartogenin (KGN), a biocompound, to enhance the healing of injured TBJs. We first determined the effects of KGN on the proliferation and chondrogenic differentiation of rabbit bone marrow stromal cells (BMSCs) and patellar tendon stem/progenitor cells (PTSCs) in vitro. KGN enhanced cell proliferation in both cell types in a concentration-dependent manner and induced chondrogenic differentiation of stem cells, as demonstrated by high expression levels of chondrogenic markers aggrecan, collagen II and Sox-9. Besides, KGN induced the formation of cartilage-like tissues in cell cultures, as observed through the staining of abundant proteoglycans, collagen II and osteocalcin. When injected into intact rat patellar tendons in vivo, KGN induced cartilage-like tissue formation in the injected area. Similarly, when KGN was injected into experimentally injured rat Achilles TBJs, wound healing in the TBJs was enhanced, as evidenced by the formation of extensive cartilage-like tissues. These results suggest that KGN may be used as an effective cell-free clinical therapy to enhance the healing of injured TBJs.

  5. Kartogenin induces cartilage-like tissue formation in tendon-bone junction.

    Science.gov (United States)

    Zhang, Jianying; Wang, James H-C

    2014-01-01

    Tendon-bone junctions (TBJs) are frequently injured, especially in athletic settings. Healing of TBJ injuries is slow and is often repaired with scar tissue formation that compromises normal function. This study explored the feasibility of using kartogenin (KGN), a biocompound, to enhance the healing of injured TBJs. We first determined the effects of KGN on the proliferation and chondrogenic differentiation of rabbit bone marrow stromal cells (BMSCs) and patellar tendon stem/progenitor cells (PTSCs) in vitro. KGN enhanced cell proliferation in both cell types in a concentration-dependent manner and induced chondrogenic differentiation of stem cells, as demonstrated by high expression levels of chondrogenic markers aggrecan, collagen II and Sox-9. Besides, KGN induced the formation of cartilage-like tissues in cell cultures, as observed through the staining of abundant proteoglycans, collagen II and osteocalcin. When injected into intact rat patellar tendons in vivo, KGN induced cartilage-like tissue formation in the injected area. Similarly, when KGN was injected into experimentally injured rat Achilles TBJs, wound healing in the TBJs was enhanced, as evidenced by the formation of extensive cartilage-like tissues. These results suggest that KGN may be used as an effective cell-free clinical therapy to enhance the healing of injured TBJs. PMID:25419468

  6. Mesenchymal stem cells as a potent cell source for articular cartilage regeneration

    Institute of Scientific and Technical Information of China (English)

    Mohamadreza; Baghaban; Eslaminejad; Elham; Malakooty; Poor

    2014-01-01

    Since articular cartilage possesses only a weak capac-ity for repair, its regeneration potential is considered one of the most important challenges for orthopedic surgeons. The treatment options, such as marrow stimulation techniques, fail to induce a repair tissue with the same functional and mechanical properties of native hyaline cartilage. Osteochondral transplantation is considered an effective treatment option but is as-sociated with some disadvantages, including donor-site morbidity, tissue supply limitation, unsuitable mechani-cal properties and thickness of the obtained tissue. Although autologous chondrocyte implantation results in reasonable repair, it requires a two-step surgical pro-cedure. Moreover, chondrocytes expanded in culture gradually undergo dedifferentiation, so lose morpho-logical features and specialized functions. In the search for alternative cells, scientists have found mesenchymal stem cells(MSCs) to be an appropriate cellular mate-rial for articular cartilage repair. These cells were origi-nally isolated from bone marrow samples and further investigations have revealed the presence of the cells in many other tissues. Furthermore, chondrogenic dif-ferentiation is an inherent property of MSCs noticedat the time of the cell discovery. MSCs are known to exhibit homing potential to the damaged site at which they differentiate into the tissue cells or secrete a wide spectrum of bioactive factors with regenerative proper-ties. Moreover, these cells possess a considerable im-munomodulatory potential that make them the general donor for therapeutic applications. All of these topics will be discussed in this review.

  7. Experimental pharmacological investigation of the antiarthrotic effects of the cartilage and bone marrow extract Rumalon

    Energy Technology Data Exchange (ETDEWEB)

    Kalbhen, D.A.

    1981-08-05

    On the basis of animal experiments, the authors have developed a model of arthrosis which is compatible in its radiological, macroscopic, biochemical, and histological aspects with the pathophysiology of human arthrosis and has been tried in the testing of the antiarthrotic properties of pharmaceuticals. Biochemically induced gonarthroses of experimental animals were used for studies of the effects of a cartilage and bone marrow extract (Rumalon) and a cartilage extract and its high-molecular component DAK-16 on the frequency and progression of degenerative joint diseases. As test parameters, measurements of the articular space, X-ray findings, and macroscopic findings were quantitatively evaluated. The animal experiments show that the inhibitive effects of steroidal and nonsteroidal antirheumatics on the synthesis of the cartilage matrix can be prevented or reduced by simultaneous administration of chondroprotective pharmaceuticals; this may be important on the clinical sector. This antagonism between antiphlogistic agents and Rumalon, which has been observed also in fibroblast cultures and wound healing experiments, is of interest especially for the treatment of activated arthroses.

  8. Experimental pharmacological investigation of the antiarthrotic effects of the cartilage and bone marrow extract Rumalon

    International Nuclear Information System (INIS)

    On the basis of animal experiments, the authors have developed a model of arthrosis which is compatible in its radiological, macroscopic, biochemical, and histological aspects with the pathophysiology of human arthrosis and has been tried in the testing of the antiarthrotic properties of pharmaceuticals. Biochemically induced gonarthroses of experimental animals were used for studies of the effects of a cartilage and bone marrow extract (Rumalon) and a cartilage extract and its high-molecular component DAK-16 on the frequency and progression of degenerative joint diseases. As test parameters, measurements of the articular space, X-ray findings, and macroscopic findings were quantitatively evaluated. The animal experiments show that the inhibitive effects of steroidal and nonsteroidal antirheumatics on the synthesis of the cartilage matrix can be prevented or reduced by simultaneous administration of chondroprotective pharmaceuticals; this may be important on the clinical sector. This antagonism between antiphlogistic agents and Rumalon, which has been observed also in fibroblast cultures and wound healing experiments, is of interest especially for the treatment of activated arthroses. (orig./MG)

  9. REGENERATION OF ARTICULAR CARTILAGE UNDER THE IMPLANTATION OF BONE MATRIX

    Directory of Open Access Journals (Sweden)

    Yuri M. Iryanov, Nikolay A. Kiryanov, Olga V. Dyuriagina , Tatiana Yu. Karaseva, Evgenii A. Karasev

    2015-07-01

    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.

  10. Follistatin Alleviates Synovitis and Articular Cartilage Degeneration Induced by Carrageenan

    Science.gov (United States)

    Yamada, Jun; Abula, Kahaer; Inoue, Makiko; Sekiya, Ichiro; Muneta, Takeshi

    2014-01-01

    Activins are proinflammatory cytokines which belong to the TGFβ superfamily. Follistatin is an extracellular decoy receptor for activins. Since both activins and follistatin are expressed in articular cartilage, we hypothesized that activin-follistatin signaling participates in the process of joint inflammation and cartilage degeneration. To test this hypothesis, we examined the effects of follistatin in a carrageenan-induced mouse arthritis model. Synovitis induced by intra-articular injection of carrageenan was significantly alleviated by preinjection with follistatin. Macrophage infiltration into the synovial membrane was significantly reduced in the presence of follistatin. In addition, follistatin inhibited proteoglycan erosion induced by carrageenan in articular cartilage. These data indicate that activin-follistatin signaling is involved in joint inflammation and cartilage homeostasis. Our data suggest that follistatin can be a new therapeutic target for inflammation-induced articular cartilage degeneration. PMID:25574420

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

    Directory of Open Access Journals (Sweden)

    Robert Mark Wellard

    2014-05-01

    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.

  12. 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)

    2011-07-01

    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)

  13. Biochemical effects on long-term frozen human costal cartilage

    International Nuclear Information System (INIS)

    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)

  14. Radiological observation of determination of sex by costal cartilage calcification

    International Nuclear Information System (INIS)

    The difference of patterns of costal cartilage calcification in male and female had been first described by Fischer in 1955. Thereafter several reports were published, but specific clinical significance was not found. During the period from January, 1978 to December, 1978, we, in the Department of Radiology, Jeonbug National University, studied 2164 cases that showed the entire 12 pairs of ribs. Among these we detected 1494 cases of costal cartilage calcification and frequent sites of calcification. Patterns of costal cartilage calcification were classified into six groups- type l: central, type II: marginal, type III: junctional type, type IV: railroad, type V: diffuse, type VI: mixed. Results are as follows; 1. In a total of 2164 cases, calcification of costal cartilage was present in 1494 cases(69.0%). Of 1181 males 780 cases(66.0%) showed calcification, and of 983 females 714 cases (72.6%) showed calcification. 2. In 439 cases of males, except for 341 cases that showed calcification within the first costal cartilage, patterns of costal cartilage calcification were as follows: marginal type in 265 cases (60.4%), junctional type in 134 cases (30.5%), mixed type in 21 cases (0.5%), central type in 17 cases(3.8%), and railroad type in 2 cases (0.5%). Diffuse type was not present. 3. In 492 cases of females, except of 222 cases that showed calcification within the first costal cartilage, patterns of costal cartilage calcification were as follows; central type in 336 cases (68.3%), junctional type in 94 cases(19.1%), mixed type in 24 cases (4.9%), railroad type in 19 cases (3.9%), and diffuse type in 14 cases (2.8%). 4. When central calcification was observed, predictive value to female was 94.7%. When marginal calcification was observed, predictive value to male was 987.4%. 5. Males frequently showed calcification in upper costal cartilages, and females in lower costal cartilages.

  15. Selective laser sintered poly-ε-caprolactone scaffold hybridized with collagen hydrogel for cartilage tissue engineering

    International Nuclear Information System (INIS)

    Selective laser sintering (SLS), an additive manufacturing (AM) technology, can be used to produce tissue engineering scaffolds with pre-designed macro and micro features based on computer-aided design models. An in-house SLS machine was built and 3D poly-ε-caprolactone (PCL) scaffolds were manufactured using a layer-by-layer design of scaffold struts with varying orientations (0°/45°/0°/45°, 0°/90°/0°/90°, 0°/45°/90°/135°), producing scaffolds with pores of different shapes and distribution. To better enhance the scaffold properties, chondrocytes were seeded in collagen gel and loaded in scaffolds for cartilage tissue engineering. Gel uptake and dynamic mechanical analysis demonstrated the better suitability of the 0°/90°/0°/90° scaffolds for reconstructive cartilage tissue engineering purposes. Chondrocytes were then seeded onto the 0°/90°/0°/90° scaffolds in collagen I hydrogel (PCL/COL1) and compared to medium-suspended cells in terms of their cartilage-like tissue engineering parameters. PCL/COL1 allowed better cell proliferation when compared to PCL or two-dimensional tissue culture polystyrene. Scanning electron microscopy and confocal microscopy observations demonstrated a similar trend for extracellular matrix production and cell survival. Glycosaminoglycan and collagen II quantification also demonstrated the superior matrix secretion properties of PCL/COL1 hybrid scaffolds. Collagen-gel-suspended chondrocytes loaded in SLS-manufactured PCL scaffolds may provide a means of producing tissue-engineered cartilage with customized shapes and designs via AM technology. (paper)

  16. An ovine in vitro model for chondrocyte-based scaffold-assisted cartilage grafts

    Directory of Open Access Journals (Sweden)

    Endres Michaela

    2012-11-01

    Full Text Available Abstract Background Scaffold-assisted autologous chondrocyte implantation is an effective clinical procedure for cartilage repair. From the regulatory point of view, the ovine model is one of the suggested large animal models for pre-clinical studies. The aim of our study was to evaluate the in vitro re-differentiation capacity of expanded ovine chondrocytes in biomechanically characterized polyglycolic acid (PGA/fibrin biomaterials for scaffold-assisted cartilage repair. Methods Ovine chondrocytes harvested from adult articular cartilage were expanded in monolayer and re-assembled three-dimensionally in PGA-fibrin scaffolds. De- and re-differentiation of ovine chondrocytes in PGA-fibrin scaffolds was assessed by histological and immuno-histochemical staining as well as by real-time gene expression analysis of typical cartilage marker molecules and the matrix-remodelling enzymes matrix metalloproteinases (MMP -1, -2 and −13 as well as their inhibitors. PGA scaffolds characteristics including degradation and stiffness were analysed by electron microscopy and biomechanical testing. Results Histological, immuno-histochemical and gene expression analysis showed that dedifferentiated chondrocytes re-differentiate in PGA-fibrin scaffolds and form a cartilaginous matrix. Re-differentiation was accompanied by the induction of type II collagen and aggrecan, while MMP expression decreased in prolonged tissue culture. Electron microscopy and biomechanical tests revealed that the non-woven PGA scaffold shows a textile structure with high tensile strength of 3.6 N/mm2 and a stiffness of up to 0.44 N/mm2, when combined with gel-like fibrin. Conclusion These data suggest that PGA-fibrin is suited as a mechanically stable support structure for scaffold-assisted chondrocyte grafts, initiating chondrogenic re-differentiation of expanded chondrocytes.

  17. Designed composites for mimicking compressive mechanical properties of articular cartilage matrix.

    Science.gov (United States)

    Zhu, Youjia; Wu, Hua; Sun, Shaofa; Zhou, Ting; Wu, Jingjing; Wan, Ying

    2014-08-01

    Collagen, chitosan-polycaprolactone (CH-PCL) copolymer with PCL content of around 40wt% and chondroitin sulfate (CS) were mixed together at various ratios to prepare collagen/CH-PCL/CS composites and the resulting composites were used to build stratified porous scaffolds that are potentially applicable for articular cartilage repair. The ternary composites were designed in such a way that collagen content in the scaffolds decreased from the top layer to the bottom layer while the content of CH-PCL and CS altered in a reversed trend in order to reach partial similarity to cartilage matrix in the composition of main components. Porous structures inside collagen/CH-PCL/CS scaffolds were constructed using a low-temperature deposition processing technique and graded average pore-size and porosity for the scaffolds were established. Such produced scaffolds were further crosslinked using 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide under optimized conditions, and the obtained scaffolds showed well-defined elastic compressive properties. Compressive modulus (E) and stress at 10% strain (σ10) of full scaffolds in wet state reached about 2.8MPa and 0.3MPa, respectively, and meanwhile, E and σ10 of layers inside hydrated scaffolds changed in a gradient-increased manner from the top layer to the bottom layer with significant differences between contiguous layers, which partially mimics compressive mechanical properties of cartilage matrix. In addition, in vitro culture of cell-scaffold constructs exhibited that scaffolds were able to well support the ingrowth and migration of seeded cells, and cells also showed relatively uniform distribution throughout the scaffolds. These results suggest that the presently developed collagen/CH-PCL/CS scaffolds have promising potential for applications in articular cartilage repair. PMID:24793172

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

    Directory of Open Access Journals (Sweden)

    Amir Ghaffar SHAHRIARI

    2012-05-01

    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.

  19. Poroelasticity of cartilage at the nanoscale.

    Science.gov (United States)

    Nia, Hadi Tavakoli; Han, Lin; Li, Yang; Ortiz, Christine; Grodzinsky, Alan

    2011-11-01

    Atomic-force-microscopy-based oscillatory loading was used in conjunction with finite element modeling to quantify and predict the frequency-dependent mechanical properties of the superficial zone of young bovine articular cartilage at deformation amplitudes, δ, of ~15 nm; i.e., at macromolecular length scales. Using a spherical probe tip (R ~ 12.5 μm), the magnitude of the dynamic complex indentation modulus, |E*|, and phase angle, φ, between the force and tip displacement sinusoids, were measured in the frequency range f ~ 0.2-130 Hz at an offset indentation depth of δ(0) ~ 3 μm. The experimentally measured |E*| and φ corresponded well with that predicted by a fibril-reinforced poroelastic model over a three-decade frequency range. The peak frequency of phase angle, f(peak), was observed to scale linearly with the inverse square of the contact distance between probe tip and cartilage, 1/d(2), as predicted by linear poroelasticity theory. The dynamic mechanical properties were observed to be independent of the deformation amplitude in the range δ = 7-50 nm. Hence, these results suggest that poroelasticity was the dominant mechanism underlying the frequency-dependent mechanical behavior observed at these nanoscale deformations. These findings enable ongoing investigations of the nanoscale progression of matrix pathology in tissue-level disease. PMID:22067171

  20. Floret primordia differentiation from in vitro cultured sunflower capitula

    OpenAIRE

    Pellegrini C.N.; Hernandez L.F.

    2004-01-01

    In vitro culture has become a useful tool for studies concerning the assessment of physiological and biochemical factors controlling floral morphogenesis. For sunflower, in vitro techniques starting from different explants yield good results on culture initiation and plant regeneration. Although inflorescence generation on cultured shoots is sometimes achieved there are no reports about in vitro florets developed from cultured young capitula. This work describes a protocol for the in vitro cu...

  1. Abnormal cartilage from the mandibular condyle of stumpy (stm) mutant mice.

    OpenAIRE

    Johnson, D.R.

    1983-01-01

    The mammalian mandibular condyle is composed of secondary cartilage and may thus be susceptible to genes causing achondroplasia and which result in abnormal++ primary cartilage formation. This paper describes the secondary cartilage in the mandible of the stumpy achondroplastic mutation in the mouse: both primary and secondary cartilage are affected by the gene.

  2. Deginerative changes of femoral articular cartilage in the knee : comparative study of specimen sonography and pathology

    International Nuclear Information System (INIS)

    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

  3. Self-Assembled Infrapatellar Fat-Pad Progenitor Cells on a Poly-ε-Caprolactone Film For Cartilage Regeneration.

    Science.gov (United States)

    Prabhakar, Alisha; Lynch, Amy P; Ahearne, Mark

    2016-04-01

    Cartilage defects resulting from osteoarthritis (OA) or physical injury can severely reduce the quality of life for sufferers. Current treatment options are costly and not always effective in producing stable hyaline cartilage. Here we investigated a new treatment option that could potentially repair and regenerate damaged cartilage tissue. This novel approach involves the application of infrapatellar fat-pad derived chondroprogenitor cells onto a mechanically stable biodegradable polymer film that can be easily implanted into a defect site. Poly-ε-caprolactone (PCL) films were fabricated via solvent casting in either acetone or chloroform. The hydrophobicity, mechanical properties, and surface morphology of the films were examined. Progenitor cells from infrapatellar fat-pad were isolated, expanded, and then seeded onto the films. The cells were allowed to self-assemble on films, and these were then cultured in a chemically defined chondrogenic media for 28 days. The self-assembled tissue was characterized via histological staining, gene expression analysis, immunohistochemistry, and biochemical analysis. Chondrogenic differentiation was induced to generate a cartilaginous matrix upon the films. Despite differences between in the appearance, surface morphology, and mechanical properties of the films cast in chloroform or acetone, both methods produced tissues rich in sulfated glycosaminoglycan and collagen, although the extracellular matrix produced on chloroform-cast films appeared to contain more collagen type II and less collagen type I than acetone-cast films. These self-assembled constructs have the potential to be implanted into defect sites as a potential treatment for cartilage defect regeneration. PMID:26516689

  4. In vitro and In vivo Evaluation of the Developed PLGA/HAp/Zein Scaffolds for Bone-Cartilage Interface Regeneration

    Institute of Scientific and Technical Information of China (English)

    LIN Yong Xin; DING Zhi Yong; ZHOU Xiao Bin; LI Si Tao; XIE De Ming; LI Zhi Zhong; SUN Guo Dong

    2015-01-01

    Objective To investigate the effect of electronspun PLGA/HAp/Zein scaffolds on the repair of cartilage defects. Methods The PLGA/HAp/Zein composite scaffolds were fabricated by electrospinning method. The physiochemical properties and biocompatibility of the scaffolds were separately characterized by scanning electron microscope (SEM), transmission electron microscope (TEM), and fourier transform infrared spectroscopy (FTIR), human umbilical cord mesenchymal stem cells (hUC-MSCs) culture and animal experiments. Results The prepared PLGA/HAp/Zein scaffolds showed fibrous structure with homogenous distribution. hUC-MSCs could attach to and grow well on PLGA/HAp/Zein scaffolds, and there was no significant difference between cell proliferation on scaffolds and that without scaffolds (P>0.05). The PLGA/HAp/Zein scaffolds possessed excellent ability to promote in vivo cartilage formation. Moreover, there was a large amount of immature chondrocytes and matrix with cartilage lacuna on PLGA/HAp/Zein scaffolds. Conclusion The data suggest that the PLGA/HAp/Zein scaffolds possess good biocompatibility, which are anticipated to be potentially applied in cartilage tissue engineering and reconstruction.

  5. EFFECTIVENESS OF AUXIN INDUCED IN VITRO ROOT CULTURE IN CHICORY

    OpenAIRE

    Nandagopal, S.; B Ranjitha Kumari

    2007-01-01

    An efficient protocol has been developed for the root culture of (Cichorium intybus L. cv. Focus), the leaf and hypocotyl explants from 25 days old in vitro raised seedlings were cultured on half-strength Murashige and Skoog (MS) medium supplemented with different concentrations and combinations of Indole-3-acetic acid (IAA), Indole-3-butyric acid (IBA), α-Napthalenacetic acid (NAA). 0.5 mg/l NAA and 0.1 mg/l IBA induced highest percentage of rooting from matured leaf explants, under total da...

  6. Biocompatibility Assessment of Novel Collagen-Sericin Scaffolds Improved with Hyaluronic Acid and Chondroitin Sulfate for Cartilage Regeneration

    OpenAIRE

    Sorina Dinescu; Bianca Gălăţeanu; Mădălina Albu; Adriana Lungu; Eugen Radu; Anca Hermenean; Marieta Costache

    2013-01-01

    Cartilage tissue engineering (CTE) applications are focused towards the use of implantable biohybrids consisting of biodegradable scaffolds combined with in vitro cultured cells. Hyaluronic acid (HA) and chondroitin sulfate (CS) were identified as the most potent prochondrogenic factors used to design new biomaterials for CTE, while human adipose-derived stem cells (ASCs) were proved to display high chondrogenic potential. In this context, our aim was not only to build novel 3D porous scaffol...

  7. Trophic action of epidermal growth factor on human duodenal mucosa cultured in vitro.

    OpenAIRE

    Challacombe, D N; Wheeler, E. E.

    1991-01-01

    The action of epidermal growth factor on the human duodenal mucosa has been studied by estimating the crypt cell production rate in cultured explants, using a stathmokinetic technique with crypt microdissection. The addition of epidermal growth factor (400 ng/ml) to paired explants from five patients caused an almost fivefold increase in the crypt cell production rate, showing that epidermal growth factor has a trophic action on the human duodenal mucosa in vitro.

  8. Plant Regeneration from Unfertilized Ovaries of Sugar Beet ( Beta vulgaris L.) Cultured In Vitro

    OpenAIRE

    GÜREL, Ekrem

    1998-01-01

    A. method is described for plant regeneration from unfertilized ovaries isolated from a diploid male sterile sugar beet ( Beta vulgaris L.) breeding line that was developed at the Sugar Institute, Ankara, Turkey. Ovary explants were cultured on Murashige & Skoog (MS) medium containing 2.0 mg/l benzylaminopurine (BAP). Two treatments were tested by incubating all of the explants in darkness for 15 days, and then transferring one half to light and keeping the other half in darkness throu...

  9. Micropropagation of Woody Legume (Albizia lebbeck) Through Tissue Culture

    OpenAIRE

    A.N.K. Mamun; M.N. Matin; Bari, M. A.; N.A. Siddique; R.S. Sultana; M.H. Rahman; Musa, A.S.M.

    2004-01-01

    The woody legume, Albizia lebbeck, commonly cultivated for social forestry in rural area of Bangladesh was selected for micropropagation by tissue culture technique. Cotyledon, nodal segment of in vitro grown seedlings and nodal segments of field grown mature tree were used as explants. With proper manipulation of cytokinin and auxin combinations and concentrations, it was possible to induce callus from three types of explants of Albizia lebbeck. Different combinations of BA with NAA and KIN ...

  10. ERK activation is required for hydrostatic pressure induced-tensile changes in engineered articular cartilage

    OpenAIRE

    DuRaine, G D; Athanasiou, K.A.

    2012-01-01

    The objective of this study was to identify the ERK 1/2 involvement in the changes in compressive and tensile mechanical properties associated with hydrostatic pressure treatment of self-assembled cartilage constructs. In study 1, ERK 1/2 phosphorylation was detected by immunoblot following application of hydrostatic pressure (1 hour of static 10MPa) applied at day 10-14 of self-assembly culture. In study 2, ERK 1/2 activation was blocked during hydrostatic pressure application on days 10-14....

  11. Plant regeneration of bananas Ambon kuning and Barangan mutant lines were carried out by using female organ and shoot-tip as explants source

    International Nuclear Information System (INIS)

    Plant regeneration of bananas Ambon Kuning and Barangan mutant lines were carried out by using female organ and shoot-tip as explants source. Female organ was taken from heart of banana stem, while shoot-tip taken from sucker in banana plantation at Pasar Jumat, Jakarta. Those explants were cultured on MS medium containing 3 mg/l BAP, 0.5 mg/l IAA and supplemented by 100 tyrosin and 80 mg/l adenin hemisulphate. Observation showed that 180 and 42 buds were obtained from JBR 02 mutant lines respectively, while 84 and 79 buds for JAK 01 and JAK 02 respectively. The highest shoot formation was 1.013 shoots were obtained from BRC variety and lowest one was JBR 01 mutant line. statistical data analysis indicated that shoot formation between BRC variety and another mutant lines were significant difference using LSD test at level 0.05. Plantlet formation derived from female organ as well as shoot-tip showed that BRC variety produced number of plantlets per bottle was higher that another one. (author)

  12. Plant regeneration from pulse-treated longitudinally sliced half cotyledon node explants of Turkish ochrus chickling [Lathyrus ochrus (L. D.C.

    Directory of Open Access Journals (Sweden)

    Saglam S.

    2012-01-01

    Full Text Available The forage legume ochrus chickling [Lathyrus ochrus (L. D.C.] which is distributed in the Mediterranean region, is gaining importance in terms of economy and agriculture in Turkey. However, the full potential of the legume has yet to be realized due to the presence of neurotoxin, β-N-oxalyl-L-a, β-diaminopropionic acid (ODAP causing lathyrism. This study aimed to develop an efficient micropropagation system using longitudinally sliced cotyledon node explants for use in Agrobacterium-mediated genetic transformation in the future. The results show that the maximum number of shoots per explant was achieved on MS medium solidified with 8 g/l isubgol gelled medium containing 0.30 mg/l BA-0.2 mg/l NAA. Well-developed shoots were rooted by pulse treatment with 50 mg/l IBA and culturing on an 8 g/l isubgol gel solidified MS medium. The results showed 60% rooting in the treated shoots. The rooted plantlets were transferred to pots containing sand and organic matter and acclimatized.

  13. The identification of matrix Gla protein in cartilage.

    Science.gov (United States)

    Hale, J E; Fraser, J D; Price, P A

    1988-04-25

    The vitamin K-dependent bone protein matrix gamma-carboxyglutamic acid (Gla) protein (MGP) has been identified by radioimmunoassay in the guanidine extract of rat cartilage. MGP was present in all cartilages tested at levels comparable to the MGP level in bone. Western blot analysis indicated that the molecular weight of cartilage MGP is the same as bone MGP, and Northern blot analysis revealed that MGP mRNA from cartilage is the same size as the MGP mRNA from bone. The structurally related vitamin K-dependent protein bone Gla protein could not be detected in cartilage by radioimmunoassay or by Northern blot analysis. The discovery that MGP is synthesized by growth plate cartilage could provide an explanation for the excessive growth plate mineralization disorder seen in rats treated with the vitamin K antagonist warfarin and the punctate mineralization of the growth plate seen in infants whose mothers received warfarin in the first trimester of pregnancy (the fetal warfarin syndrome). Both disorders appear to be caused by the inactivation of a vitamin K-dependent mineralization inhibitor in cartilage, an inhibitor which we suggest is MGP. PMID:3258600

  14. Critical temperature transitions in laser-mediated cartilage reshaping

    Science.gov (United States)

    Wong, Brian J.; Milner, Thomas E.; Kim, Hong H.; Telenkov, Sergey A.; Chew, Clifford; Kuo, Timothy C.; Smithies, Derek J.; Sobol, Emil N.; Nelson, J. Stuart

    1998-07-01

    In this study, we attempted to determine the critical temperature [Tc] at which accelerated stress relaxation occurred during laser mediated cartilage reshaping. During laser irradiation, mechanically deformed cartilage tissue undergoes a temperature dependent phase transformation which results in accelerated stress relaxation. When a critical temperature is attained, cartilage becomes malleable and may be molded into complex new shapes that harden as the tissue cools. Clinically, reshaped cartilage tissue can be used to recreate the underlying cartilaginous framework of structures such as the ear, larynx, trachea, and nose. The principal advantages of using laser radiation for the generation of thermal energy in tissue are precise control of both the space-time temperature distribution and time- dependent thermal denaturation kinetics. Optimization of the reshaping process requires identification of the temperature dependence of this phase transformation and its relationship to observed changes in cartilage optical, mechanical, and thermodynamic properties. Light scattering, infrared radiometry, and modulated differential scanning calorimetry (MDSC) were used to measure temperature dependent changes in the biophysical properties of cartilage tissue during fast (laser mediated) and slow (conventional calorimetric) heating. Our studies using MDSC and laser probe techniques have identified changes in cartilage thermodynamic and optical properties suggestive of a phase transformation occurring near 60 degrees Celsius.

  15. Isolation of carrot plant lines with altered carotene contents from gamma irradiated explants

    International Nuclear Information System (INIS)

    Dietary vitamin A is mainly obtained from carotenes of vegetables and fruits. Carrot (Daucus carota L.) is one of the major sources of carotene. Carrot cultivars have been obtained mainly through classical breeding, and genetic selection has permitted the creation of new varieties with high carotene contents. The fact that in several crops agronomically important mutants/variants have been generated by in vitro culture techniques prompted us to combine gamma irradiation and in vitro somatic embryogenesis to obtain regenerants with variations in carotene content in carrot. To test the effect of gamma rays on somatic embryogenesis and on the carotene level, aseptically germinated seedlings of 8 carrot varieties were exposed to 5; 10 and 500 Gy before culturing petiole segments on LN1 medium. Non-irradiated petioles produced calli with somatic embryos, while irradiated explants reacted differently according to radiation dose. After 4 weeks of culture on LN1 medium, petiole segments of different varieties irradiated with 5 and 10 Gy gave more callus with embryos than those with non-irradiated segments. However, after the subculture on LN medium, the development of embryos into plantlets was rare. It was also noted that after irradiation with 5 Gy, the petiole segments gave voluminous calli. Further, in variety 'Chantenay', the irradiated calli were deep orange while non-irradiated calli were green. However, embryo formation was not observed in these calli. This orange coloration suggests an appreciable synthesis of carotene in the calli. Gamma rays, probably produced cell lines with different colors and carotene content. Of the 8 cultivars tested, normal plantlets of 3 varieties were regenerated from somatic embryos irradiated with 10 Gy, and were transferred to greenhouse to develop roots. For each assay, the carotene analysis was carried out on 2 roots, and compared with plants produced from non-irradiated somatic embryos. Carotene level in the plants, derived from

  16. Cytotoxicity of unsaturated fatty acids in fresh human tumor explants: concentration thresholds and implications for clinical efficacy

    Directory of Open Access Journals (Sweden)

    Scheim David E

    2009-12-01

    Full Text Available Abstract Background Unsaturated fatty acids (UFAs exhibit in vitro cytotoxicity against many malignant cell lines and yield decreased cancer incidence and reduced tumor growth in animal models. But clinical and animal studies to date have achieved response using only localized delivery methods such as intratumoral infusion. To explore possibilities for enhanced clinical efficacy, fresh surgical explants of tumors from 22 patients with five malignancies were exposed to γ-linolenic acid (GLA and α-linolenic acid (ALA and analyzed with an in vitro chemosensitivity testing system, the Fluorescent Cytoprint Assay (FCA. A total of 282 micro-organ cultures derived from these malignant tumors were exposed to GLA and ALA at different concentrations. Results GLA and ALA exhibited greater than 90% cytotoxicity at a sharp concentration threshold between 500 μM and 1 mM against all but two malignant micro-organ cultures tested in 5-10% serum. In tests using 30-40% serum, GLA and ALA killed tumor at concentrations of 2 mM and above. Conclusions The concentration threshold of 500 μM to 2 mM exhibited for antitumor activity by GLA and ALA is much higher than that observed in most previously reported cell culture studies but consistent with physiological concentrations found to kill tumor clinically and in animals. A mechanism of antitumor activity by unsaturated fatty acids through selective destabilization of the malignant plasma membrane is considered. An oral regimen is proposed for phase I clinical testing that could push the area under the curve for serum concentration of unbound unsaturated fatty acids over time to much higher levels than previously achieved for systemic administration and into the range that could yield antitumor response.

  17. Regeneração in vitro de urucum (Bixa orellana L. a partir de diferentes tipos de explantes In vitro regeneration of annatto (Bixa orellana L. from various explants

    Directory of Open Access Journals (Sweden)

    Jane Fiuza Rodrigues Portela de Carvalho

    2005-12-01

    Full Text Available Este trabalho teve como objetivo avaliar a regeneração in vitro de plantas de urucum (Bixa orellana L. a partir de diferentes tipos de explantes. Para definir o meio de cultura adequado para indução de brotações, diferentes concentrações e, ou, combinações da auxina AIA e das citocininas BAP e ZEA foram testadas. As melhores respostas de regeneração para segmentos de hipocótilo, nós cotiledonares e hipocótilos invertidos foram observadas em meios suplementados de ZEA (2,28 µM e AIA (0,30 µM, ZEA (4,56 µM e ZEA (4,56 µM, respectivamente. O meio de enraizamento mais eficaz foi o MS, com a metade de sua concentração salina e 5 µM de AIB. Análises citológicas, realizadas antes da aclimatação, confirmaram a estabilidade cromossômica das plantas cultivadas in vitro, não sendo detectado variação com relação ao número de cromossomos metafásicos (2n = 14.The present work aimed the establishment of a regeneration protocol for annatto (Bixa orellana L. from different juvenile explants. In order to promote shoot induction, different concentrations and/or combinations of IAA and the cytokinins BAP and ZEA were assessed. Better regeneration responses were achieved when segmented hypocotyl, cotiledonary nodes and inverted hypocotyl were cultured onto MS-based medium supplemented with ZEA (2.28 µM and IAA (0.30 µM, ZEA (4.56 µM or ZEA (4.56 µM, respectively. Rooting of elongated shoots displayed higher frequencies when half-strength MS medium with IBA (5 µM was used. No genetic variation was detected among regenerants as revealed by cytological analysis based on metaphasic chromosome countings (2n = 14.

  18. Experimental articular cartilage repair in the Göttingen minipig

    DEFF Research Database (Denmark)

    Christensen, Bjørn Borsøe; Foldager, Casper Bindzus; Olesen, Morten Lykke;

    2015-01-01

    BACKGROUND: A gold standard treatment for articular cartilage injuries is yet to be found, and a cost-effective and predictable large animal model is needed to bridge the gap between in vitro studies and clinical studies. Ideally, the animal model should allow for testing of clinically relevant...... treatments and the biological response should be reproducible and comparable to humans. This allows for a reliable translation of results to clinical studies.This study aimed at verifying the Göttingen minipig as a pre-clinical model for articular cartilage repair by testing existing clinical cartilage...

  19. Premature Calcifications of Costal Cartilages: A New Perspective Premature Calcifications of Costal Cartilages: A New Perspective

    International Nuclear Information System (INIS)

    Calcifications of the costal cartilages occur, as a rule, not until the age of 30 years. The knowledge of the clinical significance of early and extensive calcifications is still incomplete. Materials and Methods. A search was made to find patients below the age of 30 years who showed distinct calcifications of their lower costal cartilages by viewing 360 random samples of intravenous pyelograms and abdominal plain films. The histories, and clinical and laboratory findings of these patients were analyzed. Results. Nineteen patients fulfilled the criteria of premature calcifications of costal cartilages (CCCs). The patients had in common that they were frequently referred to a hospital and were treated by several medical disciplines. Nevertheless many complaints of the patients remained unsolved. Premature CCCs were often associated with rare endocrine disorders, inborn errors of metabolism, and abnormal hematologic findings. Among the metabolic disorders there were 2 proven porphyrias and 7 patients with a suspected porphyria but with inconclusive laboratory findings. Conclusion. Premature CCCs are unlikely to be a normal variant in skeletal radiology. The findings in this small group of patients call for more intensive studies, especially in regard to the putative role of a porphyria

  20. Establishment of an efficient protocol for micropropagation of some pakistani cultivars of date palm (phonex dactylifera l.) using novel inflorescence explants

    International Nuclear Information System (INIS)

    An efficient protocol for rapid and large scale In vitro propagation of some Pakistani cultivars of date palm has been established using inflorescence explants at Date Palm Research Institute (DPRI), Shah Abdul Latif University (SALU), Khairpur, Pakistan. Immature inflorescences of desired cultivars of date palm detached from mother palms followed by surface sterilization with low torrent of current tap water and then 30 percentage NaOCl/sub 2/ solution, the outer cover were removed in order to get spike explants and cut into the 2-3 cm small pieces and cultured on modified MS medium supplemented with 0.1 mg l-1 2, 4-D + 0.1 mg l-1 IAA + 5.0 mg l-1 NAA for initiation and establishment of cultures. The obtained somatic embryos were subjected to multiplication medium involved 0.1 mg l-1 NAA + 0.05 mg l-1BA. Rooting was achieved using quarter strength MS medium containing 0.1 mg l-1NAA without activated charcoal (AC) initially and then with 3 g l-1 AC. Strong rooted plantlets with 2-3 leaves were transferred to pots contained sand and peat moss mixture (1:1 v/v) with more than 95 percentage success in acclimatization. The acclimatized plants with at least one compound leaf were shifted to the open field conditions at SALU campus for further studying morphological and fruit characterization to ensure the true-to-type nature of tissue culture derived plantlets. High multiplication efficiency and survival percentage with no any somaclonal variation ensured the efficacy of the protocol developed for the production of elite cultivars of date palm of Pakistan and can be used to optimize production of other cultivars of date palm worldwide. (author)

  1. Three-dimensional evaluation of cartilage thickness and cartilage volume in the knee joint with MR imaging: reproducibility in volunteers

    International Nuclear Information System (INIS)

    Objective: To determine the reproductibility of three-dimensional volume and thickness measurements of the knee joint cartilage with MRI in volunteers. Methods: The knees of 7 healthy individuals (ages 23 to 58 yrs.) were sagitally imaged with a resolution of 2x0.31x0.31 mm3, using a fat-suppressed FLASH-3 D sequence. The knee was repositioned in between replicate acquisitions, 6 data sets being obtained in each case. After semiautomatic segmentation and three-dimensional reconstruction of the cartilage, the thickness was determined independent of the original section orientation. The coefficient of variation for repeated volume measurements and the deviations of the maximal cartilage thickness values were calculated subsequently. Results: The mean variation of the cartilage volumes of the replicate measurements was 1.4% (±0.8%) in the patella, 1.7% (±1.5%) in the femur, 3.0% (±1.2%) in the medial tibial plateau and 3.5% (±2.0%) in the lateral tibial plateau. The comparison of the distribution patterns of cartilage thickness yielded a high degree of agreement. Only in rare cases deviations of more than 0.5 mm were observed. Conclusions: The results show that the presented method for determining the quantitative distribution of articular cartilage yields a high degree of precision. It offers new possibilities in screening risk groups, monitoring the course of degenerative joint disease and the investigation of functional adaptation of the cartilage to mechanical loading. (orig.)

  2. Butterfly cartilage graft versus fat graft myringoplasty

    Directory of Open Access Journals (Sweden)

    Sonika Kanotra

    2016-01-01

    Full Text Available Aim: The aim of the study was to compare the graft take up rates of two minimally invasive techniques of butterfly cartilage graft (BCG and fat graft myringoplasty (FGM. Materials and Methods: Two groups of 30 patients each with small dry central perforations of the tympanic membrane (T.M. were randomly subjected to either of the two techniques of myringoplasty. Statistical Analysis Used: The results were compared using the Chi-square test. A value of <0.05 was taken as statistically significant. Results: The graft take up rate was 93.3% with BCG and 83.3% with fat graft. Conclusions: The BCG scores over FGM in small perforations of the T.M.

  3. Polylactide fibrous scaffolds for cartilage implant engineering

    Czech Academy of Sciences Publication Activity Database

    Mulinková, Katarína; Machová, Luďka; Lesný, P.; Kubies, Dana; Rypáček, František

    Prague: Czech Society for New Materials and Technologies, 2005. Poster Session II. [European Congress on Advanced Materials and Processes. 5.9.2005-8.9.2005, Prague] R&D Projects: GA MZd ND7448 Keywords : biodegradable polymers * polylactide fibres * cartilage engineering Subject RIV: FJ - Surgery incl. Transplants http://webdb.dgm.de/dgm_lit/prg/FMPro?-db=w%5fprogram&- format =prog%5fpaper%5fresults.htm&-lay=standard&TB=%3d%3d688&tgb%5fsymposium%5fund%5fnr=B14%20Engineering%20and%20Design%20of%20Biomedical%20Materials&-max=20&-skip=20&-token.0=688&-token.1=B14%20Engineering%20and%20Design%20of%20Biomedical%20Materials&-find=

  4. The development of the collagen fibre network in tissue-engineered cartilage constructs in vivo. Engineered cartilage reorganises fibre network

    Directory of Open Access Journals (Sweden)

    H Paetzold

    2012-04-01

    Full Text Available For long term durability of tissue-engineered cartilage implanted in vivo, the development of the collagen fibre network orientation is essential as well as the distribution of collagen, since expanded chondrocytes are known to synthesise collagen type I. Typically, these properties differ strongly between native and tissue-engineered cartilage. Nonetheless, the clinical results of a pilot study with implanted tissue-engineered cartilage in pigs were surprisingly good. The purpose of this study was therefore to analyse if the structure and composition of the artificial cartilage tissue changes in the first 52 weeks after implantation. Thus, collagen network orientation and collagen type distribution in tissue-engineered cartilage-carrier-constructs implanted in the knee joints of Göttinger minipigs for 2, 26 or 52 weeks have been further investigated by processing digitised microscopy images of histological sections. The comparison to native cartilage demonstrated that fibre orientation over the cartilage depth has a clear tendency towards native cartilage with increasing time of implantation. After 2 weeks, the collagen fibres of the superficial zone were oriented parallel to the articular surface with little anisotropy present in the middle and deep zones. Overall, fibre orientation and collagen distribution within the implants were less homogenous than in native cartilage tissue. Despite a relatively low number of specimens, the consistent observation of a continuous approximation to native tissue is very promising and suggests that it may not be necessary to engineer the perfect tissue for implantation but rather to provide an intermediate solution to help the body to heal itself.

  5. Rod and cone photoreceptor cells produce ROS in response to stress in a live retinal explant system.

    LENUS (Irish Health Repository)

    Bhatt, Lavinia

    2010-01-01

    PURPOSE: The production of reactive oxygen species (ROS) can lead to oxidative stress, which is a strong contributory factor to many ocular diseases. In this study, the removal of trophic factors is used as a model system to investigate the effects of stress in the retina. The aims were to determine if both rod and cone photoreceptor cells produce ROS when they are deprived of trophic factor support and to demonstrate if the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) enzymes are responsible for this ROS production. METHODS: Retinas were explanted from mice aged between postnatal days 8-10 and cultured overnight. The following morning, confocal microscopy combined with various fluorescent probes was used to detect the production of ROS. Each time peanut agglutinin (PNA), a cone photoreceptor marker, was used to facilitate orientation of the retina. Dihydroethidium and dihydrorhodamine 123 (DHR123) were used to determine which cells produce ROS. Subsequently, western blots of retinal serial sections were used to detect the presence of Noxs in the different retinal layers. The Nox inhibitor apocynin was then tested to determine if it altered the production of ROS within these cells. RESULTS: Live retinal explants, viewed at high magnifications using confocal microscopy, displayed an increase in the fluorescent products of dihydroethidium and DHR123 upon serum removal when compared to controls. DHR123 fluorescence, once oxidized, localized to mitochondria and was found in the same focal plane as the PNA staining. This showed that cones and rods produced ROS when stressed. Retinal serial sectioning established that the photoreceptor layer expressed Nox4, dual oxidase (Duox) 1, and Duox2 at varying levels. Finally, the Nox inhibitor apocynin decreased the burst stimulated by the stress of serum removal. CONCLUSIONS: Confocal microscopy and PNA staining allowed differentiation of cell types within the outermost layers of the retina, demonstrating

  6. Long-term gene therapy with Del1 fragment using nonviral vectors in mice with explanted tumors

    Directory of Open Access Journals (Sweden)

    Kitano H

    2016-01-01

    Full Text Available Hisataka Kitano,1 Atsushi Mamiya,1 Tomomi Ishikawa,2 Kayo Egoshi,3 Shinichiro Kokubun,2 Chiaki Hidai2 1Division of Dental Surgery, School of Medicine, Nihon University, Tokyo, Japan; 2Division of Physiology, Department of Biomedical Science, School of Medicine, Nihon University, Tokyo, Japan; 3School of Medicine, Nihon University, Tokyo, Japan Abstract: Cancer gene therapy using nonviral vectors is useful for long periods of treatment because such vectors are both safe and inexpensive, and thus can be used repeatedly. It has been reported that gene therapy with an E3C1 fragment of Del1 in a mouse explanted tumor model improved prognosis. The present study aimed to analyze the long-term effects of repeated nonviral gene transfer of E3C1. Mice with explanted tumors of SCCKN cells, a human squamous carcinoma, were treated with a plasmid encoding E3C1. Plasmids were injected locally every week using a transfection reagent. Control mice treated with mock DNA started to be euthanized on day 18, because the tumors had grown to over 15% of the body weight, and all of them had died by day 43. On the other hand, the tumors in two of ten mice treated with E3C1 had disappeared. The other eight mice started to be euthanized on day 46 and eight of ten mice had been euthanized by day 197. After 18 days of therapy, the tumor volume of control mice was 2,804±829 mm3 and that of the E3C1 mice was 197±159 mm3. Histochemical studies showed enhanced apoptosis in the E3C1-treated tumors, as compared with controls. Changes in cell morphology and decreased polymerized actin induced by E3C1 indicated disturbed cell adhesion to the matrix. In in vitro studies of SCCKN cells, prolonged administration of an E3C1 recombinant protein to cultured cells reduced adhesion-independent growth of cancer cells, as compared with control cells. These data suggest that E3C1 treatment induces anoikis. Keywords: cancer gene therapy, nonviral vector, apoptosis, anoikis, Del1

  7. Comparative effects of plant growth regulators on leaf and stem explants of Labisia pumila var. alata

    OpenAIRE

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

    2013-01-01

    Objective: 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. Metho...

  8. T cells fail to develop in the human skin-cell explants system; an inconvenient truth

    OpenAIRE

    Vanderlocht Joris; Lumeij Stijn BJ; Tonnaer Siebe; van der Stegen Sjoukje JC; Huijskens Mirelle JAJ; Van Elssen Catharina HMJ; Meek Bob; Kirkland Mark A; Hesselink Reinout; Germeraad Wilfred TV; Bos Gerard MJ

    2011-01-01

    Abstract Background Haplo-identical hematopoietic stem cell (HSC) transplantation is very successful in eradicating haematological tumours, but the long post-transplant T-lymphopenic phase is responsible for high morbidity and mortality rates. Clark et al. have described a skin-explant system capable of producing host-tolerant donor-HSC derived T-cells. Because this T-cell production platform has the potential to replenish the T-cell levels following transplantation, we set out to validate th...

  9. Ni2+ treatment causes cement gland formation in ectoderm explants of Xenopus laevis embryo

    Institute of Scientific and Technical Information of China (English)

    HUANGYONG; XIAOYANDING

    1999-01-01

    We found T-type calcium channel blocker Ni2+ can efficiently induce the formation of cement gland in Xenopus laevis animal cap explants.Nother T-typer specific calcium channel blocker Amiloride can also induce the formation of cement gland,while L-type specific calcium channel blocker Nifedipine as no inductive effect.These results may offer us an new approach to study the differentiation of cement gland through the change of intracelluar calcium concentration.

  10. Cartilage (Bovine and Shark) (PDQ®)—Health Professional Version

    Science.gov (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.

  11. Now approaches to the treatment of articular cartilage lesions

    Directory of Open Access Journals (Sweden)

    M. Coviello

    2011-01-01

    Full Text Available Various approaches to the treatment of cartilage defects have been proposed in the literature; reparative and regenerative methods and, more recently, the Maioregen technique are currently available.

  12. Endobronchial Cartilage Rupture: A Rare Cause of Lobar Collapse.

    Science.gov (United States)

    Dasa, Osama; Siddiqui, Nauman; Ruzieh, Mohammed; Javaid, Toseef

    2016-01-01

    Endobronchial cartilage rupture is a rare clinical condition, which can present in patients with severe emphysema with sudden onset shortness of breath. We present a case of a 62-year-old male who presented to our emergency department with sudden onset shortness of breath. Chest X-ray showed lung hyperinflation and a right lung field vague small density. Chest Computed Tomography confirmed the presence of right middle lobe collapse. Bronchoscopy revealed partial right middle lobe atelectasis and an endobronchial cartilage rupture. Endobronchial cartilage rupture is a rare condition that can present as sudden onset shortness of breath due to lobar collapse in patients with emphysema and can be triggered by cough. Bronchoscopic findings include finding a collapsed lung lobe and a visible ruptured endobronchial cartilage. A high index of suspicion, chest imaging, and early bronchoscopy can aid in the diagnosis and help prevent complications. PMID:27525149

  13. Namaste (counterbalancing technique: Overcoming warping in costal cartilage

    Directory of Open Access Journals (Sweden)

    Kapil S Agrawal

    2015-01-01

    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.

  14. The sulphation of chondroitin sulphate in embryonic chicken cartilage

    Science.gov (United States)

    Robinson, H. C.

    1969-01-01

    1. Whole tissue preparations and subcellular fractions from embryonic chicken cartilage were used to measure the rate of incorporation of inorganic sulphate into chondroitin sulphate in vitro. 2. In cartilage from 14-day-old embryos, [35S]sulphate is incorporated to an equal extent into chondroitin 4-sulphate and chondroitin 6-sulphate at a rate of 1·5nmoles of sulphate/hr./mg. dry wt. of cartilage. 3. Microsomal and soluble enzyme preparations from embryonic cartilage catalyse the transfer of sulphate from adenosine 3′-phosphate 5′-sulphatophosphate into both chondroitin 4-sulphate and chondroitin 6-sulphate. 4. The effects of pH, ionic strength, adenosine 3′-phosphate 5′-sulphatophosphate concentration and acceptor chondroitin sulphate concentration on the soluble sulphotransferase activity were examined. These factors all influence the activity of the sulphotransferase, and pH and incubation time also influence the percentage of chondroitin 4-sulphate formed. PMID:5807213

  15. Cartilage oligomeric matrix protein in patients with juvenile idiopathic arthritis

    DEFF Research Database (Denmark)

    Bjørnhart, Birgitte; Juul, Anders; Nielsen, Susan;

    2009-01-01

    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 with...

  16. Tailored PVA/ECM Scaffolds for Cartilage Regeneration

    Directory of Open Access Journals (Sweden)

    Elena Stocco

    2014-01-01

    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.

  17. Starch-modified magnetite nanoparticles for impregnation into cartilage

    International Nuclear Information System (INIS)

    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

  18. In vitro regeneration from petiole explants of non-toxic Jatropha curcas

    KAUST Repository

    Kumar, Nitish

    2011-01-01

    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. Heparin exerts anti-apoptotic effects on uterine explants by targeting the endocannabinoid system.

    Science.gov (United States)

    Salazar, Ana Inés; Vercelli, Claudia; Schiariti, Victoria; Davio, Carlos; Correa, Fernando; Franchi, Ana María

    2016-09-01

    Miscarriage caused by Gram-negative bacteria infecting the female genital tract is one of the most common complications of human pregnancy. Intraperitoneal administration of LPS to 7-days pregnant mice induces embryo resorption after 24 h. Here, we show that LPS induced apoptosis on uterine explants from 7-days pregnant mice and that CB1 receptor was involved in this effect. On the other hand, heparin has been widely used for the prevention of pregnancy loss in women with frequent miscarriage with or without thrombophilia. Besides its anticoagulant properties, heparin exerts anti-inflammatory, immunomodulatory and anti-apoptotic effects. Here, we sought to investigate whether the administration of heparin prevented LPS-induced apoptosis in uterine explants from 7-days pregnant mice. We found that heparin enhanced cell survival in LPS-treated uterine explants and that this effect was mediated by increasing uterine FAAH activity. Taken together, our results point towards a novel mechanism involved in the protective effects of heparin. PMID:27364950

  20. Articular cartilage repair and the evolving role of regenerative medicine

    Directory of Open Access Journals (Sweden)

    Pieter K Bos

    2010-10-01

    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