GROWTH AND ROOTING SYSTEM OF ACACIA MANGIUM OBTAINED BY TISSUE CULTURE

Directory of Open Access Journals (Sweden)

SUPRIYANTO

1991-01-01

Full Text Available Since 1980/1981, the government of Indonesia through the Ministry of Forestry has started to reforest logged-over, alang-alang, unproductive areas and to convert them to Forest Industry Plantation. The target is 300 000 ha per year. It means, 750 million seedlings should be provided per year (planting distance 2 m x 2 m. The tree species to be planted in forest industry plantation should have shorter life cycle (8 - 10 years, good stem-form, good rooting system, and should be fast growing. Acacia mangium has been selected as one of the important tree species for forest industry plantation due to its growth, quality of fiber wood (pulp and paper industry and rooting system (produce a lot of secondary root and nitrogen fixater (Soebardjo 1986. The reforestation of logged-over Dipterocarp forests in Malaysia with A. mangium has also been considered (Appanah and Weinland 1989. Generally, reforestation with A. mangium is done with seedlings obtained by seed germination. A. mangium produce a lot of seeds but its production is still limited by the season, while the conventional method of vegetative propagation through cuttings gave very low percentage of rooted-cuttings (1% (Umboh and Syamsul Yani 1989. The micropropagation of A. mangium through tissue culture is a promising method. The production of A. mangium plantlets through that method has been done at the Forest Genetic Laboratory, Tropical Forest Biology, SEAMEO BIOTROP (Situmorang 1988, Umboh 1988, Umboh et al. 1989, 1990. These rooted-plantlets (plantlings were first put in the green house (acclimatization before planting in the field. Field tests of some agricultural plants have been done but information on forest trees species is still lacking because the production of plantlings through tissue culture is still limited as there are still problems of their rooting. In fact, the progress of reproducing woody plants by tissue culture has been much slower than with herbaceous plants. The major

Substituted Indoleacetic Acids Tested in Tissue Cultures

DEFF Research Database (Denmark)

Engvild, Kjeld Christensen

1978-01-01

Monochloro substituted IAA inhibited shoot induction in tobacco tissue cultures about as much as IAA. Dichloro substituted IAA inhibited shoot formation less. Other substituted IAA except 5-fluoro- and 5-bromoindole-3-acetic acid were less active than IAA. Callus growth was quite variable...

Human meniscal proteoglycan metabolism in long-term tissue culture

NARCIS (Netherlands)

Verbruggen, G.; Verdonk, R.; Veys, E. M.; van Daele, P.; de Smet, P.; van den Abbeele, K.; Claus, B.; Baeten, D.

1996-01-01

For the purpose of human meniscal allografting, menisci have been maintained viable in in vitro culture. The influence of long-term tissue culture on the extracellular matrix metabolism of the meniscus has been studied. Fetal calf serum (FCS) was used as a supplement for the growth factors necessary

Advances in tissue engineering through stem cell-based co-culture.

Science.gov (United States)

Paschos, Nikolaos K; Brown, Wendy E; Eswaramoorthy, Rajalakshmanan; Hu, Jerry C; Athanasiou, Kyriacos A

2015-05-01

Stem cells are the future in tissue engineering and regeneration. In a co-culture, stem cells not only provide a target cell source with multipotent differentiation capacity, but can also act as assisting cells that promote tissue homeostasis, metabolism, growth and repair. Their incorporation into co-culture systems seems to be important in the creation of complex tissues or organs. In this review, critical aspects of stem cell use in co-culture systems are discussed. Direct and indirect co-culture methodologies used in tissue engineering are described, along with various characteristics of cellular interactions in these systems. Direct cell-cell contact, cell-extracellular matrix interaction and signalling via soluble factors are presented. The advantages of stem cell co-culture strategies and their applications in tissue engineering and regenerative medicine are portrayed through specific examples for several tissues, including orthopaedic soft tissues, bone, heart, vasculature, lung, kidney, liver and nerve. A concise review of the progress and the lessons learned are provided, with a focus on recent developments and their implications. It is hoped that knowledge developed from one tissue can be translated to other tissues. Finally, we address challenges in tissue engineering and regenerative medicine that can potentially be overcome via employing strategies for stem cell co-culture use. Copyright © 2014 John Wiley & Sons, Ltd.

The basic design and requirement for plant tissue culture laboratory in MINT

International Nuclear Information System (INIS)

Azraf Azman; Rosli Darmawan; Rusli Ibrahim; Mohd Nazir Basiran; Azhar Mohamad; Mohamed Najli Mohamed Yasin; Shuhaimi Shamsuddin

2005-01-01

The production of multiple species plantlets involves a relatively complex process and it is a highly specialized operation. Tissue culture technology is rapidly becoming a commercialized method for propagating new cultivars, rare species and difficult-to-propagate plant. Not only are skills and knowledge essential but the laboratory itself also plays an important role to ensure the successful growth of the plantlets. To produce quality plantlets, plant tissue culture laboratories should fulfill the basic requirements. The laboratory should have proper building and layout which comprise of media preparation and washing room, sterilization or autoclave room, transfer room and culture or growth room. The scope of this paper is to compare these fundamental requirements with the plant tissue culture laboratory in MINT. All the basic needs and differences will be discussed and the proposal for corrective actions will be presented. (Author)

Plant Tissue Culture

Indian Academy of Sciences (India)

Admin

Plant tissue culture is a technique of culturing plant cells, tissues and organs on ... working methods (Box 2) and discovery of the need for B vita- mins and auxins for ... Kotte (Germany) reported some success with growing isolated root tips.

Oxygen and tissue culture affect placental gene expression.

Science.gov (United States)

Brew, O; Sullivan, M H F

2017-07-01

Placental explant culture is an important model for studying placental development and functions. We investigated the differences in placental gene expression in response to tissue culture, atmospheric and physiologic oxygen concentrations. Placental explants were collected from normal term (38-39 weeks of gestation) placentae with no previous uterine contractile activity. Placental transcriptomic expressions were evaluated with GeneChip ® Human Genome U133 Plus 2.0 arrays (Affymetrix). We uncovered sub-sets of genes that regulate response to stress, induction of apoptosis programmed cell death, mis-regulation of cell growth, proliferation, cell morphogenesis, tissue viability, and protection from apoptosis in cultured placental explants. We also identified a sub-set of genes with highly unstable pattern of expression after exposure to tissue culture. Tissue culture irrespective of oxygen concentration induced dichotomous increase in significant gene expression and increased enrichment of significant pathways and transcription factor targets (TFTs) including HIF1A. The effect was exacerbated by culture at atmospheric oxygen concentration, where further up-regulation of TFTs including PPARA, CEBPD, HOXA9 and down-regulated TFTs such as JUND/FOS suggest intrinsic heightened key biological and metabolic mechanisms such as glucose use, lipid biosynthesis, protein metabolism; apoptosis, inflammatory responses; and diminished trophoblast proliferation, differentiation, invasion, regeneration, and viability. These findings demonstrate that gene expression patterns differ between pre-culture and cultured explants, and the gene expression of explants cultured at atmospheric oxygen concentration favours stressed, pro-inflammatory and increased apoptotic transcriptomic response. Copyright © 2017 Elsevier Ltd. All rights reserved.

Culture methods of allograft musculoskeletal tissue samples in Australian bacteriology laboratories.

Science.gov (United States)

Varettas, Kerry

2013-12-01

Samples of allograft musculoskeletal tissue are cultured by bacteriology laboratories to determine the presence of bacteria and fungi. In Australia, this testing is performed by 6 TGA-licensed clinical bacteriology laboratories with samples received from 10 tissue banks. Culture methods of swab and tissue samples employ a combination of solid agar and/or broth media to enhance micro-organism growth and maximise recovery. All six Australian laboratories receive Amies transport swabs and, except for one laboratory, a corresponding biopsy sample for testing. Three of the 6 laboratories culture at least one allograft sample directly onto solid agar. Only one laboratory did not use a broth culture for any sample received. An international literature review found that a similar combination of musculoskeletal tissue samples were cultured onto solid agar and/or broth media. Although variations of allograft musculoskeletal tissue samples, culture media and methods are used in Australian and international bacteriology laboratories, validation studies and method evaluations have challenged and supported their use in recovering fungi and aerobic and anaerobic bacteria.

Usefulness of fibroblast culture for testing of cattle tissues polluted with heavy metals

International Nuclear Information System (INIS)

Weglarz, L.; Drozdz, M.Wa.; Wardas, M.; Kula, B.; Pawlaczyk-Szpilowa, M.

1990-01-01

Cattle tissues (liver, kidney, brain, and lung) that had been polluted with heavy metals were tested for their ability to alter fibroblast culture growth, cellular protein and DNA content, and fibroblast DNA synthesis. At 72 hr of incubation a significant increase in cellular DNA and [14C]thymidine incorporation was noted in the primary cultures as well as in the subcultures compared to controls. Fibroblast cultures also displayed growth inhibition and reduction in protein content. The measurement of basic biochemical parameters of the fibroblast culture may represent a sensitive means of assessing rapidly the activity of heavy metals deposited in the tissues of cattle as a result of their grazing on polluted soil

Heritability of regeneration in tissue cultures of sweet potato (Ipomoea batatas L.).

Science.gov (United States)

Templeton-Somers, K M; Collins, W W

1986-03-01

A population of open-pollinated progeny from 12 parents, and the 12 parents, was surveyed for in vitro growth and regeneration characteristics. Four different tissue culture procedures involving different media and the use of different explants to initiate the cultures were used. Petiole explants from young leaves were used as explants for initiation of callus cultures. These were evaluated for callus growth rate, friability, and callus color and texture, before transferring to each of three different regeneration media for evaluation of morphogenetic potential. Small shoot tips also were used to initiate callus cultures, which were evaluated for the same growth characteristics and transferred to growth-regulator free regeneration media. Regeneration occurred through root or shoot regeneration or through embryogenesis. Tissue culture treatment effects, as well as genotypic effects, were highly significant in determining: the types of callus produced, callus growth rates, color and texture on the two types of media used for the second and third subcultures. The family x treatment interaction was generally not statistically significant, affecting only callus color. Estimates of narrow sense heritability for callus growth rate in both the second and third subcultures were high enough (0.35 and 0.63, respectively) for the evaluation of parental lines for selection procedures. These characteristics were also the only early culture callus traits that were consistently correlated with later morphogenesis of the cultures. They were negatively correlated with root or shoot regeneration. The occurence of somatic embryogenesis was not correlated with early callus growth characteristics. Genetic and treatment effects were highly significant in the evaluation of morphogenetic potential, through root or shoot regeneration, or through embryogenesis. Regeneration of all types was of low frequency for all procedures, expressed in ≦ 11% of the cultures of the total population.

Growth factor effects on costal chondrocytes for tissue engineering fibrocartilage

Science.gov (United States)

Johns, D.E.; Athanasiou, K.A.

2010-01-01

Tissue engineered fibrocartilage could become a feasible option for replacing tissues like the knee meniscus or temporomandibular joint disc. This study employed five growth factors insulin-like growth factor-I, transforming growth factor-β1, epidermal growth factor, platelet-derived growth factor-BB, and basic fibroblast growth factor in a scaffoldless approach with costal chondrocytes, attempting to improve biochemical and mechanical properties of engineered constructs. Samples were quantitatively assessed for total collagen, glycosaminoglycans, collagen type I, collagen type II, cells, compressive properties, and tensile properties at two time points. Most treated constructs were worse than the no growth factor control, suggesting a detrimental effect, but the IGF treatment tended to improve the constructs. Additionally, the 6wk time point was consistently better than 3wks, with total collagen, glycosaminoglycans, and aggregate modulus doubling during this time. Further optimization of the time in culture and exogenous stimuli will be important in making a more functional replacement tissue. PMID:18597118

A method for establishing human primary gastric epithelial cell culture from fresh surgical gastric tissues.

Science.gov (United States)

Aziz, Faisal; Yang, Xuesong; Wen, Qingping; Yan, Qiu

2015-08-01

At present, biopsy specimens, cancer cell lines and tissues obtained by gastric surgery are used in the study and analysis of gastric cancer, including the molecular mechanisms and proteomics. However, fibroblasts and other tissue components may interfere with these techniques. Therefore, the present study aimed to develop a procedure for the isolation of viable human gastric epithelial cells from gastric surgical tissues. A method was developed to culture human gastric epithelial cells using fresh, surgically excised tissues and was evaluated using immunocytochemistry, periodic acid-Schiff (PAS) staining and cell viability assays. Low cell growth was observed surrounding the gastric tissue on the seventh day of tissue explant culture. Cell growth subsequently increased, and at 12 days post-explant a high number of pure epithelial cells were detected. The gastric cancer cells exhibited rapid growth with a doubling time of 13-52 h, as compared to normal cells, which had a doubling time of 20-53 h. Immunocytochemical analyses of primary gastric cells revealed positive staining for cytokeratin 18 and 19, which indicated that the culture was comprised of pure epithelial cells and contained no fibroblasts. Furthermore, PAS staining demonstrated that the cultured gastric cells produced neutral mucin. Granulin and carbohydrate antigen 724 staining confirmed the purity of gastric cancer and normal cells in culture. This method of cell culture indicated that the gastric cells in primary culture consisted of mucin-secreting gastric epithelial cells, which may be useful for the study of gastric infection with Helicobacter pylori and gastric cancer.

The effect of plant growth regulators on optimization of tissue culture ...

African Journals Online (AJOL)

USER

2010-04-05

Apr 5, 2010 ... ISSN 1684–5315 © 2010 Academic Journals ... tissue culture system in Malaysian upland rice ... Scientists believe that using new cultivars which have potential ..... providing the financial support and to Firouzeh Ashjazadeh ...

Yield improvement strategies for the production of secondary metabolites in plant tissue culture: silymarin from Silybum marianum tissue culture.

Science.gov (United States)

AbouZid, S

2014-01-01

Plant cell culture can be a potential source for the production of important secondary metabolites. This technology bears many advantages over conventional agricultural methods. The main problem to arrive at a cost-effective process is the low productivity. This is mainly due to lack of differentiation in the cultured cells. Many approaches have been used to maximise the yield of secondary metabolites produced by cultured plant cells. Among these approaches: choosing a plant with a high biosynthetic capacity, obtaining efficient cell line for growth and production of metabolite of interest, manipulating culture conditions, elicitation, metabolic engineering and organ culture. This article gives an overview of the various approaches used to maximise the production of pharmaceutically important secondary metabolites in plant cell cultures. Examples of using these different approaches are shown for the production of silymarin from Silybum marianum tissue culture.

Plant tissue culture techniques

Directory of Open Access Journals (Sweden)

Rolf Dieter Illg

1991-01-01

Full Text Available Plant cell and tissue culture in a simple fashion refers to techniques which utilize either single plant cells, groups of unorganized cells (callus or organized tissues or organs put in culture, under controlled sterile conditions.

21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Tissue culture media for human ex vivo tissue and cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell culture...

Pathogen and biological contamination management in plant tissue culture: phytopathogens, vitro pathogens, and vitro pests.

Science.gov (United States)

Cassells, Alan C

2012-01-01

The ability to establish and grow plant cell, organ, and tissue cultures has been widely exploited for basic and applied research, and for the commercial production of plants (micro-propagation). Regardless of whether the application is for research or commerce, it is essential that the cultures be established in vitro free of biological contamination and be maintained as aseptic cultures during manipulation, growth, and storage. The risks from microbial contamination are spurious experimental results due to the effects of latent contaminants or losses of valuable experimental or commercial cultures. Much of the emphasis in culture contamination management historically focussed on the elimination of phytopathogens and the maintenance of cultures free from laboratory contamination by environmental bacteria, fungi (collectively referred to as "vitro pathogens", i.e. pathogens or environmental micro-organisms which cause culture losses), and micro-arthropods ("vitro pests"). Microbial contamination of plant tissue cultures is due to the high nutrient availability in the almost universally used Murashige and Skoog (Physiol Plant 15:473-497, 1962) basal medium or variants of it. In recent years, it has been shown that many plants, especially perennials, are at least locally endophytically colonized intercellularly by bacteria. The latter, and intracellular pathogenic bacteria and viruses/viroids, may pass latently into culture and be spread horizontally and vertically in cultures. Growth of some potentially cultivable endophytes may be suppressed by the high salt and sugar content of the Murashige and Skoog basal medium and suboptimal temperatures for their growth in plant tissue growth rooms. The management of contamination in tissue culture involves three stages: disease screening (syn. disease indexing) of the stock plants with disease and endophyte elimination where detected; establishment and pathogen and contaminant screening of established initial cultures

Cultured human foreskin fibroblasts produce a factor that stimulates their growth with properties similar to basic fibroblast growth factor

International Nuclear Information System (INIS)

Story, M.T.

1989-01-01

To determine if fibroblasts could be a source of fibroblast growth factor (FGF) in tissue, cells were initiated in culture from newborn human foreskin. Fibroblast cell lysates promoted radiolabeled thymidine uptake by cultured quiescent fibroblasts. Seventy-nine percent of the growth-promoting activity of lysates was recovered from heparin-Sepharose. The heparin-binding growth factor reacted on immunoblots with antiserum to human placenta-derived basic FGF and competed with iodinated basic FGF for binding to antiserum to (1-24)bFGF synthetic peptide. To confirm that fibroblasts were the source of the growth factor, cell lysates were prepared from cells incubated with radiolabeled methionine. Heparin affinity purified material was immunoprecipitated with basic FGF antiserum and electrophoresed. Radiolabeled material was detected on gel autoradiographs in the same molecular weight region as authentic iodinated basic FGF. The findings are consistant with the notion that cultured fibroblasts express basic FGF. As these cells also respond to the mitogen, it is possible that the regulation of their growth is under autocrine control. Fibroblasts may be an important source of the growth factor in tissue

Addressing the instability of DNA nanostructures in tissue culture.

Science.gov (United States)

Hahn, Jaeseung; Wickham, Shelley F J; Shih, William M; Perrault, Steven D

2014-09-23

DNA nanotechnology is an advanced technique that could contribute diagnostic, therapeutic, and biomedical research devices to nanomedicine. Although such devices are often developed and demonstrated using in vitro tissue culture models, these conditions may not be compatible with DNA nanostructure integrity and function. The purpose of this study was to characterize the sensitivity of 3D DNA nanostructures produced via the origami method to the in vitro tissue culture environment and identify solutions to prevent loss of nanostructure integrity. We examined whether the physiological cation concentrations of cell culture medium and the nucleases present in fetal bovine serum (FBS) used as a medium supplement result in denaturation and digestion, respectively. DNA nanostructure denaturation due to cation depletion was design- and time-dependent, with one of four tested designs remaining intact after 24 h at 37 °C. Adjustment of medium by addition of MgSO4 prevented denaturation. Digestion of nanostructures by FBS nucleases in Mg(2+)-adjusted medium did not appear design-dependent and became significant within 24 h and when medium was supplemented with greater than 5% FBS. We estimated that medium supplemented with 10% FBS contains greater than 256 U/L equivalent of DNase I activity in digestion of DNA nanostructures. Heat inactivation at 75 °C and inclusion of actin protein in medium inactivated and inhibited nuclease activity, respectively. We examined the impact of medium adjustments on cell growth, viability, and phenotype. Adjustment of Mg(2+) to 6 mM did not appear to have a detrimental impact on cells. Heat inactivation was found to be incompatible with in vitro tissue culture, whereas inclusion of actin had no observable effect on growth and viability. In two in vitro assays, immune cell activation and nanoparticle endocytosis, we show that using conditions compatible with cell phenotype and nanostructure integrity is critical for obtaining reliable

A Method to Preclude Moisture Condensation in Plated Tissue Cultures

Science.gov (United States)

Alex M. Diner

1992-01-01

Excessive condensate normally accumulates in in vitro-illuminated petri dishes containing plant tissue cultures, causing avariety of problems. A dark-colored rubber net-mesh placed over the petri dishes prevented such condensation, even when charcoal-supplemented media are used under high light intensity in a growth chamber.

Methods of Monitoring Cell Fate and Tissue Growth in Three-Dimensional Scaffold-Based Strategies for In Vitro Tissue Engineering.

Science.gov (United States)

Leferink, Anne M; van Blitterswijk, Clemens A; Moroni, Lorenzo

2016-08-01

In the field of tissue engineering, there is a need for methods that allow assessing the performance of tissue-engineered constructs noninvasively in vitro and in vivo. To date, histological analysis is the golden standard to retrieve information on tissue growth, cellular distribution, and cell fate on tissue-engineered constructs after in vitro cell culture or on explanted specimens after in vivo applications. Yet, many advances have been made to optimize imaging techniques for monitoring tissue-engineered constructs with a sub-mm or μm resolution. Many imaging modalities have first been developed for clinical applications, in which a high penetration depth has been often more important than lateral resolution. In this study, we have reviewed the current state of the art in several imaging approaches that have shown to be promising in monitoring cell fate and tissue growth upon in vitro culture. Depending on the aimed tissue type and scaffold properties, some imaging methods are more applicable than others. Optical methods are mostly suited for transparent materials such as hydrogels, whereas magnetic resonance-based methods are mostly applied to obtain contrast between hard and soft tissues regardless of their transparency. Overall, this review shows that the field of imaging in scaffold-based tissue engineering is developing at a fast pace and has the potential to overcome the limitations of destructive endpoint analysis.

Tissue culture of ornamental cacti

Directory of Open Access Journals (Sweden)

Eugenio Pérez-Molphe-Balch

2015-12-01

Full Text Available Cacti species are plants that are well adapted to growing in arid and semiarid regions where the main problem is water availability. Cacti have developed a series of adaptations to cope with water scarcity, such as reduced leaf surface via morphological modifications including spines, cereous cuticles, extended root systems and stem tissue modifications to increase water storage, and crassulacean acid metabolism to reduce transpiration and water loss. Furthermore, seeds of these plants very often exhibit dormancy, a phenomenon that helps to prevent germination when the availability of water is reduced. In general, cactus species exhibit a low growth rate that makes their rapid propagation difficult. Cacti are much appreciated as ornamental plants due to their great variety and diversity of forms and their beautiful short-life flowers; however, due to difficulties in propagating them rapidly to meet market demand, they are very often over-collected in their natural habitats, which leads to numerous species being threatened, endangered or becoming extinct. Therefore, plant tissue culture techniques may facilitate their propagation over a shorter time period than conventional techniques used for commercial purposes; or may help to recover populations of endangered or threatened species for their re-introduction in the wild; or may also be of value to the preservation and conservation of the genetic resources of this important family. Herein we present the state-of-the-art of tissue culture techniques used for ornamental cacti and selected suggestions for solving a number of the problems faced by members of the Cactaceae family.

Effects of Apollo 12 lunar material on lipid levels of tobacco tissue and slash pine cultures

Science.gov (United States)

Weete, J. D.

1972-01-01

Investigations of the lipid components of pine tissues (Pinus elloitii) are discussed, emphasizing fatty acids and steroids. The response by slash pine tissue cultures to growth in contact with Apollo lunar soil, earth basalt, and Iowa soil is studied. Tissue cultures of tobacco grown for 12 weeks in contact with lunar material from Apollo 12 flight contained 21 to 35 percent more total pigment than control tissues. No differences were noted in the fresh or dry weight of the experimental and control samples.

Multiple shoot cultures of Atropa belladonna: Effect of physico-chemical factors on growth and alkaloid formation

International Nuclear Information System (INIS)

Benjamin, B.D.; Roja, P.C.; Heble, M.R.; Chandha, M.S.

1987-01-01

Multiple shoot cultures were established from shoot tip and axillary meristem of the plant Atropa belladonna. The cultures were initially raised on agar medium and subsequently maintained on liquid medium of Murashige and Skoog (1962) supplemented with BA. These cultures were subjected to different doses of -y-irradiation. Recovery from the radiation effects was observed in tissues subjected to 29 Gy during four successive passages. Plant growth regulators influenced the growth and morphogenetic events of the tissues. The precursors of tropane alkaloids marginally increased the alkaloid synthesis during the stationary phase of growth. Shoot cultures, established from different field grown plants varying in alkaloid content, were morphologically similar and did not exhibit the parental characteristics with respect to alkaloid formation

The interplay between tissue growth and scaffold degradation in engineered tissue constructs

KAUST Repository

O’Dea, R. D.

2012-09-18

In vitro tissue engineering is emerging as a potential tool to meet the high demand for replacement tissue, caused by the increased incidence of tissue degeneration and damage. A key challenge in this field is ensuring that the mechanical properties of the engineered tissue are appropriate for the in vivo environment. Achieving this goal will require detailed understanding of the interplay between cell proliferation, extracellular matrix (ECM) deposition and scaffold degradation. In this paper, we use a mathematical model (based upon a multiphase continuum framework) to investigate the interplay between tissue growth and scaffold degradation during tissue construct evolution in vitro. Our model accommodates a cell population and culture medium, modelled as viscous fluids, together with a porous scaffold and ECM deposited by the cells, represented as rigid porous materials. We focus on tissue growth within a perfusion bioreactor system, and investigate how the predicted tissue composition is altered under the influence of (1) differential interactions between cells and the supporting scaffold and their associated ECM, (2) scaffold degradation, and (3) mechanotransduction-regulated cell proliferation and ECM deposition. Numerical simulation of the model equations reveals that scaffold heterogeneity typical of that obtained from μCT scans of tissue engineering scaffolds can lead to significant variation in the flow-induced mechanical stimuli experienced by cells seeded in the scaffold. This leads to strong heterogeneity in the deposition of ECM. Furthermore, preferential adherence of cells to the ECM in favour of the artificial scaffold appears to have no significant influence on the eventual construct composition; adherence of cells to these supporting structures does, however, lead to cell and ECM distributions which mimic and exaggerate the heterogeneity of the underlying scaffold. Such phenomena have important ramifications for the mechanical integrity of

Aromatase inhibitor (anastrozole) affects growth of endometrioma cells in culture.

Science.gov (United States)

Badawy, Shawky Z A; Brown, Shereene; Kaufman, Lydia; Wojtowycz, Martha A

2015-05-01

To study the effects of aromatase inhibitor (anastrozole) on the growth and estradiol secretion of endometrioma cells in culture. Endometrioma cells are grown in vitro until maximum growth before used in this study. This was done in the research laboratory for tissue culture, in an academic hospital. Testosterone at a concentration of 10 μg/mL was added as a substrate for the intracellular aromatase. In addition, aromatase inhibitor was added at a concentration of 200 and 300 μg/mL. The effect on cell growth and estradiol secretion is evaluated using Student's t-test. The use of testosterone increased estradiol secretion by endometrioma cells in culture. The use of aromatase inhibitor significantly inhibited the growth of endometrioma cells, and estradiol secretion. Aromatase inhibitor (anastrozole) may be an effective treatment for endometriosis due to inhibition of cellular aromatase. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Prevention of pink-pigmented methylotrophic bacteria (Methylohacterium mesophilicum) contamination of plant tissue cultures.

Science.gov (United States)

Chanprame, S; Todd, J J; Widholm, J M

1996-12-01

Pink-pigmented facultative methylotrophic bacteria (PPFMs) have been found on the surfaces of leaves of most plants tested. We found PPFMs on the leaf surfaces of all 40 plants (38 species) tested and on soybean pods by pressing onto AMS medium with methanol as the sole carbon source. The abundance ranged from 0.5 colony forming unit (cfu) /cm(2) to 69.4 cfu/cm(2) on the leaf surfaces. PPFMs were found in homogenized leaf tissues of only 4 of the species after surface disinfestation with 1.05% sodium hypochlorite and were rarely found in cultures initiated from surface disinfested Datura innoxia leaves or inside surface disinfested soybean pods. Of 20 antibiotics tested for PPFM growth inhibition, rifampicin was the most effective and of seven others which also inhibited PPFM growth, cefotaxime should be the most useful due to the expected low plant cell toxicity. These antibiotics could be used in concert with common surface sterilization procedures to prevent the introduction or to eliminate PPFM bacteria in tissue cultures. Thus, while PPFMs are present on the surfaces of most plant tissues, surface disinfestation alone can effectively remove them so that uncontaminated tissue cultures can be initiated in most cases.

Growth of plant root cultures in liquid- and gas-dispersed reactor environments.

Science.gov (United States)

McKelvey, S A; Gehrig, J A; Hollar, K A; Curtis, W R

1993-01-01

The growth of Agrobacterium transformed "hairy root" cultures of Hyoscyamus muticus was examined in various liquid- and gas-dispersed bioreactor configurations. Reactor runs were replicated to provide statistical comparisons of nutrient availability on culture performance. Accumulated tissue mass in submerged air-sparged reactors was 31% of gyratory shake-flask controls. Experiments demonstrate that poor performance of sparged reactors is not due to bubble shear damage, carbon dioxide stripping, settling, or flotation of roots. Impaired oxygen transfer due to channeling and stagnation of the liquid phase are the apparent causes of poor growth. Roots grown on a medium-perfused inclined plane grew at 48% of gyratory controls. This demonstrates the ability of cultures to partially compensate for poor liquid distribution through vascular transport of nutrients. A reactor configuration in which the medium is sprayed over the roots and permitted to drain down through the root tissue was able to provide growth rates which are statistically indistinguishable (95% T-test) from gyratory shake-flask controls. In this type of spray/trickle-bed configuration, it is shown that distribution of the roots becomes a key factor in controlling the rate of growth. Implications of these results regarding design and scale-up of bioreactors to produce fine chemicals from root cultures are discussed.

Screening Test of Greenhouse Seeding Exercise Matrix for Tissue Culture Seeding of Dendrobium Officinale Kimura et Migo

Directory of Open Access Journals (Sweden)

Zhou Yuan

2015-01-01

Full Text Available The Dendrobium officinale Kimura et Migo has a high demand on planting matrix, while its tissue culture seeding has much more demands on planting matrix. To find out a seeding exercise matrix to enhance the survival rate of tissue culture seeding of Dendrobium officinale Kimura et Migo more efficiently, this article carries out a screening test of greenhouse seeding exercise matrix material for tissue culture seeding of Dendrobium officinale Kimura et Migo. The test adopts full random test design, mainly for screening test of five matrix materials, namely pine bark, camphor tree bark, fern root, peanut shell and longan bark. Compare the impact of prepared seeding exercise matrix on the survival rate and growth trend (including plant height, growth rate and bud growth rate. The test result shows that: The seeding exercise matrix prepared by fern root is the most efficient, and the survival rate, plant height, growth rate and bud growth rate have achieved 100%, 4.5cm, 43.67% and 54.33% respectively. The main reason may be that the seeding exercise matrix C prepared by fern root is fairly loose and has a great water permeability, which is conducive to the growth of Dendrobium officinale Kimura et Migo.

Adhesive protein interactions with chitosan: consequences for valve endothelial cell growth on tissue-engineering materials.

Science.gov (United States)

Cuy, Janet L; Beckstead, Benjamin L; Brown, Chad D; Hoffman, Allan S; Giachelli, Cecilia M

2003-11-01

Stable endothelialization of a tissue-engineered heart valve is essential for proper valve function, although adhesive characteristics of the native valve endothelial cell (VEC) have rarely been explored. This research evaluated VEC adhesive qualities and attempted to enhance VEC growth on the biopolymer chitosan, a novel tissue-engineering scaffold material with promising biological and chemical properties. Aortic VEC cultures were isolated and found to preferentially adhere to fibronectin, collagen types IV and I over laminin and osteopontin in a dose-dependent manner. Seeding of VEC onto comparison substrates revealed VEC growth and morphology to be preferential in the order: tissue culture polystyrene > gelatin, poly(DL-lactide-co-glycolide), chitosan > poly(hydroxy alkanoate). Adhesive protein precoating of chitosan did not significantly enhance VEC growth, despite equivalent protein adsorption as to polystyrene. Initial cell adhesion to protein-precoated chitosan, however, was higher than for polystyrene. Composite chitosan/collagen type IV films were investigated as an alternative to simple protein precoatings, and were shown to improve VEC growth and morphology over chitosan alone. These findings suggest potential manipulation of chitosan properties to improve amenability to valve tissue-engineering applications. Copyright 2003 Wiley Periodicals, Inc.

Hormonal effect on polyphenol accumulation in Cassia tissues cultured in vitro

Energy Technology Data Exchange (ETDEWEB)

Shah, R R; Subbaiah, K V; Mehta, A R

1976-06-01

Effects of auxin and kinetin on growth and production of phenolic compounds in cultured Cassia fistula L. tissues were examined. Initiation of polyphenols was largely determined by the auxin concentration in the medium. Growth of the cells in relation to accumulation of polyphenols was studied at different auxin and kinetin concentrations. The accumulation of phenolic materials was essentially restricted to the most rapid phase of the growth cycle. Progressive changes in the pattern of peroxidase activity were followed and their relationship with polyphenol synthesis is examined.

Influence of postmortem time on the outcome of blood cultures among cadaveric tissue donors.

Science.gov (United States)

Saegeman, V; Verhaegen, J; Lismont, D; Verduyckt, B; De Rijdt, T; Ectors, N

2009-02-01

Tissue banks provide tissues of human cadaver donors for transplantation. The maximal time limit for tissue retrieval has been set at 24 h postmortem. This study aimed at evaluating the evidence for this limit from a microbiological point of view. The delay of growth in postmortem blood cultures, the identification of the species isolated and clinical/environmental factors were investigated among 100 potential tissue donors. No significant difference was found in the rate of donors with grown blood cultures within (25/65=38%) compared with after (24/65=37%) 24 h of death. Coagulase-negative staphylococci and gastro-intestinal microorganisms were isolated within and after 24 h of death. Two factors--antimicrobial therapy and "delay before body cooling"--were significantly inversely related with donors' blood culture results. From a microbiological point of view, there is no evidence for avoiding tissue retrieval among donors after 24 h of death.

Effects of epidermal growth factor on neural crest cells in tissue culture

International Nuclear Information System (INIS)

Erickson, C.A.; Turley, E.A.

1987-01-01

Epidermal growth factor (EGF) stimulates the release of hyaluronic acid (HA) and chondroitin sulfate proteoglycan (CSPG) from quail trunk neural crest cultures in a dose-dependent fashion. It also promotes the expression of cell-associated heparan sulfate proteoglycan (HSPG) as detected by immunofluorescence and immunoprecipitation of the 3 H-labeled proteoglycan. Furthermore, EGF stimulates [ 3 H]thymidine incorporation into total cell DNA. These results raise the possibility that EGF or an analogous growth factor is involved in regulation of neural crest cell morphogenesis

The plant tissue culture

International Nuclear Information System (INIS)

Crocomo, O.J.; Sharp, W.R.

1973-01-01

Progress in the field of plant tissue culture at the Plant Biochemistry Sector, Centro de Energia na Agricultura (CENA), Piracicaba, S.P., Brazil, pertains to the simplification of development in 'Phaseolus vulgaris' by dividing the organism into its component organs, tissues, and cells and the maintenance of these components on defined culture media 'in vitro'. This achievement has set the stage for probing the basis for the stability of the differentiated states and/or the reentry of mature differentiated cells into the mitotic cell cycle and their subsequent redifferentiation. Data from such studies at the cytological and biochemical level have been invaluable in the elucidation of the control mechanisms responsible for expression of the cellular phenotype. Unlimited possibilities exist for the application of tissue culture in the vegetative propagation of 'Phaseolus' and other important cultivars in providing genocopies or a large scale and/or readily obtaining plantlets from haploid cell lines or from protoplast (wall-less cells) hybridization products following genetic manipulation. These tools are being applied in this laboratory for the development and selection of high protein synthesizing 'Phaseolus' cultivars

Structure and component alteration of rabbit Achilles tendon in tissue culture.

Science.gov (United States)

Hosaka, Yoshinao; Ueda, Hiromi; Yamasaki, Tadatsugu; Suzuki, Daisuke; Matsuda, Naoya; Takehana, Kazushige

2005-12-01

The aim of this study was to investigate alterations of cultured tendon tissues to determine whether tissue culture is a useful method for biological analyses of the tendon. Tendon tissues for tissue culture were isolated from Achilles tendons of rabbits. The tendon segments were placed one segment per well and incubated in growth medium consisting of Dullbecco's modified Eagle's medium supplemented with 5% fetal bovine serum at 37 degrees C in a humidified atmosphere with 5% CO(2) for various periods. The alignment of collagen fibrils was preserved for 48 h, but tendon structure has disintegrated at 96 h. Alcian blue staining and gelatine zymography revealed that proteoglycan markedly diminished and that matrix metalloproteinase (MMPs) activity was upregulated sharply at 72 and 96 h. The ratio of collagen fibrils with large diameter had increased and the mean diameter and mass average diameter value had reached maximum at 48 h. The values then decreased and mean diameters at 72 and 96 h were significantly different from that at 48 h. At 96 h, the ratio of collagen fibrils with small diameters had increased and collagen fibrils with large diameters had disappeared. These findings indicate that structural alteration is possible to be induced by disintegration of collagen fibrils and disappearance of glycosaminoglycans from extracellular matrix (ECM), subsequent of upregulation of MMPs activity. Although the study period is limited, the tissue culture method is available for investigating cell-ECM interaction in tendons.

A Protocol for Rapid, Measurable Plant Tissue Culture Using Stem Disc Meristem Micropropagation of Garlic ("Allium Sativum L.")

Science.gov (United States)

Peat, Gerry; Jones, Meriel

2012-01-01

Plant tissue culture is becoming an important technique for the mass propagation of plants. Problems with existing techniques, such as slow growth and contamination, have restricted the practical work in plant tissue culture carried out in schools. The new protocol using garlic meristematic stem discs explained in this article addresses many of…

Three-dimensional hydrogel cell culture systems for modeling neural tissue

Science.gov (United States)

Frampton, John

Two-dimensional (2-D) neural cell culture systems have served as physiological models for understanding the cellular and molecular events that underlie responses to physical and chemical stimuli, control sensory and motor function, and lead to the development of neurological diseases. However, the development of three-dimensional (3-D) cell culture systems will be essential for the advancement of experimental research in a variety of fields including tissue engineering, chemical transport and delivery, cell growth, and cell-cell communication. In 3-D cell culture, cells are provided with an environment similar to tissue, in which they are surrounded on all sides by other cells, structural molecules and adhesion ligands. Cells grown in 3-D culture systems display morphologies and functions more similar to those observed in vivo, and can be cultured in such a way as to recapitulate the structural organization and biological properties of tissue. This thesis describes a hydrogel-based culture system, capable of supporting the growth and function of several neural cell types in 3-D. Alginate hydrogels were characterized in terms of their biomechanical and biochemical properties and were functionalized by covalent attachment of whole proteins and peptide epitopes. Methods were developed for rapid cross-linking of alginate hydrogels, thus permitting the incorporation of cells into 3-D scaffolds without adversely affecting cell viability or function. A variety of neural cell types were tested including astrocytes, microglia, and neurons. Cells remained viable and functional for longer than two weeks in culture and displayed process outgrowth in 3-D. Cell constructs were created that varied in cell density, type and organization, providing experimental flexibility for studying cell interactions and behavior. In one set of experiments, 3-D glial-endothelial cell co-cultures were used to model blood-brain barrier (BBB) structure and function. This co-culture system was

Demonstration of the economic feasibility of plant tissue culture for jojoba (Simmondsia chinensis) and Euphorbia spp

Energy Technology Data Exchange (ETDEWEB)

Sluis, C.

1980-09-01

The economic feasibility of plant tissue culture was demonstrated as applied to two plants: jojoba (Simmondsia chinensis) and Euphorbia spp. The gopher weed (Euphorbia lathyris) was selected as the species of Euphorbia to research due to the interest in this plant as a potential source of hydrocarbon-like compounds. High yield female selections of jojoba were chosen from native stands and were researched to determine the economic feasibility of mass producing these plants via a tissue culture micropropagation program. The female jojoba selection was successfully mass produced through tissue culture. Modifications in initiation techniques, as well as in multiplication media and rooting parameters, were necessary to apply the tissue culture system, which had been developed for juvenile seedling tissue, to mature jojobas. Since prior attempts at transfer of tissue cultured plantlets were unsuccessful, transfer research was a major part of the project and has resulted in a system for transfer of rooted jojoba plantlets to soil. Euphorbia lathyris was successfully cultured using shoot tip cultures. Media and procedures were established for culture initiation, multiplication of shoots, callus induction and growth, and root initiation. Well-developed root systems were not attained and root initiation percentages should be increased if the system is to become commercially feasible.

A multiscale analysis of nutrient transport and biological tissue growth in vitro

KAUST Repository

O'Dea, R. D.

2014-10-15

© The authors 2014. In this paper, we consider the derivation of macroscopic equations appropriate to describe the growth of biological tissue, employing a multiple-scale homogenization method to accommodate explicitly the influence of the underlying microscale structure of the material, and its evolution, on the macroscale dynamics. Such methods have been widely used to study porous and poroelastic materials; however, a distinguishing feature of biological tissue is its ability to remodel continuously in response to local environmental cues. Here, we present the derivation of a model broadly applicable to tissue engineering applications, characterized by cell proliferation and extracellular matrix deposition in porous scaffolds used within tissue culture systems, which we use to study coupling between fluid flow, nutrient transport, and microscale tissue growth. Attention is restricted to surface accretion within a rigid porous medium saturated with a Newtonian fluid; coupling between the various dynamics is achieved by specifying the rate of microscale growth to be dependent upon the uptake of a generic diffusible nutrient. The resulting macroscale model comprises a Darcy-type equation governing fluid flow, with flow characteristics dictated by the assumed periodic microstructure and surface growth rate of the porous medium, coupled to an advection-reaction equation specifying the nutrient concentration. Illustrative numerical simulations are presented to indicate the influence of microscale growth on macroscale dynamics, and to highlight the importance of including experimentally relevant microstructural information to correctly determine flow dynamics and nutrient delivery in tissue engineering applications.

Laboratory Workflow Analysis of Culture of Periprosthetic Tissues in Blood Culture Bottles.

Science.gov (United States)

Peel, Trisha N; Sedarski, John A; Dylla, Brenda L; Shannon, Samantha K; Amirahmadi, Fazlollaah; Hughes, John G; Cheng, Allen C; Patel, Robin

2017-09-01

Culture of periprosthetic tissue specimens in blood culture bottles is more sensitive than conventional techniques, but the impact on laboratory workflow has yet to be addressed. Herein, we examined the impact of culture of periprosthetic tissues in blood culture bottles on laboratory workflow and cost. The workflow was process mapped, decision tree models were constructed using probabilities of positive and negative cultures drawn from our published study (T. N. Peel, B. L. Dylla, J. G. Hughes, D. T. Lynch, K. E. Greenwood-Quaintance, A. C. Cheng, J. N. Mandrekar, and R. Patel, mBio 7:e01776-15, 2016, https://doi.org/10.1128/mBio.01776-15), and the processing times and resource costs from the laboratory staff time viewpoint were used to compare periprosthetic tissues culture processes using conventional techniques with culture in blood culture bottles. Sensitivity analysis was performed using various rates of positive cultures. Annualized labor savings were estimated based on salary costs from the U.S. Labor Bureau for Laboratory staff. The model demonstrated a 60.1% reduction in mean total staff time with the adoption of tissue inoculation into blood culture bottles compared to conventional techniques (mean ± standard deviation, 30.7 ± 27.6 versus 77.0 ± 35.3 h per month, respectively; P < 0.001). The estimated annualized labor cost savings of culture using blood culture bottles was $10,876.83 (±$337.16). Sensitivity analysis was performed using various rates of culture positivity (5 to 50%). Culture in blood culture bottles was cost-effective, based on the estimated labor cost savings of $2,132.71 for each percent increase in test accuracy. In conclusion, culture of periprosthetic tissue in blood culture bottles is not only more accurate than but is also cost-saving compared to conventional culture methods. Copyright © 2017 American Society for Microbiology.

Potency testing of mesenchymal stromal cell growth expanded in human platelet lysate from different human tissues.

Science.gov (United States)

Fazzina, R; Iudicone, P; Fioravanti, D; Bonanno, G; Totta, P; Zizzari, I G; Pierelli, L

2016-08-25

Mesenchymal stromal cells (MSCs) have been largely investigated, in the past decade, as potential therapeutic strategies for various acute and chronic pathological conditions. MSCs isolated from different sources, such as bone marrow (BM), umbilical cord tissue (UCT) and adipose tissue (AT), share many biological features, although they may show some differences on cumulative yield, proliferative ability and differentiation potential. The standardization of MSCs growth and their functional amplification is a mandatory objective of cell therapies. The aim of this study was to evaluate the cumulative yield and the ex vivo amplification potential of MSCs obtained from various sources and different subjects, using defined culture conditions with a standardized platelet lysate (PL) as growth stimulus. MSCs isolated from BM, UCT and AT and expanded in human PL were compared in terms of cumulative yield and growth potential per gram of starting tissue. MSCs morphology, phenotype, differentiation potential, and immunomodulatory properties were also investigated to evaluate their biological characteristics. The use of standardized PL-based culture conditions resulted in a very low variability of MSC growth. Our data showed that AT has the greater capacity to generate MSC per gram of initial tissue, compared to BM and UCT. However, UCT-MSCs replicated faster than AT-MSCs and BM-MSCs, revealing a greater proliferation capacity of this source irrespective of its lower MSC yield. All MSCs exhibited the typical MSC phenotype and the ability to differentiate into all mesodermal lineages, while BM-MSCs showed the most prominent immunosuppressive effect in vitro. The adoption of standardized culture conditions may help researchers and clinicians to reveal particular characteristics and inter-individual variability of MSCs sourced from different tissues. These data will be beneficial to set the standards for tissue collection and MSCs clinical-scale expansion both for cell banking

Comparison of Fluorescence Microscopy and Different Growth Media Culture Methods for Acanthamoeba Keratitis Diagnosis.

Science.gov (United States)

Peretz, Avi; Geffen, Yuval; Socea, Soergiu D; Pastukh, Nina; Graffi, Shmuel

2015-08-01

Acanthamoeba keratitis (AK), a potentially blinding infection of the cornea, is caused by a free-living protozoan. Culture and microscopic examination of corneal scraping tissue material is the conventional method for identifying Acanthamoeba. In this article, we compared several methods for AK diagnosis of 32 patients: microscopic examination using fluorescent dye, specific culture on growth media-non-nutrient agar (NNA), culture on liquid growth media-peptone yeast glucose (PYG), and TYI-S-33. AK was found in 14 patients. Thirteen of the specimens were found AK positive by fluorescence microscopic examination, 11 specimens were found AK positive on PYG growth media, and 9 specimens were found AK positive on TYI-S-33 growth media. Only five specimens were found AK positive on NNA growth media. Therefore, we recommend using fluorescence microscopy technique and culture method, especially PYG liquid media. © The American Society of Tropical Medicine and Hygiene.

Influence of epidermal growth factor (EGF) and hydrocortisone on the co-culture of mature adipocytes and endothelial cells for vascularized adipose tissue engineering.

Science.gov (United States)

Huber, Birgit; Czaja, Alina Maria; Kluger, Petra Juliane

2016-05-01

The composition of vascularized adipose tissue is still an ongoing challenge as no culture medium is available to supply adipocytes and endothelial cells appropriately. Endothelial cell medium is typically supplemented with epidermal growth factor (EGF) as well as hydrocortisone (HC). The effect of EGF on adipocytes is discussed controversially. Some studies say it inhibits adipocyte differentiation while others reported of improved adipocyte lipogenesis. HC is known to have lipolytic activities, which might result in mature adipocyte dedifferentiation. In this study, we evaluated the influence of EGF and HC on the co-culture of endothelial cells and mature adipocytes regarding their cell morphology and functionality. We showed in mono-culture that high levels of HC promoted dedifferentiation and proliferation of mature adipocytes, whereas EGF seemed to have no negative influence. Endothelial cells kept their typical cobblestone morphology and showed a proliferation rate comparable to the control independent of EGF and HC concentration. In co-culture, HC promoted dedifferentiation of mature adipocytes, which was shown by a higher glycerol release. EGF had no negative impact on adipocyte morphology. No negative impact on endothelial cell morphology and functionality could be seen with reduced EGF and HC supplementation in co-culture with mature adipocytes. Taken together, our results demonstrate that reduced levels of HC are needed for co-culturing mature adipocytes and endothelial cells. In co-culture, EGF had no influence on mature adipocytes. Therefore, for the composition of vascularized adipose tissue constructs, the media with low levels of HC and high or low levels of EGF can be used. © 2016 International Federation for Cell Biology.

Human colon tissue in organ culture: calcium and multi-mineral-induced mucosal differentiation.

Science.gov (United States)

Dame, Michael K; Veerapaneni, Indiradevi; Bhagavathula, Narasimharao; Naik, Madhav; Varani, James

2011-01-01

We have recently shown that a multi-mineral extract from the marine red algae, Lithothamnion calcareum, suppresses colon polyp formation and inflammation in mice. In the present study, we used intact human colon tissue in organ culture to compare responses initiated by Ca(2+) supplementation versus the multi-mineral extract. Normal human colon tissue was treated for 2 d in culture with various concentrations of calcium or the mineral-rich extract. The tissue was then prepared for histology/immunohistochemistry, and the culture supernatants were assayed for levels of type I procollagen and type I collagen. At higher Ca(2+) concentrations or with the mineral-rich extract, proliferation of epithelial cells at the base and walls of the mucosal crypts was suppressed, as visualized by reduced Ki67 staining. E-cadherin, a marker of differentiation, was more strongly expressed at the upper third of the crypt and at the luminal surface. Treatment with Ca(2+) or with the multi-mineral extract influenced collagen turnover, with decreased procollagen and increased type I collagen. These data suggest that calcium or mineral-rich extract has the capacity to (1) promote differentiation in human colon tissue in organ culture and (2) modulate stromal function as assessed by increased levels of type I collagen. Taken together, these data suggest that human colon tissue in organ culture (supporting in vivo finding in mice) will provide a valuable model for the preclinical assessment of agents that regulate growth and differentiation in the colonic mucosa.

Progress in planta transformation without tissue culture

International Nuclear Information System (INIS)

Gu Yunhong; Chinese Academy of Sciences, Hefei; Qin Guangyong; Huo Yuping; Yu Zengliang

2004-01-01

With the development of planta genetic engineering, more emphases have been laid on convenient and high efficient genetic transformation methods. And transformation without tissue culture is a prospective direction of it. In this paper, traditional transformation methods and the methods of non-tissue culture were summarized. With the exploration and application of Arabidopsis transformation mechanism, with the use of ion beam-mediated transformation invented by Chinese scientists and the development of other transformation methods, transformation methods without tissue culture and planta genetic engineering could be improved rapidly. (authors)

[Regeneration of autologous tissue-engineered cartilage by using basic-fibroblast growth factor in vitro culture].

Science.gov (United States)

Ding, Xiao-bang; Cheng, Ning-xin; Chen, Bing; Xia, Wan-yao; Cui, Lei; Liu, Wei; Cao, Yi-lin

2004-05-01

To investigate the effect of the basic fibroblast growth factor (b-FGF) to regenerate an autologous tissue-engineered cartilage in vitro. The Cells were harvested from the elastic auricular cartilage of swine,and were plated at the concentration of 1 x 10(4) cells/cm2 , studied in vitro at two different media enviroments: Group I contained Ham's F-12 with supplements and b-FGF, Group II contained Ham's F-12 only with supplements. The passage 2 cells (after 12.75 +/- 1.26 days) were harvested and mixed with 30% pluronic F-127/Ham's F-12 at the concentration of 50 x 10(6) cells/ml. It was injected subcutaneously at 0.5 ml per implant. The implants were harvested 8 weeks after the vivo culture and examined with the histological stains. The chondrocytes displayed morphologically similar to the fibroblasts in the media containing basic-FGF. The number of cell doublings (after 12.75 +/- 1.26 days) in vitro culture was as the following: Group I, 70; Group II, 5.4. Eight 8 weeks after the vivo autologous implantation, the average weight (g) and volume (cm3) in each group was as the following: Group I, 0.371 g/0.370 cm3 Group II, 0.179 g/0.173 cm3 (P < 0.01). With the b-FGF in vitro culture, the cells were expanded by 70 times after 2 weeks. Histologically, all of the engineered cartilage in the two groups were similar to the native elastic cartilage. These results indicate that the basic-FGF could be used positively to enhance the quality and quantity of the seeding cells for the generation of the well-engineered cartilage.

Morphological, biochemical and genetic influence of mutagen treatments on medicinal plant tissue cultures

International Nuclear Information System (INIS)

Onisei, T.; Toth, E.; Tesio, B.; Floria, F.

1994-01-01

Gamma rays and/or alkylant agents have been applied on callus tissue, young regenerants and cell suspension in order to establish their effect on morphogenesis, regeneration ability and biosynthetic potential. Growth dynamics, morpho-anatomic variables, secondary metabolite production, cell cytogenetics, enzyme specific activities, isoperoxidase and isoesterase patterns were analyzed in relation to the morphogenetic response of Atropa belladonna, Datura innoxia, Lavandula angustifolia, Chamomilla recutita, Digitalis lanata and Vinca minor tissue cultures. The effects of gamma-ray doses varied from one species to another; 10 to 20 Gy were generally able to stimulate growth and plant regeneration (via organogenesis and somatic embryogenesis), while 10 to 50 Gy enhanced secondary metabolite biosynthesis both in callus and cell suspension culture. Semnificative increase of secondary metabolite production was obtained when treatments with EMS (0.1-0.2%) have been applied to young regenerants. Many differences in biological features and biochemical behaviour were registered 20 days and one year, respectively, after treatment. (author)

Effect of lunar materials on plant tissue culture.

Science.gov (United States)

Walkinshaw, C. H.; Venketeswaran, S.; Baur, P. S.; Croley, T. E.; Scholes, V. E.; Weete, J. D.; Halliwell, R. S.; Hall, R. H.

1973-01-01

Lunar material collected during the Apollo 11, 12, 14, and 15 missions has been used to treat 12 species of higher plant tissue cultures. Biochemical and morphological studies have been conducted on several of these species. Tobacco tissue cultures treated with 0.22 g of lunar material exhibited increased greening more complex chloroplasts, less cytoplasmic vacuolation and greater vesiculation. Pine tissue cultures reacted to treatment by an increased deposition of tannin-like materials. The percentage of dry weight and soluble protein was increased in cultures treated with either lunar or terrestrial rock materials.

Dynamics of anisotropic tissue growth

Energy Technology Data Exchange (ETDEWEB)

Bittig, Thomas; Juelicher, Frank [Max Planck Institute for the Physics of Complex Systems, Noethnitzer Strasse 38, 01187 Dresden (Germany); Wartlick, Ortrud; Kicheva, Anna; Gonzalez-Gaitan, Marcos [Department of Biochemistry and Department of Molecular Biology, Geneva University, Sciences II, Quai Ernest-Ansermet 30, 1211 Geneva 4 (Switzerland)], E-mail: Marcos.Gonzalez@biochem.unige.ch, E-mail: julicher@pks.mpg.de

2008-06-15

We study the mechanics of tissue growth via cell division and cell death (apoptosis). The rearrangements of cells can on large scales and times be captured by a continuum theory which describes the tissue as an effective viscous material with active stresses generated by cell division. We study the effects of anisotropies of cell division on cell rearrangements and show that average cellular trajectories exhibit anisotropic scaling behaviors. If cell division and apoptosis balance, there is no net growth, but for anisotropic cell division the tissue undergoes spontaneous shear deformations. Our description is relevant for the study of developing tissues such as the imaginal disks of the fruit fly Drosophila melanogaster, which grow anisotropically.

Participation of cob tissue in the transport of medium components into maize kernels cultured in vitro

International Nuclear Information System (INIS)

Felker, F.C.

1990-01-01

Maize (Zea mays L.) kernels cultured in vitro while still attached to cob pieces have been used as a model system to study the physiology of kernel development. In this study, the role of the cob tissue in uptake of medium components into kernels was examined. Cob tissue was essential for in vitro kernel growth, and better growth occurred with larger cob/kernel ratios. A symplastically transported fluorescent dye readily permeated the endosperm when supplied in the medium, while an apoplastic dye did not. Slicing the cob tissue to disrupt vascular connections, but not apoplastic continuity, greatly reduced [ 14 C]sucrose uptake into kernels. [ 14 C]Sucrose uptake by cob and kernel tissue was reduced 31% and 68%, respectively, by 5 mM PCMBS. L-[ 14 C]glucose was absorbed much more slowly than D-[ 14 C]glucose. These and other results indicate that phloem loading of sugars occurs in the cob tissue. Passage of medium components through the symplast cob tissue may be a prerequisite for uptake into the kernel. Simple diffusion from the medium to the kernels is unlikely. Therefore, the ability of substances to be transported into cob tissue cells should be considered in formulating culture medium

Macroporous Hydrogel Scaffolds for Three-Dimensional Cell Culture and Tissue Engineering.

Science.gov (United States)

Fan, Changjiang; Wang, Dong-An

2017-10-01

Hydrogels have been promising candidate scaffolds for cell delivery and tissue engineering due to their tissue-like physical properties and capability for homogeneous cell loading. However, the encapsulated cells are generally entrapped and constrained in the submicron- or nanosized gel networks, seriously limiting cell growth and tissue formation. Meanwhile, the spatially confined settlement inhibits attachment and spreading of anchorage-dependent cells, leading to their apoptosis. In recent years, macroporous hydrogels have attracted increasing attention in use as cell delivery vehicles and tissue engineering scaffolds. The introduction of macropores within gel scaffolds not only improves their permeability for better nutrient transport but also creates space/interface for cell adhesion, proliferation, and extracellular matrix deposition. Herein, we will first review the development of macroporous gel scaffolds and outline the impact of macropores on cell behaviors. In the first part, the advantages and challenges of hydrogels as three-dimensional (3D) cell culture scaffolds will be described. In the second part, the fabrication of various macroporous hydrogels will be presented. Third, the enhancement of cell activities within macroporous gel scaffolds will be discussed. Finally, several crucial factors that are envisaged to propel the improvement of macroporous gel scaffolds are proposed for 3D cell culture and tissue engineering.

Smallholder adoption and economic impacts of tissue culture ...

African Journals Online (AJOL)

This study was conducted with an objective of determining the correlates of adoption of tissue culture banana technology and its impacts on household incomes in Kenya. The results show that while some households have opted not to adopt tissue culture banana biotechnology, almost all the adopters are growing tissue ...

Walnut tissue culture: research and field applications

Science.gov (United States)

2004-01-01

Vitrotech Biotecnologia Vegetal began researching propagating Juglans regia (English walnut) and various Juglans hybrids by tissue culture in 1993 and has operated on a commercial scale since 1996. Since this time, more than one and a half million walnuts of different species have been propagated and field planted. Tissue cultured...

Biotransformations with plant tissue cultures.

Science.gov (United States)

Carew, D P; Bainbridge, T

1976-01-01

Suspension cultures of Catharanthus roseus, Apocynum cannabinum and Conium maculatum were examined for their capacity to transform aniline, anisole, acetanilide, benzoic acid and coumarin. None of the cultures transformed acetanilide but each produced acetanilide when fed aniline. All three cultures converted benzoic acid to its para-hydroxy derivative. Coumarin was selectively hydroxylated at the 7-position by Catharanthus and Conium and anisole was O-demethylated only by older Catharanthus tissue.

Effects of glucose irradiated by high doses of 60cobalt gamma rays, and of some products of glucose radiolysis on the growth of Jerusalem Artichoke tissue and potato shoots culture in vitro

International Nuclear Information System (INIS)

Manant, Pierre

1975-01-01

Glucose, irradiated in dry conditions by gamma rays from 5.10 5 to 10 7 rad, and incorporated into culture medium, inhibits growth and, simultaneously, increases rhizogenesis of Jerusalem Artichoke tissue in culture. Tuberisation of potato shoots grown in vitro is delayed and partially inhibited. Some substances which result from radiolysis of sugars give the same results, but only at higher concentrations [fr

Tissue culture as a plant production technique for horticultural crops ...

African Journals Online (AJOL)

Over 100 years ago, Haberlandt envisioned the concept of plant tissue culture and provided the groundwork for the cultivation of plant cells, tissues and organs in culture. Initially plant tissue cultures arose as a research tool and focused on attempts to culture and study the development of small, isolated cells and segments ...

Development of an Autonomous, Dual Chamber Bioreactor for the Growth of 3-Dimensional Epithelial-Stromal Tissues in Microgravity

Science.gov (United States)

Patel, Zarana S.; Wettergreen, Matthew A.; Huff, Janice L.

2014-01-01

We are developing a novel, autonomous bioreactor that can provide for the growth and maintenance in microgravity of 3-D organotypic epithelial-stromal cultures that require an air-liquid interface. These complex 3-D tissue models accurately represent the morphological features, differentiation markers, and growth characteristics observed in normal human epithelial tissues, including the skin, esophagus, lung, breast, pancreas, and colon. However, because of their precise and complex culture requirements, including that of an air-liquid interface, these 3-D models have yet to be utilized for life sciences research aboard the International Space Station. The development of a bioreactor for these cultures will provide the capability to perform biological research on the ISS using these realistic, tissue-like human epithelial-stromal cell models and will contribute significantly to advances in fundamental space biology research on questions regarding microgravity effects on normal tissue development, aging, cancer, and other disease processes. It will also allow for the study of how combined stressors, such as microgravity with radiation and nutritional deficiencies, affect multiple biological processes and will provide a platform for conducting countermeasure investigations on the ISS without the use of animal models. The technology will be autonomous and consist of a cell culture chamber that provides for air-liquid, liquid-liquid, and liquid-air exchanges within the chambers while maintaining the growth and development of the biological samples. The bioreactor will support multiple tissue types and its modular design will provide for incorporation of add-on capabilities such as microfluidics drug delivery, media sampling, and in situ biomarker analysis. Preliminary flight testing of the hardware will be conducted on a parabolic platform through NASA's Flight Opportunities Program.

Application of Hanging Drop Technique for Kidney Tissue Culture.

Science.gov (United States)

Wang, Shaohui; Wang, Ximing; Boone, Jasmine; Wie, Jin; Yip, Kay-Pong; Zhang, Jie; Wang, Lei; Liu, Ruisheng

2017-01-01

The hanging drop technique is a well-established method used in culture of animal tissues. However, this method has not been used in adult kidney tissue culture yet. This study was to explore the feasibility of using this technique for culturing adult kidney cortex to study the time course of RNA viability in the tubules and vasculature, as well as the tissue structural integrity. In each Petri dish with the plate covered with sterile buffer, a section of mouse renal cortex was cultured within a drop of DMEM culture medium on the inner surface of the lip facing downward. The tissue were then harvested at each specific time points for Real-time PCR analysis and histological studies. The results showed that the mRNA level of most Na+ related transporters and cotransporters were stably maintained within 6 hours in culture, and that the mRNA level of most receptors found in the vasculature and glomeruli were stably maintained for up to 9 days in culture. Paraffin sections of the cultured renal cortex indicated that the tubules began to lose tubular integrity after 6 hours, but the glomeruli and vasculatures were still recognizable up to 9 days in culture. We concluded that adult kidney tissue culture by hanging drop method can be used to study gene expressions in vasculature and glomeruli. © 2017 The Author(s). Published by S. Karger AG, Basel.

Application of Hanging Drop Technique for Kidney Tissue Culture

Directory of Open Access Journals (Sweden)

Shaohui Wang

2017-05-01

Full Text Available Background/Aims: The hanging drop technique is a well-established method used in culture of animal tissues. However, this method has not been used in adult kidney tissue culture yet. This study was to explore the feasibility of using this technique for culturing adult kidney cortex to study the time course of RNA viability in the tubules and vasculature, as well as the tissue structural integrity. Methods: In each Petri dish with the plate covered with sterile buffer, a section of mouse renal cortex was cultured within a drop of DMEM culture medium on the inner surface of the lip facing downward. The tissue were then harvested at each specific time points for Real-time PCR analysis and histological studies. Results: The results showed that the mRNA level of most Na+ related transporters and cotransporters were stably maintained within 6 hours in culture, and that the mRNA level of most receptors found in the vasculature and glomeruli were stably maintained for up to 9 days in culture. Paraffin sections of the cultured renal cortex indicated that the tubules began to lose tubular integrity after 6 hours, but the glomeruli and vasculatures were still recognizable up to 9 days in culture. Conclusions: We concluded that adult kidney tissue culture by hanging drop method can be used to study gene expressions in vasculature and glomeruli.

Picroside I and Picroside II from Tissue Cultures of Picrorhiza kurroa

Science.gov (United States)

Ganeshkumar, Yamjala; Ramarao, Ajmera; Veeresham, Ciddi

2017-01-01

Background: Picrorhiza kurroa (PK) belongs to Scrophulariaceae family and is a representative endemic, medicinal herb, widely distributed throughout the higher altitudes of alpine Himalayas from west to east, between 3000 and 4500 m above mean sea level. Objective: The objective of the present study is to assess the production of picroside I and picroside II from tissue cultures of PK. Materials and Methods: Auxiliary shoot tips of PK were incubated in Murashige and Skoog medium supplemented with indole-3-butyric acid and kinetin phytohormones. The callus produced was collected at different time intervals and was processed for extraction of picroside I and picroside II followed by thin layer chromatography and high-performance liquid chromatography HPLC analysis. Results: The maximum growth index was found to be 5.109 ± 0.159 at 16-week-old callus culture. The estimation of picroside-I and picroside-II was carried out by (HPLC) analysis; quantity of secondary metabolite found to be 16.37 ± 0.0007 mg/g for PK-I and 6.34 ± 0.0012 mg/g for PK-II. Conclusion: This is the first attempt to produce the Picroside-I and II in large amount by the tissue culture technique. It can be observed that the method of callus culture can be used in production of secondary metabolites Picroside-I and II from PK SUMMARY Picrorhiza kurroa is a high value medicinal herb due to rich source of hepatoprotective metabolites, Picroside-I and Picroside-II. The medicinal importance of P. kurroa is due to its pharmacological properties like hepatoprotective, antioxidant (particularly in liver), antiallergic and antiasthamatic, anticancer activity particularly in liver and immunomodulatory. Shoot apices which were produced a good response was inoculated on selected medium i.e., on MS medium containing 2, 4 D (mg/l) + KN (1mg/l) for induction of callus. The initiation of callus was observed after 4weeks and it was light green and fragile Maximum growth was observed with 3% w/v of sucrose

The use of animal tissues alongside human tissue: Cultural and ethical considerations.

Science.gov (United States)

Kaw, Anu; Jones, D Gareth; Zhang, Ming

2016-01-01

Teaching and research facilities often use cadaveric material alongside animal tissues, although there appear to be differences in the way we handle, treat, and dispose of human cadaveric material compared to animal tissue. This study sought to analyze cultural and ethical considerations and provides policy recommendations on the use of animal tissues alongside human tissue. The status of human and animal remains and the respect because of human and animal tissues were compared and analyzed from ethical, legal, and cultural perspectives. The use of animal organs and tissues is carried out within the context of understanding human anatomy and function. Consequently, the interests of human donors are to be pre-eminent in any policies that are enunciated, so that if any donors find the presence of animal remains unacceptable, the latter should not be employed. The major differences appear to lie in differences in our perceptions of their respective intrinsic and instrumental values. Animals are considered to have lesser intrinsic value and greater instrumental value than humans. These differences stem from the role played by culture and ethical considerations, and are manifested in the resulting legal frameworks. In light of this discussion, six policy recommendations are proposed, encompassing the nature of consent, respect for animal tissues as well as human remains, and appropriate separation of both sets of tissues in preparation and display. © 2015 Wiley Periodicals, Inc.

Lemongrass-Incorporated Tissue Conditioner Against Candida albicans Culture

Science.gov (United States)

Amornvit, Pokpong; Srithavaj, Theerathavaj

2014-01-01

Background: Tissue conditioner is applied popularly with dental prosthesis during wound healing process but it becomes a reservoir of oral microbiota, especially Candida species after long-term usage. Several antifungal drugs have been mixed with this material to control fungal level. In this study, lemongrass essential oil was added into COE-COMFORT tissue conditioner before being determined for anti-Candida efficacy. Materials and Methods: Lemongrass (Cymbopogon citratus) essential oil was primarily determined for antifungal activity against C. albicans American type culture collection (ATCC) 10231 and MIC (minimum inhibitory concentration) value by agar disk diffusion and broth microdilution methods, respectively. COE-COMFORT tissue conditioner was prepared as recommended by the manufacturer after a fixed volume of the oil at its MIC or higher concentrations were mixed thoroughly in its liquid part. Antifungal efficacy of the tissue conditioner with/without herb was finally analyzed. Results: Lemongrass essential oil displayed potent antifungal activity against C. albicans ATCC 10231and its MIC value was 0.06% (v/v). Dissimilarly, the tissue conditioner containing the oil at MIC level did not cease the growth of the tested fungus. Both reference and clinical isolates of C. albicans were completely inhibited after exposed to the tissue conditioner containing at least 0.25% (v/v) of the oil (approximately 4-time MIC). The tissue conditioner without herb or with nystatin was employed as negative or positive control, respectively. Conclusion: COE-COMFORT tissue conditioner supplemented with lemongrass essential oil obviously demonstrated another desirable property as in vitro anti-Candida efficacy to minimize the risk of getting Candidal infection. PMID:25177638

Use of fibroblast growth factor 2 for expansion of chondrocytes and tissue engineering

Science.gov (United States)

Vunjak-Novakovic, Gordana (Inventor); Martin, Ivan (Inventor); Freed, Lisa E. (Inventor); Langer, Robert (Inventor)

2003-01-01

The present invention provides an improved method for expanding cells for use in tissue engineering. In particular the method provides specific biochemical factors to supplement cell culture medium during the expansion process in order to reproduce events occurring during embryonic development with the goal of regenerating tissue equivalents that resemble natural tissues both structurally and functionally. These specific biochemical factors improve proliferation of the cells and are capable of de-differentiation mature cells isolated from tissue so that the differentiation potential of the cells is preserved. The bioactive molecules also maintain the responsiveness of the cells to other bioactive molecules. Specifically, the invention provides methods for expanding chondrocytes in the presence of fibroblast growth factor 2 for use in regeneration of cartilage tissue.

Citrus tissue culture employing vegetative explants.

Science.gov (United States)

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

2001-11-01

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

Tissue culture of black pepper (piper nigrum l.) in Pakistan

International Nuclear Information System (INIS)

Hussain, A.; Naz, S.; Nazir, H.; Shinwari, Z.K.

2011-01-01

Black pepper (Piper nigrum L.) the 'King of Spices' is a universal table condiment. It is extensively used in Pakistani cuisines and herbal medicines and imported in bulk from neighboring countries. The black pepper vine is generally cultivated by seed because other vegetative propagation methods are slow and time consuming. Therefore the tissue culture technique is considered more efficient and reliable method for rapid and mass propagation of this economically important plant. The present study was initiated to develop protocol for micro-propagation of black pepper vine. The stem, leaf and shoot tip explants from mature vine were cultured on MS medium supplemented with different concentrations of plant growth regulators (2,4-D, BA, IBA). Best callus was produced on MS medium with 1.5 mg/l BA by shoot tip explant. Shoot regeneration was excellent on MS medium with 0.5 mg/l BA. The plantlets formed were rooted best on 1.5 mg/l IBA. The rooted plants were transplanted in soil medium and acclimatized in growth room. The plants raised were test planted under the local conditions of Hattar. (author)

Production of mutants by irradiation of in vitro-cultured tissues of coconut and banana and their mass propagation by the tissue culture technique

International Nuclear Information System (INIS)

Guzman, E.V. de; Rosario, A.G. del; Pagcaliwagan, P.C.

1982-01-01

Regeneration of buds/shoots as well as plantlets was induced from banana shoot tip explants cultured in highly modified Murashige and Skoog's medium supplemented with coconut water and benzyladenine. Initially shoot regeneration was sparse, but on further subculture became profuse. Gamma irradiation at low dosage (1.0 kR) was stimulating to explant growth and bud formation with the two types of explants used. With Bungulan stimulation was observed even at 2.5 kR. Several morphological aberrations were exhibited by shoots of 'irradiated' in vitro plants growing in potted soil. A highly and continuously proliferating tissue strain has been isolated from a subculture which was ultimately derived from an irradiated explant. Its continued proliferation is dependent on an external supply of coconut water and benzyladenine. In vitro-produced plants have been established under field conditions. The 'irradiated' plants are comparable with, and some seem to be better than, the unirradiated controls with respect to height, girth, sucker production and number of hands and fingers per bunch. Higher doses of irradiation are required to produce an adverse effect on growth of coconut embryos during the liquid culture than when growing in solid medium. (author)

Lactococcus lactis Metabolism and Gene Expression during Growth on Plant Tissues

Science.gov (United States)

Golomb, Benjamin L.

2014-01-01

Lactic acid bacteria have been isolated from living, harvested, and fermented plant materials; however, the adaptations these bacteria possess for growth on plant tissues are largely unknown. In this study, we investigated plant habitat-specific traits of Lactococcus lactis during growth in an Arabidopsis thaliana leaf tissue lysate (ATL). L. lactis KF147, a strain originally isolated from plants, exhibited a higher growth rate and reached 7.9-fold-greater cell densities during growth in ATL than the dairy-associated strain L. lactis IL1403. Transcriptome profiling (RNA-seq) of KF147 identified 853 induced and 264 repressed genes during growth in ATL compared to that in GM17 laboratory culture medium. Genes induced in ATL included those involved in the arginine deiminase pathway and a total of 140 carbohydrate transport and metabolism genes, many of which are involved in xylose, arabinose, cellobiose, and hemicellulose metabolism. The induction of those genes corresponded with L. lactis KF147 nutrient consumption and production of metabolic end products in ATL as measured by gas chromatography-time of flight mass spectrometry (GC-TOF/MS) untargeted metabolomic profiling. To assess the importance of specific plant-inducible genes for L. lactis growth in ATL, xylose metabolism was targeted for gene knockout mutagenesis. Wild-type L. lactis strain KF147 but not an xylA deletion mutant was able to grow using xylose as the sole carbon source. However, both strains grew to similarly high levels in ATL, indicating redundancy in L. lactis carbohydrate metabolism on plant tissues. These findings show that certain strains of L. lactis are well adapted for growth on plants and possess specific traits relevant for plant-based food, fuel, and feed fermentations. PMID:25384484

Ex Vivo Growth of Bioengineered Ligaments and Other Tissues

Science.gov (United States)

Altman, Gregory; Kaplan, David L.; Martin, Ivan; Vunjak-Novakovic, Gordana

2005-01-01

A method of growing bioengineered tissues for use in surgical replacement of damaged anterior cruciate ligaments has been invented. An anterior cruciate ligament is one of two ligaments (the other being the posterior cruciate ligament) that cross in the middle of a knee joint and act to prevent the bones in the knee from sliding forward and backward relative to each other. Anterior cruciate ligaments are frequently torn in sports injuries and traffic accidents, resulting in pain and severe limitations on mobility. By making it possible to grow replacement anterior cruciate ligaments that structurally and functionally resemble natural ones more closely than do totally synthetic replacements, the method could create new opportunities for full or nearly full restoration of functionality in injured knees. The method is also adaptable to the growth of bioengineered replacements for other ligaments (e.g., other knee ligaments as well as those in the hands, wrists, and elbows) and to the production of tissues other than ligaments, including cartilage, bones, muscles, and blood vessels. The method is based on the finding that the histomorphological properties of a bioengineered tissue grown in vitro from pluripotent cells within a matrix are affected by the direct application of mechanical force to the matrix during growth generation. This finding provides important new insights into the relationships among mechanical stress, biochemical and cell-immobilization methods, and cell differentiation, and is applicable to the production of the variety of tissues mentioned above. Moreover, this finding can be generalized to nonmechanical (e.g., chemical and electromagnetic) stimuli that are experienced in vivo by tissues of interest and, hence, the method can be modified to incorporate such stimuli in the ex vivo growth of replacements for the various tissues mentioned above. In this method, a three-dimensional matrix made of a suitable material is seeded with pluripotent stem

Non-exponential growth of Mycobacterium leprae Thai-53 strain cultured in vitro.

Science.gov (United States)

Amako, Kazunobu; Iida, Ken-Ichiro; Saito, Mitsumasa; Ogura, Yoshitoshi; Hayashi, Tetsuya; Yoshida, Shin-Ichi

2016-12-01

In this study, attempts were made to culture this bacterium in media supplemented with a variety of biological materials to determine why cultivation of Mycobacterium leprae in vitro has not this far been successful. A slight increase in the number of cells in medium supplemented with human blood plasma and an extract of nude mouse tissue as observed after more than 3 months of cultivation at 30 °C. To ascertain whether this increase was real growth, the growth was analyzed by droplet digital PCR, which showed a slow increase in the copy number of cell-associated DNA and the release of a large amount of DNA into the culture medium from bacterial cells during cultivation. These results were supported by electron microscopic examination of M. leprae in infected mouse tissues, which showed that most of the replicated bacteria had degenerated and only a few cells survived. Based on these results, it was postulated that many of the replicated cells degenerate during M. leprae growth and that only a few cells remain to participate in the next growth stage. This means that, unlike other cultivable bacteria, the growth of M. leprae is not exponential and the number of cells therefore increase extremely slowly. Thus, accurate judging of the success of M. leprae cultivation requires observation of growth over a long period of time and careful measurement of the increase in number of viable cells. © 2016 The Authors. Microbiology and Immunology published by The Societies and John Wiley & Sons Australia, Ltd.

The growth of tissue engineering.

Science.gov (United States)

Lysaght, M J; Reyes, J

2001-10-01

This report draws upon data from a variety of sources to estimate the size, scope, and growth rate of the contemporary tissue engineering enterprise. At the beginning of 2001, tissue engineering research and development was being pursued by 3,300 scientists and support staff in more than 70 startup companies or business units with a combined annual expenditure of over $600 million. Spending by tissue engineering firms has been growing at a compound annual rate of 16%, and the aggregate investment since 1990 now exceeds $3.5 billion. At the beginning of 2001, the net capital value of the 16 publicly traded tissue engineering startups had reached $2.6 billion. Firms focusing on structural applications (skin, cartilage, bone, cardiac prosthesis, and the like) comprise the fastest growing segment. In contrast, efforts in biohybrid organs and other metabolic applications have contracted over the past few years. The number of companies involved in stem cells and regenerative medicine is rapidly increasing, and this area represents the most likely nidus of future growth for tissue engineering. A notable recent trend has been the emergence of a strong commercial activity in tissue engineering outside the United States, with at least 16 European or Australian companies (22% of total) now active.

Hybrid cellular automaton modeling of nutrient modulated cell growth in tissue engineering constructs.

Science.gov (United States)

Chung, C A; Lin, Tze-Hung; Chen, Shih-Di; Huang, Hsing-I

2010-01-21

Mathematic models help interpret experimental results and accelerate tissue engineering developments. We develop in this paper a hybrid cellular automata model that combines the differential nutrient transport equation to investigate the nutrient limited cell construct development for cartilage tissue engineering. Individual cell behaviors of migration, contact inhibition and cell collision, coupled with the cell proliferation regulated by oxygen concentration were carefully studied. Simplified two-dimensional simulations were performed. Using this model, we investigated the influence of cell migration speed on the overall cell growth within in vitro cell scaffolds. It was found that intense cell motility can enhance initial cell growth rates. However, since cell growth is also significantly modulated by the nutrient contents, intense cell motility with conventional uniform cell seeding method may lead to declined cell growth in the final time because concentrated cell population has been growing around the scaffold periphery to block the nutrient transport from outside culture media. Therefore, homogeneous cell seeding may not be a good way of gaining large and uniform cell densities for the final results. We then compared cell growth in scaffolds with various seeding modes, and proposed a seeding mode with cells initially residing in the middle area of the scaffold that may efficiently reduce the nutrient blockage and result in a better cell amount and uniform cell distribution for tissue engineering construct developments.

Advanced tissue culture used by Twyfords to build up jojoba clones

Energy Technology Data Exchange (ETDEWEB)

1983-01-01

Twyford Plant Laboratories Ltd. in the UK, using their own advanced methods of plant tissue culture, have built up a bank of 30 different male and female clones of jojoba, the arid land crop whose seeds produced a liquid wax which - amongst other uses - can be substituted for sperm whale oil. The technique involves growing microscopic parts of a parent plant on a medium containing all the necessary growth hormones, salts, vitamins and other nutrients. Growth takes place under artificial light in an all-electric controlled, air-conditioned environment. No other method is so successful for rapidly multiplying plants, particularly those that do not breed true from seed. These include most fruits and some flowers and vegetables.

[Chromosome variability in the tissue culture of rare Gentiana species].

Science.gov (United States)

Tvardovs'ka, M O; Strashniuk, N M; Mel'nyk, V M; Adonin, V I; Kunakh, V A

2008-01-01

Cytogenetic analysis of plants and tissue culture of Gentiana lutea, G. punctata, G. acaulis has been carried out. Culturing in vitro was found to result in the changes of chromosome number in the calluses of the species involved. Species specificity for variation of the cultured cell genomes was shown. Contribution of the original plant genotypes to the cytogenetic structure of the tissue culture was established. Gentiana callus tissues (except for in vitro culture of G. punctata, derived from plant of Breskul'ska population) were found to exhibit modal class with the cells of diploid and nearly diploid chromosome sets.

Evaluation of viability and proliferative activity of human urothelial cells cultured onto xenogenic tissue-engineered extracellular matrices.

LENUS (Irish Health Repository)

Davis, Niall F

2011-04-01

To evaluate the viability and proliferative activity of human urothelial cells (HUCs) cultured on tissue-engineered extracellular matrix scaffolds and to assess the potential of extracellular matrixes to support the growth of HUCs in their expected in vivo urine environment.

Influence of ionizing radiation on synthesis and molecular heterogeneity of catalase in tissue culture of Rauwolfia serpentina

International Nuclear Information System (INIS)

Komov, V.P.; Bespalova, E.V.; Strelkova, M.A.

1998-01-01

Changes in activity and molecular heterogeneity of catalase in tissue culture of Rauwolfia serpentina following irradiation in early growth period at the doses of 8 and 50 Gy has been studied. Ionizing radiation accelerate the synthesis and degradation rates of catalase and total protein. A comparative study of changes in enzyme and protein turnover during growth on irradiated and non-irradiated medium has been made [ru

Beauveria bassiana (Balsamo) Vuillemin as an endophyte in tissue culture banana (Musa spp.).

Science.gov (United States)

Akello, Juliet; Dubois, Thomas; Gold, Clifford S; Coyne, Daniel; Nakavuma, Jessica; Paparu, Pamela

2007-09-01

Beauveria bassiana is considered a virulent pathogen against the banana weevil Cosmopolites sordidus. However, current field application techniques for effective control against this pest remain a limitation and an alternative method for effective field application needs to be investigated. Three screenhouse experiments were conducted to determine the ability of B. bassiana to form an endophytic relationship with tissue culture banana (Musa spp.) plants and to evaluate the plants for possible harmful effects resulting from this relationship. Three Ugandan strains of B. bassiana (G41, S204 and WA) were applied by dipping the roots and rhizome in a conidial suspension, by injecting a conidial suspension into the plant rhizome and by growing the plants in sterile soil mixed with B. bassiana-colonized rice substrate. Four weeks after inoculation, plant growth parameters were determined and plant tissue colonization assessed through re-isolation of B. bassiana. All B. bassiana strains were able to colonize banana plant roots, rhizomes and pseudostem bases. Dipping plants in a conidial suspension achieved the highest colonization with no negative effect on plant growth or survival. Beauveria bassiana strain G41 was the best colonizer (up to 68%, 79% and 41% in roots, rhizome and pseudostem base, respectively) when plants were dipped. This study demonstrated that, depending on strain and inoculation method, B. bassiana can form an endophytic relationship with tissue culture banana plants, causing no harmful effects and might provide an alternative method for biological control of C. sordidus.

Co-culture systems-based strategies for articular cartilage tissue engineering.

Science.gov (United States)

Zhang, Yu; Guo, Weimin; Wang, Mingjie; Hao, Chunxiang; Lu, Liang; Gao, Shuang; Zhang, Xueliang; Li, Xu; Chen, Mingxue; Li, Penghao; Jiang, Peng; Lu, Shibi; Liu, Shuyun; Guo, Quanyi

2018-03-01

Cartilage engineering facilitates repair and regeneration of damaged cartilage using engineered tissue that restores the functional properties of the impaired joint. The seed cells used most frequently in tissue engineering, are chondrocytes and mesenchymal stem cells. Seed cells activity plays a key role in the regeneration of functional cartilage tissue. However, seed cells undergo undesirable changes after in vitro processing procedures, such as degeneration of cartilage cells and induced hypertrophy of mesenchymal stem cells, which hinder cartilage tissue engineering. Compared to monoculture, which does not mimic the in vivo cellular environment, co-culture technology provides a more realistic microenvironment in terms of various physical, chemical, and biological factors. Co-culture technology is used in cartilage tissue engineering to overcome obstacles related to the degeneration of seed cells, and shows promise for cartilage regeneration and repair. In this review, we focus first on existing co-culture systems for cartilage tissue engineering and related fields, and discuss the conditions and mechanisms thereof. This is followed by methods for optimizing seed cell co-culture conditions to generate functional neo-cartilage tissue, which will lead to a new era in cartilage tissue engineering. © 2017 Wiley Periodicals, Inc.

Critical Point in Self-Organized Tissue Growth

Science.gov (United States)

Aguilar-Hidalgo, Daniel; Werner, Steffen; Wartlick, Ortrud; González-Gaitán, Marcos; Friedrich, Benjamin M.; Jülicher, Frank

2018-05-01

We present a theory of pattern formation in growing domains inspired by biological examples of tissue development. Gradients of signaling molecules regulate growth, while growth changes these graded chemical patterns by dilution and advection. We identify a critical point of this feedback dynamics, which is characterized by spatially homogeneous growth and proportional scaling of patterns with tissue length. We apply this theory to the biological model system of the developing wing of the fruit fly Drosophila melanogaster and quantitatively identify signatures of the critical point.

Gastric tissue biopsy and culture

Science.gov (United States)

... symptoms may include: Loss of appetite or weight loss Nausea and vomiting Pain in the upper part of the belly Black stools Vomiting blood or coffee ground-like material A gastric tissue biopsy and culture can help detect: Cancer Infections, most commonly Helicobacter ...

Tenascin-Y, a component of distinctive connective tissues, supports muscle cell growth.

Science.gov (United States)

Hagios, C; Brown-Luedi, M; Chiquet-Ehrismann, R

1999-12-15

Chicken tenascin-Y is an extracellular matrix protein most closely related to the mammalian tenascin-X. It is highly expressed in the connective tissue of skeletal muscle (C. Hagios, M. Koch, J. Spring, M. Chiquet, and R. Chiquet-Ehrismann, 1996, J. Cell Biol. 134, 1499-1512). Here we demonstrate the presence of tenascin-Y in specific areas of the connective tissues in developing lung, kidney, and skin. In skin tenascin-Y shows a complementary expression pattern to tenascin-C, whereas in the lung and kidney the sites of expression are partly overlapping. Tenascin-Y is also present in embryonic skeletal muscle where it is expressed in the developing connective tissue in between the muscle fibers. This connective tissue is also the major site of alpha5 integrin expression. We purified recombinantly expressed tenascin-Y and tested its effect on cell adhesion and its influence on muscle cell growth and differentiation. C2C12 myoblasts were able to adhere to tenascin-Y and showed extensive formation of actin-rich processes without generation of stress fibers. Furthermore, we found that tenascin-Y influenced cell morphology of chick embryo fibroblasts over prolonged times in culture and that it supports primary muscle cell growth and restricts muscle cell differentiation. Copyright 1999 Academic Press.

Development of human nervous tissue upon differentiation of embryonic stem cells in three-dimensional culture.

Science.gov (United States)

Preynat-Seauve, Olivier; Suter, David M; Tirefort, Diderik; Turchi, Laurent; Virolle, Thierry; Chneiweiss, Herve; Foti, Michelangelo; Lobrinus, Johannes-Alexander; Stoppini, Luc; Feki, Anis; Dubois-Dauphin, Michel; Krause, Karl Heinz

2009-03-01

Researches on neural differentiation using embryonic stem cells (ESC) require analysis of neurogenesis in conditions mimicking physiological cellular interactions as closely as possible. In this study, we report an air-liquid interface-based culture of human ESC. This culture system allows three-dimensional cell expansion and neural differentiation in the absence of added growth factors. Over a 3-month period, a macroscopically visible, compact tissue developed. Histological coloration revealed a dense neural-like neural tissue including immature tubular structures. Electron microscopy, immunochemistry, and electrophysiological recordings demonstrated a dense network of neurons, astrocytes, and oligodendrocytes able to propagate signals. Within this tissue, tubular structures were niches of cells resembling germinal layers of human fetal brain. Indeed, the tissue contained abundant proliferating cells expressing markers of neural progenitors. Finally, the capacity to generate neural tissues on air-liquid interface differed for different ESC lines, confirming variations of their neurogenic potential. In conclusion, this study demonstrates in vitro engineering of a human neural-like tissue with an organization that bears resemblance to early developing brain. As opposed to previously described methods, this differentiation (a) allows three-dimensional organization, (b) yields dense interconnected neural tissue with structurally and functionally distinct areas, and (c) is spontaneously guided by endogenous developmental cues.

Allelopathy of small everlasting (Antennaria microphylla) : Phytotoxicity to leafy spurge (Euphorbia esula) in tissue culture.

Science.gov (United States)

Hogan, M E; Manners, G D

1990-03-01

Media and media extracts from callus cultures of small everlasting (Antennaria microphylla) inhibited leafy spurge (Euphorbia esula L.) callus tissue and suspension culture growth (50 and 70% of control, respectively) and were phytotoxic in lettuce and leafy spurge root elongation bioassays (64 and 77% of control, respectively). Hydroquinone, a phytotoxic compound previously isolated from small everlasting, was also biosynthesized by callus and suspension cultures of this species. Exogenously supplied hydroquinone (0.5 mM) was toxic to leafy spurge suspension culture cells and was only partially biotransformed to its nontoxic water-soluble monoglucoside, arbutin, by these cells. This report confirms the chronic involvement of hydroquinone in the allelopathic interaction between small everlasting and leafy spurge.

Improved Diagnosis of Prosthetic Joint Infection by Culturing Periprosthetic Tissue Specimens in Blood Culture Bottles

Directory of Open Access Journals (Sweden)

Trisha N. Peel

2016-01-01

Full Text Available Despite known low sensitivity, culture of periprosthetic tissue specimens on agars and in broths is routine. Culture of periprosthetic tissue samples in blood culture bottles (BCBs is potentially more convenient, but it has been evaluated in a limited way and has not been widely adopted. The aim of this study was to compare the sensitivity and specificity of inoculation of periprosthetic tissue specimens into blood culture bottles with standard agar and thioglycolate broth culture, applying Bayesian latent class modeling (LCM in addition to applying the Infectious Diseases Society of America (IDSA criteria for prosthetic joint infection. This prospective cohort study was conducted over a 9-month period (August 2013 to April 2014 at the Mayo Clinic, Rochester, MN, and included all consecutive patients undergoing revision arthroplasty. Overall, 369 subjects were studied; 117 (32% met IDSA criteria for prosthetic joint infection, and 82% had late chronic infection. Applying LCM, inoculation of tissues into BCBs was associated with a 47% improvement in sensitivity compared to the sensitivity of conventional agar and broth cultures (92.1 versus 62.6%, respectively; this magnitude of change was similar when IDSA criteria were applied (60.7 versus 44.4%, respectively; P = 0.003. The time to microorganism detection was shorter with BCBs than with standard media (P < 0.0001, with aerobic and anaerobic BCBs yielding positive results within a median of 21 and 23 h, respectively. Results of our study demonstrate that the semiautomated method of periprosthetic tissue culture in blood culture bottles is more sensitive than and as specific as agar and thioglycolate broth cultures and yields results faster.

Versatile electrochemial sensor for tissue culturing and sample handling

DEFF Research Database (Denmark)

Bakmand, Tanya; Kwasny, Dorota; Al Atraktchi, Fatima Al-Zahraa

2014-01-01

Culturing of organtypic brain tissues is a routine procedure in neural research. The visual inspection of the medium is the only way of determining the state of the tissue. At the end of culturing, post-processing techniques such as HPLC can be used to measure the concentration of the secreted...

Epigenetics in plant tissue culture

NARCIS (Netherlands)

Smulders, M.J.M.; Klerk, de G.J.M.

2011-01-01

Plants produced vegetatively in tissue culture may differ from the plants from which they have been derived. Two major classes of off-types occur: genetic ones and epigenetic ones. This review is about epigenetic aberrations. We discuss recent studies that have uncovered epigenetic modifications at

Evaluation of the effects of different culture media on the myogenic differentiation potential of adipose tissue- or bone marrow-derived human mesenchymal stem cells.

Science.gov (United States)

Stern-Straeter, Jens; Bonaterra, Gabriel Alejandro; Juritz, Stephanie; Birk, Richard; Goessler, Ulrich Reinhart; Bieback, Karen; Bugert, Peter; Schultz, Johannes; Hörmann, Karl; Kinscherf, Ralf; Faber, Anne

2014-01-01

The creation of functional muscles/muscle tissue from human stem cells is a major goal of skeletal muscle tissue engineering. Mesenchymal stem cells (MSCs) from fat/adipose tissue (AT-MSCs), as well as bone marrow (BM-MSCs) have been shown to bear myogenic potential, which makes them candidate stem cells for skeletal muscle tissue engineering applications. The aim of this study was to analyse the myogenic differentiation potential of human AT-MSCs and BM-MSCs cultured in six different cell culture media containing different mixtures of growth factors. The following cell culture media were used in our experiments: mesenchymal stem cell growth medium (MSCGM)™ as growth medium, MSCGM + 5-azacytidine (5-Aza), skeletal muscle myoblast cell growth medium (SkGM)-2 BulletKit™, and 5, 30 and 50% conditioned cell culture media, i.e., supernatant of human satellite cell cultures after three days in cell culture mixed with MSCGM. Following the incubation of human AT-MSCs or BM-MSCs for 0, 4, 8, 11, 16 or 21 days with each of the cell culture media, cell proliferation was measured using the alamarBlue® assay. Myogenic differentiation was evaluated by quantitative gene expression analyses, using quantitative RT-PCR (qRT-PCR) and immunocytochemical staining (ICC), using well-defined skeletal markers, such as desmin (DES), myogenic factor 5 (MYF5), myosin, heavy chain 8, skeletal muscle, perinatal (MYH8), myosin, heavy chain 1, skeletal muscle, adult (MYH1) and skeletal muscle actin-α1 (ACTA1). The highest proliferation rates were observed in the AT-MSCs and BM-MSCs cultured with SkGM-2 BulletKit medium. The average proliferation rate was higher in the AT-MSCs than in the BM-MSCs, taking all six culture media into account. qRT-PCR revealed the expression levels of the myogenic markers, ACTA1, MYH1 and MYH8, in the AT-MSC cell cultures, but not in the BM-MSC cultures. The muscle-specific intermediate filament, DES, was only detected (by ICC) in the AT-MSCs, but not in the BM

Metabolomics reveals the heterogeneous secretome of two entomopathogenic fungi to ex vivo cultured insect tissues.

Directory of Open Access Journals (Sweden)

Charissa de Bekker

Full Text Available Fungal entomopathogens rely on cellular heterogeneity during the different stages of insect host infection. Their pathogenicity is exhibited through the secretion of secondary metabolites, which implies that the infection life history of this group of environmentally important fungi can be revealed using metabolomics. Here metabolomic analysis in combination with ex vivo insect tissue culturing shows that two generalist isolates of the genus Metarhizium and Beauveria, commonly used as biological pesticides, employ significantly different arrays of secondary metabolites during infectious and saprophytic growth. It also reveals that both fungi exhibit tissue specific strategies by a distinguishable metabolite secretion on the insect tissues tested in this study. In addition to showing the important heterogeneous nature of these two entomopathogens, this study also resulted in the discovery of several novel destruxins and beauverolides that have not been described before, most likely because previous surveys did not use insect tissues as a culturing system. While Beauveria secreted these cyclic depsipeptides when encountering live insect tissues, Metarhizium employed them primarily on dead tissue. This implies that, while these fungi employ comparable strategies when it comes to entomopathogenesis, there are most certainly significant differences at the molecular level that deserve to be studied.

Metabolic Profile of Pancreatic Acinar and Islet Tissue in Culture

Science.gov (United States)

Suszynski, Thomas M.; Mueller, Kathryn; Gruessner, Angelika C.; Papas, Klearchos K.

2016-01-01

The amount and condition of exocrine impurities may affect the quality of islet preparations especially during culture. In this study, the objective was to determine the oxygen demandand viability of islet and acinar tissue post-isolation and whether they change disproportionately while in culture. We compare the OCR normalized to DNA (OCR/DNA, a measure of fractional viability in units nmol/min/mg DNA), and percent change in OCR and DNA recoveries between adult porcine islet and acinar tissue from the same preparation (paired) over a 6-9 days of standard culture. Paired comparisons were done to quantify differences in OCR/DNA between islet and acinar tissue from the same preparation, at specified time points during culture; the mean (± standard error) OCR/DNA was 74.0 (±11.7) units higher for acinar (vs. islet) tissue on the day of isolation (n=16, p<0.0001), but 25.7 (±9.4) units lower after 1 day (n=8, p=0.03), 56.6 (±11.5) units lower after 2 days (n=12, p=0.0004), and 65.9 (±28.7) units lower after 8 days (n=4, p=0.2) in culture. DNA and OCR recoveries decreased at different rates for acinar versus islet tissue over 6-9 days in culture (n=6). DNA recovery decreased to 24±7% for acinar and 75±8% for islets (p=0.002). Similarly, OCR recovery decreased to 16±3% for acinar and remained virtually constant for islets (p=0.005). Differences in the metabolic profile of acinarand islet tissue should be considered when culturing impure islet preparations. OCR-based measurements may help optimize pre-IT culture protocols. PMID:25131082

Revision washout decreases implant capsule tissue culture positivity: a multicenter study.

Science.gov (United States)

Henry, Gerard D; Carson, Culley C; Wilson, Steven K; Wiygul, Jeremy; Tornehl, Chris; Cleves, Mario A; Simmons, Caroline J; Donatucci, Craig F

2008-01-01

Positive cultures, visible biofilm and confocal micrography confirm bacterial presence on clinically uninfected inflatable penile prostheses at revision surgery. Salvage irrigation has been proved to rescue patients with clinically infected inflatable penile prostheses. Similar washout at revision for noninfectious reasons significantly lowers subsequent infection rates. We investigated a larger series of patients for positive culture rates and evaluated implant capsule tissue culture rates before and after revision washout. At 4 institutions a total of 148 patients with inflatable penile prostheses underwent revision surgery for noninfectious reasons between June 2001 and September 2005. Swab cultures of the fluid around the pump and visible biofilm were obtained. Also, in 65 patients a wedge of tissue from the capsule that forms around the pump was cultured. After implant removal revision washout of the implant spaces was performed and a second wedge of tissue was cultured. Of the 148 patients 97 (66%) had positive bacterial swab cultures of the fluid around the pump or biofilm. A total of 124 isolates were cultured. Of the 65 implant capsule tissue cultures obtained before washout 28 (43%) were positive for bacteria, while 16 (25%) obtained after revision washout were positive. Positive cultures and visible bacterial biofilm are present on clinically uninfected inflatable penile prostheses at revision surgery in most patients. Revision washout appears to decrease the bacterial load on implant capsule tissue at revision surgery of inflatable penile prostheses for noninfectious reasons.

Effect of aging on GHRF-induced growth hormone release from anterior pituitary cells in primary culture

International Nuclear Information System (INIS)

Spik, K.W.; Boyd, R.L.; Sonntag, W.E.

1991-01-01

Five criteria were developed to validate the primary cell culture model for comparison of GRF-induced release of growth hormone in pituitary tissue from aging animals. Pituitaries from young (5-mo), middle-aged (14-mo), and old (24-mo) male Fischer 344 rats were dispersed using either trypsin/trypsin inhibitor or dispase and compared with respect to the number of pituitary cells recovered, cell viability, 3H-leucine incorporation into total protein, time course for recovery of optimal response to GRF, and the dose-relationship for GRF-induced release of growth hormone 2, 4, and 6 days after dispersal. Results indicated that direct comparison of cellular responses between tissues from young, middle-aged, and old rats in primary cell culture is confounded by variations in time for recovery of optimal responses, the effects of the enzymes used for dispersal, and the methods used to express the data

Organotypic culture of human bone marrow adipose tissue.

Science.gov (United States)

Uchihashi, Kazuyoshi; Aoki, Shigehisa; Shigematsu, Masamori; Kamochi, Noriyuki; Sonoda, Emiko; Soejima, Hidenobu; Fukudome, Kenji; Sugihara, Hajime; Hotokebuchi, Takao; Toda, Shuji

2010-04-01

The precise role of bone marrow adipose tissue (BMAT) in the marrow remains unknown. The purpose of the present study was therefore to describe a novel method for studying BMAT using 3-D collagen gel culture of BMAT fragments, immunohistochemistry, ELISA and real-time reverse transcription-polymerase chain reaction. Mature adipocytes and CD45+ leukocytes were retained for >3 weeks. Bone marrow stromal cells (BMSC) including a small number of lipid-laden preadipocytes and CD44+/CD105+ mesenchymal stem cell (MSC)-like cells, developed from BMAT. Dexamethasone (10 micromol/L), but not insulin (20 mU/mL), significantly increased the number of preadipocytes. Dexamethasone and insulin also promoted leptin production and gene expression in BMAT. Adiponectin production by BMAT was BMAT, in which adiponectin protein secretion is normally very low, and that BMAT may exhibit a different phenotype from that of the visceral and subcutaneous adipose tissues. BMAT-osteoblast interactions were also examined, and it was found that osteoblasts inhibited the development of BMSC and reduced leptin production, while BMAT inhibited the growth and differentiation of osteoblasts. The present novel method proved to be useful for the study of BMAT biology.

TCUP: A novel hAT transposon active in maize tissue culture

Directory of Open Access Journals (Sweden)

Alan eSmith

2012-01-01

Full Text Available Transposable elements are capable of inducing heritable de novo genetic variation. The sequences capable of reactivation, and environmental factors that induce mobilization, remain poorly defined even in well-studied genomes such as maize. We treated maize tissue culture with the demethylating agent 5-aza-2-deoxcytidine and examined long-term tissue culture lines to discover silenced transposable elements that have the potential to induce heritable genetic variation. Through these screens we have identified a novel low copy number hAT transposon, Tissue Culture Up-Regulated (TCUP, which is transcribed at high levels in long-term maize Black Mexican Sweet (BMS tissue culture and up-regulated in response to treatment with 5-aza-2-deoxycytidine. Analysis of the TIGR Maize Gene Index revealed that this element is the most frequently represented EST from the BMS cell culture library and is not represented in other tissue libraries, which is the basis for its name. A full-length sequence was assembled in inbred B73 that contains the putative functional motifs required for autonomous movement of a hAT transposon. Transposon display detected movement of TCUP in two long-term tissue cultured cell lines of the genotype Hi-II AxB and BMS. This research implicates TCUP as a transposon that is capable of reactivation and which may also be particularly sensitive to the stress of the tissue culture environment. Our findings are consistent with the hypothesis that epigenetic alterations potentiate genomic responses to stress during clonal propagation of plants.

The autologus graft of epithelial tissue culture

Directory of Open Access Journals (Sweden)

Minaee B

1999-08-01

Full Text Available With the intention of research about culture and autologus graft of epithelial tissue we used 4 french Albino Rabbits with an average age of 2 months. After reproduction on the support in EMEM (Eagle's Minimum Essential Medium we used this for graft after 4 weeks. This region which grafted total replaced. After fixation of this sample and passing them through various process, histological sections were prepared. These sections were stained with H & E and masson's trichrome and studied by light microscope. We succeeded in graft. We hope in the near future by using the method of epithelium tissue culture improving to treat burned patients.

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

OpenAIRE

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 stabili...

[Research progress of co-culture system for constructing vascularized tissue engineered bone].

Science.gov (United States)

Fu, Weili; Xiang, Zhou

2014-02-01

To review the research progress of the co-culture system for constructing vascularized tissue engineered bone. The recent literature concerning the co-culture system for constructing vascularized tissue engineered bone was reviewed, including the selection of osteogenic and endothelial lineages, the design and surface modification of scaffolds, the models and dimensions of the co-culture system, the mechanism, the culture conditions, and their application progress. The construction of vascularized tissue engineered bone is the prerequisite for their survival and further clinical application in vivo. Mesenchymal stem cells (owning the excellent osteogenic potential) and endothelial progenitor cells (capable of directional differentiation into endothelial cell) are considered as attractive cell types for the co-culture system to construct vascularized tissue engineered bone. The culture conditions need to be further optimized. Furthermore, how to achieve the clinical goals of minimal invasion and autologous transplantation also need to be further studied. The strategy of the co-culture system for constructing vascularized tissue engineered bone would have a very broad prospects for clinical application in future.

Connective tissue growth factor regulates fibrosis-associated renal lymphangiogenesis

NARCIS (Netherlands)

Kinashi, Hiroshi; Falke, Lucas L.; Nguyen, Tri Q.; Bovenschen, Niels; Aten, Jan; Leask, Andrew; Ito, Yasuhiko; Goldschmeding, Roel

2017-01-01

Lymphangiogenesis is correlated with the degree of renal interstitial fibrosis. Pro-fibrotic transforming growth factor beta induces VEGF-C production, the main driver of lymphangiogenesis. Connective tissue growth factor (CTGF) is an important determinant of fibrotic tissue remodeling, but its

Connective tissue growth factor regulates fibrosis-associated renal lymphangiogenesis

NARCIS (Netherlands)

Kinashi, Hiroshi; Falke, Lucas L.; Nguyen, Tri Q.; Bovenschen, Niels; Aten, Jan; Leask, Andrew; Ito, Yasuhiko; Goldschmeding, Roel

2017-01-01

Lymphangiogenesis is correlated with the degree of renal interstitial fibrosis. Pro-fibrotic transforming growth factor β induces VEGF-C production, the main driver of lymphangiogenesis. Connective tissue growth factor (CTGF) is an important determinant of fibrotic tissue remodeling, but its

Improved Diagnosis of Prosthetic Joint Infection by Culturing Periprosthetic Tissue Specimens in Blood Culture Bottles.

Science.gov (United States)

Peel, Trisha N; Dylla, Brenda L; Hughes, John G; Lynch, David T; Greenwood-Quaintance, Kerryl E; Cheng, Allen C; Mandrekar, Jayawant N; Patel, Robin

2016-01-05

Despite known low sensitivity, culture of periprosthetic tissue specimens on agars and in broths is routine. Culture of periprosthetic tissue samples in blood culture bottles (BCBs) is potentially more convenient, but it has been evaluated in a limited way and has not been widely adopted. The aim of this study was to compare the sensitivity and specificity of inoculation of periprosthetic tissue specimens into blood culture bottles with standard agar and thioglycolate broth culture, applying Bayesian latent class modeling (LCM) in addition to applying the Infectious Diseases Society of America (IDSA) criteria for prosthetic joint infection. This prospective cohort study was conducted over a 9-month period (August 2013 to April 2014) at the Mayo Clinic, Rochester, MN, and included all consecutive patients undergoing revision arthroplasty. Overall, 369 subjects were studied; 117 (32%) met IDSA criteria for prosthetic joint infection, and 82% had late chronic infection. Applying LCM, inoculation of tissues into BCBs was associated with a 47% improvement in sensitivity compared to the sensitivity of conventional agar and broth cultures (92.1 versus 62.6%, respectively); this magnitude of change was similar when IDSA criteria were applied (60.7 versus 44.4%, respectively; P = 0.003). The time to microorganism detection was shorter with BCBs than with standard media (P Prosthetic joint infections are a devastating complication of arthroplasty surgery. Despite this, current microbiological techniques to detect and diagnose infections are imperfect. This study examined a new approach to diagnosing infections, through the inoculation of tissue samples from around the prosthetic joint into blood culture bottles. This study demonstrated that, compared to current laboratory practices, this new technique increased the detection of infection. These findings are important for patient care to allow timely and accurate diagnosis of infection. Copyright © 2016 Peel et al.

Tumor tissue slice cultures as a platform for analyzing tissue-penetration and biological activities of nanoparticles.

Science.gov (United States)

Merz, Lea; Höbel, Sabrina; Kallendrusch, Sonja; Ewe, Alexander; Bechmann, Ingo; Franke, Heike; Merz, Felicitas; Aigner, Achim

2017-03-01

The success of therapeutic nanoparticles depends, among others, on their ability to penetrate a tissue for actually reaching the target cells, and their efficient cellular uptake in the context of intact tissue and stroma. Various nanoparticle modifications have been implemented for altering physicochemical and biological properties. Their analysis, however, so far mainly relies on cell culture experiments which only poorly reflect the in vivo situation, or is based on in vivo experiments that are often complicated by whole-body pharmacokinetics and are rather tedious especially when analyzing larger nanoparticle sets. For the more precise analysis of nanoparticle properties at their desired site of action, efficient ex vivo systems closely mimicking in vivo tissue properties are needed. In this paper, we describe the setup of organotypic tumor tissue slice cultures for the analysis of tissue-penetrating properties and biological activities of nanoparticles. As a model system, we employ 350μm thick slice cultures from different tumor xenograft tissues, and analyze modified or non-modified polyethylenimine (PEI) complexes as well as their lipopolyplex derivatives for siRNA delivery. The described conditions for tissue slice preparation and culture ensure excellent tissue preservation for at least 14days, thus allowing for prolonged experimentation and analysis. When using fluorescently labeled siRNA for complex visualization, fluorescence microscopy of cryo-sectioned tissue slices reveals different degrees of nanoparticle tissue penetration, dependent on their surface charge. More importantly, the determination of siRNA-mediated knockdown efficacies of an endogenous target gene, the oncogenic survival factor Survivin, reveals the possibility to accurately assess biological nanoparticle activities in situ, i.e. in living cells in their original environment. Taken together, we establish tumor (xenograft) tissue slices for the accurate and facile ex vivo assessment of

Observations on human smooth muscle cell cultures from hyperplastic lesions of prosthetic bypass grafts: Production of a platelet-derived growth factor-like mitogen and expression of a gene for a platelet-derived growth factor receptor--a preliminary study

International Nuclear Information System (INIS)

Birinyi, L.K.; Warner, S.J.; Salomon, R.N.; Callow, A.D.; Libby, P.

1989-01-01

Prosthetic bypass grafts placed to the distal lower extremity often fail because of an occlusive tissue response in the perianastomotic region. The origin of the cells that comprise this occlusive lesion and the causes of the cellular proliferation are not known. To increase our understanding of this process we cultured cells from hyperplastic lesions obtained from patients at the time of reexploration for lower extremity graft failure, and we studied their identity and growth factor production in tissue culture. These cultures contain cells that express muscle-specific actin isoforms, shown by immunohistochemical staining, consistent with vascular smooth muscle origin. These cultures also released material that stimulated smooth muscle cell growth. A portion of this activity was similar to platelet-derived growth factor, since preincubation with antibody-to-human platelet-derived growth factor partially blocked the mitogenic effect of medium conditioned by human anastomotic hyperplastic cells. These conditioned media also contained material that competed with platelet-derived growth factor for its receptor, as measured in a radioreceptor assay. Northern blot analysis showed that these cells contain messenger RNA that encodes the A chain but not the B chain of platelet-derived growth factor. In addition, these cells contain messenger RNA that encodes a platelet-derived growth factor receptor. We conclude that cultured smooth muscle cells from human anastomotic hyperplastic lesions express genes for platelet-derived growth factor A chain and a platelet-derived growth factor receptor and secrete biologically active molecules similar to platelet-derived growth factor

Gastrointestinal Epithelial Organoid Cultures from Postsurgical Tissues.

Science.gov (United States)

Hahn, Soojung; Yoo, Jongman

2017-08-17

An organoid is a cellular structure three-dimensionally (3D) cultured from self-organizing stem cells in vitro, which has a cell population, architectures, and organ specific functions like the originating organs. Recent advances in the 3D culture of isolated intestinal crypts or gastric glands have enabled the generation of human gastrointestinal epithelial organoids. Gastrointestinal organoids recapitulate the human in vivo physiology because of all the intestinal epithelial cell types that differentiated and proliferated from tissue resident stem cells. Thus far, gastrointestinal organoids have been extensively used for generating gastrointestinal disease models. This protocol describes the method of isolating a gland or crypt using stomach or colon tissue after surgery and establishing them into gastroids or colonoids.

Tissue culture of surgically prepared temporalis fascia.

Science.gov (United States)

Walby, A P; Kerr, A G; Nevin, N C; Woods, G

1982-10-01

Temporalis fascia which is used to graft the tympanic membrane has been shown to be viable in tissue culture by a previous pilot study. This present study reports the effect on the viability of the fascia by scraping loose connective tissue from it and allowing it to dry. Pieces of fascia from 30 patients were each divided in 4 and prepared to give explants, fresh, fresh and scraped, dried, and dried and scraped. The fascia grew from 17 patients when cultured fresh, 5 when fresh and scraped, 1 when dried, and none when dried and scraped. These results are significantly different and show that the fascia is devitilized when prepared by the normal method for use in tympanoplasty.

Antiandrogenic actions of medroxyprogesterone acetate on epithelial cells within normal human breast tissues cultured ex vivo.

Science.gov (United States)

Ochnik, Aleksandra M; Moore, Nicole L; Jankovic-Karasoulos, Tanja; Bianco-Miotto, Tina; Ryan, Natalie K; Thomas, Mervyn R; Birrell, Stephen N; Butler, Lisa M; Tilley, Wayne D; Hickey, Theresa E

2014-01-01

Medroxyprogesterone acetate (MPA), a component of combined estrogen-progestin therapy (EPT), has been associated with increased breast cancer risk in EPT users. MPA can bind to the androgen receptor (AR), and AR signaling inhibits cell growth in breast tissues. Therefore, the aim of this study was to investigate the potential of MPA to disrupt AR signaling in an ex vivo culture model of normal human breast tissue. Histologically normal breast tissues from women undergoing breast surgical operation were cultured in the presence or in the absence of the native AR ligand 5α-dihydrotestosterone (DHT), MPA, or the AR antagonist bicalutamide. Ki67, bromodeoxyuridine, B-cell CLL/lymphoma 2 (BCL2), AR, estrogen receptor α, and progesterone receptor were detected by immunohistochemistry. DHT inhibited the proliferation of breast epithelial cells in an AR-dependent manner within tissues from postmenopausal women, and MPA significantly antagonized this androgenic effect. These hormonal responses were not commonly observed in cultured tissues from premenopausal women. In tissues from postmenopausal women, DHT either induced or repressed BCL2 expression, and the antiandrogenic effect of MPA on BCL2 was variable. MPA significantly opposed the positive effect of DHT on AR stabilization, but these hormones had no significant effect on estrogen receptor α or progesterone receptor levels. In a subset of postmenopausal women, MPA exerts an antiandrogenic effect on breast epithelial cells that is associated with increased proliferation and destabilization of AR protein. This activity may contribute mechanistically to the increased risk of breast cancer in women taking MPA-containing EPT.

Mary Jane Hogue (1883-1962): A pioneer in human brain tissue culture.

Science.gov (United States)

Zottoli, Steven J; Seyfarth, Ernst-August

2018-05-16

The ability to maintain human brain explants in tissue culture was a critical step in the use of these cells for the study of central nervous system disorders. Ross G. Harrison (1870-1959) was the first to successfully maintain frog medullary tissue in culture in 1907, but it took another 38 years before successful culture of human brain tissue was accomplished. One of the pioneers in this achievement was Mary Jane Hogue (1883-1962). Hogue was born into a Quaker family in 1883 in West Chester, Pennsylvania, and received her undergraduate degree from Goucher College in Baltimore, Maryland. Research with the developmental biologist Theodor Boveri (1862-1915) in Würzburg, Germany, resulted in her Ph.D. (1909). Hogue transitioned from studying protozoa to the culture of human brain tissue in the 1940s and 1950s, when she was one of the first to culture cells from human fetal, infant, and adult brain explants. We review Hogue's pioneering contributions to the study of human brain cells in culture, her putative identification of progenitor neuroblast and/or glioblast cells, and her use of the cultures to study the cytopathogenic effects of poliovirus. We also put Hogue's work in perspective by discussing how other women pioneers in tissue culture influenced Hogue and her research.

Three-Dimensional Culture Model of Skeletal Muscle Tissue with Atrophy Induced by Dexamethasone.

Science.gov (United States)

Shimizu, Kazunori; Genma, Riho; Gotou, Yuuki; Nagasaka, Sumire; Honda, Hiroyuki

2017-06-15

Drug screening systems for muscle atrophy based on the contractile force of cultured skeletal muscle tissues are required for the development of preventive or therapeutic drugs for atrophy. This study aims to develop a muscle atrophy model by inducing atrophy in normal muscle tissues constructed on microdevices capable of measuring the contractile force and to verify if this model is suitable for drug screening using the contractile force as an index. Tissue engineered skeletal muscles containing striated myotubes were prepared on the microdevices for the study. The addition of 100 µM dexamethasone (Dex), which is used as a muscle atrophy inducer, for 24 h reduced the contractile force significantly. An increase in the expression of Atrogin-1 and MuRF-1 in the tissues treated with Dex was established. A decrease in the number of striated myotubes was also observed in the tissues treated with Dex. Treatment with 8 ng/mL Insulin-like Growth Factor (IGF-I) for 24 h significantly increased the contractile force of the Dex-induced atrophic tissues. The same treatment, though, had no impact on the force of the normal tissues. Thus, it is envisaged that the atrophic skeletal muscle tissues induced by Dex can be used for drug screening against atrophy.

Establishment of primary keratinocyte culture from horse tissue biopsates

Directory of Open Access Journals (Sweden)

Jernej OGOREVC

2015-12-01

Full Text Available Primary cell lines established from skin tissue can be used in immunological, proteomic and genomic studies as in vitro skin models. The goal of our study was to establish a primary keratinocyte cell culture from tissue biopsates of two horses. The primary keratinocyte cell culture was obtained by mechanical and enzymatic dissociation and with explant culture method. The result was a heterogeneous primary culture comprised of keratinocytes and fibroblasts. To distinguish epithelial and mesenchymal cells immunofluorescent characterisation was performed, using antibodies against cytokeratin 14 and vimentin. We successfully at attained a primary cell line of keratinocytes, which could potentially be used to study equine skin diseases, as an animal model for human diseases, and for cosmetic and therapeutic product testing.

Skin equivalent tissue-engineered construct: co-cultured fibroblasts/ keratinocytes on 3D matrices of sericin hope cocoons.

Directory of Open Access Journals (Sweden)

Sunita Nayak

Full Text Available The development of effective and alternative tissue-engineered skin replacements to autografts, allografts and xenografts has became a clinical requirement due to the problems related to source of donor tissue and the perceived risk of disease transmission. In the present study 3D tissue engineered construct of sericin is developed using co-culture of keratinocytes on the upper surface of the fabricated matrices and with fibroblasts on lower surface. Sericin is obtained from "Sericin Hope" silkworm of Bombyx mori mutant and is extracted from cocoons by autoclave. Porous sericin matrices are prepared by freeze dried method using genipin as crosslinker. The matrices are characterized biochemically and biophysically. The cell proliferation and viability of co-cultured fibroblasts and keratinocytes on matrices for at least 28 days are observed by live/dead assay, Alamar blue assay, and by dual fluorescent staining. The growth of the fibroblasts and keratinocytes in co-culture is correlated with the expression level of TGF-β, b-FGF and IL-8 in the cultured supernatants by enzyme-linked immunosorbent assay. The histological analysis further demonstrates a multi-layered stratified epidermal layer of uninhibited keratinocytes in co-cultured constructs. Presence of involucrin, collagen IV and the fibroblast surface protein in immuno-histochemical stained sections of co-cultured matrices indicates the significance of paracrine signaling between keratinocytes and fibroblasts in the expression of extracellular matrix protein for dermal repair. No significant amount of pro inflammatory cytokines (TNF-α, IL-1β and nitric oxide production are evidenced when macrophages grown on the sericin matrices. The results all together depict the potentiality of sericin 3D matrices as skin equivalent tissue engineered construct in wound repair.

Survival and growth of isolated pre-antral follicles from human ovarian medulla tissue during long-term 3D culture

DEFF Research Database (Denmark)

Yin, H L; Kristensen, S G; Jiang, H

2016-01-01

during long-term culture has received only little attention. STUDY DESIGN, SIZE, DURATION: Two to ten human pre-antral follicles were encapsulated together within an alginate bead and cultured with or without ovarian interstitial tissue for either 7 days or >30 days. Follicles were cultured in either 20...... interregional project ReproHigh are thanked for having funded this study; and the Key Program of Medical Science and Technology Innovation of Nanjing Military Area Command in China (14ZX06; 11Z010). They had no role in the study design, collection and analysis of data, data interpretation or in writing...

Culture growth of testate amoebae under different silicon concentrations.

Science.gov (United States)

Wanner, Manfred; Seidl-Lampa, Barbara; Höhn, Axel; Puppe, Daniel; Meisterfeld, Ralf; Sommer, Michael

2016-10-01

Testate amoebae with self-secreted siliceous shell platelets ("idiosomes") play an important role in terrestrial silicon (Si) cycles. In this context, Si-dependent culture growth dynamics of idiosomic testate amoebae are of interest. Clonal cultures of idiosomic testate amoebae were analyzed under three different Si concentrations: low (50μmolL -1 ), moderate/site-specific (150μmolL -1 ) and high Si supply (500μmolL -1 ). Food (Saccharomyces cerevisiae) was provided in surplus. (i) Shell size of four different clones of idiosomic testate amoebae either decreased (Trinema galeata, Euglypha filifera cf.), increased (E. rotunda cf.), or did not change (E. rotunda) under the lowest Si concentration (50μmolSiL -1 ). (ii) Culture growth of idiosomic Euglypha rotunda was dependent on Si concentration. The more Si available in the culture medium, the earlier the entry into exponential growth phase. (iii) Culture growth of idiosomic Euglypha rotunda was dependent on origin of inoculum. Amoebae previously cultured under a moderate Si concentration revealed highest sustainability in consecutive cultures. Amoebae derived from cultures with high Si concentrations showed rapid culture growth which finished early in consecutive cultures. (iv) Si (diluted in the culture medium) was absorbed by amoebae and fixed in the amoeba shells resulting in decreased Si concentrations. Copyright © 2016 Elsevier GmbH. All rights reserved.

Using organotypic (raft) epithelial tissue cultures for the biosynthesis and isolation of infectious human papillomaviruses.

Science.gov (United States)

Ozbun, Michelle A; Patterson, Nicole A

2014-08-01

Papillomaviruses have a strict tropism for epithelial cells, and they are fully reliant on cellular differentiation for completion of their life cycles, resulting in the production of progeny virions. Thus, a permissive environment for full viral replication in vitro-wherein virion morphogenesis occurs under cooperative viral and cellular cues-requires the cultivation of epithelium. Presented in the first section of this unit is a protocol to grow differentiating epithelial tissues that mimic many important morphological and biochemical aspects of normal skin. The technique involves growing epidermal cells atop a dermal equivalent consisting of live fibroblasts and a collagen lattice. Epithelial stratification and differentiation ensues when the keratinocyte-dermal equivalent is placed at the air-liquid interface. The apparent floating nature of the cell-matrix in this method led to the nickname "raft" cultures. The general technique can be applied to normal low passage keratinocytes, to cells stably transfected with papillomavirus genes or genomes, or keratinocytes established from neoplastic lesions. However, infectious papillomavirus particles have only been isolated from organotypic epithelial cultures initiated with cells that maintain oncogenic human papillomavirus genomes in an extrachomosomal replicative form. The second section of this unit is dedicated to a virion isolation method that minimizes aerosol and skin exposure to these human carcinogens. Although the focus of the protocols is on the growth of tissues that yields infectious papillomavirus progeny, this culture system facilitates the investigation of these fastidious viruses during their complex replicative cycles, and raft tissues can be manipulated and harvested at any point during the process. Importantly, a single-step virus growth cycle is achieved in this process, as it is unlikely that progeny virions are released to initiate subsequent rounds of infection. Copyright © 2014 John Wiley

Micro fluidic System for Culturing and Monitoring of Neuronal Cells and Tissue

DEFF Research Database (Denmark)

Bakmand, Tanya; Waagepetersen, Helle S.

The aim of this Ph.D. project was to combine experience within cell and tissue culturing, electrochemistry and microfabrication in order to develop an in vivo-like fluidic culturing platform, challenging the traditional culturing methods. The first goal was to develope a fluidic system for cultur...... with mass production. The last part of this thesis also includes perspectives on how to expand the latest designed device to facilitate culturing of tissue and co-culturing of cells....

Effects of cyclic compression on the mechanical properties and calcification process of immature chick bone tissue in culture.

Science.gov (United States)

Maeda, Eijiro; Nakagaki, Masashi; Ichikawa, Katsuhisa; Nagayama, Kazuaki; Matsumoto, Takeo

2017-06-01

Contribution of mechanical loading to tissue growth during both the development and post-natal maturation is of a particular interest, as its understanding would be important to strategies in bone tissue engineering and regenerative medicine. The present study has been performed to investigate how immature bone responds to mechanical loading using an ex vivo culture system. A slice of the tibia, with the thickness of 3 mm, was obtained from 0-day-old chick. For the ex vivo culture experiment in conjunction with cyclic compressive loading, we developed a custom-made, bioreactor system where both the load and the deformation applied to the specimen was recorded. Cyclic compression, with an amplitude of 0.3 N corresponding to 1 to 2% compressive strain, was applied to immature bone specimen during a 3-day culture period at an overall loading rate 3-4 cycles/min, in the presence of β-glycerol phosphate and dexamethasone in culture medium. The stress-strain relationship was obtained at the beginning and the end of the culture experiment. In addition, analyses for alkaline phosphate release, cell viability and tissue calcification were also performed. It was exhibited that elastic moduli of bone slices were significantly elevated at the end of the 3-day culture in the presence of cyclic compression, which was a similar phenomenon to significant elevation of the elastic moduli of bone tissue by the maturation from 0-day old to 3-day old. By contrast, no significant changes in the moduli were observed in the absence of cyclic compression or in deactivated, cell-free samples. The increases in the moduli were coincided with the increase in calcified area in the bone samples. It was confirmed that immature bone can respond to compressive loading in vitro and demonstrate the growth of bone matrix, similar to natural, in vivo maturation. The elevation of the elastic moduli was attributable to the increased calcified area and the realignment of collagen fibers parallel to

Effects of cyclic compression on the mechanical properties and calcification process of immature chick bone tissue in culture

Directory of Open Access Journals (Sweden)

Eijiro Maeda

2017-06-01

Full Text Available Contribution of mechanical loading to tissue growth during both the development and post-natal maturation is of a particular interest, as its understanding would be important to strategies in bone tissue engineering and regenerative medicine. The present study has been performed to investigate how immature bone responds to mechanical loading using an ex vivo culture system. A slice of the tibia, with the thickness of 3 mm, was obtained from 0-day-old chick. For the ex vivo culture experiment in conjunction with cyclic compressive loading, we developed a custom-made, bioreactor system where both the load and the deformation applied to the specimen was recorded. Cyclic compression, with an amplitude of 0.3 N corresponding to 1 to 2% compressive strain, was applied to immature bone specimen during a 3-day culture period at an overall loading rate 3–4 cycles/min, in the presence of β-glycerol phosphate and dexamethasone in culture medium. The stress-strain relationship was obtained at the beginning and the end of the culture experiment. In addition, analyses for alkaline phosphate release, cell viability and tissue calcification were also performed. It was exhibited that elastic moduli of bone slices were significantly elevated at the end of the 3-day culture in the presence of cyclic compression, which was a similar phenomenon to significant elevation of the elastic moduli of bone tissue by the maturation from 0-day old to 3-day old. By contrast, no significant changes in the moduli were observed in the absence of cyclic compression or in deactivated, cell-free samples. The increases in the moduli were coincided with the increase in calcified area in the bone samples. It was confirmed that immature bone can respond to compressive loading in vitro and demonstrate the growth of bone matrix, similar to natural, in vivo maturation. The elevation of the elastic moduli was attributable to the increased calcified area and the realignment of collagen

Ex vivo culture of patient tissue & examination of gene delivery.

LENUS (Irish Health Repository)

Rajendran, Simon

2012-01-31

This video describes the use of patient tissue as an ex vivo model for the study of gene delivery. Fresh patient tissue obtained at the time of surgery is sliced and maintained in culture. The ex vivo model system allows for the physical delivery of genes into intact patient tissue and gene expression is analysed by bioluminescence imaging using the IVIS detection system. The bioluminescent detection system demonstrates rapid and accurate quantification of gene expression within individual slices without the need for tissue sacrifice. This slice tissue culture system may be used in a variety of tissue types including normal and malignant tissue and allows us to study the effects of the heterogeneous nature of intact tissue and the high degree of variability between individual patients. This model system could be used in certain situations as an alternative to animal models and as a complementary preclinical mode prior to entering clinical trial.

Effect of radiation and other cytotoxic agents on the growth of cells cultured from normal and tumor tissues from the female genital tract

International Nuclear Information System (INIS)

Mothersill, C.; Seymour, C.B.; Bonnar, J.

1990-01-01

A technique is presented which allows the response of human gynecological tissue to radiation and cytotoxic drugs to be assessed using a tissue culture explant system. The technique is simple to use and gives results in line with those obtained for human tissues by more complex culture methods. Data are presented showing how the explant technique developed by the group for other tissues can be adapted to yield acceptable results for normal tissue response to radiation. The potential of the technique for use in predictive testing of individual tumor response is then assessed in five cases of gynecological malignancy. It is clear that variations in sensitivity to different radio- and chemotherapy agents and combinations can be detected. The results obtained require clinical validation and it is hoped that this will come over the next few years from evaluation of patient response to treatment using individually optimized, rather than empirical therapy

Gene therapy with growth factors for periodontal tissue engineering–A review

Science.gov (United States)

Gupta, Shipra; Mahendra, Aneet

2012-01-01

The treatment of oral and periodontal diseases and associated anomalies accounts for a significant proportion of the healthcare burden, with the manifestations of these conditions being functionally and psychologically debilitating. A challenge faced by periodontal therapy is the predictable regeneration of periodontal tissues lost as a consequence of disease. Growth factors are critical to the development, maturation, maintenance and repair of oral tissues as they establish an extra-cellular environment that is conducive to cell and tissue growth. Tissue engineering principles aim to exploit these properties in the development of biomimetic materials that can provide an appropriate microenvironment for tissue development. The aim of this paper is to review emerging periodontal therapies in the areas of materials science, growth factor biology and cell/gene therapy. Various such materials have been formulated into devices that can be used as vehicles for delivery of cells, growth factors and DNA. Different mechanisms of drug delivery are addressed in the context of novel approaches to reconstruct and engineer oral and tooth supporting structure. Key words: Periodontal disease, gene therapy, regeneration, tissue repair, growth factors, tissue engineering. PMID:22143705

21 CFR 864.2240 - Cell and tissue culture supplies and equipment.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Cell and tissue culture supplies and equipment. 864.2240 Section 864.2240 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Cell And Tissue Culture Products...

Assessment of growth factor treatment on fibrochondrocyte and chondrocyte co-cultures for TMJ fibrocartilage engineering.

Science.gov (United States)

Kalpakci, Kerem N; Kim, Eric J; Athanasiou, Kyriacos A

2011-04-01

Treatments for patients suffering from severe temporomandibular joint (TMJ) dysfunction are limited, motivating the development of strategies for tissue regeneration. In this study, co-cultures of fibrochondrocytes (FCs) and articular chondrocytes (ACs) were seeded in agarose wells, and supplemented with growth factors, to engineer tissue with biomechanical properties and extracellular matrix composition similar to native TMJ fibrocartilage. In the first phase, growth factors were applied alone and in combination, in the presence or absence of serum, while in the second phase, the best overall treatment was applied at intermittent dosing. Continuous treatment of AC/FC co-cultures with TGF-β1 in serum-free medium resulted in constructs with glycosaminoglycan/wet weight ratios (12.2%), instantaneous compressive moduli (790 kPa), relaxed compressive moduli (120 kPa) and Young's moduli (1.87 MPa) that overlap with native TMJ disc values. Among co-culture groups, TGF-β1 treatment increased collagen deposition ∼20%, compressive stiffness ∼130% and Young's modulus ∼170% relative to controls without growth factor. Serum supplementation, though generally detrimental to functional properties, was identified as a powerful mediator of FC construct morphology. Finally, both intermittent and continuous TGF-β1 treatment showed positive effects, though continuous treatment resulted in greater enhancement of construct functional properties. This work proposes a strategy for regeneration of TMJ fibrocartilage and its future application will be realized through translation of these findings to clinically viable cell sources. Copyright © 2011 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Different culture media affect growth characteristics, surface marker distribution and chondrogenic differentiation of human bone marrow-derived mesenchymal stromal cells.

Science.gov (United States)

Hagmann, Sebastien; Moradi, Babak; Frank, Sebastian; Dreher, Thomas; Kämmerer, Peer Wolfgang; Richter, Wiltrud; Gotterbarm, Tobias

2013-07-30

Bone marrow-derived mesenchymal stromal cells (BM-MSCs) play an important role in modern tissue engineering, while distinct variations of culture media compositions and supplements have been reported. Because MSCs are heterogeneous regarding their regenerative potential and their surface markers, these parameters were compared in four widely used culture media compositions. MSCs were isolated from bone marrow and expanded in four established cell culture media. MSC yield/1000 MNCs, passage time and growth index were observed. In P4, typical MSC surface markers were analysed by fluorescence cytometry. Additionally, chondrogenic, adipogenic and osteogenic differentiation potential were evaluated. Growth index and P0 cell yield varied importantly between the media. The different expansion media had a significant influence on the expression of CD10, CD90, CD105, CD140b CD146 and STRO-1. While no significant differences were observed regarding osteogenic and adipogenic differentiation, chondrogenic differentiation was superior in medium A as reflected by GAG/DNA content. The choice of expansion medium can have a significant influence on growth, differentiation potential and surface marker expression of mesenchymal stromal cells, which is of fundamental importance for tissue engineering procedures.

Biomaterials in co-culture systems: towards optimizing tissue integration and cell signaling within scaffolds.

Science.gov (United States)

Battiston, Kyle G; Cheung, Jane W C; Jain, Devika; Santerre, J Paul

2014-05-01

Most natural tissues consist of multi-cellular systems made up of two or more cell types. However, some of these tissues may not regenerate themselves following tissue injury or disease without some form of intervention, such as from the use of tissue engineered constructs. Recent studies have increasingly used co-cultures in tissue engineering applications as these systems better model the natural tissues, both physically and biologically. This review aims to identify the challenges of using co-culture systems and to highlight different approaches with respect to the use of biomaterials in the use of such systems. The application of co-culture systems to stimulate a desired biological response and examples of studies within particular tissue engineering disciplines are summarized. A description of different analytical co-culture systems is also discussed and the role of biomaterials in the future of co-culture research are elaborated on. Understanding the complex cell-cell and cell-biomaterial interactions involved in co-culture systems will ultimately lead the field towards biomaterial concepts and designs with specific biochemical, electrical, and mechanical characteristics that are tailored towards the needs of distinct co-culture systems. Copyright © 2014 Elsevier Ltd. All rights reserved.

Tissue engineering of bladder using vascular endothelial growth factor gene-modified endothelial progenitor cells.

Science.gov (United States)

Chen, Bai-Song; Xie, Hua; Zhang, Sheng-Li; Geng, Hong-Quan; Zhou, Jun-Mei; Pan, Jun; Chen, Fang

2011-12-01

This study assessed the use of vascular endothelial growth factor (VEGF) gene-modified endothelial progenitor cells (EPCs) seeded onto bladder acellular matrix grafts (BAMGs), to enhance the blood supply in tissue-engineered bladders in a porcine model. Autologous porcine peripheral EPCs were isolated, cultured, expanded, characterized, and modified with the VEGF gene using an adenovirus vector. The expression of VEGF was examined using reverse transcriptase polymerase chain reaction (RT-PCR) and an enzyme-linked immunosorbent assay (ELISA). VEGF gene modified EPCs were seeded onto BAMG and cultured for 3 days before implantation into pigs for bladder tissue engineering. A partial bladder cystectomy was performed in 12 pigs. The experimental group (6 pigs) received VEGF gene-modified EPC-seeded BAMG. The control group (6 pigs) received BAMG without seeded EPCs. The resulting tissue-engineered bladders were subject to a general and histological analysis. Microvessel density (MVD) was assessed using immunohistochemistry. The ex vivo transfection efficiency of EPCs was greater than 60%-70% when concentrated adenovirus was used. The genetically modified cells expressed both VEGF and green fluorescent protein (GFP). Scanning electron microscopy (SEM) and Masson's trichrome staining of cross sections of the cultured cells seeded to BAMG showed cell attachment and proliferation on the surface of the BAMG. Histological examination revealed bladder regeneration in a time-dependent fashion. Significant increases in MVD were observed in the experimental group, in comparison with the control group. VEGF-modified EPCs significantly enhanced neovascularization, compared with BAMG alone. These results indicate that EPCs, combined with VEGF gene therapy, may be a suitable approach for increasing blood supply in the tissue engineering of bladders. Thus, a useful strategy to achieve a tissue-engineered bladder is indicated.

Towards modular bone tissue engineering using Ti-Co-doped phosphate glass microspheres: cytocompatibility and dynamic culture studies.

Science.gov (United States)

Peticone, Carlotta; De Silva Thompson, David; Owens, Gareth J; Kim, Hae-Won; Micheletti, Martina; Knowles, Jonathan C; Wall, Ivan

2017-09-01

The production of large quantities of functional vascularized bone tissue ex vivo still represent an unmet clinical challenge. Microcarriers offer a potential solution to scalable manufacture of bone tissue due to their high surface area-to-volume ratio and the capacity to be assembled using a modular approach. Microcarriers made of phosphate bioactive glass doped with titanium dioxide have been previously shown to enhance proliferation of osteoblast progenitors and maturation towards functional osteoblasts. Furthemore, doping with cobalt appears to mimic hypoxic conditions that have a key role in promoting angiogenesis. This characteristic could be exploited to meet the clinical requirement of producing vascularized units of bone tissue. In the current study, the human osteosarcoma cell line MG-63 was cultured on phosphate glass microspheres doped with 5% mol titanium dioxide and different concentrations of cobalt oxide (0%, 2% and 5% mol), under static and dynamic conditions (150 and 300 rpm on an orbital shaker). Cell proliferation and the formation of aggregates of cells and microspheres were observed over a period of two weeks in all glass compositions, thus confirming the biocompatibility of the substrate and the suitability of this system for the formation of compact micro-units of tissue. At the concentrations tested, cobalt was not found to be cytotoxic and did not alter cell metabolism. On the other hand, the dynamic environment played a key role, with moderate agitation having a positive effect on cell proliferation while higher agitation resulting in impaired cell growth. Finally, in static culture assays, the capacity of cobalt doping to induce vascular endothelial growth factor (VEGF) upregulation by osteoblastic cells was observed, but was not found to increase linearly with cobalt oxide content. In conclusion, Ti-Co phosphate glasses were found to support osteoblastic cell growth and aggregate formation that is a necessary precursor to tissue

Low cost options for tissue culture technology in developing countries. Proceedings of a technical meeting

Energy Technology Data Exchange (ETDEWEB)

NONE

2004-02-01

Tissue culture technology is used for the production of doubled haploids, cryopreservation, propagating new plant varieties, conserving rare and endangered plants, difficult-to-propagate plants, and to produce secondary metabolites and transgenic plants. The production of high quality planting material of crop plants and fruit trees, propagated from vegetative parts, has created new opportunities in global trading, benefited growers, farmers, and nursery owners, and improved rural employment. However, there are still major opportunities to produce and distribute high quality planting material, e.g. crops like banana, date palm, cassava, pineapple, plantain, potato, sugarcane, sweet potato, yams, ornamentals, fruit and forest trees. The main advantage of tissue culture technology lies in the production of high quality and uniform planting material that can be multiplied on a year-round basis under disease-free conditions anywhere irrespective of the season and weather. However, the technology is capital, labor and energy intensive. Although, labor is cheap in many developing countries, the resources of trained personnel and equipment are often not readily available. In addition, energy, particularly electricity, and clean water are costly. The energy requirements for tissue culture technology depend on day temperature, day-length and relative humidity, and they have to be controlled during the process of propagation. Individual plant species also differ in their growth requirements. Hence, it is necessary to have low cost options for weaning, hardening of micropropagated plants and finally growing them in the field. This publication describes options for reducing costs to establish and operate tissue culture facilities and primarily focus on plant micropropagation. It includes papers on the basics of tissue culture technology, low cost options for the design of laboratories, use of culture media and containers, energy and labor saving, integration and adoption of

Low cost options for tissue culture technology in developing countries. Proceedings of a technical meeting

International Nuclear Information System (INIS)

2004-02-01

Tissue culture technology is used for the production of doubled haploids, cryopreservation, propagating new plant varieties, conserving rare and endangered plants, difficult-to-propagate plants, and to produce secondary metabolites and transgenic plants. The production of high quality planting material of crop plants and fruit trees, propagated from vegetative parts, has created new opportunities in global trading, benefited growers, farmers, and nursery owners, and improved rural employment. However, there are still major opportunities to produce and distribute high quality planting material, e.g. crops like banana, date palm, cassava, pineapple, plantain, potato, sugarcane, sweet potato, yams, ornamentals, fruit and forest trees. The main advantage of tissue culture technology lies in the production of high quality and uniform planting material that can be multiplied on a year-round basis under disease-free conditions anywhere irrespective of the season and weather. However, the technology is capital, labor and energy intensive. Although, labor is cheap in many developing countries, the resources of trained personnel and equipment are often not readily available. In addition, energy, particularly electricity, and clean water are costly. The energy requirements for tissue culture technology depend on day temperature, day-length and relative humidity, and they have to be controlled during the process of propagation. Individual plant species also differ in their growth requirements. Hence, it is necessary to have low cost options for weaning, hardening of micropropagated plants and finally growing them in the field. This publication describes options for reducing costs to establish and operate tissue culture facilities and primarily focus on plant micropropagation. It includes papers on the basics of tissue culture technology, low cost options for the design of laboratories, use of culture media and containers, energy and labor saving, integration and adoption of

Plant regeneration from petiole segments of some species in tissue culture

Directory of Open Access Journals (Sweden)

Krystyna Klimaszewska

2013-12-01

Full Text Available The regeneration ability of 21 plant species belonging to 14 families was tested. The method of tissue culture in vitro was applied, on basic MS medium with an addition of growth regulators from the auxin and cytokinin groups. From among the investigated plant groups Peperomia scandens and Caladium × hortulanum were capable of plant regeneration, Passiilora coerulea regenerated shoots, Hedera helix, Begonia glabra, Coleus blumei, Fuchsia hybrida, Passiflora suberosa and Peperomia eburnea formed callus and roots, Kalanchoe blossfeldiana, Pelargonium grandiflorum, P. peltatum, P. radula, Coleus shirensis and Magnolia soulangeana produced callus, Philodendron scandens, Rhododendron smirnovii, Hibiscus rosa-sinensis, Coprosma baueri, Cestrum purpureum and Solanum rantonnetii did not exhibit any regeneration reactions.

Influence of Cryopreservation Solution on the In Vitro Culture of Skin Tissues Derived from Collared Peccary (Pecari tajacu Linnaeus, 1758).

Science.gov (United States)

Borges, Alana A; Lira, Gabriela P O; Nascimento, Lucas E; Queiroz Neta, Luiza B; Santos, Maria V O; Oliveira, Moacir F; Silva, Alexandre R; Pereira, Alexsandra F

2018-04-01

Skin vitrification is a promising and alternative tool for the conservation of biodiversity, especially for wild mammals, such as collared peccaries. Several factors can affect the success of this procedure, such as the cryoprotectant solution used. Therefore, this study was carried out to compare the efficiency of various vitrification solutions for recovery of viable cells after in vitro culture of cryopreserved skin tissues derived from the collared peccary, aiming to study the application in biobanking, where cellular use is not immediately required. Then, Dulbecco's modified Eagle's medium (DMEM) composed of 2.2 g/L sodium bicarbonate and 10% fetal bovine serum (FBS) was supplemented with 3.0 M ethylene glycol (EG) or 3.0 M dimethyl sulfoxide (DMSO) or 1.5 M EG plus 1.5 M DMSO with or without sucrose (SUC; 0.25 M) to produce six solutions for solid-surface vitrification. After warming, skin tissues were cultured in vitro and recovered cells were analyzed for morphology, adhesion, subconfluence, and proliferative activity for developing the growth curve and determining the population doubling time (PDT), and viability by Trypan Blue. The vitrification did not alter the ability of the tissues to adhere to the culture dish, as well as the day of all explants with cell growth, subconfluence samples, subconfluence total time, and PDT (p > 0.05). Moreover, independent of the cryoprotectant solution used, the vitrification altered the day of all attached explants (p  0.05). Additionally, for viability after the third passage, only the EG-SUC group maintained the cell quality (88.3%), when compared with the nonvitrified (97.8%, p > 0.05). In conclusion, DMEM with 10% FBS, 3.0 M EG, and 0.25 M sucrose was the most efficient solution for vitrifying collared peccary skin tissues, leading to the in vitro culture of viable cells.

IN VITRO PROPAGATION OF DENDROBIUM AND PHALAENOPSIS THROUGH TISSUE CULTURE FOR CONSERVATION

Directory of Open Access Journals (Sweden)

Lita Soetopo

2012-06-01

Full Text Available The studies were focused on developing an efficient and effective propagation protocol for orchid species from genera Dendrobioum and Phalaenopsis through tissue culture. The Materials used were explants from adventive shoot tip, floral stalk buds and PLBs derived from seeds. The results indicated growth and development of adventive shoot tip explants of Dendrobium: a high survival percentage for explant with green color was shown by D. racianum, followed by D. laxiflorum, D. pseudo-conantum, D. strebloceras, D. lineale, and D. veratrifolium. However, plantlets regeneration occurred only on D. pseudoconantum, and D. strebloceras. Explant regeneration from seed derived protocorm-like bodies on D. spectabile occurred 40 days after inoculation transfer and subculture. High survival percentage of explant from floral stalk shoot was shown by P. amabilis. There were several plantlets surviving in acclimatisation. Explant regeneration from seed derived from protocorm-like bodies on P. hieroglypha occurred 40 days after inoculation and subculture. It was suggested that for ex situ conservation on certain species of Dendrobium and Phalaenopsis in the category of rare germplasms, tissue culture could be applied effectively and efficiently by using explant from adventive shoot tip, floral stalk buds and seed derived protocorm-like body explant for vegetative seed multiplication.

Propagation of Aquilaria malaccensis seedlings through tissue culture techniques

International Nuclear Information System (INIS)

Salahbiah Abdul Majid; Zaiton Ahmad; Mohd Rafaie Abdul Salam; Nurhayati Irwan; Affrida Abu Hassan; Rusli Ibrahim

2010-01-01

Aquilaria malaccensis or karas is the principal source of gaharu resin, which is used in many cultures for incense, perfumes and traditional medicines. The species is mainly propagated conventionally through seeds, cuttings and graftings. Propagation by seeds is usually a reliable method for other forest species, but for karas, this technique is inadequate to meet the current demand of seedling supplies. This is principally due to its low seed viability, low germination rate, delayed rooting of seedlings, long life-cycle and rare seed production. Tissue culture has several advantages over conventional propagation, especially for obtaining large number of uniform and high-yielding plantlets or clones. This paper presents the current progress on mass-propagation of Aquilaria malaccensis seedlings through tissue culture technique at Nuclear Malaysia. (author)

Bridging the gap between cell culture and live tissue

Directory of Open Access Journals (Sweden)

Stefan Przyborski

2017-11-01

Full Text Available Traditional in vitro two-dimensional (2-D culture systems only partly imitate the physiological and biochemical features of cells in their original tissue. In vivo, in organs and tissues, cells are surrounded by a three-dimensional (3-D organization of supporting matrix and neighbouring cells, and a gradient of chemical and mechanical signals. Furthermore, the presence of blood flow and mechanical movement provides a dynamic environment (Jong et al., 2011. In contrast, traditional in vitro culture, carried out on 2-D plastic or glass substrates, typically provides a static environment, which, however is the base of the present understanding of many biological processes, tissue homeostasis as well as disease. It is clear that this is not an exact representation of what is happening in vivo and the microenvironment provided by in vitro cell culture models are significantly different and can cause deviations in cell response and behaviour from those distinctive of in vivo tissues. In order to translate the present basic knowledge in cell control, cell repair and regeneration from the laboratory bench to the clinical application, we need a better understanding of the cell and tissue interactions. This implies a detailed comprehension of the natural tissue environment, with its organization and local signals, in order to more closely mimic what happens in vivo, developing more physiological models for efficient in vitro systems. In particular, it is imperative to understand the role of the environmental cues which can be mainly divided into those of a chemical and mechanical nature.

Culture-Independent Identification of Mycobacterium avium Subspecies paratuberculosis in Ovine Tissues: Comparison with Bacterial Culture and Histopathological Lesions

Directory of Open Access Journals (Sweden)

Kamal R. Acharya

2017-12-01

Full Text Available Johne’s disease is a chronic debilitating enteropathy of ruminants caused by Mycobacterium avium subspecies paratuberculosis (MAP. Current abattoir surveillance programs detect disease via examination of gross lesions and confirmation by histopathological and/or tissue culture, which is time-consuming and has relatively low sensitivity. This study aimed to investigate whether a high-throughput quantitative PCR (qPCR test is a viable alternative for tissue testing. Intestine and mesenteric lymph nodes were sourced from sheep experimentally infected with MAP and the DNA extracted using a protocol developed for tissues, comprised enzymatic digestion of the tissue homogenate, chemical and mechanical lysis, and magnetic bead-based DNA purification. The extracted DNA was tested by adapting a previously validated qPCR for fecal samples, and the results were compared with culture and histopathology results of the corresponding tissues. The MAP tissue qPCR confirmed infection in the majority of sheep with gross lesions on postmortem (37/38. Likewise, almost all tissue culture (61/64 or histopathology (52/58 positives were detected with good to moderate agreement (Cohen’s kappa statistic and no significant difference to the reference tests (McNemar’s Chi-square test. Higher MAP DNA quantities corresponded to animals with more severe histopathology (odds ratio: 1.82; 95% confidence interval: 1.60, 2.07. Culture-independent strain typing on tissue DNA was successfully performed. This MAP tissue qPCR method had a sensitivity equivalent to the reference tests and is thus a viable replacement for gross- and histopathological examination of tissue samples in abattoirs. In addition, the test could be validated for testing tissue samples intended for human consumption.

Growth versus metabolic tissue replacement in mouse tissues determined by stable carbon and nitrogen isotope analysis

Science.gov (United States)

Macavoy, S. E.; Jamil, T.; Macko, S. A.; Arneson, L. S.

2003-12-01

Stable isotope analysis is becoming an extensively used tool in animal ecology. The isotopes most commonly used for analysis in terrestrial systems are those of carbon and nitrogen, due to differential carbon fractionation in C3 and C4 plants, and the approximately 3‰ enrichment in 15N per trophic level. Although isotope signatures in animal tissues presumably reflect the local food web, analysis is often complicated by differential nutrient routing and fractionation by tissues, and by the possibility that large organisms are not in isotopic equilibrium with the foods available in their immediate environment. Additionally, the rate at which organisms incorporate the isotope signature of a food through both growth and metabolic tissue replacement is largely unknown. In this study we have assessed the rate of carbon and nitrogen isotopic turnover in liver, muscle and blood in mice following a diet change. By determining growth rates, we were able to determine the proportion of tissue turnover caused by growth versus that caused by metabolic tissue replacement. Growth was found to account for approximately 10% of observed tissue turnover in sexually mature mice (Mus musculus). Blood carbon was found to have the shortest half-life (16.9 days), followed by muscle (24.7 days). Liver carbon turnover was not as well described by the exponential decay equations as other tissues. However, substantial liver carbon turnover was observed by the 28th day after diet switch. Surprisingly, these tissues primarily reflect the carbon signature of the protein, rather than carbohydrate, source in their diet. The nitrogen signature in all tissues was enriched by 3 - 5‰ over their dietary protein source, depending on tissue type, and the isotopic turnover rates were comparable to those observed in carbon.

Studies on salt and drought tolerance of lavender using tissue culture and gamma rays

International Nuclear Information System (INIS)

Essam, K.E.; El-Sharnoby, M.E.

2005-01-01

The present study was carried out to investigate the propagation and chemical composition of Lavandula spica using different cytokinins (BA, Ki and 2ip) on MS medium,, besides medium strength. Also, GA3 concentrations, auxin types, gamma irradiation (0, 2, 4, 6 and 8 Krad), different concentrations of mannitol (10, 20, 40, 80 and 100 mg/l) as well as different salinity concentrations of CaCl 2 or NaCl or both at levels 250, 500, 750 and 1000 ppm were used in the study. Maximum proliferation parameter was produced on MS medium supplemented with 1 mg/l BA than other cytokinin types. Growing the explant of Lavandula. spica on MS medium, containing 3 mg/l BA, gave the highest proliferation, growth and greening parameters. However, using MS medium at half strength supplemented with 3 mg/l BA resulted in significant increase in both shoot elongation and greening parameters as compared with the other medium strengths, while 4 mg/l GA induced the best shoot elongation. Adding 1 mg/l IBA enhanced rooting and also the obtained results showed that increasing gamma irradiation decreased growth and proliferation parameters. Moreover, increasing drought stress induced an adverse effect on tissue culture parameter while some chemical analysis parameters were increased to maximum degree of their tolerance to drought stress. Also, using different salinity treatment by NaCl, CaCl 2 and their combinations showed adverse effects on tissue culture parameters chemical composition

Development of molecular markers for zebrafish (Danio rerio) ovarian follicle growth assessment following in-vitro culture in cryopreservation studies.

Science.gov (United States)

Anil, Siji; Rawson, David; Zhang, Tiantian

2018-05-29

Development of in vitro culture protocol for early stage ovarian follicles of zebrafish is important since cryopreserved early stage ovarian follicles would need to be matured in vitro following cryopreservation before they can be fertilised. Development of molecular markers for zebrafish (Danio rerio) ovarian follicle growth assessment following in vitro culture of early stage zebrafish ovarian follicles in ovarian tissue fragments is reported here for the first time although some work has been reported for in vitro culture of isolated early stage zebrafish ovarian follicles. The main aim of the present study was to develop molecular markers in an optimised in vitro culture protocol for stage I and stage II zebrafish ovarian follicles in ovarian tissue fragments. The effect of concentration of the hormones human chorionic gonadotropin and follicle stimulating hormones, and additives such as Foetal Bovine Serum and Bovine Serum Albumin were studied. The results showed that early stage zebrafish ovarian fragments containing stage I and stage II follicles which are cultured in vitro for 24 h in 20% FBS and 100mIU/ml FSH in 90% L-15 medium at 28 °C can grow to the size of stage II and stage III ovarian follicles respectively. More importantly the follicle growth from stage I to stage II and from stage II to stage III were confirmed using molecular markers such as cyp19a1a (also known as P450aromA) and vtg1 genes respectively. However, no follicle growth was observed following cryopreservation and in vitro culture. Copyright © 2018 Elsevier Inc. All rights reserved.

On the genetic control of planar growth during tissue morphogenesis in plants.

Science.gov (United States)

Enugutti, Balaji; Kirchhelle, Charlotte; Schneitz, Kay

2013-06-01

Tissue morphogenesis requires extensive intercellular communication. Plant organs are composites of distinct radial cell layers. A typical layer, such as the epidermis, is propagated by stereotypic anticlinal cell divisions. It is presently unclear what mechanisms coordinate cell divisions relative to the plane of a layer, resulting in planar growth and maintenance of the layer structure. Failure in the regulation of coordinated growth across a tissue may result in spatially restricted abnormal growth and the formation of a tumor-like protrusion. Therefore, one way to approach planar growth control is to look for genetic mutants that exhibit localized tumor-like outgrowths. Interestingly, plants appear to have evolved quite robust genetic mechanisms that govern these aspects of tissue morphogenesis. Here we provide a short summary of the current knowledge about the genetics of tumor formation in plants and relate it to the known control of coordinated cell behavior within a tissue layer. We further portray the integuments of Arabidopsis thaliana as an excellent model system to study the regulation of planar growth. The value of examining this process in integuments was established by the recent identification of the Arabidopsis AGC VIII kinase UNICORN as a novel growth suppressor involved in the regulation of planar growth and the inhibition of localized ectopic growth in integuments and other floral organs. An emerging insight is that misregulation of central determinants of adaxial-abaxial tissue polarity can lead to the formation of spatially restricted multicellular outgrowths in several tissues. Thus, there may exist a link between the mechanisms regulating adaxial-abaxial tissue polarity and planar growth in plants.

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

International Nuclear Information System (INIS)

Hall, E.R.; Falkler, W.A. Jr.; Suzuki, J.B.

1990-01-01

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

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

Energy Technology Data Exchange (ETDEWEB)

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

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.

Trickle-bed root culture bioreactor design and scale-up: growth, fluid-dynamics, and oxygen mass transfer.

Science.gov (United States)

Ramakrishnan, Divakar; Curtis, Wayne R

2004-10-20

Trickle-bed root culture reactors are shown to achieve tissue concentrations as high as 36 g DW/L (752 g FW/L) at a scale of 14 L. Root growth rate in a 1.6-L reactor configuration with improved operational conditions is shown to be indistinguishable from the laboratory-scale benchmark, the shaker flask (mu=0.33 day(-1)). These results demonstrate that trickle-bed reactor systems can sustain tissue concentrations, growth rates and volumetric biomass productivities substantially higher than other reported bioreactor configurations. Mass transfer and fluid dynamics are characterized in trickle-bed root reactors to identify appropriate operating conditions and scale-up criteria. Root tissue respiration goes through a minimum with increasing liquid flow, which is qualitatively consistent with traditional trickle-bed performance. However, liquid hold-up is much higher than traditional trickle-beds and alternative correlations based on liquid hold-up per unit tissue mass are required to account for large changes in biomass volume fraction. Bioreactor characterization is sufficient to carry out preliminary design calculations that indicate scale-up feasibility to at least 10,000 liters.

Effect of gamma-radiation on callus initiation and oraganogenesis in the tissue culture of Nicotiana tabaccum L

International Nuclear Information System (INIS)

Shin, S. H.; Kim, J. G.; Song, H. S.

2004-01-01

It is generally agreed that ionizing radiations stimulate cell division, growth and development in various organisms including animals and plants. Differentiating tissues are the most sensitive to radiation. The present experiment was carried out to investigate the effects of ionizing radiation on callus initiation and organogenesis from the stem in the culture of Nicotiana tabaccum L. cv. When the stem segments were cultured on a Murashige and Skoog (MS) medium with 2 mg/L kinetin, with 1 mg/L 2,4-Dichlorophenoxyacetic acid (2,4-D), with 2 mg/L kinetin and 1 mg/L 2,4-D, the shoots and callus were differentiated 14 days after cultivation. Callus was especially formed on the MS medium with 2,4-D and/or kinetin and the formation was promoted by 1 Gy and 5 Gy of gamma radiation. The formation of the shoot clusters on the MS medium with 2 mg/L kinetin were prominent in the 5 Gy-irradiated groups. It is concluded that that gamma radiation enhanced the callus initiation and organogenesis in the tissue culture of Nicotiana tabaccum L

Discarded human fetal tissue and cell cultures for transplantation research

International Nuclear Information System (INIS)

Hay, R.J.; Phillips, T.; Thompson, A.; Vilner, L.; Cleland, M.; Tchaw-ren Chen; Zabrenetzky, V.

1999-01-01

A feasibility study has been performed to explore the utility of various tissues from discarded human abortuses for transplantation and related research. Specifically, aborted fetuses plus parental blood samples and all relevant clinical data were obtained through a local hospital complex. Whenever possible, pancreas, skin and skeletal muscle, heart, liver, kidney, cartilage and lung tissues were removed, dissociated and subfractionated for cryopreservation, characterization and cultivation trials in vitro. Existing protocols for these manipulations were compared and improved upon as required. Clonal culture, cell aggregate maintenance techniques and use of feeder cell populations have been utilized where appropriate to develop quantitative comparative data. Histological and biochemical assays were applied both to evaluate separation/cultivation methods and to identify optimal culture conditions for maintaining functional cells. Immunochemical and molecular biological procedures were applied to study expression of Major Histocompatibility Vomplex (MHC) class 1 and 11 molecules on cell lines derived. Tissue and cell culture populations were examined for infections with bacteria, ftingi, mycoplasma, HIV, CMV, hepatitis B and other viruses. Only 1% of the abortuses tested were virally infected. Cytogenetic analyses confin-ned the normal diploid status in the vast majority (>98%) of lines tested. A total of over 250 abortuses have been obtained and processed. Only 25 were found to be contaminated with bacteria or fungi and unsuitable for further cultivation trials. A total of over 200 cell populations were isolated, characterized and cryopreserved for further study. Included were kidney, lung, liver and epidermal epithelia: cartilage-derived cells from the spine and epiphyses plus myogenic myoblasts. Selected lines have been immortalized using HPV I 6E6/E7 sequences. Epithelia from the liver and pancreas and cardiac myocytes were the most problematic in that initial

The effect of radiation in combination with carcinogens on the growth of normal urothelium in explant culture

International Nuclear Information System (INIS)

Mothersill, C.; O'Brien, A.

1990-01-01

Radiation is known to be carcinogenic to humans but attempts to demonstrate the process using human tissue culture models have met with little success. In the present study explants were established from urothelium and exposed to radiation and a range of chemical carcinogens, suspected promotor or metabolic agents. The resulting outgrowth was monitored for growth rate, proliferating epithelial fraction and development and differentiation of endothelial cells in culture. The results indicate that enhanced growth of epithelial cells can be seen when cultures are irradiated in the presence of various nitrosamines, benzo(a)pyrene or aniline. Radiation alone reduced the overall growth area measured but several proliferative foci developed on the resulting outgrowth. Their ultrastructural appearance reveals that they carry severe mitochondrial damage and exposure of treated cultures to metabolic inhibitors confirms that their respiration is defective. Endothelial cells proliferated over the surface of the epithelial monolayer and both the number and the degree of differentiation of the endothelial cells increased with increasing dose up to 10 Gy. While the cultures are not immortalised by the treatment, it appears that the epithelial cells have an extended lifespan (division capacity) and that a subpopulation has undergone a number of premalignant changes. Changes in endothelial cell proliferation also occur. (orig.)

Low technology tissue culture materials for initiation and ...

African Journals Online (AJOL)

Low technology tissue culture materials for initiation and multiplication of banana plants. ... African Crop Science Journal ... locally available macronutrients, micronutrients, sugar, equipment and facility reduced the cost of consumable material

A structure-based extracellular matrix expansion mechanism of fibrous tissue growth.

Science.gov (United States)

Kalson, Nicholas S; Lu, Yinhui; Taylor, Susan H; Starborg, Tobias; Holmes, David F; Kadler, Karl E

2015-05-20

Embryonic growth occurs predominately by an increase in cell number; little is known about growth mechanisms later in development when fibrous tissues account for the bulk of adult vertebrate mass. We present a model for fibrous tissue growth based on 3D-electron microscopy of mouse tendon. We show that the number of collagen fibrils increases during embryonic development and then remains constant during postnatal growth. Embryonic growth was explained predominately by increases in fibril number and length. Postnatal growth arose predominately from increases in fibril length and diameter. A helical crimp structure was established in embryogenesis, and persisted postnatally. The data support a model where the shape and size of tendon is determined by the number and position of embryonic fibroblasts. The collagen fibrils that these cells synthesise provide a template for postnatal growth by structure-based matrix expansion. The model has important implications for growth of other fibrous tissues and fibrosis.

Renal cell carcinoma primary cultures maintain genomic and phenotypic profile of parental tumor tissues

International Nuclear Information System (INIS)

Cifola, Ingrid; Magni, Fulvio; Signorini, Stefano; Battaglia, Cristina; Perego, Roberto A; Bianchi, Cristina; Mangano, Eleonora; Bombelli, Silvia; Frascati, Fabio; Fasoli, Ester; Ferrero, Stefano; Di Stefano, Vitalba; Zipeto, Maria A

2011-01-01

Clear cell renal cell carcinoma (ccRCC) is characterized by recurrent copy number alterations (CNAs) and loss of heterozygosity (LOH), which may have potential diagnostic and prognostic applications. Here, we explored whether ccRCC primary cultures, established from surgical tumor specimens, maintain the DNA profile of parental tumor tissues allowing a more confident CNAs and LOH discrimination with respect to the original tissues. We established a collection of 9 phenotypically well-characterized ccRCC primary cell cultures. Using the Affymetrix SNP array technology, we performed the genome-wide copy number (CN) profiling of both cultures and corresponding tumor tissues. Global concordance for each culture/tissue pair was assayed evaluating the correlations between whole-genome CN profiles and SNP allelic calls. CN analysis was performed using the two CNAG v3.0 and Partek software, and comparing results returned by two different algorithms (Hidden Markov Model and Genomic Segmentation). A very good overlap between the CNAs of each culture and corresponding tissue was observed. The finding, reinforced by high whole-genome CN correlations and SNP call concordances, provided evidence that each culture was derived from its corresponding tissue and maintained the genomic alterations of parental tumor. In addition, primary culture DNA profile remained stable for at least 3 weeks, till to third passage. These cultures showed a greater cell homogeneity and enrichment in tumor component than original tissues, thus enabling a better discrimination of CNAs and LOH. Especially for hemizygous deletions, primary cultures presented more evident CN losses, typically accompanied by LOH; differently, in original tissues the intensity of these deletions was weaken by normal cell contamination and LOH calls were missed. ccRCC primary cultures are a reliable in vitro model, well-reproducing original tumor genetics and phenotype, potentially useful for future functional approaches

Computational modeling of adherent cell growth in a hollow-fiber membrane bioreactor for large-scale 3-D bone tissue engineering.

Science.gov (United States)

Mohebbi-Kalhori, Davod; Behzadmehr, Amin; Doillon, Charles J; Hadjizadeh, Afra

2012-09-01

The use of hollow-fiber membrane bioreactors (HFMBs) has been proposed for three-dimensional bone tissue growth at the clinical scale. However, to achieve an efficient HFMB design, the relationship between cell growth and environmental conditions must be determined. Therefore, in this work, a dynamic double-porous media model was developed to determine nutrient-dependent cell growth for bone tissue formation in a HFMB. The whole hollow-fiber scaffold within the bioreactor was treated as a porous domain in this model. The domain consisted of two interpenetrating porous regions, including a porous lumen region available for fluid flow and a porous extracapillary space filled with a collagen gel that contained adherent cells for promoting long-term growth into tissue-like mass. The governing equations were solved numerically and the model was validated using previously published experimental results. The contributions of several bioreactor design and process parameters to the performance of the bioreactor were studied. The results demonstrated that the process and design parameters of the HFMB significantly affect nutrient transport and thus cell behavior over a long period of culture. The approach presented here can be applied to any cell type and used to develop tissue engineering hollow-fiber scaffolds.

21 CFR 864.2220 - Synthetic cell and tissue culture media and components.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Synthetic cell and tissue culture media and components. 864.2220 Section 864.2220 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Cell And Tissue Culture...

Growth and accumulation of flavan-3-ol in Camellia sinensis through callus culture and suspension culture method

Directory of Open Access Journals (Sweden)

Sutini Sutini

2017-02-01

Full Text Available This study was aimed to assess flavan-3-ol biomass in C. sinensis through callus cultures and suspension cultures derived from leaf explants. Callus initiation of both cultures were using Murashige and Skoog medium were enriched with plant growth regulators Naphtha-lene Acetic Acid 3.0 mg/L and kinetin 2.0 mg/L. The procedures in this study were: (1 callus initiation by cutting the leaves of C. sinen-sis shoots then planted on Murashige and Skoog medium that were enriched with plant growth regulators, (2 sub callus culture on fresh medium that enriched with the same growth regulators, (3 suspension culture initiation of liquid callus, (4 growth examination of callus and suspension cultures in week 12, (5 examination of qualitative-quantitative content of flavan-3-olin suspension cultures at week 4. The results show that suspension cultures contain biomass flavan-3-ol that increase in the same manner of the increase of callus age and weight

Mathematical Model of Growth Factor Driven Haptotaxis and Proliferation in a Tissue Engineering Scaffold

KAUST Repository

Pohlmeyer, J. V.

2013-01-29

Motivated by experimental work (Miller et al. in Biomaterials 27(10):2213-2221, 2006, 32(11):2775-2785, 2011) we investigate the effect of growth factor driven haptotaxis and proliferation in a perfusion tissue engineering bioreactor, in which nutrient-rich culture medium is perfused through a 2D porous scaffold impregnated with growth factor and seeded with cells. We model these processes on the timescale of cell proliferation, which typically is of the order of days. While a quantitative representation of these phenomena requires more experimental data than is yet available, qualitative agreement with preliminary experimental studies (Miller et al. in Biomaterials 27(10):2213-2221, 2006) is obtained, and appears promising. The ultimate goal of such modeling is to ascertain initial conditions (growth factor distribution, initial cell seeding, etc.) that will lead to a final desired outcome. © 2013 Society for Mathematical Biology.

Does education engender cultural values that matter for economic growth?

OpenAIRE

Prosper F. Bangwayo-Skeete; Afaf H. Rahim; Precious Zikhali

2009-01-01

Empirical research has shown that cultural values matter for economic growth and has specifically identified the achievement motivation as an aspect of culture that engenders economic growth. If specific cultural values engender economic growth, how then can societies promote them? This paper attempts to answer this question using the 2005 wave of the World Values Survey data for 43 countries. We test the contention that education significantly impacts the relative importance an individual pl...

Effect of x-ray irradiation on maize inbred line B73 tissue cultures and regenerated plants

International Nuclear Information System (INIS)

Wang, A.S.; Cheng, D.S.K.; Milcic, J.B.; Yang, T.C.

1988-01-01

In order to enhance variation induced by the tissue culture process and to obtain agronomically desirable mutants, friable embryogenic tissue cultures of maize (Zea mays L.) inbred line B73 were x-ray irradiated with 11 doses [0-8.4 kilorads (kR)]. Reductions in callus growth rate and embryogenic callus formation occurred with increasing x-ray doses 20 d and 3 months after irradiation. Callus irradiated with 0.8 kR showed a significant increase in growth rate and a 20% increase in embryogenic callus 9 months after irradiation. A total of 230 R 0 plants were regenerated for evaluation. Pollen fertility and seed set of R 0 plants decreased with increasing x-ray dosage. Days to anthesis and plant height of R 0 plants varied among x-ray treatments but were generally reduced with higher dosages. The number of chromosomal aberrations increased with x-ray dosage. The R 1 seeds taken from R 0 plants were also grown and tested for mutant segregation. Plants regenerated from irradiated calli had a two- to 10-fold increase in mutations over plants regenerated from unirradiated control callus. Germination frequency of seeds from R 0 plants decreased with increasing x-ray dosage. Although chlorophyll mutants were most frequently observed, a number of vigorous plants with earlier anthesis date were also recovered

Renal cell carcinoma primary cultures maintain genomic and phenotypic profile of parental tumor tissues.

Science.gov (United States)

Cifola, Ingrid; Bianchi, Cristina; Mangano, Eleonora; Bombelli, Silvia; Frascati, Fabio; Fasoli, Ester; Ferrero, Stefano; Di Stefano, Vitalba; Zipeto, Maria A; Magni, Fulvio; Signorini, Stefano; Battaglia, Cristina; Perego, Roberto A

2011-06-13

Clear cell renal cell carcinoma (ccRCC) is characterized by recurrent copy number alterations (CNAs) and loss of heterozygosity (LOH), which may have potential diagnostic and prognostic applications. Here, we explored whether ccRCC primary cultures, established from surgical tumor specimens, maintain the DNA profile of parental tumor tissues allowing a more confident CNAs and LOH discrimination with respect to the original tissues. We established a collection of 9 phenotypically well-characterized ccRCC primary cell cultures. Using the Affymetrix SNP array technology, we performed the genome-wide copy number (CN) profiling of both cultures and corresponding tumor tissues. Global concordance for each culture/tissue pair was assayed evaluating the correlations between whole-genome CN profiles and SNP allelic calls. CN analysis was performed using the two CNAG v3.0 and Partek software, and comparing results returned by two different algorithms (Hidden Markov Model and Genomic Segmentation). A very good overlap between the CNAs of each culture and corresponding tissue was observed. The finding, reinforced by high whole-genome CN correlations and SNP call concordances, provided evidence that each culture was derived from its corresponding tissue and maintained the genomic alterations of parental tumor. In addition, primary culture DNA profile remained stable for at least 3 weeks, till to third passage. These cultures showed a greater cell homogeneity and enrichment in tumor component than original tissues, thus enabling a better discrimination of CNAs and LOH. Especially for hemizygous deletions, primary cultures presented more evident CN losses, typically accompanied by LOH; differently, in original tissues the intensity of these deletions was weaken by normal cell contamination and LOH calls were missed. ccRCC primary cultures are a reliable in vitro model, well-reproducing original tumor genetics and phenotype, potentially useful for future functional approaches

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

African Journals Online (AJOL)

To achieve the best explants and media for spinach tissue culture, the effects of two different plant growth regulators, two explants and cultivars on adventitious shoot regeneration were tested. The Analysis of Variance (ANOVA) showed that the effects of plant growth regulators on spinach tissue culture were significant; ...

Mouse embryonic stem cell culture for generation of three-dimensional retinal and cortical tissues.

Science.gov (United States)

Eiraku, Mototsugu; Sasai, Yoshiki

2011-12-15

Generation of compound tissues with complex structures is a major challenge in cell biology. In this article, we describe a protocol for mouse embryonic stem cell (ESC) culture for in vitro generation of three-dimensional retinal tissue, comparing it with the culture protocol for cortical tissue generation. Dissociated ESCs are reaggregated in a 96-well plate with reduced cell-plate adhesion and cultured as floating aggregates. Retinal epithelium is efficiently generated when ESC aggregates are cultured in serum-free medium containing extracellular matrix proteins, spontaneously forming hemispherical vesicles and then progressively transforming into a shape reminiscent of the embryonic optic cup in 9-10 d. In long-term culture, the ESC-derived optic cup generates a fully stratified retinal tissue consisting of all major neural retinal components. In contrast, the cortical differentiation culture can be started without exogenous extracellular matrix proteins, and it generates stratified cortical epithelia consisting of four distinct layers in 13 d.

Endothelial cells stimulate growth of normal and cancerous breast epithelial cells in 3D culture

Directory of Open Access Journals (Sweden)

Magnusson Magnus K

2010-07-01

Full Text Available Abstract Background Epithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer. However, research on the regulatory role of the endothelial component in the normal and malignant breast gland has largely been neglected. The aim of the study was to investigate the effects of endothelial cells on growth and differentiation of human breast epithelial cells in a three-dimensional (3D co-culture assay. Methods Breast luminal and myoepithelial cells and endothelial cells were isolated from reduction mammoplasties. Primary cells and established normal and malignant breast cell lines were embedded in reconstituted basement membrane in direct co-culture with endothelial cells and by separation of Transwell filters. Morphogenic and phenotypic profiles of co-cultures was evaluated by phase contrast microscopy, immunostaining and confocal microscopy. Results In co-culture, endothelial cells stimulate proliferation of both luminal- and myoepithelial cells. Furthermore, endothelial cells induce a subpopulation of luminal epithelial cells to form large acini/ducts with a large and clear lumen. Endothelial cells also stimulate growth and cloning efficiency of normal and malignant breast epithelial cell lines. Transwell and gradient co-culture studies show that endothelial derived effects are mediated - at least partially - by soluble factors. Conclusion Breast endothelial cells - beside their role in transporting nutrients and oxygen to tissues - are vital component of the epithelial microenvironment in the breast and provide proliferative signals to the normal and malignant breast epithelium. These growth promoting effects of endothelial cells should be taken into consideration in breast cancer biology.

A stress driven growth model for soft tissue considering biological availability

International Nuclear Information System (INIS)

Oller, S; Bellomo, F J; Nallim, L G; Armero, F

2010-01-01

Some of the key factors that regulate growth and remodeling of tissues are fundamentally mechanical. However, it is important to take into account the role of bioavailability together with the stresses and strains in the processes of normal or pathological growth. In this sense, the model presented in this work is oriented to describe the growth of soft biological tissue under 'stress driven growth' and depending on the biological availability of the organism. The general theoretical framework is given by a kinematic formulation in large strain combined with the thermodynamic basis of open systems. The formulation uses a multiplicative decomposition of deformation gradient, splitting it in a growth part and visco-elastic part. The strains due to growth are incompatible and are controlled by an unbalanced stresses related to a homeostatic state. Growth implies a volume change with an increase of mass maintaining constant the density. One of the most interesting features of the proposed model is the generation of new tissue taking into account the contribution of mass to the system controlled through biological availability. Because soft biological tissues in general have a hierarchical structure with several components (usually a soft matrix reinforced with collagen fibers), the developed growth model is suitable for the characterization of the growth of each component. This allows considering a different behavior for each of them in the context of a generalized theory of mixtures. Finally, we illustrate the response of the model in case of growth and atrophy with an application example.

Microfluidic 3D cell culture: potential application for tissue-based bioassays

Science.gov (United States)

Li, XiuJun (James); Valadez, Alejandra V.; Zuo, Peng; Nie, Zhihong

2014-01-01

Current fundamental investigations of human biology and the development of therapeutic drugs, commonly rely on two-dimensional (2D) monolayer cell culture systems. However, 2D cell culture systems do not accurately recapitulate the structure, function, physiology of living tissues, as well as highly complex and dynamic three-dimensional (3D) environments in vivo. The microfluidic technology can provide micro-scale complex structures and well-controlled parameters to mimic the in vivo environment of cells. The combination of microfluidic technology with 3D cell culture offers great potential for in vivo-like tissue-based applications, such as the emerging organ-on-a-chip system. This article will review recent advances in microfluidic technology for 3D cell culture and their biological applications. PMID:22793034

Which Dimensions of Culture Matter for Long-Run Growth?

OpenAIRE

Yuriy Gorodnichenko; Gerard Roland

2011-01-01

We present empirical evidence that, among a variety of cultural dimensions, the individualism-collectivism dimension, based on Hofstede's (2001) data, is the most important and robustly significant effect of culture on long run growth. Other dimensions that have a significant effect, albeit less robust, are generally strongly correlated with individualism and convey similar information. We found no significant or robust effect on growth from cultural dimensions that are independent from the i...

Zastosowanie peptonu Peptobak-Bacutil tu hodowli merystematycznej tkanki Cymbidium Sw. [Application of the peptone Peptobak-Bacutil medium in the culture of meristem-tissue of Cymbidium Sw.

Directory of Open Access Journals (Sweden)

K. Kukułczanka

2015-06-01

Full Text Available In was found that peptone of Peptobak-Bacutil brand (produced in Poland fulfills perfectly the requirements in cultures in vitro of the meristem tissues of Cymbidium in the modified Tsuchiya's medium in a concentration of 2 g per 1. A decrease in the concentration to 1 g per 1. affects the growth intensity of the tissue, but an increase of concentration up to 3 g per 1. retards the transformation of protocorms into the plant-stage. Growth of the meristem tissue of Cymbidium in the medium consolidated with agar is weaker and the number of protocorms produced 2,3 times lower than in liquid media.

Protein and Glycoprotein Patterns Related to Morphogenesis in Mammillaria gracillis Pfeiff. Tissue Culture

Directory of Open Access Journals (Sweden)

Biljana Balen

2002-01-01

Full Text Available As plants with Crassulacean Acid Metabolism (CAM, cacti are highly affected by artificial environmental conditions in tissue culture. Plants of Mammillaria gracillis Pfeiff. (Cactaceae propagated in vitro produced callus spontaneously. This habituated callus regenerated normal and hyperhydric shoots without the addition of growth regulators. In order to compare habituated callus with the tumorous one, cactus cells were transformed with two strains of Agrobacterium tumefaciens: the wild strain B6S3 (tumour line TW and the rooty mutant GV3101 (tumour line TR. Gene expression in cactus plants, habituated callus, regenerated shoots and two tumour lines was analysed at the level of cellular and extracellular protein and glycoprotein profiles. Proteins were separated by SDS-polyacrylamide gel electrophoresis and 2-D PAGE electrophoresis and silver stained. Concavalin A-peroxidase staining detected glycoproteins with D-manose in their glycan component on protein blots. Developmentally specific protein patterns of Mammillaria gracillis tissue lines were detected. The 2-D PAGE electrophoresis revealed some tissue specific protein groups. The cellular glycoprotein of 42 kDa detected by ConA was highly expressed in undifferentiated tissues (habituated callus, TW and TR tumours and in hyperhydric regenerants. Tumours produced extracellular proteins of 33, 23 and 22 kDa. The N glycosylation of cellular and extracellular proteins was related to specific developmental stage of cactus tissue.

Sensitivity of Pigment Content of Banana and Orchid Tissue Culture Exposed to Extremely Low Frequency Electromagnetic Fiel

OpenAIRE

Prihatini, Riry; Saleh, Norihan Mohamad

2016-01-01

Natural exposure of extremely low frequency electromagnetic field (ELF-EMF) occurs in the environment and acts as one of the abiotic factors that affect the growth and development of organisms. This study was conducted to determine the effect of ELF-EMF on the tissue cultured banana and slipper orchid chlorophyll content as one of the indicators in measuring plant photosynthetic capacity. Four days old banana (Musa sp. cv. Berangan) corm and seven days old slipper orchid (Paphiopedilum rothsc...

Smallholder adoption and economic impacts of tissue culture ...

African Journals Online (AJOL)

PRECIOUS

2009-12-01

Dec 1, 2009 ... ISSN 1684–5315 © 2009 Academic Journals. Full Length ... Key words: Biotechnology, adoption, tissue culture bananas, Kenya. INTRODUCTION ... Recent studies about the agronomic and economic impacts of biotech- ..... accused scientist for 'playing God', others have supported biotechnologies.

Seasonal variations in the pattern of RNA metabolism of tuber tissue in response to excision and culture

International Nuclear Information System (INIS)

Macleod, A.J.; Mills, E.D.; Yeoman, M.M.

1979-01-01

Between December 1975 and June 1976 explants excised from Jerusalem artichoke tubers were cultured in the presence and in the absence of 2,4-D, the cells in the tissue dividing only in the presence of 2,4-D, in which the length of the first cell cycle increased nonlinearly from 18 hours to 40 hours as the tubers aged in storage at 4 0 C. Simultaneously the amount of RNA in the tissue declined linearly from 8 to 5 μg RNA per explant. Detailed examination of the RNA metabolism in dividing and in non-dividing cells during February and June 1976 revealed superimposed but independent responses to wounding during excision and to stimulation into growth by 2,4-D. The responses to wounding involved only a very low level of metabolic activity, were complete within a few hours of excision and changed very little with the storage of the tubers. Tissue treated with 2,4-D showed a much higher level of metabolic activity including the periodic accumulation of RNA coupled to its discontinuous synthesis. The features of these growth-related responses changed considerably during the investigation. (author)

Sensing in tissue bioreactors

Science.gov (United States)

Rolfe, P.

2006-03-01

Specialized sensing and measurement instruments are under development to aid the controlled culture of cells in bioreactors for the fabrication of biological tissues. Precisely defined physical and chemical conditions are needed for the correct culture of the many cell-tissue types now being studied, including chondrocytes (cartilage), vascular endothelial cells and smooth muscle cells (blood vessels), fibroblasts, hepatocytes (liver) and receptor neurones. Cell and tissue culture processes are dynamic and therefore, optimal control requires monitoring of the key process variables. Chemical and physical sensing is approached in this paper with the aim of enabling automatic optimal control, based on classical cell growth models, to be achieved. Non-invasive sensing is performed via the bioreactor wall, invasive sensing with probes placed inside the cell culture chamber and indirect monitoring using analysis within a shunt or a sampling chamber. Electroanalytical and photonics-based systems are described. Chemical sensing for gases, ions, metabolites, certain hormones and proteins, is under development. Spectroscopic analysis of the culture medium is used for measurement of glucose and for proteins that are markers of cell biosynthetic behaviour. Optical interrogation of cells and tissues is also investigated for structural analysis based on scatter.

Strategies on process engineering of chondrocyte culture for cartilage tissue regeneration.

Science.gov (United States)

Mallick, Sarada Prasanna; Rastogi, Amit; Tripathi, Satyavrat; Srivastava, Pradeep

2017-04-01

The current work is an attempt to study the strategies for cartilage tissue regeneration using porous scaffold in wavy walled airlift bioreactor (ALBR). Novel chitosan, poly (L-lactide) and hyaluronic acid based composite scaffold were prepared. The scaffolds were cross-linked with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide, N-hydroxysuccinimide and chondroitin sulfate to obtain interconnected 3D microstructure showing excellent biocompatibility, higher cellular differentiation and increased stability. The surface morphology and porosity of the scaffolds were analyzed using scanning electron microscopy (SEM) and mercury intrusion porosimeter and optimized for chondrocyte regeneration. The study shows that the scaffolds were highly porous with pore size ranging from 48 to 180 µm and the porosities in the range 80-92%. Swelling and in vitro degradation studies were performed for the composite scaffolds; by increasing the chitosan: HA ratio in the composite scaffolds, the swelling property increases and stabilizes after 24 h. There was controlled degradation of composite scaffolds for 4 weeks. The uniform chondrocyte distribution in the scaffold using various growth modes in the shake flask and ALBR was studied by glycosaminoglycans (GAG) quantification, MTT assay and mixing time evaluation. The cell culture studies demonstrated that efficient designing of ALBR increases the cartilage regeneration as compared to using a shake flask. The free chondrocyte microscopy and cell attachment were performed by inverted microscope and SEM, and from the study it was confirmed that the cells uniformly attached to the scaffold. This study focuses on optimizing strategies for the culture of chondrocyte using suitable scaffold for improved cartilage tissue regeneration.

Developing a Plant Culture Medium Composed of Vinasse Originating from Haematococcus Pluvialis Culture

International Nuclear Information System (INIS)

Gollo, A. L.; Silva, A. L. L. D.; Lima, K. K. D. D.; Camara, M. C.; Rodrigues, C.; Vandenberghe, L. P. D. S.; Soccol, V. T.; Soccol, C. R.; Biasi, L. A.

2016-01-01

The mineral nutrients in vinasse provide support for algal and plant growth. Algal culture releases organic compounds into its liquid culture medium. These organic and inorganic substances can be useful for formulating a plant tissue culture medium, because tissue culture medium is composed of organic and inorganic components. Therefore, the aims of this study were to develop a plant culture medium by using the vinasse that is employed for Haematococcus pluvialis culture (algal filtrate); to investigate the possible beneficial effects of the biocompounds in the micropropagation of Nidularium procerum (Bromeliaceae), to evaluate quercetin content, total phenolics content in vinasse and to evaluate the cytotoxicity of the media by performing a bioassay with Artemia salina. The vinasse that originated from H. pluvialis culture can be used to formulate plant tissue culture at a 3% dilution, and its mineral nutrients can support In vitro plant growth, but some nutrients must be supplemented to enhance its efficiency. An efficient micropropagation protocol was developed for N. procerum. The micropropagated plants were suitable for transfer to the field (they were acclimatized). This culture medium provides a way to reuse wastewater, gives a rational alternative to vinasse disposal and adds value to what is currently considered to be an undesirable residue. Moreover, this process can reduce the production costs of clonal seedlings and/or bioactive compounds in biofactories. There was no apparent biostimulatory effect by the algal filtrate on morphogenesis; however, it did increase quercetin production. The H. pluvialis culture that was grown in the vinasse decreased the cytotoxicity and phenolic compound contents, which prevented explant tissue necrosis and represented a treatment for this residue for safer disposal in the environment. (author)

Characterization of the Embryogenic Tissue of the Norway Spruce Including a Transition Layer between the Tissue and the Culture Medium by Magnetic Resonance Imaging

Directory of Open Access Journals (Sweden)

Kořínek R.

2017-02-01

Full Text Available The paper describes the visualization of the cells (ESEs and mucilage (ECMSN in an embryogenic tissue via magnetic resonance imaging (MRI relaxometry measurement combined with the subsequent multi-parametric segmentation. The computed relaxometry maps T1 and T2 show a thin layer (transition layer between the culture medium and the embryogenic tissue. The ESEs, mucilage, and transition layer differ in their relaxation times T1 and T2; thus, these times can be used to characterize the individual parts within the embryogenic tissue. The observed mean values of the relaxation times T1 and T2 of the ESEs, mucilage, and transition layer are as follows: 1469 ± 324 and 53 ± 10 ms, 1784 ± 124 and 74 ± 8 ms, 929 ± 164 and 32 ± 4.7 ms, respectively. The multi-parametric segmentation exploiting the T1 and T2 relaxation times as a classifier shows the distribution of the ESEs and mucilage within the embryogenic tissue. The discussed T1 and T2 indicators can be utilized to characterize both the growth-related changes in an embryogenic tissue and the effect of biotic/abiotic stresses, thus potentially becoming a distinctive indicator of the state of any examined embryogenic tissue.

Growth hormone and adipose tissue: beyond the adipocyte.

Science.gov (United States)

Berryman, Darlene E; List, Edward O; Sackmann-Sala, Lucila; Lubbers, Ellen; Munn, Rachel; Kopchick, John J

2011-06-01

The last two decades have seen resurgence in research focused on adipose tissue. In part, the enhanced interest stems from an alarming increase in obesity rates worldwide. However, an understanding that this once simple tissue is significantly more intricate and interactive than previously realized has fostered additional attention. While few would argue that growth hormone (GH) radically alters fat mass, newer findings revealing the complexity of adipose tissue requires that GH's influence on this tissue be reexamined. Therefore, the objective of this review is to describe the more recent understanding of adipose tissue and to summarize our current knowledge of how GH may influence and contribute to these newer complexities of this tissue with special focus on the available data from mice with altered GH action. Copyright © 2011 Elsevier Ltd. All rights reserved.

The Big Bang of tissue growth: Apical cell constriction turns into tissue expansion.

Science.gov (United States)

Janody, Florence

2018-03-05

How tissue growth is regulated during development and cancer is a fundamental question in biology. In this issue, Tsoumpekos et al. (2018. J. Cell Biol. https://doi.org/10.1083/jcb.201705104) and Forest et al. (2018. J. Cell Biol. https://doi.org/10.1083/jcb.201705107) identify Big bang (Bbg) as an important growth regulator of the Drosophila melanogaster wing imaginal disc. © 2018 Janody.

Immunolocalization of transforming growth factor alpha in normal human tissues

DEFF Research Database (Denmark)

Christensen, M E; Poulsen, Steen Seier

1996-01-01

anchorage-independent growth of normal cells and was, therefore, considered as an "oncogenic" growth factor. Later, its immunohistochemical presence in normal human cells as well as its biological effects in normal human tissues have been demonstrated. The aim of the present investigation was to elucidate...... the distribution of the growth factor in a broad spectrum of normal human tissues. Indirect immunoenzymatic staining methods were used. The polypeptide was detected with a polyclonal as well as a monoclonal antibody. The polyclonal and monoclonal antibodies demonstrated almost identical immunoreactivity. TGF......-alpha was found to be widely distributed in cells of normal human tissues derived from all three germ layers, most often in differentiated cells. In epithelial cells, three different kinds of staining patterns were observed, either diffuse cytoplasmic, cytoplasmic in the basal parts of the cells, or distinctly...

The establishment of an in vitro gene bank in Dianthus spiculifolius Schur and D. glacialis ssp. gelidus (Schott Nym. et Kotschy Tutin: I. The initiation of a tissue collection and the characterization of the cultures in minimal growth conditions

Directory of Open Access Journals (Sweden)

Mihaela Holobiuc

2009-12-01

Full Text Available In the last decades the plants have to cope with the warming of the climate. As a consequence of this process more than half of the plant species could become vulnerable or threatened until 2080. Romania has a high plant diversity, with endemic and endangered plant species, the measures of biodiversity conservation being necessary. The integrated approach of biodiversity conservation involves both in situ and ex situ strategies. Among ex situ methods of conservation, besides the traditional ones (including field and botanic collection and seed banks, in vitro tissues techniques offer a viable alternative. The germplasm collections can efficiently preserve the species (of economic, scientific and conservative importance, in the same time being a source of plant material for international exchanges and for reintroduction in the native habitats.The "in vitro gene banking" term refers to in vitro tissues cultures from many accessions of a target species and involves the collection of plant material from field or from native habitats, the elaboration of sterilization, micropropagation and maintaining protocols. These collections have to be maintained in optimal conditions, morphologically and genetically characterized. The aim of our work was to characterize the response of the plant material to the minimal in vitro growth protocol for medium-term cultures achievement as a prerequisite condition for an active gene bank establishment in two rare Caryophyllaceae taxa: Dianthus spiculifolius and D. glacialis ssp. gelidus. Among different factors previously tested for medium-term preservation in Dianthus genus, mannitol proved to be more efficient for minimal cultures achievement. In vitro, the cultures were evaluated concerning their growth, regenerability and enzyme activity (POX, SOD, CAT as a response to the preservation conditions in the incipient phase of the initiation of the in vitro collection. The two species considered in this study showed a

Mouse pancreas tissue slice culture facilitates long-term studies of exocrine and endocrine cell physiology in situ.

Science.gov (United States)

Marciniak, Anja; Selck, Claudia; Friedrich, Betty; Speier, Stephan

2013-01-01

Studies on pancreatic cell physiology rely on the investigation of exocrine and endocrine cells in vitro. Particularly, in the case of the exocrine tissue these studies have suffered from a reduced functional viability of acinar cells in culture. As a result not only investigations on dispersed acinar cells and isolated acini were limited in their potential, but also prolonged studies on pancreatic exocrine and endocrine cells in an intact pancreatic tissue environment were unfeasible. To overcome these limitations, we aimed to establish a pancreas tissue slice culture platform to allow long-term studies on exocrine and endocrine cells in the intact pancreatic environment. Mouse pancreas tissue slice morphology was assessed to determine optimal long-term culture settings for intact pancreatic tissue. Utilizing optimized culture conditions, cell specificity and function of exocrine acinar cells and endocrine beta cells were characterized over a culture period of 7 days. We found pancreas tissue slices cultured under optimized conditions to have intact tissue specific morphology for the entire culture period. Amylase positive intact acini were present at all time points of culture and acinar cells displayed a typical strong cell polarity. Amylase release from pancreas tissue slices decreased during culture, but maintained the characteristic bell-shaped dose-response curve to increasing caerulein concentrations and a ca. 4-fold maximal over basal release. Additionally, endocrine beta cell viability and function was well preserved until the end of the observation period. Our results show that the tissue slice culture platform provides unprecedented maintenance of pancreatic tissue specific morphology and function over a culture period for at least 4 days and in part even up to 1 week. This analytical advancement now allows mid -to long-term studies on the cell biology of pancreatic disorder pathogenesis and therapy in an intact surrounding in situ.

Mouse pancreas tissue slice culture facilitates long-term studies of exocrine and endocrine cell physiology in situ.

Directory of Open Access Journals (Sweden)

Anja Marciniak

Full Text Available Studies on pancreatic cell physiology rely on the investigation of exocrine and endocrine cells in vitro. Particularly, in the case of the exocrine tissue these studies have suffered from a reduced functional viability of acinar cells in culture. As a result not only investigations on dispersed acinar cells and isolated acini were limited in their potential, but also prolonged studies on pancreatic exocrine and endocrine cells in an intact pancreatic tissue environment were unfeasible. To overcome these limitations, we aimed to establish a pancreas tissue slice culture platform to allow long-term studies on exocrine and endocrine cells in the intact pancreatic environment. Mouse pancreas tissue slice morphology was assessed to determine optimal long-term culture settings for intact pancreatic tissue. Utilizing optimized culture conditions, cell specificity and function of exocrine acinar cells and endocrine beta cells were characterized over a culture period of 7 days. We found pancreas tissue slices cultured under optimized conditions to have intact tissue specific morphology for the entire culture period. Amylase positive intact acini were present at all time points of culture and acinar cells displayed a typical strong cell polarity. Amylase release from pancreas tissue slices decreased during culture, but maintained the characteristic bell-shaped dose-response curve to increasing caerulein concentrations and a ca. 4-fold maximal over basal release. Additionally, endocrine beta cell viability and function was well preserved until the end of the observation period. Our results show that the tissue slice culture platform provides unprecedented maintenance of pancreatic tissue specific morphology and function over a culture period for at least 4 days and in part even up to 1 week. This analytical advancement now allows mid -to long-term studies on the cell biology of pancreatic disorder pathogenesis and therapy in an intact surrounding in situ.

Cultural diversity and economic growth

DEFF Research Database (Denmark)

Ager, Philipp; Brückner, Markus

2013-01-01

We exploit the large inflow of immigrants to the US during the 1870–1920 period to examine the effects that within-county changes in the cultural composition of the US population had on output growth. We construct measures of fractionalization and polarization to distinguish between the different...

Culture of three-dimensional tissue model and its application in bystander-effect research

International Nuclear Information System (INIS)

Wu Ruqun; Xu An; Wu Lijun; Hu Burong

2012-01-01

Compared with the cultured monolayer (2D) cells, three-dimensional (3D) tissue could be more similar to the environment in vivo including the physical support, chemical factors, cell-cell and cell-matrix interaction and so on. With the development of three-dimensional cell culture techniques (TDCC), 3D tissue is widely used in the areas of bystander effect research. This review focuses on introducing the TDCC method and its application in bystander-effect research. First, the development process of 3D tissue culture method was introduced. Secondly, the induction of radiation induced bystander effects both in 2D cell and 3D tissue and its mechanisms were reviewed. Finally, because heavy ion (carbon ion beam) has been developed as a useful tool to cure solid cancer, and the 3D tissue model is an ideal material to study the damages on body after being irradiated and to understand the underlying mechanisms, future study about heavy ion radiation inducing bystander effect in 3D tissue was discussed. (authors)

Standard Operating Procedure (SOP) for Rapid and Efficient Production of Stevia Tissue Culture Seedlings

International Nuclear Information System (INIS)

Norazlina Noordin; Peng, C.S.; Rusli Ibrahim

2015-01-01

Stevia rebaudiana Bertoni is a non-caloric natural sweetener which is 300 times sweeter than cane sugar. Extracts from stevia leaves has vast application in food and beverages based industries, can be added to tea and coffee, cooked or baked goods, processed foods and confectionary goods. Recently, stevia attained awareness owing to its natural, non-caloric sweetness by diet/ health conscious and diabetic persons (Arpita et al., 2011). This natural sweetener has high commercial value in global market, it was estimated that global market value for stevia is be around USD11 billion by year 2015. Although stevia is being largely popularized in Malaysia and other countries but large-scale propagation procedures for the continuous supply of planting materials in commercial plantation has yet to be established, optimized and standardized. Furthermore, propagation through stevia seeds is often very difficult due to self-incompatibility which results in sterile seeds (Sakaguchi et al., 1982). Tissue culture is the only rapid process for the mass propagation of stevia and there have been few reports of in vitro growth of stevia (Miyagaya et al., 1986) and in vitro micropropagation from shoot tip and leaf (Uddin et al., 2006). Hence, study was carried out to establish a suitable protocol for in vitro propagation of S. rebaudiana Bertoni that can be further up-scaled for mass propagation of stevia seedlings. The established Standard Operating Procedure (SOP) will ensure rapid and efficient production of stevia tissue culture seedlings for continuous supply of planting materials for commercial stevia plantations in Malaysia. Preparation of growth medium, multiplication of shoots, rooting of plant lets and hardening of ex-vitro rooted plant lets is discussed in this paper. (author)

Mass spectrometric characterization of elements and molecules in cell cultures and tissues

International Nuclear Information System (INIS)

Arlinghaus, H.F.; Kriegeskotte, C.; Fartmann, M.; Wittig, A.; Sauerwein, W.; Lipinsky, D.

2006-01-01

Time-of-flight secondary ion mass spectrometry (ToF-SIMS) and laser post-ionization secondary neutral mass spectrometry (laser-SNMS) have been used to image and quantify targeted compounds, intrinsic elements and molecules with subcellular resolution in single cells of both cell cultures and tissues. Special preparation procedures for analyzing cell cultures and tissue materials were developed. Cancer cells type MeWo, incubated with boronated compounds, were sandwiched between two substrates, cryofixed, freeze-fractured and freeze-dried. Also, after injection with boronated compounds, different types of mouse tissues were extracted, prepared on a special specimen carrier and plunged with high velocity into LN 2 -cooled propane for cryofixation. After trimming, these tissue blocks were freeze-dried. The measurements of the K/Na ratio demonstrated that for both cell cultures and tissue materials the special preparation techniques used were appropriate for preserving the chemical and structural integrity of the living cell. The boron images show inter- and intracellular boron signals with different intensities. Molecular images show distinct features partly correlated with the cell structure. A comparison between laser-SNMS and ToF-SIMS showed that especially laser-SNMS is particularly well-suited for identifying specific cell structures and imaging ultratrace element concentrations in tissues

Adipose tissue-derived mesenchymal stem cell yield and growth characteristics are affected by the tissue-harvesting procedure

NARCIS (Netherlands)

Oedayrajsingh-Varma, M. J.; van Ham, S. M.; Knippenberg, M.; Helder, M. N.; Klein-Nulend, J.; Schouten, T. E.; Ritt, M. J. P. F.; van Milligen, F. J.

2006-01-01

Adipose tissue contains a stromal vascular fraction that can be easily isolated and provides a rich source of adipose tissue-derived mesenchymal stem cells (ASC). These ASC are a potential source of cells for tissue engineering. We studied whether the yield and growth characteristics of ASC were

Identification of Stevioside Using Tissue Culture-Derived Stevia (Stevia rebaudiana) Leaves

Science.gov (United States)

Karim, Md. Ziaul; Uesugi, Daisuke; Nakayama, Noriyuki; Hossain, M. Monzur; Ishihara, Kohji; Hamada, Hiroki

2015-01-01

Stevioside is a natural sweetener from Stevia leaf, which is 300 times sweeter than sugar. It helps to reduce blood sugar levels dramatically and thus can be of benefit to diabetic people. Tissue culture is a very potential modern technology that can be used in large-scale disease-free stevia production throughout the year. We successfully produced stevia plant through in vitro culture for identification of stevioside in this experiment. The present study describes a potential method for identification of stevioside from tissue culture-derived stevia leaf. Stevioside in the sample was identified using HPLC by measuring the retention time. The percentage of stevioside content in the leaf samples was found to be 9.6%. This identification method can be used for commercial production and industrialization of stevia through in vitro culture across the world. PMID:28008268

In vitro cholesteatoma growth and secretion of cytokines.

Science.gov (United States)

Helgaland, Tore; Engelen, Bart; Olsnes, Carla; Aarstad, Hans Jørgen; Vassbotn, Flemming S

2010-07-01

Our results show a significant difference between skin and cholesteatoma biology in vitro. Cholesteatoma disease is a process of destruction characterized by uncontrolled growth of squamous epithelial cells in the middle ear or temporal bone. The pathophysiology behind the cholesteatoma development is controversial, and the mechanisms driving the cholesteatoma growth, migration and destructive properties is still unclear. We aimed to provide a method to study the effect of various compounds on cholesteatoma and skin tissue growth, as well as to further investigate the biological differences between normal skin and cholesteatoma tissue. We have established a method to study cholesteatoma biopsy tissue in vitro. Cholesteatoma tissues from patients undergoing surgery for chronic otitis were grown in culture medium and compared to growth patterns and behaviour of normal retroauricular skin. Conditioned medium was analysed for various secreted cytokines. We found a radial outgrowth of keratinocyte epithelium from the circular biopsies. After 5 days of culture we found a significant growth of both cholesteatoma and skin-derived cells. Cholesteatoma samples showed higher growth rate as compared with skin control cultures from the same patient. Moreover, the cholesteatoma cells showed higher production of monocyte chemoattractant protein-1 (MCP-1) and interleukin (IL)-6 as compared with normal skin.

Insulin-like growth factor I (IGF-1) Ec/Mechano Growth factor--a splice variant of IGF-1 within the growth plate.

Science.gov (United States)

Schlegel, Werner; Raimann, Adalbert; Halbauer, Daniel; Scharmer, Daniela; Sagmeister, Susanne; Wessner, Barbara; Helmreich, Magdalena; Haeusler, Gabriele; Egerbacher, Monika

2013-01-01

Human insulin-like growth factor 1 Ec (IGF-1Ec), also called mechano growth factor (MGF), is a splice variant of insulin-like growth factor 1 (IGF-1), which has been shown in vitro as well as in vivo to induce growth and hypertrophy in mechanically stimulated or damaged muscle. Growth, hypertrophy and responses to mechanical stimulation are important reactions of cartilaginous tissues, especially those in growth plates. Therefore, we wanted to ascertain if MGF is expressed in growth plate cartilage and if it influences proliferation of chondrocytes, as it does in musculoskeletal tissues. MGF expression was analyzed in growth plate and control tissue samples from piglets aged 3 to 6 weeks. Furthermore, growth plate chondrocyte cell culture was used to evaluate the effects of the MGF peptide on proliferation. We showed that MGF is expressed in considerable amounts in the tissues evaluated. We found the MGF peptide to be primarily located in the cytoplasm, and in some instances, it was also found in the nucleus of the cells. Addition of MGF peptides was not associated with growth plate chondrocyte proliferation.

Detachably assembled microfluidic device for perfusion culture and post-culture analysis of a spheroid array.

Science.gov (United States)

Sakai, Yusuke; Hattori, Koji; Yanagawa, Fumiki; Sugiura, Shinji; Kanamori, Toshiyuki; Nakazawa, Kohji

2014-07-01

Microfluidic devices permit perfusion culture of three-dimensional (3D) tissue, mimicking the flow of blood in vascularized 3D tissue in our body. Here, we report a microfluidic device composed of a two-part microfluidic chamber chip and multi-microwell array chip able to be disassembled at the culture endpoint. Within the microfluidic chamber, an array of 3D tissue aggregates (spheroids) can be formed and cultured under perfusion. Subsequently, detailed post-culture analysis of the spheroids collected from the disassembled device can be performed. This device facilitates uniform spheroid formation, growth analysis in a high-throughput format, controlled proliferation via perfusion flow rate, and post-culture analysis of spheroids. We used the device to culture spheroids of human hepatocellular carcinoma (HepG2) cells under two controlled perfusion flow rates. HepG2 spheroids exhibited greater cell growth at higher perfusion flow rates than at lower perfusion flow rates, and exhibited different metabolic activity and mRNA and protein expression under the different flow rate conditions. These results show the potential of perfusion culture to precisely control the culture environment in microfluidic devices. The construction of spheroid array chambers allows multiple culture conditions to be tested simultaneously, with potential applications in toxicity and drug screening. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Banana Musa tissue culture plants enhanced by endophytic fungi

African Journals Online (AJOL)

Mo

Merging biotechnology with biological control: Banana Musa tissue culture plants enhanced by endophytic .... While working in the laminar flow cabinet, sterile filter papers were placed in ..... University of Bonn, Bonn, Germany. Niere, B., 2001.

Research progress in plant mutation by combining ion beam irradiations and tissue culture

International Nuclear Information System (INIS)

Zhou Linbin; Li Wenjian; Qu Ying; Li Ping

2007-01-01

About a new mutation breeding method which combines plant tissue culture technique with heavy ion beam irradiations were discussed in this paper with the principles, operation steps, molecular mechanisms, etc. The mutation method developed a few advantages coming from plant tissue culture, which can produce offspring by asexual ways. Meanwhile, using this method, the study of biological effects of high energy particles with different linear energy transfer values on plant tissues or cells can be explored and optimized in theory or practice. (authors)

HYPOLIPIDEMIC EFFECT OF ARGLABIN IN HEPATOMA TISSUE CULTURE

Directory of Open Access Journals (Sweden)

A. V. Ratkin

2015-01-01

Full Text Available Objective. Investigation of hypolipidemic effect of sesquiterpene γ-lactone Arglabin in hepatoma tissue culture (HTC.Materials and methods. In this study we’ve evaluated the effect of sesquiterpene γ-lactone Arglabin and gemfibrozil (reference drug on the lipid content in the hepatoma tissue culture (HTC which were incubated with a fat emulsion “Lipofundin” by fluorescent method with vital dye Nile Red. The cell viability was investigated using the MTT-test and staining by Trypan blue.Results. Cultivation of cell cultures of rat’s hepatoma cell line HTC with Arglabin and gemfibrozil in concentrations from 10 to 50 μmol and from 0.25 to 0.5 mmol, respectively, had no cytotoxic effect. HTC cell viability did not change compared with the corresponding rate in the control culture. Experimental hyperlipidemia in hepatoma culture was induced by the addition in the incubation medium of fat emulsion “Lipofundin” in a final concentration of 0.05 %. The fluorescence intensity of Nile Red in the cells was increased 4-fold (p < 0.05, which indicates a significant accumulation of lipids in the cytosol of cells. In these steady-state Arglabin and gemfibrozil at concentrations 75–100 μM and 0.25–1.0 mM, respectively, reduced the content of lipid in cells. Conclusion. In the model of hyperlipidemia induced by lipofundin, sesquiterpene γ-lactone Arglabin prevents the accumulation of lipids in the HTC cell line, as evidenced by a decrease in Nile Red fluorescence. However hypolipidemic effect of Arglabin is associated with cytotoxic effects, which is typical for anticancer drugs.

Proteolytic processing of connective tissue growth factor in normal ocular tissues and during corneal wound healing.

Science.gov (United States)

Robinson, Paulette M; Smith, Tyler S; Patel, Dilan; Dave, Meera; Lewin, Alfred S; Pi, Liya; Scott, Edward W; Tuli, Sonal S; Schultz, Gregory S

2012-12-13

Connective tissue growth factor (CTGF) is a fibrogenic cytokine that is up-regulated by TGF-β and mediates most key fibrotic actions of TGF-β, including stimulation of synthesis of extracellular matrix and differentiation of fibroblasts into myofibroblasts. This study addresses the role of proteolytic processing of CTGF in human corneal fibroblasts (HCF) stimulated with TGF-β, normal ocular tissues and wounded corneas. Proteolytic processing of CTGF in HCF cultures, normal animal eyes, and excimer laser wounded rat corneas were examined by Western blot. The identity of a 21-kDa band was determined by tandem mass spectrometry, and possible alternative splice variants of CTGF were assessed by 5' Rapid Amplification of cDNA Ends (RACE). HCF stimulated by TGF-β contained full length 38-kDa CTGF and fragments of 25, 21, 18, and 13 kDa, while conditioned medium contained full length 38- and a 21-kDa fragment of CTGF that contained the middle "hinge" region of CTGF. Fragmentation of recombinant CTGF incubated in HCF extracts was blocked by the aspartate protease inhibitor, pepstatin. Normal mouse, rat, and rabbit whole eyes and rabbit ocular tissues contained abundant amounts of C-terminal 25- and 21-kDa fragments and trace amounts of 38-kDa CTGF, although no alternative transcripts were detected. All forms of CTGF (38, 25, and 21 kDa) were detected during healing of excimer ablated rat corneas, peaking on day 11. Proteolytic processing of 38-kDa CTGF occurs during corneal wound healing, which may have important implications in regulation of corneal scar formation.

HTCC-Modified Nanoclay for Tissue Engineering Applications: A Synergistic Cell Growth and Antibacterial Efficiency

Directory of Open Access Journals (Sweden)

Majid Aliabadi

2013-01-01

Full Text Available This paper deals with the synthesis of a biocompatible chitosan ammonium salt N-(2-hydroxy propyl-3-trimethylammonium chitosan chloride (HTCC and using it in montmorillonite ion-exchange process. HTCC-modified montmorillonite (Mt with different chemical ratios was successfully synthesized, and their characteristics have been verified by XRD and FTIR analyses. Produced samples have been evaluated in terms of antibacterial efficiency and biocompatibility (cell culture test. Antibacterial efficiency of synthesized HTCC/Mt samples has been confirmed against both gram negative bacteria (Escherichia coli and gram positive bacteria (Staphylococcus aureus. The results disclosed that the antibacterial efficiency of HTCC-modified montmorillonite was unexpectedly even more than HTCC. This excellent synergistic effect has been referred to entrapping bacteria between the intercalated structures of HTCC-modified montmorillonite. Then HTCC on clay layers can seriously attack and damage the entrapped bacteria. An extraordinary biocompatibility, cell attachment, and cell growth even more than tissue culture polystyrene (TCPS have been recorded in the case of this novel kind of modified clay. Due to existing concerns about serious and chronic infections after implant placement, this natural-based bioactive and antibacterial modified clay can be used in electrospun nanofibers and other polymeric implants with promising mechanical properties for tissue engineering applications.

Mixed-culture transcriptome analysis reveals the molecular basis of mixed-culture growth in Streptococcus thermophilus and Lactobacillus bulgaricus.

Science.gov (United States)

Sieuwerts, Sander; Molenaar, Douwe; van Hijum, Sacha A F T; Beerthuyzen, Marke; Stevens, Marc J A; Janssen, Patrick W M; Ingham, Colin J; de Bok, Frank A M; de Vos, Willem M; van Hylckama Vlieg, Johan E T

2010-12-01

Many food fermentations are performed using mixed cultures of lactic acid bacteria. Interactions between strains are of key importance for the performance of these fermentations. Yogurt fermentation by Streptococcus thermophilus and Lactobacillus bulgaricus (basonym, Lactobacillus delbrueckii subsp. bulgaricus) is one of the best-described mixed-culture fermentations. These species are believed to stimulate each other's growth by the exchange of metabolites such as folic acid and carbon dioxide. Recently, postgenomic studies revealed that an upregulation of biosynthesis pathways for nucleotides and sulfur-containing amino acids is part of the global physiological response to mixed-culture growth in S. thermophilus, but an in-depth molecular analysis of mixed-culture growth of both strains remains to be established. We report here the application of mixed-culture transcriptome profiling and a systematic analysis of the effect of interaction-related compounds on growth, which allowed us to unravel the molecular responses associated with batch mixed-culture growth in milk of S. thermophilus CNRZ1066 and L. bulgaricus ATCC BAA-365. The results indicate that interactions between these bacteria are primarily related to purine, amino acid, and long-chain fatty acid metabolism. The results support a model in which formic acid, folic acid, and fatty acids are provided by S. thermophilus. Proteolysis by L. bulgaricus supplies both strains with amino acids but is insufficient to meet the biosynthetic demands for sulfur and branched-chain amino acids, as becomes clear from the upregulation of genes associated with these amino acids in mixed culture. Moreover, genes involved in iron uptake in S. thermophilus are affected by mixed-culture growth, and genes coding for exopolysaccharide production were upregulated in both organisms in mixed culture compared to monocultures. The confirmation of previously identified responses in S. thermophilus using a different strain combination

Effect of Different Concentrations of Growth Regulators on Gardenia jasminoides cv. Veitchii Micropropagation by Tissue Culture Technique

Directory of Open Access Journals (Sweden)

G. R. Abdullah

2003-01-01

Full Text Available Micropropagation techniques were set up for Gardenia jasminoides c.v. veitchi. Many plantlets were obtained by culturing shoot cuttings in MS nutrient media, 30 g/L Sucrose, 7 g/L Agar Agar, and different concentrations of BAP and IAA. The best concentration was 1mg /L BAP with 0.5 mg/L IAA. This concentration gave the best sprout growth suitable for rooting in primary and secondary culture by reculturing the stuck cutting every 6 weeks and for many times. We also obtained a high rooting percentage up to 98 % of natural rooting in rooting media different from propagation media by reducing mineral salt concentration to half, Sucrose to 20gm/L, and 2gm/L active charcoal, and 1mg/L IAA. Plantlets were transferred to greenhous and subjected for hardening. This technique gave 22 plantlets from one cutting in one year.

Nanotechnology and picotechnology to increase tissue growth: a summary of in vivo studies

Directory of Open Access Journals (Sweden)

Alpaslan E

2014-05-01

Full Text Available Ece Alpaslan,1 Thomas J Webster1,21Department of Chemical Engineering, College of Engineering, Northeastern University, Boston, MA, USA; 2Center of Excellence for Advanced Materials Research, King Abdulaziz University, Jeddah, Saudi ArabiaAbstract: The aim of tissue engineering is to develop functional substitutes for damaged tissues or malfunctioning organs. Since only nanomaterials can mimic the surface properties (ie, roughness of natural tissues and have tunable properties (such as mechanical, magnetic, electrical, optical, and other properties, they are good candidates for increasing tissue growth, minimizing inflammation, and inhibiting infection. Recently, the use of nanomaterials in various tissue engineering applications has demonstrated improved tissue growth compared to what has been achieved until today with our conventional micron structured materials. This short report paper will summarize some of the more relevant advancements nanomaterials have made in regenerative medicine, specifically improving bone and bladder tissue growth. Moreover, this short report paper will also address the continued potential risks and toxicity concerns, which need to be accurately addressed by the use of nanomaterials. Lastly, this paper will emphasize a new field, picotechnology, in which researchers are altering electron distributions around atoms to promote surface energy to achieve similar increased tissue growth, decreased inflammation, and inhibited infection without potential nanomaterial toxicity concerns.Keywords: nanomaterials, tissue engineering, toxicity

[Comparative study on alkaloids of tissue-culture seedling and wild plant of Dendrobium huoshanense ].

Science.gov (United States)

Chen, Nai-dong; Gao, Feng; Lin, Xin; Jin, Hui

2014-06-01

To compare the composition and content of alkaloid of Dendrobium huoshanense tissue-culture seedling and wild plant. A comparative evaluation on the quality was carried out by HPLC and TLC methods including the composition and the content of alkaloids. Remarkable variation existed in the two kinds of Dendrobium huoshanense. For the tissue-culture plant, only two alkaloids were checked out by both HPLC and TLC while four alkaloids were observed in the wild plant. The alkaloid content of tissue-culture seedling and wild plant was(0. 29 ± 0. 11)%o and(0. 43 ± 0. 15) %o,respectively. Distinguished difference is observed in both composition and content of alkaloids from the annual shoots of different provenances of Dendrobium huoshanense. It suggested that the quality of tissue-culture seedling of Dendrobium huoshanense might be inconsistent with the wild plant. Furthermore, the established alkaloids-knock-out HPLC method would provide a new research tool on quality control of Chinese medicinal materials which contain unknown alkaloids.

Modeling biological tissue growth: discrete to continuum representations.

Science.gov (United States)

Hywood, Jack D; Hackett-Jones, Emily J; Landman, Kerry A

2013-09-01

There is much interest in building deterministic continuum models from discrete agent-based models governed by local stochastic rules where an agent represents a biological cell. In developmental biology, cells are able to move and undergo cell division on and within growing tissues. A growing tissue is itself made up of cells which undergo cell division, thereby providing a significant transport mechanism for other cells within it. We develop a discrete agent-based model where domain agents represent tissue cells. Each agent has the ability to undergo a proliferation event whereby an additional domain agent is incorporated into the lattice. If a probability distribution describes the waiting times between proliferation events for an individual agent, then the total length of the domain is a random variable. The average behavior of these stochastically proliferating agents defining the growing lattice is determined in terms of a Fokker-Planck equation, with an advection and diffusion term. The diffusion term differs from the one obtained Landman and Binder [J. Theor. Biol. 259, 541 (2009)] when the rate of growth of the domain is specified, but the choice of agents is random. This discrepancy is reconciled by determining a discrete-time master equation for this process and an associated asymmetric nonexclusion random walk, together with consideration of synchronous and asynchronous updating schemes. All theoretical results are confirmed with numerical simulations. This study furthers our understanding of the relationship between agent-based rules, their implementation, and their associated partial differential equations. Since tissue growth is a significant cellular transport mechanism during embryonic growth, it is important to use the correct partial differential equation description when combining with other cellular functions.

A multiscale description of growth and transport in biological tissues

Directory of Open Access Journals (Sweden)

Grillo A.

2007-01-01

Full Text Available We study a growing biological tissue as an open biphasic mixture with mass exchange between phases. The solid phase is identified with the matrix of a porous medium, while the fluid phase is comprised of water, together with all the dissolved chemical substances coexisting in the pore space. We assume that chemical substances evolve according to transport mechanisms determined by kinematical and constitutive relations, and we propose to consider growth as a process able to influence transport by continuously varying the thermo-mechanic state of the tissue. By focusing on the case of anisotropic growth, we show that such an influence occurs through a continuous rearrangement of the tissue material symmetries. In order to illustrate this interaction, we restrict ourselves to diffusion-dominated transport, and we assume that the time-scales associated with growth and the transport process of interest are largely separated. This allows for performing an asymptotic analysis of the "field equations" of the system. In this framework, we provide a formal solution of the transport equation in terms of its associated Green's function, and we show how the macroscopic concentration of a given chemical substance is "modulated" by anisotropic growth. .

Plasmid-dependent attachment of Agrobacterium tumefaciens to plant tissue culture cells.

Science.gov (United States)

Matthysse, A G; Wyman, P M; Holmes, K V

1978-11-01

Kinetic, microscopic, and biochemical studies show that virulent Ti (tumor inducing)-plasmid-containing strains of Agrobacterium attach to normal tobacco and carrot tissue culture cells. Kinetic studies showed that virulent strains of A. tumefaciens attach to the plant tissue culture cells in increasing numbers during the first 1 to 2 h of incubation of the bacteria with the plant cells. Five Ti-plasmid-containing virulent Agrobacterium strains showed greater attachment to tobacco cells than did five avirulent strains. Light and scanning electron microscopic observations confirmed that virulent strains showed little attachment. Bacterial attachment was blocked by prior incubation of the plant cells with lipopolysaccharide extracted from A. tumefaciens, but not from A. radiobacter, suggesting that bacterial lipopolysaccharide is one of the components involved in the attachment process. At least one other bacterial product may be required for attachment in tissue culture because the virulent A. tumefaciens NT1, which lacks the Ti plasmid, does not itself attach to tobacco cells, but its lipopolysaccharide does inhibit the attachment of virulent strains.

Enhancement of erythroid colony growth in culture by hemin

International Nuclear Information System (INIS)

Porter, P.N.; Meints, R.H.; Mesner, K.

1979-01-01

Hemin was found to enhance the growth of murine erythroid colonies in culture. In the presence of 100 mU/ml erythropoietin (EPO), the addition of hemin (0.05-0.2 mM) resulted in the growth of twice as many colonies as were obtained with EPO alone. Hemin also significantly increased erythroid colony formation in culture in the absence of added EPO. Hemoblobin synthesis as measured by the incorporation of 59 Fe into cyclohexanone extractable heme was augmented in culture by hemin. Neither Δ-aminolevulinic acid, a hemin precursor, nor FeCl 3 increased colony number. (author)

The Effect of Plant Growth Regulators and Different Explants on the Response of Tissue Culture and Cell Suspension Cultures of German Chamomile (Matricaria chamomilla L.

Directory of Open Access Journals (Sweden)

L. Koohi,

2014-07-01

Full Text Available German chamomile (Matricaria chamomilla L. is one of the most important medicinal plants that its essential oils used in different medicinal industries. In this study which was carried out in 2013 growing season at the Faculty of Agricultural Sciences of the University of Mohaghegh Ardabili, the in vitro response of leaf and hypocotyl explants of German Chamomile in B5 medium supplemented with different levels of plant growth regulators including 2,4-D, naphthalene acetic acid (NAA, kinetin and 6-benzylaminopurine (BAP were investigated in a factorial experiment based on completely randomized design (CRD.In addition, cell suspension cultures were established and characterized. Hypocotyl and leaf explants exhibited cell proliferation and produced callus within 1-2 weeks. The highest fresh weight of the callus (264.1 mg was produced by leaf explants in the medium supplemented with 0.5 mg/l 2,4-D and 1 mg/l BAP. However, the leaf explants cultured on medium containing 1.5 mg/l 2,4-D showed the lowest cell proliferation and callus yield (40.42 mg. The highest percentage of root induction from leaf explants (58.73% was observed on the medium containing 4 mg/l 2,4-D and 1 mg/l Kin, and from hypocotyl explants (48.61% was observed on medium supplemented with 1.5 mg/l NAA. The 42.22% of calli derived from hypocotyl explants on B5 medium supplemented with 4 mg/l NAA and 3 mg/l BAP, were friable. Cell suspension cultures of German chamomile were established by transferring of hypocotyl-derived friable calli into the MS medium supplemented with 1.5 mg/l 2,4-D and 1 mg/l kinetin. The growth curve of cell proliferations started 4 days after culture and continued to grow until day 13th, where the cells entered stationary phase.

Growth Inhibition of Osteosarcoma Cell Lines in 3D Cultures: Role of Nitrosative and Oxidative Stress.

Science.gov (United States)

Gorska, Magdalena; Krzywiec, Pawel Bieniasz; Kuban-Jankowska, Alicja; Zmijewski, Michal; Wozniak, Michal; Wierzbicka, Justyna; Piotrowska, Anna; Siwicka, Karolina

2016-01-01

3D cell cultures have revolutionized the understanding of cell behavior, allowing culture of cells with the possibility of resembling in vivo intercellular signaling and cell-extracellular matrix interaction. The effect of limited oxygen penetration into 3D culture of highly metastatic osteosarcoma 143B cells in terms of expression of nitro-oxidative stress markers was investigated and compared to standard 2D cell culture. Human osteosarcoma (143B cell line) cells were cultured as monolayers, in collagen and Matrigel. Cell viability, gene expression of nitro-oxidative stress markers, and vascular endothelial growth factor were determined using Trypan blue assay, quantitative polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Three-dimensional environments modify nitro-oxidative stress and influence gene expression and cell proliferation of OS 143B cells. Commercial cell lines might not constitute a good model of 3D cultures for bone tissue engineering, as they are highly sensitive to hypoxia, and hypoxic conditions can induce oxidation of the cellular environment. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Correlated responses in tissue weights measured in vivo by computer tomography in Dorset Down sheep selected for lean tissue growth

International Nuclear Information System (INIS)

Nsoso, S.J.; Young, M.J.; Beatson, P.R.

2003-01-01

The aim of this study was to estimate correlated responses in lean, fat and bone weights in vivo in Dorset Down sheep selected for lean tissue growth. Over the period 1986-1992 inclusive, the lean tissue growth line had been selected using two economic indices for an increased aggregate breeding value incorporating predicted lean and fat weights with positive and negative economic weightings, respectively. The control line was selected for no change in lean tissue growth each year. Animals were born and run on pasture all year round. X-ray computer tomography was used to estimate the weights of lean, fat and bone in vivo in the 1994-born sheep, aged 265-274 days and selected randomly into 12 rams and 12 ewes from the selected line and 10 rams and 9 ewes from the control line. The lean tissue growth line had significantly greater responses in lean weight (+0.65 + 0.10 kg) and lean percentage (+1.19 + 0.17%) and significantly lesser fat weight (-0.36 + 0.08 kg) and fat percentage (-1.88 + 0.20%) compared to the control line. There was a significant increase in bone weight (+0.27 + 0.03 kg) and bone percentage (+0.69 + 0.09%) in the lean tissue growth line compared to the control line. Responses differed significantly between sexes of the lean tissue growth line, rams having a greater response in weight of lean (+1.22 + 0.20 vs. +0.08 + 0.22 kg) and bone (+0.45 + 0.06 vs. +0.09 + 0.07 kg), and a lesser response in weight of fat (-0.03 + 0.15 vs. -0.70 + 0.16 kg) than the ewes. Selection led to significant changes in lean (increase) and fat weights (decrease), and bone weight increased. Although responses in the lean tissue growth line differed significantly between sexes, there were confounding factors due to differences in management and lack of comparison at equal stage of development. Therefore, to assess real genetic differences further studies should be conducted taking these factors into consideration

Chemical Growth Regulators for Guayule Plants

Science.gov (United States)

Dastoor, M. N.; Schubert, W. W.; Petersen, G. R.

1982-01-01

Test Tubes containing Guayule - tissue cultures were used in experiments to test effects of chemical-growth regulators. The shoots grew in response to addition of 2-(3,4-dichlorophenoxy)-triethylamine (triethylamine (TEA) derivative) to agar medium. Preliminary results indicate that a class of compounds that promotes growth in soil may also promote growth in a culture medium. Further experiments are needed to define the effect of the TEA derivative.

Identification of Stevioside Using Tissue Culture-Derived Stevia () Leaves

OpenAIRE

Ziaul Karim Md.; Daisuke Uesugi; Noriyuki Nakayama; M. Monzur Hossain; Kohji Ishihara; Hiroki Hamada

2015-01-01

Stevioside is a natural sweetener from Stevia leaf, which is 300 times sweeter than sugar. It helps to reduce blood sugar levels dramatically and thus can be of benefit to diabetic people. Tissue culture is a very potential modern technology that can be used in large-scale disease-free stevia production throughout the year. We successfully produced stevia plant through in vitro culture for identification of stevioside in this experiment. The present study describes a potential method for iden...

NMR-based metabolomics of mammalian cell and tissue cultures

International Nuclear Information System (INIS)

Aranibar, Nelly; Borys, Michael; Mackin, Nancy A.; Ly, Van; Abu-Absi, Nicholas; Abu-Absi, Susan; Niemitz, Matthias; Schilling, Bernhard; Li, Zheng Jian; Brock, Barry; Russell, Reb J.; Tymiak, Adrienne; Reily, Michael D.

2011-01-01

NMR spectroscopy was used to evaluate growth media and the cellular metabolome in two systems of interest to biomedical research. The first of these was a Chinese hamster ovary cell line engineered to express a recombinant protein. Here, NMR spectroscopy and a quantum mechanical total line shape analysis were utilized to quantify 30 metabolites such as amino acids, Krebs cycle intermediates, activated sugars, cofactors, and others in both media and cell extracts. The impact of bioreactor scale and addition of anti-apoptotic agents to the media on the extracellular and intracellular metabolome indicated changes in metabolic pathways of energy utilization. These results shed light into culture parameters that can be manipulated to optimize growth and protein production. Second, metabolomic analysis was performed on the superfusion media in a common model used for drug metabolism and toxicology studies, in vitro liver slices. In this study, it is demonstrated that two of the 48 standard media components, choline and histidine are depleted at a faster rate than many other nutrients. Augmenting the starting media with extra choline and histidine improves the long-term liver slice viability as measured by higher tissues levels of lactate dehydrogenase (LDH), glutathione and ATP, as well as lower LDH levels in the media at time points out to 94 h after initiation of incubation. In both models, media components and cellular metabolites are measured over time and correlated with currently accepted endpoint measures.

NMR-based metabolomics of mammalian cell and tissue cultures

Energy Technology Data Exchange (ETDEWEB)

Aranibar, Nelly; Borys, Michael; Mackin, Nancy A.; Ly, Van; Abu-Absi, Nicholas; Abu-Absi, Susan [Bristol-Myers Squibb Company (United States); Niemitz, Matthias [PERCH Solutions Ltd. (Finland); Schilling, Bernhard; Li, Zheng Jian; Brock, Barry; Russell, Reb J.; Tymiak, Adrienne; Reily, Michael D., E-mail: michael.reily@bms.com [Bristol-Myers Squibb Company (United States)

2011-04-15

NMR spectroscopy was used to evaluate growth media and the cellular metabolome in two systems of interest to biomedical research. The first of these was a Chinese hamster ovary cell line engineered to express a recombinant protein. Here, NMR spectroscopy and a quantum mechanical total line shape analysis were utilized to quantify 30 metabolites such as amino acids, Krebs cycle intermediates, activated sugars, cofactors, and others in both media and cell extracts. The impact of bioreactor scale and addition of anti-apoptotic agents to the media on the extracellular and intracellular metabolome indicated changes in metabolic pathways of energy utilization. These results shed light into culture parameters that can be manipulated to optimize growth and protein production. Second, metabolomic analysis was performed on the superfusion media in a common model used for drug metabolism and toxicology studies, in vitro liver slices. In this study, it is demonstrated that two of the 48 standard media components, choline and histidine are depleted at a faster rate than many other nutrients. Augmenting the starting media with extra choline and histidine improves the long-term liver slice viability as measured by higher tissues levels of lactate dehydrogenase (LDH), glutathione and ATP, as well as lower LDH levels in the media at time points out to 94 h after initiation of incubation. In both models, media components and cellular metabolites are measured over time and correlated with currently accepted endpoint measures.

Migration assay on primary culture isolated from patient's primary breast cancer tissue

Directory of Open Access Journals (Sweden)

ED Yuliana

2014-12-01

Full Text Available Background: Migration is an essential component of breast cancer metastasis, which studyhas been concentrated on culture of established breast cancer cell lines that do not accuratelyrepresent the sophistication and heterogeneity of patient's breast cancer. An attempt toperform migration assay using Boyden Chamber Assay (BCA on primary culture originatingfrom patient's breast cancer tissue was developed to accommodate upcoming study of breastcancer migration in lndonesian patients.Methods: Pathologically proven primary breast cancer tissue samples were obtained fromCiptomangunkusumo Hospital during core (n=4 and incisional (n=3 biopsies of stage llAup to stage lllA breast cancer patients. Following biopsy, the breast cancer tissue samplesunderwent processings to isolate the cancer cells. These cancer cells were -then resuspendedwithin Dulbecco's modified Eagle's medium (DMEM ahd cultured in 12-well plate. The growthof primary culture were observed and compared between the core biopsy and the incisionalbiopsy specimens. Optimization of BCA method was later performed to investigate themigration of the breast cancer primary culture towards different experirnental conditions, whichwere control, Fetal Bovine Serum (FBS, and Stromal Derived Factor-l (SDF-1. Two differentnumber of breast cancer cells were tested for the optimization of the BCA, which were 1 x 105and3x105cells.Results: None of the culture performed on core biopsy specimens grew, while one out ofthree incisional biopsy specimens grew until confluence. The one primary culture that grewwas later assesed using BCA to assess its migration index towards different experimentalconditions. Using 1 x 10s breast cancer cells in the BCA , the result of the absorbance level ofmigrated cells showed that the migration towards SDF-1 (0.529 nearly doubled the migrationtowards controlmedium (0.239 and FBS (0.209. Meanwhile, the absorbance levelwas simiiarbetween the control medium (1.050, FBS (1 .103

Immobilization and Application of Electrospun Nanofiber Scaffold-based Growth Factor in Bone Tissue Engineering.

Science.gov (United States)

Chen, Guobao; Lv, Yonggang

2015-01-01

Electrospun nanofibers have been extensively used in growth factor delivery and regenerative medicine due to many advantages including large surface area to volume ratio, high porosity, excellent loading capacity, ease of access and cost effectiveness. Their relatively large surface area is helpful for cell adhesion and growth factor loading, while storage and release of growth factor are essential to guide cellular behaviors and tissue formation and organization. In bone tissue engineering, growth factors are expected to transmit signals that stimulate cellular proliferation, migration, differentiation, metabolism, apoptosis and extracellular matrix (ECM) deposition. Bolus administration is not always an effective method for the delivery of growth factors because of their rapid diffusion from the target site and quick deactivation. Therefore, the integration of controlled release strategy within electrospun nanofibers can provide protection for growth factors against in vivo degradation, and can manipulate desired signal at an effective level with extended duration in local microenvironment to support tissue regeneration and repair which normally takes a much longer time. In this review, we provide an overview of growth factor delivery using biomimetic electrospun nanofiber scaffolds in bone tissue engineering. It begins with a brief introduction of different kinds of polymers that were used in electrospinning and their applications in bone tissue engineering. The review further focuses on the nanofiber-based growth factor delivery and summarizes the strategies of growth factors loading on the nanofiber scaffolds for bone tissue engineering applications. The perspectives on future challenges in this area are also pointed out.

Effect of sucrose concentration and gamma irradiation on growth and essential oil composition of spearmint plant through tissue culture

International Nuclear Information System (INIS)

El-Sharnouby, M.E.

2007-01-01

In vitro culture of spearmint plant (Mentha spicata L) using different sucrose concentrations and different gamma irradiation treatments was investigated. The shoot tips of spearmint plant were cultured on MS medium without hormones and supplemented with different concentrations of sucrose (10, 20, 30 and 40 g/l) then exposed to different gamma irradiation treatments (2,4,6 and 8 Krad) to determine their effects on growth and chemical composition in different sub-culturing media . The data showed that culturing shoots of Mentha spicata on MS medium containing 10 g/l sucrose produced the highest values of callus than other treatments and the maximum number of shoots was produced on MS medium supplemented with 20 g/l sucrose. Irradiation of spearmint shoots at 8 Krad when cultured on MS medium containing 30 or 40 g/l sucrose caused minimum number of shoots, whereas the longest shoots were produced with MS medium containing 20 g/l sucrose after irradiation at 60 Gy gamma dose. Treating shoots of Mentha spicata by gamma irradiation at 8 Krad and culturing on MS medium containing 30 g/l sucrose produced all sub-cultures in shortest length of shoots. Moreover, adding 40 g/l of sucrose in MS medium gave the highest number of leaves than other treatments. Exposing shoots of spearmint plant to gamma irradiation at 8 Krad decreased the number of leaves when culturing on MS medium containing 10 or 30 g/l sucrose. Furthermore, the selected samples showed many differences on spearmint oil composition and proline content regarding sucrose levels and gamma irradiation doses

Ligament Tissue Engineering Using a Novel Porous Polycaprolactone Fumarate Scaffold and Adipose Tissue-Derived Mesenchymal Stem Cells Grown in Platelet Lysate.

Science.gov (United States)

Wagner, Eric R; Bravo, Dalibel; Dadsetan, Mahrokh; Riester, Scott M; Chase, Steven; Westendorf, Jennifer J; Dietz, Allan B; van Wijnen, Andre J; Yaszemski, Michael J; Kakar, Sanjeev

2015-11-01

Surgical reconstruction of intra-articular ligament injuries is hampered by the poor regenerative potential of the tissue. We hypothesized that a novel composite polymer "neoligament" seeded with progenitor cells and growth factors would be effective in regenerating native ligamentous tissue. We synthesized a fumarate-derivative of polycaprolactone fumarate (PCLF) to create macro-porous scaffolds to allow cell-cell communication and nutrient flow. Clinical grade human adipose tissue-derived human mesenchymal stem cells (AMSCs) were cultured in 5% human platelet lysate (PL) and seeded on scaffolds using a dynamic bioreactor. Cell growth, viability, and differentiation were examined using metabolic assays and immunostaining for ligament-related markers (e.g., glycosaminoglycans [GAGs], alkaline phosphatase [ALP], collagens, and tenascin-C). AMSCs seeded on three-dimensional (3D) PCLF scaffolds remain viable for at least 2 weeks with proliferating cells filling the pores. AMSC proliferation rates increased in PL compared to fetal bovine serum (FBS) (p ligament and tenogenic growth factor fibroblast growth factor 2 (FGF-2), especially when cultured in the presence of PL (p engineering and ligament regeneration.

Production of virus-free orchid Cymbidium aloifolium (L.) Sw. by various tissue culture techniques.

Science.gov (United States)

Pradhan, Shreeti; Regmi, Tripti; Ranjit, Mukunda; Pant, Bijaya

2016-10-01

Orchids are affected by many viruses resulting in poor growth, yield and quality, and an overall decline in population. Cymbidium mosaic virus (CymMV) is one of the common orchid viruses found in Cymbidium species but it infects different orchid genera. In this study Cymbidium aloifolium was propagated in vitro using MS medium at different strength (1.0, ½, and ¼) with or without 0.5 mg/l BAP (6-benzylaminopurine) and 0.5 mg/l NAA (Naphthalene acetic acid). To provide disease-free planting material, source plant for in vitro propagation needs to be screened for pathogenic viruses. In the present study, in vivo -grown source (mother) plants and tissue culture-derived plants of C. aloifolium were tested for CymMV virus using Double antibody sandwich enzyme linked immunosorbent assay (DAS-ELISA). All the tissue cultured plants were found to be 100% virus-free whereas the in vivo grown source plants were highly affected by CymMV virus (83.33%). The virus-free in vitro plantlets were multiplied in large scale and then acclimatized on earthen pot containing a mixture of cocopeat, litter and clay in the ratio of 3:2:1. Eighty five percent of acclimatized plantlets survived making this method an efficient mass production system for high quality virus-free C. aloifolium for commercial floriculture and germplasm preservation.

Production of virus-free orchid Cymbidium aloifolium (L. Sw. by various tissue culture techniques

Directory of Open Access Journals (Sweden)

Shreeti Pradhan

2016-10-01

Full Text Available Orchids are affected by many viruses resulting in poor growth, yield and quality, and an overall decline in population. Cymbidium mosaic virus (CymMV is one of the common orchid viruses found in Cymbidium species but it infects different orchid genera. In this study Cymbidium aloifolium was propagated in vitro using MS medium at different strength (1.0, ½, and ¼ with or without 0.5 mg/l BAP (6-benzylaminopurine and 0.5 mg/l NAA (Naphthalene acetic acid. To provide disease-free planting material, source plant for in vitro propagation needs to be screened for pathogenic viruses. In the present study, in vivo-grown source (mother plants and tissue culture-derived plants of C. aloifolium were tested for CymMV virus using Double antibody sandwich enzyme linked immunosorbent assay (DAS-ELISA. All the tissue cultured plants were found to be 100% virus-free whereas the in vivo grown source plants were highly affected by CymMV virus (83.33%. The virus-free in vitro plantlets were multiplied in large scale and then acclimatized on earthen pot containing a mixture of cocopeat, litter and clay in the ratio of 3:2:1. Eighty five percent of acclimatized plantlets survived making this method an efficient mass production system for high quality virus-free C. aloifolium for commercial floriculture and germplasm preservation. Keywords: Biological sciences, Plant biology

A Simplified Method for Three-Dimensional (3-D Ovarian Tissue Culture Yielding Oocytes Competent to Produce Full-Term Offspring in Mice.

Directory of Open Access Journals (Sweden)

Carolyn M Higuchi

Full Text Available In vitro growth of follicles is a promising technology to generate large quantities of competent oocytes from immature follicles and could expand the potential of assisted reproductive technologies (ART. Isolated follicle culture is currently the primary method used to develop and mature follicles in vitro. However, this procedure typically requires complicated, time-consuming procedures, as well as destruction of the normal ovarian microenvironment. Here we describe a simplified 3-D ovarian culture system that can be used to mature multilayered secondary follicles into antral follicles, generating developmentally competent oocytes in vitro. Ovaries recovered from mice at 14 days of age were cut into 8 pieces and placed onto a thick Matrigel drop (3-D culture for 10 days of culture. As a control, ovarian pieces were cultured on a membrane filter without any Matrigel drop (Membrane culture. We also evaluated the effect of activin A treatment on follicle growth within the ovarian pieces with or without Matrigel support. Thus we tested four different culture conditions: C (Membrane/activin-, A (Membrane/activin+, M (Matrigel/activin-, and M+A (Matrigel/activin+. We found that the cultured follicles and oocytes steadily increased in size regardless of the culture condition used. However, antral cavity formation occurred only in the follicles grown in the 3-D culture system (M, M+A. Following ovarian tissue culture, full-grown GV oocytes were isolated from the larger follicles to evaluate their developmental competence by subjecting them to in vitro maturation (IVM and in vitro fertilization (IVF. Maturation and fertilization rates were higher using oocytes grown in 3-D culture (M, M+A than with those grown in membrane culture (C, A. In particular, activin A treatment further improved 3-D culture (M+A success. Following IVF, two-cell embryos were transferred to recipients to generate full-term offspring. In summary, this simple and easy 3-D ovarian

Insulin-like growth factor I (IGF-1 Ec/Mechano Growth factor--a splice variant of IGF-1 within the growth plate.

Directory of Open Access Journals (Sweden)

Werner Schlegel

Full Text Available Human insulin-like growth factor 1 Ec (IGF-1Ec, also called mechano growth factor (MGF, is a splice variant of insulin-like growth factor 1 (IGF-1, which has been shown in vitro as well as in vivo to induce growth and hypertrophy in mechanically stimulated or damaged muscle. Growth, hypertrophy and responses to mechanical stimulation are important reactions of cartilaginous tissues, especially those in growth plates. Therefore, we wanted to ascertain if MGF is expressed in growth plate cartilage and if it influences proliferation of chondrocytes, as it does in musculoskeletal tissues. MGF expression was analyzed in growth plate and control tissue samples from piglets aged 3 to 6 weeks. Furthermore, growth plate chondrocyte cell culture was used to evaluate the effects of the MGF peptide on proliferation. We showed that MGF is expressed in considerable amounts in the tissues evaluated. We found the MGF peptide to be primarily located in the cytoplasm, and in some instances, it was also found in the nucleus of the cells. Addition of MGF peptides was not associated with growth plate chondrocyte proliferation.

Increased and correlated expression of connective tissue growth factor and transforming growth factor beta 1 in surgically removed periodontal tissues with chronic periodontitis.

Science.gov (United States)

Mize, T W; Sundararaj, K P; Leite, R S; Huang, Y

2015-06-01

Both gingival tissue destruction and regeneration are associated with chronic periodontitis, although the former overwhelms the latter. Studies have shown that transforming growth factor beta 1 (TGF-β1), a growth factor largely involved in tissue regeneration and remodeling, is upregulated in chronic periodontitis. However, the gingival expression of connective tissue growth factor (CTGF or CCN2), a TGF-β1-upregulated gene, in patients with periodontitis remains undetermined. Although both CTGF/CCN2 and TGF-b1 increase the production of extracellular matrix, they have many different biological functions. Therefore, it is important to delineate the impact of periodontitis on gingival CTGF/CCN2 expression. Periodontal tissue specimens were collected from seven individuals without periodontitis (group 1) and from 14 with periodontitis (group 2). The expression of CTGF and TGFβ1 mRNAs were quantified using real-time PCR. Analysis using the nonparametric Mann-Whitney U-test showed that the levels of expression of both CTGF/CCN2 and TGFβ1 mRNAs were significantly increased in individuals with periodontitis compared with individuals without periodontitis. Furthermore, analysis using a nonparametric correlation (Spearman r) test showed a positive correlation between TGFβ1 and CTGF/CCN2 mRNAs. The gingival expression levels of CTGF/CCN2 and TGFβ1 mRNAs in individuals with periodontitis are upregulated and correlated. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

A thin-layer liquid culture technique for the growth of Helicobacter pylori.

Science.gov (United States)

Joo, Jung-Soo; Park, Kyung-Chul; Song, Jae-Young; Kim, Dong-Hyun; Lee, Kyung-Ja; Kwon, Young-Cheol; Kim, Jung-Min; Kim, Kyung-Mi; Youn, Hee-Shang; Kang, Hyung-Lyun; Baik, Seung-Chul; Lee, Woo-Kon; Cho, Myung-Je; Rhee, Kwang-Ho

2010-08-01

Several attempts have been successful in liquid cultivation of Helicobaccter pylori. However, there is a need to improve the growth of H. pylori in liquid media in order to get affluent growth and a simple approach for examining bacterial properties. We introduce here a thin-layer liquid culture technique for the growth of H. pylori. A thin-layer liquid culture system was established by adding liquid media to a 90-mm diameter Petri dish. Optimal conditions for bacterial growth were investigated and then viability, growth curve, and released proteins were examined. Maximal growth of H. pylori was obtained by adding 3 mL of brucella broth supplemented with 10% horse to a Petri dish. H. pylori grew in both DMEM and RPMI-1640 supplemented with 10% fetal bovine serum and 0.5% yeast extract. Serum-free RPMI-1640 supported the growth of H. pylori when supplemented with dimethyl-beta-cyclodextrin (200 microg/mL) and 1% yeast extract. Under optimal growth, H. pylori grew exponentially for 28 hours, reaching a density of 3.4 OD(600) with a generation time of 3.3 hours. After 24 hours, cultures at a cell density of 1.0 OD(600) contained 1.3 +/- 0.1 x 10(9 )CFU/mL. gamma-Glutamyl transpeptidase, nuclease, superoxide dismutase, and urease were not detected in culture supernatants at 24 hours in thin-layer liquid culture, but were present at 48 hours, whereas alcohol dehydrogenase, alkylhydroperoxide reductase, catalase, and vacuolating cytotoxin were detected at 24 hours. Thin-layer liquid culture technique is feasible, and can serve as a versatile liquid culture technique for investigating bacterial properties of H. pylori.

Substrate specific hydrolysis of aromatic and aromatic-aliphatic esters in orchid tissue cultures

Directory of Open Access Journals (Sweden)

Agnieszka Mironowicz

2014-01-01

Full Text Available We found that tissue cultures of higher plants were able, similarly as microorganisms, to transform low-molecular-weight chemical compounds. In tissue cultures of orchids (Cymbidium 'Saint Pierre' and Dendrobium phalaenopsis acetates of phenols and aromatic-aliphatic alcohols were hydrolyzed, whereas methyl esters of aromatic and aromatic-aliphatic acids did not undergo this reaction. Acetates of racemic aromatic-aliphatic alcohols were hydrolyzed with distinct enantiospecificity.

Clonogenic growth of human breast cancer cells co-cultured in direct contact with serum-activated fibroblasts

International Nuclear Information System (INIS)

Samoszuk, Michael; Tan, Jenny; Chorn, Guillaume

2005-01-01

Accumulating evidence suggests that fibroblasts play a pivotal role in promoting the growth of breast cancer cells. The objective of the present study was to characterize and validate an in vitro model of the interaction between small numbers of human breast cancer cells and human fibroblasts. We measured the clonogenic growth of small numbers of human breast cancer cells co-cultured in direct contact with serum-activated, normal human fibroblasts. Using DNA microarrays, we also characterized the gene expression profile of the serum-activated fibroblasts. In order to validate the in vivo relevance of our experiments, we then analyzed clinical samples of metastatic breast cancer for the presence of myofibroblasts expressing α-smooth muscle actin. Clonogenic growth of human breast cancer cells obtained directly from in situ and invasive tumors was dramatically and consistently enhanced when the tumor cells were co-cultured in direct contact with serum-activated fibroblasts. This effect was abolished when the cells were co-cultured in transwells separated by permeable inserts. The fibroblasts in our experimental model exhibited a gene expression signature characteristic of 'serum response' (i.e. myofibroblasts). Immunostaining of human samples of metastatic breast cancer tissue confirmed that myofibroblasts are in direct contact with breast cancer cells. Serum-activated fibroblasts promote the clonogenic growth of human breast cancer cells in vitro through a mechanism that involves direct physical contact between the cells. This model shares many important molecular and phenotypic similarities with the fibroblasts that are naturally found in breast cancers

Isolation, Characterization and Growth Kinetic Comparison of Bone Marrow and Adipose Tissue Mesenchymal Stem Cells of Guinea Pig.

Science.gov (United States)

Aliborzi, Ghaem; Vahdati, Akbar; Mehrabani, Davood; Hosseini, Seyed Ebrahim; Tamadon, Amin

2016-05-30

Mesenchymal stem cells (MSCs) from different sources have different characteristics. Moreover, MSCs are not isolated and characterized in Guinea pig for animal model of cell therapy. was the isolating of bone marrow MSCs (BM-MSCs) and adipose tissue MSCs (AT-MSCs) from Guinea pig and assessing their characteristics. In this study, bone marrow and adipose tissue were collected from three Guinea pigs and cultured and expanded through eight passages. BM-MSCs and AT-MSCs at passages 2, 5 and 8 were seeded in 24-well plates in triplicate. Cells were counted from each well 1~7 days after seeding to determine population doubling time (PDT) and cell growth curves. Cells of passage 3 were cultured in osteogenic and adipogenic differentiation media. BM-MSCs and AT-MSCs attached to the culture flask and displayed spindle-shaped morphology. Proliferation rate of AT-MSCs in the analyzed passages was more than BM-MSCs. The increase in the PDT of MSCs occurs with the increase in the number of passages. Moreover, after culture of BM-MSCs and AT-MSCs in differentiation media, the cells differentiated toward osteoblasts and adipocytes as verified by Alizarin Red staining and Oil Red O staining, respectively. BM-MSCs and AT-MSCs of Guinea pig could be valuable source of multipotent stem cells for use in experimental and preclinical studies in animal models.

Self-Condensation Culture Enables Vascularization of Tissue Fragments for Efficient Therapeutic Transplantation

Directory of Open Access Journals (Sweden)

Yoshinobu Takahashi

2018-05-01

Full Text Available Summary: Clinical transplantation of tissue fragments, including islets, faces a critical challenge because of a lack of effective strategies that ensure efficient engraftment through the timely integration of vascular networks. We recently developed a complex organoid engineering method by “self-condensation” culture based on mesenchymal cell-dependent contraction, thereby enabling dissociated heterotypic lineages including endothelial cells to self-organize in a spatiotemporal manner. Here, we report the successful adaptation of this method for generating complex tissues from diverse tissue fragments derived from various organs, including pancreatic islets. The self-condensation of human and mouse islets with endothelial cells not only promoted functionalization in culture but also massively improved post-transplant engraftment. Therapeutically, fulminant diabetic mice were more efficiently treated by a vascularized islet transplant compared with the conventional approach. Given the general limitations of post-transplant vascularization associated with 3D tissue-based therapy, our approach offers a promising means of enhancing efficacy in the context of therapeutic tissue transplantation. : Takahashi et al. report on generating vascularized islet tissue from humans and mice. After transplantation, vascularized islets significantly improve survival of diabetic mice, demonstrating the quick normalization of blood glucose compared with conventional islet transplantation. Keywords: tissue engineering, tissue-based therapy, vascularization, islet transplantation, organoid

Monitoring Dynamic Interactions between Breast Cancer Cells and Human Bone Tissue in a Co-Culture Model

Science.gov (United States)

Contag, Christopher H.; Lie, Wen-Rong; Bammer, Marie C.; Hardy, Jonathan W.; Schmidt, Tobi L.; Maloney, William J.; King, Bonnie L.

2015-01-01

Purpose Bone is a preferential site of breast cancer metastasis and models are needed to study this process at the level of the microenvironment. We have used bioluminescence imaging (BLI) and multiplex biomarker immunoassays to monitor dynamic breast cancer cell behaviors in co-culture with human bone tissue. Procedures Femur tissue fragments harvested from hip replacement surgeries were co-cultured with luciferase-positive MDA-MB-231-fLuc cells. BLI was performed to quantify breast cell division and track migration relative to bone tissue. Breast cell colonization of bone tissues was assessed with immunohistochemistry. Biomarkers in co-culture supernatants were profiled with MILLIPLEX® immunoassays. Results BLI demonstrated increased MDA-MB-231-fLuc proliferation (pbones, and revealed breast cell migration toward bone. Immunohistochemistry illustrated MDA-MB-231-fLuc colonization of bone, and MILLIPLEX® profiles of culture supernatants suggested breast/bone crosstalk. Conclusions Breast cell behaviors that facilitate metastasis occur reproducibly in human bone tissue co-cultures and can be monitored and quantified using BLI and multiplex immunoassays. PMID:24008275

Air exposure induced characteristics of dry eye in conjunctival tissue culture.

Directory of Open Access Journals (Sweden)

Hui Lin

Full Text Available There are several animal models illustrating dry eye pathophysiology. Current study would like to establish an ex vivo tissue culture model for characterizing dry eye. Human conjunctival explants were cultured under airlift or submerged conditions for up to 2 weeks, and only airlifted conjunctival cultures underwent increased epithelial stratification. Starting on day 4, the suprabasal cells displayed decreased K19 expression whereas K10 keratin became evident in airlift group. Pax6 nuclear expression attenuated already at 2 days, while its perinuclear and cytoplasmic expression gradually increased. MUC5AC and MUC19 expression dramatically decreased whereas the full thickness MUC4 and MUC16 expression pattern disappeared soon after initiating the airlift condition. Real time PCR showed K16, K10 and MUC16 gene up-regulated while K19, MUC5AC, MUC19 and MUC4 down-regulated on day 8 and day 14. On day 2 was the appearance of apoptotic epithelial and stromal cells appeared. The Wnt signaling pathway was transiently activated from day 2 to day 10. The inflammatory mediators IL-1β, TNF-α, and MMP-9 were detected in the conditioned media after 6 to 8 days. In conclusion, airlifted conjunctival tissue cultures demonstrated Wnt signaling pathway activation, coupled with squamous metaplasia, mucin pattern alteration, apoptosis and upregulation of proinflammatory cytokine expression. These changes mimic the pathohistological alterations described in dry eye. This correspondence suggests that insight into the pathophysiology of dry eye may be aided through the use of airlifted conjunctival tissue cultures.

The use of tissue culture techniques to detect irradiated vegetables

International Nuclear Information System (INIS)

Al-Safadi, B.; Sharabi, N.E.; Nabulsi, I

2001-01-01

the ability of two tissue culture methods, callus and vegetable growth induction, to detect irradiated vegetables was evaluated. Potato tubers, carrot roots, garlic cloves and onion bulbs were subjected to various gamma radiation doses (0, 25, 100, 150, 250, 500, 750, and 1000 Gy). Irradiated vegetables were cultured in vitro and in vivo (pots). Gamma irradiation significantly reduced callus-forming ability especially in carrot and potato where no callus was observed in doses higher than 50 Gy. Length of shoots and roots growing from irradiated garlic and onion explants was considerably reduced starting from the 25 Gy dose. No roots were formed on garlic explants at any irradiation dose. Garlic leaves growing from irradiated explants were spotted with purple to brown spots. The intensity of these spots increased as gamma ray dosage increased. In the pot experiment, potato plant appeared in the control only. On the contrary, a complete sprouting of garlic and onion was seen in all irradiation treatments. It was not possible to distinguish between the various irradiation treatments and the control 3 days after planting in pots. The two in vitro techniques, tested in our study, may effectively be used to detect irradiated vegetables and estimate the range of doses used. The callus formation method is more useful for potato and carrot, since regeneration of shoots in vitro from these two plants takes along time, making this method unpractical. The other technique is very useful in the case of onion and garlic since it is rapid. The two techniques can be used with most of the vegetables that can be cultured in vitro. (Author)

Flowering of Woody Bamboo in Tissue Culture Systems

Directory of Open Access Journals (Sweden)

Jin-Ling Yuan

2017-09-01

Full Text Available Flowering and subsequent seed set are not only normal activities in the life of most plants, but constitute the very reason for their existence. Woody bamboos can take a long time to flower, even over 100 years. This makes it difficult to breed bamboo, since flowering time cannot be predicted and passing through each generation takes too long. Another unique characteristic of woody bamboo is that a bamboo stand will often flower synchronously, both disrupting the supply chain within the bamboo industry and affecting local ecology. Therefore, an understanding of the mechanism that initiates bamboo flowering is important not only for biology research, but also for the bamboo industry. Induction of flowering in vitro is an effective way to both shorten the flowering period and control the flowering time, and has been shown for several species of bamboo. The use of controlled tissue culture systems allows investigation into the mechanism of bamboo flowering and facilitates selective breeding. Here, after a brief introduction of flowering in bamboo, we review the research on in vitro flowering of bamboo, including our current understanding of the effects of plant growth regulators and medium components on flower induction and how in vitro bamboo flowers can be used in research.

Role of Insulin-like growth factors in initiation of follicle growth in normal and polycystic human ovaries.

Science.gov (United States)

Stubbs, Sharron A; Webber, Lisa J; Stark, Jaroslav; Rice, Suman; Margara, Raul; Lavery, Stuart; Trew, Geoffrey H; Hardy, Kate; Franks, Stephen

2013-08-01

Polycystic ovary syndrome (PCOS), the commonest cause of anovulatory infertility, is characterized by disordered follicle development including increased activation and accelerated growth of preantral follicles. Data from experimental animals and preliminary results from studies of human ovarian tissue suggest that IGFs affect preantral follicle development. Our objectives were to investigate the expression of the type-1 IGF receptor (IGFR-1) in the human ovary and to determine whether IGFs are involved in stimulating the transition of follicles from primordial to primary stage in normal and polycystic ovaries. We used archived ovarian tissue for protein expression studies and small cortical biopsies for follicle isolation and for tissue culture. This was a laboratory-based study, using clinical tissue samples. A total of 54 women, 33 with normal ovaries and 21 with polycystic ovaries, were classified by reference to menstrual cycle history and ultrasonography. We evaluated expression of IGFR-1 mRNA in isolated preantral follicles and of IGFR-1 protein in archived ovarian tissue samples from normal and polycystic ovaries and effects of exogenous IGF-1 on preantral follicle development and survival in cultured fragments of normal and polycystic ovaries. IGFR-1 mRNA and protein was expressed in preantral follicles at all stages of development and enhanced expression was noted in PCOS follicles during early preantral development. IGF-1 stimulated initiation of follicle growth in normal tissue but had little effect on preantral follicle growth in polycystic ovaries in which, characteristically, there was a higher proportion of follicles that had entered the growing phase even before culture. IGFs are plausible candidates in regulation of initiation of human follicle growth, and accelerated preantral follicle growth in PCOS may be due to increased activity of endogenous IGFs.

Growth-supporting activity for Legionella pneumophila in tap water cultures and implication of hartmannellid amoebae as growth factors.

Science.gov (United States)

Wadowsky, R M; Butler, L J; Cook, M K; Verma, S M; Paul, M A; Fields, B S; Keleti, G; Sykora, J L; Yee, R B

1988-11-01

Photosynthetic cyanobacteria, heterotrophic bacteria, free-living amoebae, and ciliated protozoa may support growth of Legionella pneumophila. Studies were done with two tap water cultures (WS1 and WS2) containing L. pneumophila and associated microbiota to characterize growth-supporting activity and assess the relative importance of the microbiota in supporting multiplication of L. pneumophila. The water cultures were incubated in the dark at 35 degrees C. The growth-supporting factor(s) was separated from each culture by filtration through 1-micron-pore-size membrane filters. The retentate was then suspended in sterile tap water. Multiplication of L. pneumophila occurred when both the retentate suspension and the filtrate from either culture were inoculated into sterile tap water. L. pneumophila did not multiply in tap water inoculated with only the filtrate, even though filtration did not reduce the concentration of L. pneumophila or heterotrophic bacteria in either culture. Growth-supporting activity of the retentate suspension from WS1 was inactivated at 60 degrees C but unaffected at 0, 25, and 45 degrees C after 30-min incubations. Filtration experiments indicated that the growth-supporting factor(s) in WS1 was 2 to 5 micron in diameter. Ciliated protozoa were not detected in either culture. Hartmannellid amoebae were conclusively demonstrated in WS2 but not in WS1. L. pneumophila multiplied in tap water inoculated with the amoebae (10(3)/ml) and the 1-micron filtrate of WS2. No multiplication occurred in tap water inoculated with the filtrate only. Growth-supporting activity for L. pneumophila may be present in plumbing systems; hartmannellid amoebae appear to be important determinants of multiplication of L. pneumophila in some tap water cultures.

Selection of seed lots of Pinus taeda L. for tissue culture

Directory of Open Access Journals (Sweden)

Diego Pascoal Golle

2014-06-01

Full Text Available The aim of this work was to identify the fungi genera associated with three Pinus taeda L. seed lots and to assess the sanitary and physiological quality of these lots for use as selection criteria for tissue culture and evaluate the in vitro establishment of explants from seminal origin in different nutritive media. It was possible to discriminate the lots on the sanitary and physiological quality, as well as to establish in vitro plants of Pinus taeda from cotyledonary nodes obtained from aseptic seed germination of a selected lot by the sanitary and physiological quality higher. The nutritive media MS, ½ MS and WPM were equally suitable for this purpose. For the sanitary analysis the fungal genera Fusarium, Penicillium and Trichoderma were those of the highest sensitivity. For the physiological evaluation were important the variables: abnormal seedlings, strong normal seedlings; length, fresh and dry weight of strong normal seedlings. The analyzes were favorable to choose lots of seeds for in vitro culture and all culture media were adequate for the establishment of this species in tissue culture.

Joint use of developed collagen-containing complexes and cell cultures in creating new tissue equivalents

Directory of Open Access Journals (Sweden)

K. V. Kulakova

2016-01-01

Full Text Available The purpose of the study is to assess the possibility of applying the integrated module as the basis of a celltissue equivalent for treatment of wounds of skin and soft tissues. In the frame of the set task the following problems were being solved: research of the spatial structure and architectonics of the surface of the developed base collagen-containing materials and their biocompatibility with cell cultures.Materials and methods. The study of a material which is a two-layer complex film, consisting of collagen and polysaccharide components was carried out. The collagen was separated from the dermis and was then impregnated with particulate demineralized bone matrix (DCM according to the original methodology. For the purposes of the study the dehydrated material was created in the form of a film. Electron microscopic examination of surfaces was performed on scanning electron microscope JEOL JSM-IT300LV in high vacuum and at low values of probe current (< 0,1 nА. Studies to assess the viability of the cells cultivated on films of collagen material (tested for cytotoxicity and the adhesive capacity were performed in vitro using strains of diploid human fibroblasts 4–6 passage. The culture condition was visually assessed using an inverted Leica microscope DM IL (Carl Zeiss, Austria, equipped with a computerizes program of control of culture growth (Leica IM 1000.Results. The data obtained in the study of the surface structure of the developed complex module showed that it seems to be promising as a basic component of the cellular-tissue system with its large number of structural formations for fixation of the cells and a well-organized barrier layer capable of vapor - permeability. Experiments in vitro confirmed the absence of toxicity of the material being studied in relation to the culture of dermal human fibroblasts, suggesting the possibility of creation on its basis of cell-tissue complex and further experimental studies in vivo

Transcriptomic comparisons between cultured human adipose tissue-derived pericytes and mesenchymal stromal cells

Directory of Open Access Journals (Sweden)

Lindolfo da Silva Meirelles

2016-03-01

Full Text Available Mesenchymal stromal cells (MSCs, sometimes called mesenchymal stem cells, are cultured cells able to give rise to mature mesenchymal cells such as adipocytes, osteoblasts, and chondrocytes, and to secrete a wide range of trophic and immunomodulatory molecules. Evidence indicates that pericytes, cells that surround and maintain physical connections with endothelial cells in blood vessels, can give rise to MSCs (da Silva Meirelles et al., 2008 [1]; Caplan and Correa, 2011 [2]. We have compared the transcriptomes of highly purified, human adipose tissue pericytes subjected to culture-expansion in pericyte medium or MSC medium, with that of human adipose tissue MSCs isolated with traditional methods to test the hypothesis that their transcriptomes are similar (da Silva Meirelles et al., 2015 [3]. Here, we provide further information and analyses of microarray data from three pericyte populations cultured in pericyte medium, three pericyte populations cultured in MSC medium, and three adipose tissue MSC populations deposited in the Gene Expression Omnibus under accession number GSE67747. Keywords: Mesenchymal stromal cells, Mesenchymal stem cells, Pericytes, Microarrays

Improved Performance in Mammalian Cell Perfusion Cultures by Growth Inhibition.

Science.gov (United States)

Wolf, Moritz K F; Closet, Aurélie; Bzowska, Monika; Bielser, Jean-Marc; Souquet, Jonathan; Broly, Hervé; Morbidelli, Massimo

2018-05-21

Mammalian cell perfusion cultures represent a promising alternative to the current fed-batch technology for the production of various biopharmaceuticals. Long-term operation at a fixed viable cell density (VCD) requires a viable culture and a constant removal of excessive cells. Product loss in the cell removing bleed stream deteriorates the process yield. In this study, the authors investigate the use of chemical and environmental growth inhibition on culture performance by either adding valeric acid (VA) to the production media or by reducing the culture temperature (33.0 °C) with respect to control conditions (36.5 °C, no VA). Low temperature significantly reduces cellular growth, thus, resulting in lower bleed rates accompanied by a reduced product loss of 11% compared to 26% under control conditions. Additionally, the cell specific productivity of the target protein improves and maintained stable leading to media savings per mass of product. VA shows initially an inhibitory effect on cellular growth. However, cells seemed to adapt to the presence of the inhibitor resulting in a recovery of the cellular growth. Cell cycle and Western blot analyses support the observed results. This work underlines the role of temperature as a key operating variable for the optimization of perfusion cultures. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Frozen and fresh ovarian tissue require different culture media to promote in vitro development of bovine preantral follicles.

Science.gov (United States)

Castro, Simone Vieira; Carvalho, Adeline Andrade; Silva, Cleidson Manoel Gomes; Santos, Francielli Weber; Campello, Cláudio Cabral; de Figueiredo, José Ricardo; Rodrigues, Ana Paula Ribeiro

2014-10-01

The aim of this study was to evaluate the efficiency of different media in the in vitro culture of bovine preantral follicles that were used either fresh or following slow freezing treatment. Frozen and fresh noncultured or cultured ovarian fragments were processed for histological, viability, and cell proliferation analyses. For cryopreservation, a solution containing 1.5 M ethylene glycol was frozen in a programmable biological freezer. After thawing, a portion of the samples was destined for frozen controls. The remainder were cultured in vitro for 5 days in three media: α-MEM, McCoy, or M199. Samples from these culture media were collected on days 1 and 5 for quantification of reactive oxygen species (ROS) and for hormonal assays. In fresh-cultured tissues, the percentage of morphologically normal follicles was significantly higher when cultured in M199 compared to that in the other media. In frozen-cultured tissues, McCoy medium was significantly superior to the other media, and was the only treatment that helped in maintaining the viability similar to fresh and frozen controls. Upon quantification of the nucleolus organizer region, we observed greater proliferation of granulosa cells in the frozen-cultured tissues with McCoy medium, and lesser proliferation in fresh-cultured tissues only with α-MEM. In frozen-cultured tissues, ROS levels were highest at day 1 and progressively reduced during culture, independent of the media used. In conclusion, under the conditions used in this study, the M199 and McCoy media are recommended for the culture of follicles derived from fresh and frozen ovarian tissues, respectively.

Differential modulation of growth and phenotypic expression of chondrocytes in sparse and confluent cultures by growth factors in cartilage

International Nuclear Information System (INIS)

Hiraki, Y.; Inoue, H.; Asada, A.; Suzuki, F.

1990-01-01

The growth-promoting actions of cartilage extracts (CE) on rabbit cultured chondrocytes were studied to assess the role of local acting growth factors in the generation and expansion of highly differentiated cells. In the present study, DNA synthesis and proteoglycan synthesis in the cultured chondrocytes were monitored by flow cytofluorometry and double-isotope autoradiography by using ( 3 H)thymidine and ( 35 S)sulfate. We report here that actions of the same set of growth factors extracted from cartilage evokes differential cellular responses depending upon cell density. Growth factors in the optimal dose of CE (2 micrograms/ml) or epidermal growth factor (EGF, 40 ng/ml) did not reveal such a cell density-dependent effect on cellular proliferation. However, growth factors in CE induced proteoglycan synthesis selectively in nonproliferating and expressing cells in confluent culture

Fabrication of myogenic engineered tissue constructs.

Science.gov (United States)

Pacak, Christina A; Cowan, Douglas B

2009-05-01

Despite the fact that electronic pacemakers are life-saving medical devices, their long-term performance in pediatric patients can be problematic owing to the restrictions imposed by a child's small size and their inevitable growth. Consequently, there is a genuine need for innovative therapies designed specifically for pediatric patients with cardiac rhythm disorders. We propose that a conductive biological alternative consisting of a collagen-based matrix containing autologously-derived cells could better adapt to growth, reduce the need for recurrent surgeries, and greatly improve the quality of life for these patients. In the present study, we describe a procedure for incorporating primary skeletal myoblast cell cultures within a hydrogel matrix to fashion a surgically-implantable tissue construct that will serve as an electrical conduit between the upper and lower chambers of the heart. Ultimately, we anticipate using this type of engineered tissue to restore atrioventricular electrical conduction in children with complete heart block. In view of that, we isolate myoblasts from the skeletal muscles of neonatal Lewis rats and plate them onto laminin-coated tissue culture dishes using a modified version of established protocols. After one to two days, cultured cells are collected and mixed with antibiotics, type 1 collagen, Matrigel, and NaHCO(3). The result is a viscous, uniform solution that can be cast into a mold of nearly any shape and size. For our tissue constructs, we employ type 1 collagen isolated from fetal lamb skin using standard procedures. Once the tissue has solidified at 37 degrees C, culture media is carefully added to the plate until the construct is submerged. The engineered tissue is then allowed to further condense through dehydration for 2 more days, at which point it is ready for in vitro assessment or surgical-implantation.

Enhanced chondrocyte culture and growth on biologically inspired nanofibrous cell culture dishes.

Science.gov (United States)

Bhardwaj, Garima; Webster, Thomas J

2016-01-01

Chondral and osteochondral defects affect a large number of people in which treatment options are currently limited. Due to its ability to mimic the natural nanofibrous structure of cartilage, this current in vitro study aimed at introducing a new scaffold, called XanoMatrix™, for cartilage regeneration. In addition, this same scaffold is introduced here as a new substrate onto which to study chondrocyte functions. Current studies on chondrocyte functions are limited due to nonbiologically inspired cell culture substrates. With its polyethylene terephthalate and cellulose acetate composition, good mechanical properties and nanofibrous structure resembling an extracellular matrix, XanoMatrix offers an ideal surface for chondrocyte growth and proliferation. This current study demonstrated that the XanoMatrix scaffolds promote chondrocyte growth and proliferation as compared with the Corning and Falcon surfaces normally used for chondrocyte cell culture. The XanoMatrix scaffolds also have greater hydrophobicity, three-dimensional surface area, and greater tensile strength, making them ideal candidates for alternative treatment options for chondral and osteochondral defects as well as cell culture substrates to study chondrocyte functions.

[18S-25S rDNA variation in tissue culture of some Gentiana L. species].

Science.gov (United States)

Mel'nyk, V M; Andrieiev, I O; Spiridonova, K V; Strashniuk, N M; Kunakh, V A

2007-01-01

18S-25S rDNA of intact plants and tissue cultures of G. acaulis, G. punctata and G. lutea have been investigated by using blot-hybridization. The decrease of rDNA amount was found in the callus cultures as compared with the plants. In contrast to other species, G. lutea showed intragenome heterogeneity of rRNA genes as well as qualitative rDNA changes in tissue culture, in particular appearance of altered repeats. The relationship between the peculiarities of rRNA gene structure and their rearrangements in in vitro culture was suggested.

The effect of gamma radiation and N-ethyl-N-nitrosourea on cultured maize callus growth and plant regeneration

International Nuclear Information System (INIS)

Moustafa, R.A.K.; Duncan, D.R.; Widholm, J.M.

1989-01-01

Regenerable maize calli of two inbred lines were exposed to 0 to 100 Gy of gamma rays or treated with 0 to 30 mM of N-ethyl-N-nitrosourea (ENU) to determine their effect on growth and plant regeneration capability. Both growth and plant regeneration capacity decreased with increasing levels of either gamma radiation or ENU; however, plant regeneration capacity was more sensitive to either agent than growth. The 50% inhibition dose (I 50 ) for callus growth (fresh-weight gain) was approximately 100 Gy of gamma radiation and 30 mM ENU. The I 50 for plant regeneration capacity of treated callus was approximately 25 Gy of gamma radiation and 2.5 mM ENU. The decrease in plant regeneration capacity correlated with a change in tissue composition of the treated callus from a hard, yellow and opaque tissue to a soft, grayish-yellow and translucent tissue. This change was quantified by measuring the reduction of MnO 4 - to MnO 2 (PR assay) by the callus. These results suggest that the effect of gamma radiation or ENU on plant regeneration capacity must be taken into consideration if these potentially mutagenic agents are to be used on maize callus cultures, for the purpose of producing useful mutations at a whole plant level. The data also suggest that the PR assay may be useful for predicting the actual plant regeneration capacity of maize callus. (author)

Upregulated epidermal growth factor receptor expression following near-infrared irradiation simulating solar radiation in a three-dimensional reconstructed human corneal epithelial tissue culture model.

Science.gov (United States)

Tanaka, Yohei; Nakayama, Jun

2016-01-01

Humans are increasingly exposed to near-infrared (NIR) radiation from both natural (eg, solar) and artificial (eg, electrical appliances) sources. Although the biological effects of sun and ultraviolet (UV) exposure have been extensively investigated, the biological effect of NIR radiation is still unclear. We previously reported that NIR as well as UV induces photoaging and standard UV-blocking materials, such as sunglasses, do not sufficiently block NIR. The objective of this study was to investigate changes in gene expression in three-dimensional reconstructed corneal epithelial tissue culture exposed to broad-spectrum NIR irradiation to simulate solar NIR radiation that reaches human tissues. DNA microarray and quantitative real-time polymerase chain reaction analysis were used to assess gene expression levels in a three-dimensional reconstructed corneal epithelial model composed of normal human corneal epithelial cells exposed to water-filtered broad-spectrum NIR irradiation with a contact cooling (20°C). The water-filter allowed 1,000-1,800 nm wavelengths and excluded 1,400-1,500 nm wavelengths. A DNA microarray with >62,000 different probes showed 25 and 150 genes that were up- or downregulated by at least fourfold and twofold, respectively, after NIR irradiation. In particular, epidermal growth factor receptor (EGFR) was upregulated by 19.4-fold relative to control cells. Quantitative real-time polymerase chain reaction analysis revealed that two variants of EGFR in human corneal epithelial tissue were also significantly upregulated after five rounds of 10 J/cm(2) irradiation (Psolar energy reaching the Earth is in the NIR region, which cannot be adequately blocked by eyewear and thus can induce eye damage with intensive or long-term exposure, protection from both UV and NIR radiation may prevent changes in gene expression and in turn eye damage.

Cell-surface glycoproteins of human sarcomas: differential expression in normal and malignant tissues and cultured cells

International Nuclear Information System (INIS)

Rettig, W.F.; Garin-Chesa, P.; Beresford, H.R.; Oettgen, H.F.; Melamed, M.R.; Old, L.J.

1988-01-01

Normal differentiation and malignant transformation of human cells are characterized by specific changes in surface antigen phenotype. In the present study, the authors have defined six cell-surface antigens of human sarcomas and normal mesenchymal cells, by using mixed hemadsorption assays and immunochemical methods for the analysis of cultured cells and immunohistochemical staining for the analysis of normal tissues and > 200 tumor specimens. Differential patterns of F19, F24, G171, G253, S5, and Thy-1 antigen expression were found to characterize (i) subsets of cultured sarcoma cell lines, (ii) cultured fibroblasts derived from various organs, (iii) normal resting and activated mesenchymal tissues, and (iv) sarcoma and nonmesenchymal tumor tissues. These results provide a basic surface antigenic map for cultured mesenchymal cells and mesenchymal tissues and permit the classification of human sarcomas according to their antigenic phenotypes

Pre-irradiation of tissue culture flasks leads to diminished stem and progenitor cell production in long-term bone marrow cultures

International Nuclear Information System (INIS)

Rooney, P.; Wright, E.G.

1993-01-01

Empty plastic tissue culture flasks were exposed to X-irradiation doses of 0.3-10.0 Gy, prior to the establishment of long-term bone marrow cultures. During the course of a 10 week culture period, all irradiated plastic flasks exhibited a dramatic decrease in the number of both haemopoietic stem cells and myeloid progenitor cells, in the non-adherent layer, when compared with controls. This decrease was not due to a decrease in the number of non-adherent cells produced. Histological examination of non-adherent cells showed an increase in mature granulocytic cells with few blast cells. Morphologically, the adherent layers of irradiated flasks demonstrated a delay in appearance or absence of fat cell production. X-irradiation of glass tissue culture flasks had no deleterious effect. (author)

Three-dimensional spheroid cell culture of umbilical cord tissue-derived mesenchymal stromal cells leads to enhanced paracrine induction of wound healing.

Science.gov (United States)

Santos, Jorge M; Camões, Sérgio P; Filipe, Elysse; Cipriano, Madalena; Barcia, Rita N; Filipe, Mariana; Teixeira, Mariana; Simões, Sandra; Gaspar, Manuela; Mosqueira, Diogo; Nascimento, Diana S; Pinto-do-Ó, Perpétua; Cruz, Pedro; Cruz, Helder; Castro, Matilde; Miranda, Joana P

2015-05-09

The secretion of trophic factors by mesenchymal stromal cells has gained increased interest given the benefits it may bring to the treatment of a variety of traumatic injuries such as skin wounds. Herein, we report on a three-dimensional culture-based method to improve the paracrine activity of a specific population of umbilical cord tissue-derived mesenchymal stromal cells (UCX®) towards the application of conditioned medium for the treatment of cutaneous wounds. A UCX® three-dimensional culture model was developed and characterized with respect to spheroid formation, cell phenotype and cell viability. The secretion by UCX® spheroids of extracellular matrix proteins and trophic factors involved in the wound-healing process was analysed. The skin regenerative potential of UCX® three-dimensional culture-derived conditioned medium (CM3D) was also assessed in vitro and in vivo against UCX® two-dimensional culture-derived conditioned medium (CM2D) using scratch and tubulogenesis assays and a rat wound splinting model, respectively. UCX® spheroids kept in our three-dimensional system remained viable and multipotent and secreted considerable amounts of vascular endothelial growth factor A, which was undetected in two-dimensional cultures, and higher amounts of matrix metalloproteinase-2, matrix metalloproteinase-9, hepatocyte growth factor, transforming growth factor β1, granulocyte-colony stimulating factor, fibroblast growth factor 2 and interleukin-6, when compared to CM2D. Furthermore, CM3D significantly enhanced elastin production and migration of keratinocytes and fibroblasts in vitro. In turn, tubulogenesis assays revealed increased capillary maturation in the presence of CM3D, as seen by a significant increase in capillary thickness and length when compared to CM2D, and increased branching points and capillary number when compared to basal medium. Finally, CM3D-treated wounds presented signs of faster and better resolution when compared to untreated and CM

Increasing cell culture population doublings for long-term growth of finite life span human cell cultures

Science.gov (United States)

Stampfer, Martha R; Garbe, James C

2015-02-24

Cell culture media formulations for culturing human epithelial cells are herein described. Also described are methods of increasing population doublings in a cell culture of finite life span human epithelial cells and prolonging the life span of human cell cultures. Using the cell culture media disclosed alone and in combination with addition to the cell culture of a compound associated with anti-stress activity achieves extended growth of pre-stasis cells and increased population doublings and life span in human epithelial cell cultures.

Human osteoblast cells: isolation, characterization, and growth on polymers for musculoskeletal tissue engineering.

Science.gov (United States)

El-Amin, Saadiq F; Botchwey, Edward; Tuli, Richard; Kofron, Michelle D; Mesfin, Addisu; Sethuraman, Swaminathan; Tuan, Rocky S; Laurencin, Cato T

2006-03-01

We performed a detailed examination of the isolation, characterization, and growth of human osteoblast cells derived from trabecular bone. We further examined the morphology, phenotypic gene expression, mineralization,and growth of these human osteoblasts on polyester polymers used for musculoskeletal tissue engineering. Polylactic-co-glycolic acid [PLAGA (85:15, 50:50, 75:25)], and poly-lactic acid (L-PLA, D,L-PLA) were examined. The osteoblastic expression of key phenotypic markers osteocalcin, alkaline phosphatase, collagen, and bone sialoprotein at 4 and 8 weeks was examined. Reverse transcription-polymerase chain reaction studies revealed that trabecular-derived osteoblasts were positive for all markers evaluated with higher levels expressed over long-term culture. These cells also revealed mineralization and maturation as evidenced by energy dispersive X-ray analysis and scanning electron microscopy. Growth studies on PLAGA at 50:50,75:25, and 85:15 ratios and PLA in the L and DL isoforms revealed that human osteoblasts actively grew, with significantly higher cell numbers attached to scaffolds composed of PLAGA 50:50 in the short term and PLAGA 85:15 in the long term compared with PLA (p < 0.05). We believe human cell adhesion among these polymeric materials may be dependent on differences in cellular integrin expression and extracellular matrix protein elaboration. (c) 2005 Wiley Periodicals, Inc.

Evaluation of the growth environment of a hydrostatic force bioreactor for preconditioning of tissue-engineered constructs.

Science.gov (United States)

Reinwald, Yvonne; Leonard, Katherine H L; Henstock, James R; Whiteley, Jonathan P; Osborne, James M; Waters, Sarah L; Levesque, Philippe; El Haj, Alicia J

2015-01-01

Bioreactors have been widely acknowledged as valuable tools to provide a growth environment for engineering tissues and to investigate the effect of physical forces on cells and cell-scaffold constructs. However, evaluation of the bioreactor environment during culture is critical to defining outcomes. In this study, the performance of a hydrostatic force bioreactor was examined by experimental measurements of changes in dissolved oxygen (O2), carbon dioxide (CO2), and pH after mechanical stimulation and the determination of physical forces (pressure and stress) in the bioreactor through mathematical modeling and numerical simulation. To determine the effect of hydrostatic pressure on bone formation, chick femur skeletal cell-seeded hydrogels were subjected to cyclic hydrostatic pressure at 0-270 kPa and 1 Hz for 1 h daily (5 days per week) over a period of 14 days. At the start of mechanical stimulation, dissolved O2 and CO2 in the medium increased and the pH of the medium decreased, but remained within human physiological ranges. Changes in physiological parameters (O2, CO2, and pH) were reversible when medium samples were placed in a standard cell culture incubator. In addition, computational modeling showed that the distribution and magnitude of physical forces depends on the shape and position of the cell-hydrogel constructs in the tissue culture format. Finally, hydrostatic pressure was seen to enhance mineralization of chick femur skeletal cell-seeded hydrogels.

Hormonal regulation of wheat growth during hydroponic culture

Science.gov (United States)

Wetherell, Donald

1988-01-01

Hormonal control of root growth has been explored as one means to alleviate the crowding of plant root systems experienced in prototype hydroponic biomass production chambers being developed by the CELSS Breadboard Project. Four plant hormones, or their chemical analogs, which have been reported to selectively inhibit root growth, were tested by adding them to the nutrient solutions on day 10 of a 25 day growth test using spring wheat in hydroponic cultures. Growth and morphological changes is both shoot and root systems were evaluated. In no case was it possible to inhibit root growth without a comparable inhibition of shoot growth. It was concluded that this approach is unlikely to prove useful for wheat.

Culture on fibrin matrices maintains the colony-forming capacity and osteoblastic differentiation of mesenchymal stem cells

International Nuclear Information System (INIS)

Colley, Helen; McArthur, Sally L; Stolzing, Alexandra; Scutt, Andy

2012-01-01

Mesenchymal stem cells (MSC) are multipotent cells capable of differentiating into a number of mesenchymal tissues including bone, cartilage, and tendon. Low numbers in vivo means exponential growth is needed in culture to enable therapeutic applications. MSC can expand rapidly in culture but usually lose their extensive capacity for differentiation that makes them therapeutically attractive. To try and maintain their capacity for differentiation and expansion in vitro, we cultured MSC on fibrin gels of different concentrations to create more physiological growth conditions for the cells. The cells were then re-plated onto tissue culture plastic and analysed. The cells that had been pre-cultured for seven days on fibrin, proliferated and maintained their differential potential to the osteogenic lineage better than tissue culture plastic expanded MSC. A concentration relationship between colony number and fibrin concentration was seen with decreasing numbers as fibrin concentration increased. These data support the concept that substrate signals significantly influence MSC growth and differentiation and that growth on a fibrin matrix could be used to maintain a stem cell phenotype during MSC expansion. (paper)

Tissue Engineering Organs for Space Biology Research

Science.gov (United States)

Vandenburgh, H. H.; Shansky, J.; DelTatto, M.; Lee, P.; Meir, J.

1999-01-01

Long-term manned space flight requires a better understanding of skeletal muscle atrophy resulting from microgravity. Atrophy most likely results from changes at both the systemic level (e.g. decreased circulating growth hormone, increased circulating glucocorticoids) and locally (e.g. decreased myofiber resting tension). Differentiated skeletal myofibers in tissue culture have provided a model system over the last decade for gaining a better understanding of the interactions of exogenous growth factors, endogenous growth factors, and muscle fiber tension in regulating protein turnover rates and muscle cell growth. Tissue engineering these cells into three dimensional bioartificial muscle (BAM) constructs has allowed us to extend their use to Space flight studies for the potential future development of countermeasures.

Chemical evaluation of strawberry plants produced by tissue culturing of gamma irradiated seedlings

International Nuclear Information System (INIS)

Maraei, R.W.

2007-01-01

studies were conducted to evaluate the influence of gamma irradiation as a supplementary factor precedes tissue culture application on strawberry seedlings (c.v.Rosa Linda). the strawberry seedling were irradiated using 8 doses of co 60 gamma rays 50.75.100.125 ,150,250, 350 and 500 gray. tissue culture technique was applied on irradiated and unirradiated strawberry seedling. different characteristics of plantlets, plant and fruit of strawberry produced from the double treatment (irradiation followed by tissue culture) were studied as well as the early, total and exportable fruit yields. data indicated that, low radiation doses 50,75 and 100 gray increased all morphological and chemical characteristics of the plantlets, plant and fruit of strawberry, whereas radiation doses higher than 100 gray decreased them significantly. moreover 350 and gray were lethal doses. radiation dose 50 gray increased the survival percentage and the length of plantlets by 1.5% and 50% respectively more than the unirradiated treatment in all multiplication stages

Studies on the reaction in tissue culture of tomato genotypes under biotic stress

Directory of Open Access Journals (Sweden)

Ewa Hanus-Fajerska

2014-01-01

Full Text Available Plant regeneration in vitro from virus-infected somatic tomato (Lycopersicon sp. tissue was performed. Regeneration experiments were started after the determination of virus presence, using enzyme-linked immunosorbent assay, in leaves used as a source of explants. Leaf explants infected with selected strains of tomato mosaic Tobamovirus or cucumber mosaic Cucumovirus respectively, were cultured on a standarised MS agar medium to induce adventitious shoots, which were afterwards excised, rooted in vitro and cultured to plants. Explants were also screened for their ability to produce callus. Diverse effects of viral infection, ranging from stimulation to inhibition of callus formation and of morphogenesis rate, were observed. The health condition of the tissue proved to affect regeneration potential of Lycopersicon esculentum, whereas wild accesions did not react in that case so distinctly. In cultivated tomato was encountered the decline in competence to reproduce shoots adventitiously in infected tissue. There was also relationship between donor plant health condition and adventitious root formation in regenerated shoots. Experiments with short-term cultures of L. esculenum reveled also that a certain number of shoots regenerated from diseased tissue can be virus-free.

Growth Factors and Tension-Induced Skeletal Muscle Growth

Science.gov (United States)

Vandenburgh, Herman H.

1994-01-01

have performed experiments to determine whether mechanical stimulation of cultured avian muscle cells alters their response to anabolic steroids or glucocorticoids. In static cultures, testosterone had no effect on muscle cell growth, but 5alpha-dihydrotestosterone and the synthetic steroid stanozolol increased cell growth by up to 18% and 30%, respectively, after a three day exposure. We completed development of a new IBM-based mechanical cell stimulator system to provide greater flexibility in operating and monitoring our experiments. Our previous long term studies on myofiber growth were designed around a perfusion system of our own design. We have recently changed to performing these studies using a modified CELLCO cartridge bioreactor system Z since it has been certified as the ground-based model for the Shuttle's Space Tissue Loss (STL) F= Cell Culture Module. The current goals of this aspect of the project are three fold: 1) to design a Z cell culture system for studying avian skeletal myofiber atrophy on the Shuttle and Space Station; 0 2) to expand the use of bioreactors to cells which do not grow in either suspension or attached to the hollow fibers; and 3) to combine the bioreactor system with our computerized mechanical cell stimulator to have a better in vitro model to study tension/gravity/stretch regulation of skeletal muscle size. Preliminary studies also reported on involved : (1) how release of tension can induce rapid atrophy of tissues cultured avian skeletal muscle cells, and (2) a mechanism to transfer and maintain avian skeletal muscle organoids in modified cartridges in the Space Tissue Loss Module.

Simple and high yielding method for preparing tissue specific extracellular matrix coatings for cell culture.

Science.gov (United States)

DeQuach, Jessica A; Mezzano, Valeria; Miglani, Amar; Lange, Stephan; Keller, Gordon M; Sheikh, Farah; Christman, Karen L

2010-09-27

The native extracellular matrix (ECM) consists of a highly complex, tissue-specific network of proteins and polysaccharides, which help regulate many cellular functions. Despite the complex nature of the ECM, in vitro cell-based studies traditionally assess cell behavior on single ECM component substrates, which do not adequately mimic the in vivo extracellular milieu. We present a simple approach for developing naturally derived ECM coatings for cell culture that provide important tissue-specific cues unlike traditional cell culture coatings, thereby enabling the maturation of committed C2C12 skeletal myoblast progenitors and human embryonic stem cells differentiated into cardiomyocytes. Here we show that natural muscle-specific coatings can (i) be derived from decellularized, solubilized adult porcine muscle, (ii) contain a complex mixture of ECM components including polysaccharides, (iii) adsorb onto tissue culture plastic and (iv) promote cell maturation of committed muscle progenitor and stem cells. This versatile method can create tissue-specific ECM coatings, which offer a promising platform for cell culture to more closely mimic the mature in vivo ECM microenvironment.

Culture of equine fibroblast-like synoviocytes on synthetic tissue scaffolds towards meniscal tissue engineering: a preliminary cell-seeding study

Directory of Open Access Journals (Sweden)

Jennifer J. Warnock

2014-04-01

Full Text Available Introduction. Tissue engineering is a new methodology for addressing meniscal injury or loss. Synovium may be an ideal source of cells for in vitro meniscal fibrocartilage formation, however, favorable in vitro culture conditions for synovium must be established in order to achieve this goal. The objective of this study was to determine cellularity, cell distribution, and extracellular matrix (ECM formation of equine fibroblast-like synoviocytes (FLS cultured on synthetic scaffolds, for potential application in synovium-based meniscal tissue engineering. Scaffolds included open-cell poly-L-lactic acid (OPLA sponges and polyglycolic acid (PGA scaffolds cultured in static and dynamic culture conditions, and PGA scaffolds coated in poly-L-lactic (PLLA in dynamic culture conditions.Materials and Methods. Equine FLS were seeded on OPLA and PGA scaffolds, and cultured in a static environment or in a rotating bioreactor for 12 days. Equine FLS were also seeded on PGA scaffolds coated in 2% or 4% PLLA and cultured in a rotating bioreactor for 14 and 21 days. Three scaffolds from each group were fixed, sectioned and stained with Masson’s Trichrome, Safranin-O, and Hematoxylin and Eosin, and cell numbers and distribution were analyzed using computer image analysis. Three PGA and OPLA scaffolds from each culture condition were also analyzed for extracellular matrix (ECM production via dimethylmethylene blue (sulfated glycosaminoglycan assay and hydroxyproline (collagen assay. PLLA coated PGA scaffolds were analyzed using double stranded DNA quantification as areflection of cellularity and confocal laser microscopy in a fluorescent cell viability assay.Results. The highest cellularity occurred in PGA constructs cultured in a rotating bioreactor, which also had a mean sulfated glycosaminoglycan content of 22.3 µg per scaffold. PGA constructs cultured in static conditions had the lowest cellularity. Cells had difficulty adhering to OPLA and the PLLA

Apollo 12 lunar material - Effects on lipid levels of tobacco tissue cultures.

Science.gov (United States)

Weete, J. D.; Walkinshaw, C. H.; Laseter, J. L.

1972-01-01

Tobacco tissue cultures grown in contact with lunar material from Apollo 12, for a 12-week period, resulted in fluctuations of both the relative and absolute concentrations of endogenous sterols and fatty acids. The experimental tissues contained higher concentrations of sterols than the controls did. The ratio of campesterol to stigmasterol was greater than 1 in control tissues, but less than 1 in the experimental tissues after 3 weeks. High relative concentrations (17.1 to 22.2 per cent) of an unidentified compound or compounds were found only in control tissues that were 3 to 9 weeks of age.

SENSITIVITY OF PIGMENT CONTENT OF BANANA AND ORCHID TISSUE CULTURE EXPOSED TO EXTREMELY LOW FREQUENCY ELECTROMAGNETIC FIEL

Directory of Open Access Journals (Sweden)

Riry Prihatini

2017-01-01

Full Text Available Natural exposure of extremely low frequency electromagnetic field (ELF-EMF occurs in the environment and acts as one of the abiotic factors that affect the growth and development of organisms. This study was conducted to determine the effect of ELF-EMF on the tissue cultured banana and slipper orchid chlorophyll content as one of the indicators in measuring plant photosynthetic capacity. Four days old banana (Musa sp. cv. Berangan corm and seven days old slipper orchid (Paphiopedilum rothschildianum cultures were exposed to 6 and 12 mT ELF-EMF generated by controllable ELF-EMF built up machine for 0.5, 1, 2 and 4 hours. After exposure, the banana and orchid cultures were incubated at 25° C for 8 and 16 weeks, respectively. The results showed that the ELF-EMF exposure had different effects on banana and slipper orchid cultures though both plant species belong to monocotyledon. The highest increase in chlorophyll content on banana was resulted by the high intensity and long duration of ELF-EMF exposure (12 mT for 4 hours, whereas on slipper orchid the modest and short duration of ELF-EMF exposure produced the most excessive chlorophyll content. Different ELF-EMF exposures (12 mT for 4 hours and 6 mT for 30 minutes had potential to be applied on each plant to improve in vitro plant (banana and slipper orchid, respectively growth. The increased chlorophyll and carotene/xanthophyll content on banana indicated that the banana was more tolerant to ELF-EMF exposure compared to slipper orchid. 

Co-culture with infrapatellar fat pad differentially stimulates proteoglycan synthesis and accumulation in cartilage and meniscus tissues.

Science.gov (United States)

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

2017-09-01

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

The diagnostic value of plasma N-terminal connective tissue growth factor levels in children with heart failure.

Science.gov (United States)

Li, Gang; Song, Xueqing; Xia, Jiyi; Li, Jing; Jia, Peng; Chen, Pengyuan; Zhao, Jian; Liu, Bin

2017-01-01

The aim of this study was to assess the diagnostic value of plasma N-terminal connective tissue growth factor in children with heart failure. Methods and results Plasma N-terminal connective tissue growth factor was determined in 61 children, including 41 children with heart failure, 20 children without heart failure, and 30 healthy volunteers. The correlations between plasma N-terminal connective tissue growth factor levels and clinical parameters were investigated. Moreover, the diagnostic value of N-terminal connective tissue growth factor levels was evaluated. Compared with healthy volunteers and children without heart failure, plasma N-terminal connective tissue growth factor levels were significantly elevated in those with heart failure (p0.05), but it obviously improved the ability of diagnosing heart failure in children, as demonstrated by the integrated discrimination improvement (6.2%, p=0.013) and net re-classification improvement (13.2%, p=0.017) indices. Plasma N-terminal connective tissue growth factor is a promising diagnostic biomarker for heart failure in children.

Effect of Plant Growth Regulators on a Shoot and Root Formation from the Leaf and Flower Culture of a Standard-type Chrysanthemum 'Jinba'

International Nuclear Information System (INIS)

Lee, J.S.; Lee, G.J.; Chung, S.J.; Kim, J.B.; Kim, D.S.; Kang, S.Y.

2008-01-01

In this study we investigated the conditions of a higher frequency for regenerated plants from different explants of a standard-type chrysanthemum 'Jinba'. In vitro culture was initiated on an MS medium containing 3% sucrose, 0.8% agar, and 5 μM benzyl adenine (BA) with naphthalene acetic acid (NAA) by using surface-sterilized leaf and flower tissues from greenhouse-grown plants. Direct shoot regeneration from the leaf and flower explants was obtained 21 to 28 days after the initial culture. Among the seven combinations of the growth regulators used for the culture, the most efficient condition for the shoot and root formation from the leaf tissue was obtained when the MS basic medium was supplemented with 0.5 mg L-¹ BA and 1.0 mg L-¹ NAA, and 0.1 mg L-¹ BA and 0.5 mg L-¹ NAA, while the culture using floret tissues was most efficient on the medium supplemented with 0.5 mg L-¹ BA and 0.5 mg L-¹ NAA, and 0.1 mg L-¹ BA and 1.0 mg L-¹ NAA. These results will provide valuable information to help set up an efficient system for a tissue culture of chrysanthemum cv. Jinba to improve one or some of its negative traits in combination with a radiation mutagenesis approach

Distribution of phospholipase C isozymes in various rat tissues and cultured cells

International Nuclear Information System (INIS)

Suh, P.G.; Ryu, S.H.; Choi, W.C.; Lee, K.Y.; Rhee, S.G.

1987-01-01

Monoclonal antibodies prepared against PLC-I or PLC-II enzyme did not cross-react with the other. Using a pair of antibodies which recognizes 2 different antigenic sites on the same molecule, radioimmunoassays were developed for the quantitation of PLC-I and PLC-II in homogenates of various tissues and cultured cells, prepared by homogenization in a 2 M KCl buffer. The contents of PLC enzymes were measured in 19 rat tissues, in human platelets and in 17 cultured cells. Results indicate that the concentration of PLC-I and PLC-II is very high in brain, PLC-I is localized mainly in brain and partly in seminal vesicles, PLC-II is found in most tissues and cells. PLC-I is highly localized even in brain: 5 different neuroblastoma did not contain PLC-I while 2 glioma and 1 astrocytoma contained significant amounts

Chronic treatment with epidermal growth factor induces growth of the rat ventral prostate

DEFF Research Database (Denmark)

Tørring, N; Jensen, L V; Wen, J G

2001-01-01

the hyperplastic growth phase of the prostate in newborn rats.MATERIAL AND METHODS: Newborn rats were treated for 8 weeks with EGF (150 microg/kg body weight per day), administered as daily subcutaneous injections. Sections of the prostate tissue were examined by a stereological technique to determine tissue......OBJECTIVE: The epidermal growth factor (EGF) system is expressed in the rat prostate, and growth factors from this system induce proliferation in prostate epithelial and stromal cell cultures. The aim of the study was to investigate the possible growth-promoting effects of the system during...... of the prostate epithelium, the stroma and the lumen following EGF treatment, in a pattern resembling physiological growth of the ventral prostate. A significant correlation (r = 0.78, p

Glioma tissue obtained by modern ultrasonic aspiration with a simple sterile suction trap for primary cell culture and pathological evaluation.

Science.gov (United States)

Schroeteler, Juliane; Reeker, Ralf; Suero Molina, Eric; Brokinkel, Benjamin; Holling, Markus; Grauer, Oliver M; Senner, Volker; Stummer, Walter; Ewelt, Christian

2014-01-01

Ultrasonic aspiration is widely used in the resection of brain tumors. Nevertheless, tumor tissue fragments obtained by ultrasonic aspiration are usually discarded. In this study, we demonstrate that these fragments are possible sources of material for histopathological study and tissue culture and compare their microscopic features and viability in tissue culture of cavitron ultrasonic surgical aspirator tissue fragments. Brain tumor tissue collected by ultrasonic aspiration (CUSA EXcel®; Integra Radionics Inc.) in a simple sterile suction trap during resection was processed for primary cell culture. Cell viability and immunohistological markers were measured by the WST-1 test, microscopy and immunofluorescent evaluation. Six gliomas are presented to demonstrate that these tissue fragments show good preservation of histological detail and tissue viability in culture. Utilization of this material may facilitate pathological interpretation by providing a more representative sample of tumor histology as well as an adequate and sterile biosource of material for tissue culture studies.

Some growth factors in neoplastic tissues of brain tumors of different histological structure

Directory of Open Access Journals (Sweden)

O. I. Kit

2016-01-01

Full Text Available Introduction. Pathologic angiogenesis is typical for angiogenic diseases including tumor growth. Vascular endothelial growth factor (VEGF, fibroblast growth factor (FGF, transforming growth factor alpha and beta (which are also known as “triggers” of angiogenesis, and other factors (Gacche, Meshram, 2013; Nijaguna et al., 2015 play a special role in its development. Evaluation of the important mechanisms of angiogenesis in physiological and pathological conditions remains to be a subject of heightened interest for the past 30 years. It is known that VEGF A is the main trigger of growing blood vessels into the tumor tissue. This is specific mitogen signal for endothelial cells that triggers the mechanisms of cell division and migration. VEGF-induced tumor vasculature has a number of structural and functional features that provide growth and progression of tumors, including increased permeability of blood vessels and their chaotic arrangement.Objective: to study in comparative aspect the level of certain growth factors in the following tissues: glioblastomas, brain metastasis of the breast cancer, meningiomas as well as corresponding peritumoral areas.Materials and methods. Tissue samples were obtained from 56 patients admitted to the surgical treatment in Rostov Research Institute of Oncology: 24 patients had glioblastomas, 19 patients had brain metastasis of the breast cancer, 13 patients with meningiomas without peritumoral edema. Histological control was carried out in all cases. Age of patients ranged from 35 to 72 years. The level of growth factor was detected in the samples of tumor tissue and regions immediately adjacent to the tumor foci (peritumoral area by the method of immunoassay and using standard test systems. The following growth factor were detected: VEGF-A and its receptors VEGF-R1 (BenderMedSystem, Austria, VEGF-C and its receptor VEGF-R3 (BenderMedSystem, Austria, EGF (Biosource, USA, IFR-1 and IFR-2 (Mediagnost, USA, TGF

Mechanotransduction mechanisms in growing spherically structured tissues

Science.gov (United States)

Littlejohns, Euan; Dunlop, Carina M.

2018-04-01

There is increasing experimental interest in mechanotransduction in multi-cellular tissues as opposed to single cells. This is driven by a growing awareness of the importance of physiologically relevant three-dimensional culture and of cell–cell and cell–gel interactions in directing growth and development. The paradigm biophysical technique for investigating tissue level mechanobiology in this context is to grow model tissues in artificial gels with well-defined mechanical properties. These studies often indicate that the stiffness of the encapsulating gel can significantly alter cellular behaviours. We demonstrate here potential mechanisms linking tissue growth with stiffness-mediated mechanotransduction. We show how tissue growth in gel systems generates points at which there is a significant qualitative change in the cellular stress and strain experienced. We show analytically how these potential switching points depend on the mechanical properties of the constraining gel and predict when they will occur. Significantly, we identify distinct mechanisms that act separately in each of the stress and strain fields at different times. These observations suggest growth as a potential physical mechanism coupling gel stiffness with cellular mechanotransduction in three-dimensional tissues. We additionally show that non-proliferating areas, in the case that the constraining gel is soft compared with the tissue, will expand and contract passively as a result of growth. Central compartment size is thus seen to not be a reliable indicator on its own for growth initiation or active behaviour.

How-To-Do-It: Using Cauliflower to Demonstrate Plant Tissue Culture.

Science.gov (United States)

Haldeman, Janice H.; Ellis, Jane P.

1988-01-01

Presents techniques used for disinfestation of plant material, preparation of equipment and media, and laboratory procedures for tissue culture using cauliflower. Details methods for preparing solutions and plant propagation by cloning. (CW)

Biomaterials for the programming of cell growth in oral tissues: The possible role of APA.

Science.gov (United States)

Salerno, Marco; Giacomelli, Luca; Larosa, Claudio

2011-01-06

Examples of programmed tissue response after the interaction of cells with biomaterials are a hot topic in current dental research. We propose here the use of anodic porous alumina (APA) for the programming of cell growth in oral tissues. In particular, APA may trigger cell growth by the controlled release of specific growth factors and/or ions. Moreover, APA may be used as a scaffold to promote generation of new tissue, due to the high interconnectivity of pores and the high surface roughness displayed by this material.

Discrepancy between growth of Coccidioides immitis in bacterial blood culture media and a radiometric growth index

International Nuclear Information System (INIS)

Ampel, N.M.; Wieden, M.A.

1988-01-01

Spherules of Coccidioides immitis grew readily after inoculation in vented trypticase soy broth, biphasic brain heart infusion media, and aerobic tryptic soy broth bottles used in a radiometric system (BACTEC). However, visible growth was not accompanied by a significant radiometric growth index. Growth of C. immitis can be visually detected in routine bacterial blood culture media while the radiometric growth index remains negative

Effects of culture systems on growth and economic performance of ...

African Journals Online (AJOL)

The effect of culture system on growth and economics performance of Orechromis niloticus (Nile tilapia) in concrete tanks was investigated. Four outdoor concrete tanks measuring 2.5 x 2 m was used for the study for 24 weeks culture period. The culture systems included the use of algae only at the stocking rates of 4 ...

Endoplasmic Reticulum-Associated rht-PA Processing in CHO Cells: Influence of Mild Hypothermia and Specific Growth Rates in Batch and Chemostat Cultures.

Directory of Open Access Journals (Sweden)

Mauricio Vergara

Full Text Available Chinese hamster ovary (CHO cells are the main host for producing recombinant proteins with human therapeutic applications mainly because of their capability to perform proper folding and glycosylation processes. In addition, mild hypothermia is one of the main strategies for maximising the productivity of these systems. However, little information is available on the effect of culture temperature on the folding and degradation processes of recombinant proteins that takes place in the endoplasmic reticulum.In order to evaluate the effect of the mild hypothermia on processing/endoplasmatic reticulum-associated degradation (ERAD processes, batch cultures of CHO cells producing recombinant human tissue plasminogen activator (rht-PA were carried out at two temperatures (37°C and 33°C and treated with specific inhibitors of glycosylation and ERAD I (Ubiquitin/Proteasome system or ERAD II (Autophagosoma/Lisosomal system pathways. The effect of mild hypothermia was analysed separately from its indirect effect on specific cell growth rate. To do this, chemostat cultures were carried out at the same incubation conditions as the batch cultures, controlling cell growth at high (0.017 h-1 and low (0.012 h-1 dilution rates. For a better understanding of the investigated phenomenon, cell behaviour was also analysed using principal component analysis (PCA.Results suggest that rht-PA is susceptible to degradation by both ERAD pathways studied, revealing that processing and/or ERAD processes are sensitive to temperature cultivation in batch culture. Moreover, by isolating the effect of culture temperature from the effect of cell growth rate verifyed by using chemostat cultures, we have found that processing and/or ERAD processes are more sensitive to reduction in specific growth rate than low temperature, and that temperature reduction may have a positive effect on protein processing. Interestingly, PCA indicated that the integrated performance displayed by CHO

Protective ventilation reduces Pseudomonas aeruginosa growth in lung tissue in a porcine pneumonia model.

Science.gov (United States)

Sperber, Jesper; Nyberg, Axel; Lipcsey, Miklos; Melhus, Åsa; Larsson, Anders; Sjölin, Jan; Castegren, Markus

2017-08-31

Mechanical ventilation with positive end expiratory pressure and low tidal volume, i.e. protective ventilation, is recommended in patients with acute respiratory distress syndrome. However, the effect of protective ventilation on bacterial growth during early pneumonia in non-injured lungs is not extensively studied. The main objectives were to compare two different ventilator settings on Pseudomonas aeruginosa growth in lung tissue and the development of lung injury. A porcine model of severe pneumonia was used. The protective group (n = 10) had an end expiratory pressure of 10 cm H 2 O and a tidal volume of 6 ml x kg -1 . The control group (n = 10) had an end expiratory pressure of 5 cm H 2 O and a tidal volume of 10 ml x kg -1 . 10 11 colony forming units of Pseudomonas aeruginosa were inoculated intra-tracheally at baseline, after which the experiment continued for 6 h. Two animals from each group received only saline, and served as sham animals. Lung tissue samples from each animal were used for bacterial cultures and wet-to-dry weight ratio measurements. The protective group displayed lower numbers of Pseudomonas aeruginosa (p protective group was unchanged (p protective ventilation with lower tidal volume and higher end expiratory pressure has the potential to reduce the pulmonary bacterial burden and the development of lung injury.

Long-term culture of human liver tissue with advanced hepatic functions.

Science.gov (United States)

Ng, Soon Seng; Xiong, Anming; Nguyen, Khanh; Masek, Marilyn; No, Da Yoon; Elazar, Menashe; Shteyer, Eyal; Winters, Mark A; Voedisch, Amy; Shaw, Kate; Rashid, Sheikh Tamir; Frank, Curtis W; Cho, Nam Joon; Glenn, Jeffrey S

2017-06-02

A major challenge for studying authentic liver cell function and cell replacement therapies is that primary human hepatocytes rapidly lose their advanced function in conventional, 2-dimensional culture platforms. Here, we describe the fabrication of 3-dimensional hexagonally arrayed lobular human liver tissues inspired by the liver's natural architecture. The engineered liver tissues exhibit key features of advanced differentiation, such as human-specific cytochrome P450-mediated drug metabolism and the ability to support efficient infection with patient-derived inoculums of hepatitis C virus. The tissues permit the assessment of antiviral agents and maintain their advanced functions for over 5 months in culture. This extended functionality enabled the prediction of a fatal human-specific hepatotoxicity caused by fialuridine (FIAU), which had escaped detection by preclinical models and short-term clinical studies. The results obtained with the engineered human liver tissue in this study provide proof-of-concept determination of human-specific drug metabolism, demonstrate the ability to support infection with human hepatitis virus derived from an infected patient and subsequent antiviral drug testing against said infection, and facilitate detection of human-specific drug hepatotoxicity associated with late-onset liver failure. Looking forward, the scalability and biocompatibility of the scaffold are also ideal for future cell replacement therapeutic strategies.

Mass Spectrometry-Based Proteomics in Molecular Diagnostics: Discovery of Cancer Biomarkers Using Tissue Culture

Directory of Open Access Journals (Sweden)

Debasish Paul

2013-01-01

Full Text Available Accurate diagnosis and proper monitoring of cancer patients remain a key obstacle for successful cancer treatment and prevention. Therein comes the need for biomarker discovery, which is crucial to the current oncological and other clinical practices having the potential to impact the diagnosis and prognosis. In fact, most of the biomarkers have been discovered utilizing the proteomics-based approaches. Although high-throughput mass spectrometry-based proteomic approaches like SILAC, 2D-DIGE, and iTRAQ are filling up the pitfalls of the conventional techniques, still serum proteomics importunately poses hurdle in overcoming a wide range of protein concentrations, and also the availability of patient tissue samples is a limitation for the biomarker discovery. Thus, researchers have looked for alternatives, and profiling of candidate biomarkers through tissue culture of tumor cell lines comes up as a promising option. It is a rich source of tumor cell-derived proteins, thereby, representing a wide array of potential biomarkers. Interestingly, most of the clinical biomarkers in use today (CA 125, CA 15.3, CA 19.9, and PSA were discovered through tissue culture-based system and tissue extracts. This paper tries to emphasize the tissue culture-based discovery of candidate biomarkers through various mass spectrometry-based proteomic approaches.

Mass Spectrometry-Based Proteomics in Molecular Diagnostics: Discovery of Cancer Biomarkers Using Tissue Culture

Science.gov (United States)

Paul, Debasish; Kumar, Avinash; Gajbhiye, Akshada; Santra, Manas K.; Srikanth, Rapole

2013-01-01

Accurate diagnosis and proper monitoring of cancer patients remain a key obstacle for successful cancer treatment and prevention. Therein comes the need for biomarker discovery, which is crucial to the current oncological and other clinical practices having the potential to impact the diagnosis and prognosis. In fact, most of the biomarkers have been discovered utilizing the proteomics-based approaches. Although high-throughput mass spectrometry-based proteomic approaches like SILAC, 2D-DIGE, and iTRAQ are filling up the pitfalls of the conventional techniques, still serum proteomics importunately poses hurdle in overcoming a wide range of protein concentrations, and also the availability of patient tissue samples is a limitation for the biomarker discovery. Thus, researchers have looked for alternatives, and profiling of candidate biomarkers through tissue culture of tumor cell lines comes up as a promising option. It is a rich source of tumor cell-derived proteins, thereby, representing a wide array of potential biomarkers. Interestingly, most of the clinical biomarkers in use today (CA 125, CA 15.3, CA 19.9, and PSA) were discovered through tissue culture-based system and tissue extracts. This paper tries to emphasize the tissue culture-based discovery of candidate biomarkers through various mass spectrometry-based proteomic approaches. PMID:23586059

Growth of melanocytes in human epidermal cell cultures

International Nuclear Information System (INIS)

Staiano-Coico, L.; Hefton, J.M.; Amadeo, C.; Pagan-Charry, I.; Madden, M.R.; Cardon-Cardo, C.

1990-01-01

Epidermal cell cultures were grown in keratinocyte-conditioned medium for use as burn wound grafts; the melanocyte composition of the grafts was studied under a variety of conditions. Melanocytes were identified by immunohistochemistry based on a monoclonal antibody (MEL-5) that has previously been shown to react specifically with melanocytes. During the first 7 days of growth in primary culture, the total number of melanocytes in the epidermal cultures decreased to 10% of the number present in normal skin. Beginning on day 2 of culture, bipolar melanocytes were present at a mean cell density of 116 +/- 2/mm2; the keratinocyte to melanocyte ratio was preserved during further primary culture and through three subpassages. Moreover, exposure of cultures to mild UVB irradiation stimulated the melanocytes to proliferate, suggesting that the melanocytes growing in culture maintained their responsiveness to external stimuli. When the sheets of cultured cells were enzymatically detached from the plastic culture flasks before grafting, melanocytes remained in the basal layer of cells as part of the graft applied to the patient

Effect of Aflatoxin B1 on Growth of Bovine Mammary Epithelial Cells in 3D and Monolayer Culture System.

Science.gov (United States)

Forouharmehr, Ali; Harkinezhad, Taher; Qasemi-Panahi, Babak

2013-01-01

Many studies have been showed transfer of aflatoxins, toxins produced by Aspergillus flvaus and Aspergillus parasiticus fungi, into milk. These toxins are transferred into the milk through digestive system by eating contaminated food. Due to the toxicity of these materials, it seems that it has side effects on the growth of mammary cells. Therefore, the present work aimed to investigate possible toxic effects of aflatoxin B1 (AFB1) on bovine mammary epithelial cells in monolayer and three-dimensional cultures. Specimens of the mammary tissue of bovine were sized out in size 2×2 cm in slaughterhouse. After disinfection and washing in sterile PBS, primary cell culture was performed by enzymatic digestion of tissue with collagenase. When proper numbers of cells were achieved in monolayer culture, cells were seeded in a 24-well culture plate for three-dimensional (3D) culture in Matrigel matrix. After 21 days of 3D culture and reaching the required number of cells, the concentrations of 15, 25 and 35 µL of AFB1 were added to the culture in quadruplicate and incubated for 8 hours. Cellular cytotoxicity was examined using standard colorimetric assay and finally, any change in the morphology of the cells was studied by microscopic technique. Microscopic investigations showed necrosis of the AFB1-exposed cells compared to the control cells. Also, bovine mammary epithelial cells were significantly affected by AFB1 in dose and time dependent manner in cell viability assays. According to the results, it seems that AFB1 can induce cytotoxicity and necrosis in bovine mammary epithelial cells.

Three-dimensional spheroid culture targeting versatile tissue bioassays using a PDMS-based hanging drop array.

Science.gov (United States)

Kuo, Ching-Te; Wang, Jong-Yueh; Lin, Yu-Fen; Wo, Andrew M; Chen, Benjamin P C; Lee, Hsinyu

2017-06-29

Biomaterial-based tissue culture platforms have emerged as useful tools to mimic in vivo physiological microenvironments in experimental cell biology and clinical studies. We describe herein a three-dimensional (3D) tissue culture platform using a polydimethylsiloxane (PDMS)-based hanging drop array (PDMS-HDA) methodology. Multicellular spheroids can be achieved within 24 h and further boosted by incorporating collagen fibrils in PDMS-HDA. In addition, the spheroids generated from different human tumor cells exhibited distinct sensitivities toward drug chemotherapeutic agents and radiation as compared with two-dimensional (2D) cultures that often lack in vivo-like biological insights. We also demonstrated that multicellular spheroids may enable key hallmarks of tissue-based bioassays, including drug screening, tumor dissemination, cell co-culture, and tumor invasion. Taken together, these results offer new opportunities not only to achieve the active control of 3D multicellular spheroids on demand, but also to establish a rapid and cost-effective platform to study anti-cancer therapeutics and tumor microenvironments.

Establishment and long-term culture of the cell lines derived from gonad tissues of Siberian sturgeon (Acipenser baerii

Directory of Open Access Journals (Sweden)

Jun Hyung Ryu

2016-06-01

Full Text Available Abstract To culture germline stem cells in vitro, establishment of the cell lines that can be used as the feeder cells is a prerequisite. In this study, we tried to establish gonad-derived cell lines in Siberian sturgeon (Acipenser baerii. Five 1-year-old A. baerii were used as a donor of gonad tissues, and gonad-dissociated cells were cultured in vitro. Subsequently, determination of growth conditions, long-term culture, characterization, and cryopreservation of the cell lines were also conducted. Five gonad-derived cell lines were stably established and cultured continuously over at least the 73th passage and 402 culture days under the media containing 20 % fetal bovine serum at 28 °C. All cell lines consisted of two main cell types based on morphology even if the ratio of the two cell types was different depending on cell lines. Despite long-term culture, all cell lines maintained diploid DNA contents and expression of several genes that are known to express in the A. baerii gonad. After freezing and thawing of the cell lines, post-thaw cell viabilities between 57.6 and 92.9 % depending on cell lines were indentified, suggesting that stable cryopreservation is possible. The results and the cell lines established in this study will contribute to the development of an in vitro system for A. baerii germline stem cell culture.

Enhancing proliferation and optimizing the culture condition for human bone marrow stromal cells using hypoxia and fibroblast growth factor-2

Directory of Open Access Journals (Sweden)

Jung-Seok Lee

2018-04-01

Full Text Available This study aimed to determine the cellular characteristics and behaviors of human bone marrow stromal cells (hBMSCs expanded in media in a hypoxic or normoxic condition and with or without fibroblast growth factor-2 (FGF-2 treatment. hBMSCs isolated from the vertebral body and expanded in these four groups were evaluated for cellular proliferation/migration, colony-forming units, cell-surface characterization, in vitro differentiation, in vivo transplantation, and gene expression. Culturing hBMSCs using a particular environmental factor (hypoxia and with the addition of FGF-2 increased the cellular proliferation rate while enhancing the regenerative potential, modulated the multipotency-related processes (enhanced chondrogenesis-related processes/osteogenesis, but reduced adipogenesis, and increased cellular migration and collagen formation. The gene expression levels in the experimental samples showed activation of the hypoxia-inducible factor-1 pathway and glycolysis in the hypoxic condition, with this not being affected by the addition of FGF-2. The concurrent application of hypoxia and FGF-2 could provide a favorable condition for culturing hBMSCs to be used in clinical applications associated with bone tissue engineering, due to the enhancement of cellular proliferation and regenerative potential. Keywords: Bone marrow stromal cells, Hypoxia, Fibroblast growth factor, Tissue regeneration, Microenvironment interactions

ALLOMETRIC GROWTH OF PRIMAL CUTS AND TISSUES IN THE PIG

Directory of Open Access Journals (Sweden)

Frank Siewerdt

1994-12-01

Full Text Available Data from 82 purebred and crossbred Large White and Duroc barrows and gilts were used to describe the growth of carcass primal cuts, of tissues, and of several organs. Pigs were allowed ad libitum to a conventional diet, which contained com and soybean meal. Pigs were weighted weekly and were slaughtered when attained a liveweight over 90kg. An allometric pattern of growth was assumed. Within the observed range of liveweight, the carcass grew slower than the whole animal. An increase of carcass weight corresponds to a similar increase of lean, but also corresponds to a larger increase of fat tissues. A suggestion to slaughter pigs near to 90kg of liveweight is presented, in order to obtain leaner carcasses.

Cost-effective nutrient sources for tissue culture of cassava ( Manihot ...

African Journals Online (AJOL)

Application of tissue culture technology is constrained by high costs making seedlings unaffordable. The objective of this study was to evaluate the possibility of using locally available fertilizers as alternative nutrient sources for cassava micropropagation. A Low Cost Medium (LCM) whereby the conventional sources of four ...

Studies on the use of gamma irradiation and tissue culture in improving brassica napus

International Nuclear Information System (INIS)

Khedr, E.K.A.

2012-01-01

The objectives of this study were to:1- Studying the effect of different doses of gamma rays on some growth and yield component traits of three Brassica napus cultivars (Serow6, Serow4 and Pactol) during four consecutive generations aiming to create new genotypes characterized with high yielding traits. 2- Studying the effect of different doses of gamma rays on in vitro biotechnology technique (tissue culture) used in improving Brassica napus. Seeds of three Brassica napus cultivars were irradiated with different gamma ray doses then sown for four consecutive seasons. Data were collected and recorded to clarify the effect gamma irradiation on some yield component traits which were days to flowering , plant height, number of main branches per plant, number of secondary branches per plant, number of pods per plant, number of seeds per pod, weight of 1000-seed, weight of grain yield/plant and oil content of seeds). Results showed that high doses of gamma radiation had enhanced all of the studied traits for each of the three tested cultivars (except the plant height trait for Serow6 and Pactol cultivars). Seven new mutant lines were selected for their superiority in one or more of the studied yield component traits. Regarding the effect of gamma rays on tissue culture techniques, the applied gamma radiation doses did not affect the percentage of seed germination of the three studied cultivars, whereas the percentage of callus induction decreased by increasing the dose of gamma rays for each of the three cultivars and in both types of explants (hypocotyl and cotyledons) used in this experiment.

Regulation by basic fibroblast growth factor of glycosaminoglycan biosynthesis in cultured vascular endothelial cells.

Science.gov (United States)

Kaji, T; Hiraga, S; Ohkawara, S; Inada, M; Yamamoto, C; Kozuka, H; Koizumi, F

1995-05-01

The alteration of glycosaminoglycans (GAGs) in cultured bovine aortic endothelial cells after exposure to basic fibroblast growth factor (bFGF) was investigated. It was found that the incorporation of [3H]glucosamine into GAGs was markedly increased by bFGF in both the cell layer and the conditioned medium; however, that of [35S]sulfate was not changed by the growth factor. These results indicated that bFGF enhanced the sugar-chain formation but did not affect their sulfation in endothelial GAG production. Similar changes were observed in either bovine aortic smooth-muscle cells and human fibroblastic IMR-90 cells to greater and lesser degrees, respectively. Characterization of GAGs in the endothelial cell layer and the conditioned medium revealed that bFGF enhanced both heparan sulfate and the other GAGs to a similar degree. The present data suggest that bFGF may be involved in the regulation of the blood coagulation system via altering GAGs of the vascular tissue when the endothelium was damaged.

Articular cartilage tissue engineering with plasma-rich in growth factors and stem cells with nano scaffolds

Science.gov (United States)

Montaser, Laila M.; Abbassy, Hadeer A.; Fawzy, Sherin M.

2016-09-01

The ability to heal soft tissue injuries and regenerate cartilage is the Holy Grail of musculoskeletal medicine. Articular cartilage repair and regeneration is considered to be largely intractable due to the poor regenerative properties of this tissue. 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. However, current methods do not perfectly restore hyaline cartilage and may lead to the apparition of fibro- or continue hypertrophic cartilage. The lack of efficient modalities of treatment has prompted research into tissue engineering combining stem cells, scaffold materials and environmental factors. The field of articular cartilage tissue engineering, which aims to repair, regenerate, and/or improve injured or diseased cartilage functionality, has evoked intense interest and holds great potential for improving cartilage therapy. Plasma-rich in growth factors (PRGF) and/or stem cells may be effective for tissue repair as well as cartilage regenerative processes. There is a great promise to advance current cartilage therapies toward achieving a consistently successful approach for addressing cartilage afflictions. Tissue engineering may be the best way to reach this objective via the use of stem cells, novel biologically inspired scaffolds and, emerging nanotechnology. In this paper, current and emergent approach in the field of cartilage tissue engineering is presented for specific application. In the next years, the development of new strategies using stem cells, in scaffolds, with supplementation of culture medium could improve the quality of new formed cartilage.

Anaerobic Cultures from Preserved Tissues of Baby Mammoth

Science.gov (United States)

Pikuta, Elena V.; Hoover, Richard B.; Fisher, Daniel

2011-01-01

Microbiological analysis of several cold-preserved tissue samples from the Siberian baby mammoth known as Lyuba revealed a number of culturable bacterial strains that were grown on anaerobic media at 4 C. Lactic acid produced by LAB (lactic acid bacteria) group, usually by members of the genera Carnobacterium and Lactosphera, appears to be a wonderful preservative that prevents other bacteria from over-dominating a system. Permafrost and lactic acid preserved the body of this one-month old baby mammoth and kept it in exceptionally good condition, resulting in this mammoth being the most complete such specimen ever recovered. The diversity of novel anaerobic isolates was expressed on morphological, physiological and phylogenetic levels. Here we discuss the specifics of the isolation of new strains, differentiation from trivial contamination, and preliminary results for the characterization of cultures.

The gene expression profile of non-cultured, highly purified human adipose tissue pericytes: Transcriptomic evidence that pericytes are stem cells in human adipose tissue

Energy Technology Data Exchange (ETDEWEB)

Silva Meirelles, Lindolfo da, E-mail: lindolfomeirelles@gmail.com [Center for Cell-Based Therapy (CEPID/FAPESP), Regional Center for Hemotherapy of Ribeirão Preto, University of São Paulo, Rua Tenente Catão Roxo 2501, 14051-140 Ribeirão Preto, SP (Brazil); Laboratory for Stem Cells and Tissue Engineering, PPGBioSaúde, Lutheran University of Brazil, Av. Farroupilha 8001, 92425-900 Canoas, RS (Brazil); Deus Wagatsuma, Virgínia Mara de; Malta, Tathiane Maistro; Bonini Palma, Patrícia Viana [Center for Cell-Based Therapy (CEPID/FAPESP), Regional Center for Hemotherapy of Ribeirão Preto, University of São Paulo, Rua Tenente Catão Roxo 2501, 14051-140 Ribeirão Preto, SP (Brazil); Araújo, Amélia Goes; Panepucci, Rodrigo Alexandre [Laboratory of Large-Scale Functional Biology (LLSFBio), Regional Center for Hemotherapy of Ribeirão Preto, University of São Paulo, Rua Tenente Catão Roxo 2501, 14051-140 Ribeirão Preto, SP (Brazil); and others

2016-12-10

Pericytes (PCs) are a subset of perivascular cells that can give rise to mesenchymal stromal cells (MSCs) when culture-expanded, and are postulated to give rise to MSC-like cells during tissue repair in vivo. PCs have been suggested to behave as stem cells (SCs) in situ in animal models, although evidence for this role in humans is lacking. Here, we analyzed the transcriptomes of highly purified, non-cultured adipose tissue (AT)-derived PCs (ATPCs) to detect gene expression changes that occur as they acquire MSC characteristics in vitro, and evaluated the hypothesis that human ATPCs exhibit a gene expression profile compatible with an AT SC phenotype. The results showed ATPCs are non-proliferative and express genes characteristic not only of PCs, but also of AT stem/progenitor cells. Additional analyses defined a gene expression signature for ATPCs, and revealed putative novel ATPC markers. Almost all AT stem/progenitor cell genes differentially expressed by ATPCs were not expressed by ATMSCs or culture-expanded ATPCs. Genes expressed by ATMSCs but not by ATPCs were also identified. These findings strengthen the hypothesis that PCs are SCs in vascularized tissues, highlight gene expression changes they undergo as they assume an MSC phenotype, and provide new insights into PC biology. - Highlights: • Non-cultured adipose tissue-derived human pericytes (ncATPCs) exhibit a distinctive gene expression signature. • ncATPCs express key adipose tissue stem cell genes previously described in vivo in mice. • ncATPCs express message for anti-proliferative and antiangiogenic molecules. • Most ncATPC-specific transcripts are absent in culture-expanded pericytes or ATMSCs • Gene expression changes ncATPCs undergo as they acquire a cultured ATMSC phenotype are pointed out.

The gene expression profile of non-cultured, highly purified human adipose tissue pericytes: Transcriptomic evidence that pericytes are stem cells in human adipose tissue

International Nuclear Information System (INIS)

Silva Meirelles, Lindolfo da; Deus Wagatsuma, Virgínia Mara de; Malta, Tathiane Maistro; Bonini Palma, Patrícia Viana; Araújo, Amélia Goes; Panepucci, Rodrigo Alexandre

2016-01-01

Pericytes (PCs) are a subset of perivascular cells that can give rise to mesenchymal stromal cells (MSCs) when culture-expanded, and are postulated to give rise to MSC-like cells during tissue repair in vivo. PCs have been suggested to behave as stem cells (SCs) in situ in animal models, although evidence for this role in humans is lacking. Here, we analyzed the transcriptomes of highly purified, non-cultured adipose tissue (AT)-derived PCs (ATPCs) to detect gene expression changes that occur as they acquire MSC characteristics in vitro, and evaluated the hypothesis that human ATPCs exhibit a gene expression profile compatible with an AT SC phenotype. The results showed ATPCs are non-proliferative and express genes characteristic not only of PCs, but also of AT stem/progenitor cells. Additional analyses defined a gene expression signature for ATPCs, and revealed putative novel ATPC markers. Almost all AT stem/progenitor cell genes differentially expressed by ATPCs were not expressed by ATMSCs or culture-expanded ATPCs. Genes expressed by ATMSCs but not by ATPCs were also identified. These findings strengthen the hypothesis that PCs are SCs in vascularized tissues, highlight gene expression changes they undergo as they assume an MSC phenotype, and provide new insights into PC biology. - Highlights: • Non-cultured adipose tissue-derived human pericytes (ncATPCs) exhibit a distinctive gene expression signature. • ncATPCs express key adipose tissue stem cell genes previously described in vivo in mice. • ncATPCs express message for anti-proliferative and antiangiogenic molecules. • Most ncATPC-specific transcripts are absent in culture-expanded pericytes or ATMSCs • Gene expression changes ncATPCs undergo as they acquire a cultured ATMSC phenotype are pointed out.

Application of Tissue Culture and Transformation Techniques in Model Species Brachypodium distachyon.

Science.gov (United States)

Sogutmaz Ozdemir, Bahar; Budak, Hikmet

2018-01-01

Brachypodium distachyon has recently emerged as a model plant species for the grass family (Poaceae) that includes major cereal crops and forage grasses. One of the important traits of a model species is its capacity to be transformed and ease of growing both in tissue culture and in greenhouse conditions. Hence, plant transformation technology is crucial for improvements in agricultural studies, both for the study of new genes and in the production of new transgenic plant species. In this chapter, we review an efficient tissue culture and two different transformation systems for Brachypodium using most commonly preferred gene transfer techniques in plant species, microprojectile bombardment method (biolistics) and Agrobacterium-mediated transformation.In plant transformation studies, frequently used explant materials are immature embryos due to their higher transformation efficiencies and regeneration capacity. However, mature embryos are available throughout the year in contrast to immature embryos. We explain a tissue culture protocol for Brachypodium using mature embryos with the selected inbred lines from our collection. Embryogenic calluses obtained from mature embryos are used to transform Brachypodium with both plant transformation techniques that are revised according to previously studied protocols applied in the grasses, such as applying vacuum infiltration, different wounding effects, modification in inoculation and cocultivation steps or optimization of bombardment parameters.

In vitro differentiation of rat spermatogonia into round spermatids in tissue culture.

Science.gov (United States)

Reda, A; Hou, M; Winton, T R; Chapin, R E; Söder, O; Stukenborg, J-B

2016-09-01

Do the organ culture conditions, previously defined for in vitro murine male germ cell differentiation, also result in differentiation of rat spermatogonia into post-meiotic germ cells exhibiting specific markers for haploid germ cells? We demonstrated the differentiation of rat spermatogonia into post-meiotic cells in vitro, with emphasis on exhibiting, protein markers described for round spermatids. Full spermatogenesis in vitro from immature germ cells using an organ culture technique in mice was first reported 5 years ago. However, no studies reporting the differentiation of rat spermatogonia into post-meiotic germ cells exhibiting the characteristic protein expression profile or into functional sperm have been reported. Organ culture of testicular fragments of 5 days postpartum (dpp) neonatal rats was performed for up to 52 days. Evaluation of microscopic morphology, testosterone levels, mRNA and protein expression as measured by RT-qPCR and immunostaining were conducted to monitor germ cell differentiation in vitro. Potential effects of melatonin, Glutamax® medium, retinoic acid and the presence of epidydimal fat tissue on the spermatogenic process were evaluated. A minimum of three biological replicates were performed for all experiments presented in this study. One-way ANOVA, ANOVA on ranks and student's t-test were applied to perform the statistical analysis. Male germ cells, present in testicular tissue pieces grown from 5 dpp rats, exhibited positive protein expression for Acrosin and Crem (cAMP (cyclic adenosine mono phosphate) response element modulator) after 52 days of culture in vitro. Intra-testicular testosterone production could be observed after 3 days of culture, while when epididymal fat tissue was added, spontaneous contractility of cultured seminiferous tubules could be observed after 21 days. However, no supportive effect of the supplementation with any factor or the co-culturing with epididymal fat tissue on germ cell differentiation in

The Influence of Bioreactor Geometry and the Mechanical Environment on Engineered Tissues

KAUST Repository

Osborne, J. M.; O’ Dea, R. D.; Whiteley, J. P.; Byrne, H. M.; Waters, S. L.

2010-01-01

A three phase model for the growth of a tissue construct within a perfusion bioreactor is examined. The cell population (and attendant extracellular matrix), culture medium, and porous scaffold are treated as distinct phases. The bioreactor system is represented by a two-dimensional channel containing a cell-seeded rigid porous scaffold (tissue construct), which is perfused with a culture medium. Through the prescription of appropriate functional forms for cell proliferation and extracellular matrix deposition rates, the model is used to compare the influence of cell density-, pressure-, and culture medium shear stress-regulated growth on the composition of the engineered tissue. The governing equations are derived in O'Dea et al. "A Three Phase Model for Tissue Construct Growth in a Perfusion Bioreactor," Math. Med. Biol., in which the long-wavelength limit was exploited to aid analysis; here, finite element methods are used to construct two-dimensional solutions to the governing equations and to investigate thoroughly their behavior. Comparison of the total tissue yield and averaged pressures, velocities, and shear stress demonstrates that quantitative agreement between the two-dimensional and long-wavelength approximation solutions is obtained for channel aspect ratios of order 10 -2 and that much of the qualitative behavior of the model is captured in the long-wavelength limit, even for relatively large channel aspect ratios. However, we demonstrate that in order to capture accurately the effect of mechanotransduction mechanisms on tissue construct growth, spatial effects in at least two dimensions must be included due to the inherent spatial variation of mechanical stimuli relevant to perfusion bioreactors, most notably, fluid shear stress, a feature not captured in the long-wavelength limit. Copyright © 2010 by ASME.

The Influence of Bioreactor Geometry and the Mechanical Environment on Engineered Tissues

KAUST Repository

Osborne, J. M.

2010-01-01

A three phase model for the growth of a tissue construct within a perfusion bioreactor is examined. The cell population (and attendant extracellular matrix), culture medium, and porous scaffold are treated as distinct phases. The bioreactor system is represented by a two-dimensional channel containing a cell-seeded rigid porous scaffold (tissue construct), which is perfused with a culture medium. Through the prescription of appropriate functional forms for cell proliferation and extracellular matrix deposition rates, the model is used to compare the influence of cell density-, pressure-, and culture medium shear stress-regulated growth on the composition of the engineered tissue. The governing equations are derived in O\\'Dea et al. "A Three Phase Model for Tissue Construct Growth in a Perfusion Bioreactor," Math. Med. Biol., in which the long-wavelength limit was exploited to aid analysis; here, finite element methods are used to construct two-dimensional solutions to the governing equations and to investigate thoroughly their behavior. Comparison of the total tissue yield and averaged pressures, velocities, and shear stress demonstrates that quantitative agreement between the two-dimensional and long-wavelength approximation solutions is obtained for channel aspect ratios of order 10 -2 and that much of the qualitative behavior of the model is captured in the long-wavelength limit, even for relatively large channel aspect ratios. However, we demonstrate that in order to capture accurately the effect of mechanotransduction mechanisms on tissue construct growth, spatial effects in at least two dimensions must be included due to the inherent spatial variation of mechanical stimuli relevant to perfusion bioreactors, most notably, fluid shear stress, a feature not captured in the long-wavelength limit. Copyright © 2010 by ASME.

Changes In Growth Culture FDA Activity Under Changing Growth Conditions

DEFF Research Database (Denmark)

Jørgensen, Per Elberg; Eriksen, Thomas Juul; Jensen, Bjørn K.

1992-01-01

The FDA hydrolysis capacities and bacterial biomass concentrations (estimated by determination of ATP content) of growth cultures prepared from activated sludge and wastewater, were measured to find out whether the FDA activity would reflect bacterial biomass under different physiological states...... of the bacteria. The FDA activity/ATP ratio was calculated for different concentrations of autoclaved sludge. A faster decay rate of ATP relative to FDA hydrolysis activity was observed, thus causing changes in the ratio. Furthermore, comparison between values obtained from pure cultures and different soils...... revealed differences up to two orders of magnitude of the ratio. Based on these results it was concluded that the FDA activity should not be applied for measurements of viable biomass in environments in which different physiological conditions occur....

[Effective productions of plant secondary metabolites having antitumor activity by plant cell and tissue cultures].

Science.gov (United States)

Taniguchi, Shoko

2005-06-01

Methods for the effective production of plant secondary metabolites with antitumor activity using plant cell and tissue cultures were developed. The factors in tannin productivity were investigated using culture strains producing different types of hydrolyzable tannins, i.e., gallotannins (mixture of galloylglucoses), ellagi-, and dehydroellagitannins. Production of ellagi- and dehydroellagitannins was affected by the concentrations and ratio of nitrogen sources in the medium. The formation of oligomeric ellagitannins in shoots of Oenothera tetraptera was correlated with the differentiation of tissues. Cultured cells of Eriobotrya japonica producing ursane- and oleanane-type triterpenes with antitumor activities were also established.

The influence of tethered epidermal growth factor on connective tissue progenitor colony formation

OpenAIRE

Marcantonio, Nicholas A.; Boehm, Cynthia A.; Rozic, Richard J.; Au, Ada; Wells, Alan; Muschler, George F.; Griffith, Linda G.

2009-01-01

Strategies to combine aspirated marrow cells with scaffolds to treat connective tissue defects are gaining increasing clinical attention and use. In situations such as large defects where initial survival and proliferation of transplanted connective tissue progenitors (CTPs) are limiting, therapeutic outcomes might be improved by using the scaffold to deliver growth factors that promote the early stages of cell function in the graft. Signaling by the epidermal growth factor receptor (EGFR) pl...

Comparison of human dermal fibroblasts (HDFs) growth rate in culture media supplemented with or without basic fibroblast growth factor (bFGF).

Science.gov (United States)

Abdian, Narges; Ghasemi-Dehkordi, Payam; Hashemzadeh-Chaleshtori, Morteza; Ganji-Arjenaki, Mahbobe; Doosti, Abbas; Amiri, Beheshteh

2015-12-01

Basic fibroblast growth factor (bFGF or FGF-2) is a member of the FGF family secreted by different kinds of cells like HDFs and it is an important nutritional factor for cell growth and differentiation. The HDFs release bFGF in culture media at very low. The present study aims to investigate the HDFs growth rate in culture media supplemented either with or without bFGF. In brief, HDFs were isolated from human foreskin sample and were cultured in vitro in media containing bFGF and lack of this factor. The cells growth rate was calculated by trypan blue. The karyotyping was performed using G-banding to investigate the chromosomal abnormality of HDFs in both groups. Total RNA of each groups were extracted and cDNA samples were synthesized then, real-time Q-PCR was used to measure the expression level of p27kip1 and cyclin D1 genes normalized to internal control gene (GAPDH). The karyotype analysis showed that HDFs cultured in media or without bFGF had normal karyotype (46 chromosomes, XY) and chromosomal abnormalities were not observed. The cell growth rates in both groups were normal with proliferated exponentially but the slope of growth curve in HDFs cultured in media containing bFGF was increased. Karyotyp test showed that bFGF does not affect on cytogenetic stability of cells. The survey of p27kip1 and cyclin D1 genes by real-time Q-PCR showed that the expression level of these genes were up-regulated when adding bFGF in culture media (p culture media with growth factor like bFGF could enhance the proliferation and differentiation capacity of cells and improve cells growth rate. Similarly, fibroblast growth factors did not induce any chromosomal abnormality in cells. Furthermore, in HDFs cultured in bFGF supplemented media, the p27kip1 and cyclin D1 genes were up-regulated and suggesting an important role for bFGF in cell-cycle regulation and progression and fibroblast division stimulation. It also suggests that the effects of bFGF on different cell types with

Regeneration and acclimatization of salt-tolerant arachis hypogaea plants through tissue culture

International Nuclear Information System (INIS)

Ghauri, E.G.

2006-01-01

Excised embryos of Arachis hypogaea were cultured on Murashige and Skoog's medium (MS medium) supplemented with different combinations of growth hormones. The highest frequency of callus proliferation (80%) was recorded on MS medium mixed with 1.0 mg/1 of 2,4-D and 0.5 mg/1 of BAP. These cultures were treated with 0.65 mg/l of trans-4-hydroxy-L-proline (HyP) a:1d various concentrations (0.1-0.5%) of NaCl. In all cases the presence of salt reduced the fresh mass of callus. Shoot regeneration in the cultures took place when transferred to MS medium supplemented with 1.0 mg/1 of kinetin (Kin) and 0.5 mg/1 of 6-benzyl aminopurine (BAP). Percentage of shoot regeneration decreased with the increase of NaCl (0.1- 0.5%) in the shoot regeneration medium. Root formation in these cultures took place when the cultures were nurtured on MS medium free of growth hormones. Regeneration, hardening and acclimatization of the salt tolerant plants was conducted. (author)

Study on Batch Culture Growth Model for Lactococcus lactis IO-1

OpenAIRE

Ishizaki, Ayaaki; Ohta, Tomomi; Kobayashi, Genta; 石崎, 文彬; 太田, 智美; 小林, 元太

1991-01-01

L-lactate fermentation employing Lactncoccus lactis IO-1 demonstrated a typical end product inhibition. By numerical analysis of fermentation results of the batch culture of this microorganism, the specific rates for cell growth, substrate consumption and product formation were clearly expressed by the end product inhibition formulae. All constants for those formulae were determined by the fermentation results. A mathematical model for batch culture growth of this microorganism in which the n...

Tissue culture-induced alteration in cytosine methylation in new rice ...

African Journals Online (AJOL)

Zizania DNA introgression could induce a large number of genetic and epigenetic changes of the new rice recombinant inbred lines genome. In this present study, we employed inter-simple sequence repeat (ISSR) to further study the genetic and epigenetic changes that are induced by tissue culture. Changes induced by ...

Culturing the Unculturable: Human Coronavirus HKU1 Infects, Replicates, and Produces Progeny Virions in Human Ciliated Airway Epithelial Cell Cultures

NARCIS (Netherlands)

Pyrc, Krzysztof; Sims, Amy C.; Dijkman, Ronald; Jebbink, Maarten; Long, Casey; Deming, Damon; Donaldson, Eric; Vabret, Astrid; Baric, Ralph; van der Hoek, Lia; Pickles, Raymond

2010-01-01

Culturing newly identified human lung pathogens from clinical sample isolates can represent a daunting task, with problems ranging from low levels of pathogens to the presence of growth suppressive factors in the specimens, compounded by the lack of a suitable tissue culture system. However, it is

Cells in human postmortem brain tissue slices remain alive for several weeks in culture

NARCIS (Netherlands)

Verwer, Ronald W. H.; Hermens, Wim T. J. M. C.; Dijkhuizen, PaulaA; ter Brake, Olivier; Baker, Robert E.; Salehi, Ahmad; Sluiter, Arja A.; Kok, Marloes J. M.; Muller, Linda J.; Verhaagen, Joost; Swaab, Dick F.

2002-01-01

Animal models for human neurological and psychiatric diseases only partially mimic the underlying pathogenic processes. Therefore, we investigated the potential use of cultured postmortem brain tissue from adult neurological patients and controls. The present study shows that human brain tissue

Methylobacterium sp. resides in unculturable state in potato tissues in vitro and becomes culturable after induction by Pseudomonas fluorescens IMGB163.

Science.gov (United States)

Podolich, O; Laschevskyy, V; Ovcharenko, L; Kozyrovska, N; Pirttilä, A M

2009-03-01

To induce growth of endophytic bacteria residing in an unculturable state in tissues of in vitro-grown potato plantlets. To isolate and identify the induced bacteria and to localize the strains in tissues of in vitro-grown potato plantlets. The inoculation of in vitro-grown potato plants with Pseudomonas fluorescens IMBG163 led to induction of another bacterium, a pink-pigmented facultative methylotroph that was identified as Methylobacterium sp. using phylogenetic 16S rDNA approach. Two molecular methods were used for localizing methylobacteria in potato plantlets: PCR and in situ hybridization (ISH/FISH). A PCR product specific for the Methylobacterium genus was found in DNA isolated from the surface-sterilized plantlet leaves. Presence of Methylobacterium rRNA was detected by ISH/FISH in leaves and stems of inoculated as well as axenic potato plantlets although the bacterium cannot be isolated from the axenic plants. Methylobacterium sp. resides in unculturable state within tissues of in vitro-grown potato plants and becomes culturable after inoculation with P. fluorescens IMBG163. In order to develop endophytic biofertilizers and biocontrol agents, a detailed knowledge of the life-style of endophytes is essential. To our knowledge, this is the first report on increase of the culturability of endophytes in response to inoculation by nonpathogenic bacteria.

Advanced cell culture technology for generation of in vivo-like tissue models

Directory of Open Access Journals (Sweden)

Stefan Przyborski

2017-06-01

Full Text Available Human tissues are mostly composed of different cell types, that are often highly organised in relation to each other. Often cells are arranged in distinct layers that enable signalling and cell-to-cell interactions. Here we describe the application of scaffold-based technology, that can be used to create advanced organotypic 3D models of various tissue types that more closely resemble in vivo-like conditions (Knight et al., 2011. The scaffold comprises a highly porous polystyrene material, engineered into a 200 micron thick membrane that is presented in various ways including multi-welled plates and well inserts, for use with conventional culture plasticware and medium perfusion systems. This technology has been applied to generate numerous unique types of co-culture model. For example: 1 a full thickness human skin construct comprising dermal fibroblasts and keratinocytes, raised to the air-liquid interface to induce cornification of the upper layers (Fig.1 (Hill et al., 2015; 2 a neuron-glial co-culture to enable the study of neurite outgrowth interacting with astroglial cells to model and investigate the glial scar found in spinal cord injury (Clarke et al., 2016; 3 formation of a sub-mucosa consisting of a polarised simple epithelium, layer of ECM proteins simulating the basement membrane, and underlying stromal tissues (e.g. intestinal mucosa. These organotypic models demonstrate the versatility of scaffold membranes and the creation of advanced in vivo-like tissue models. Creating a layered arrangement more closely simulates the true anatomy and organisation of cells within many tissue types. The addition of different cell types in a temporal and spatial fashion can be used to study inter-cellular relationships and create more physiologically relevant in vivo-like cell-based assays. Methods that are relatively straightforward to use and that recreate the organised structure of real tissues will become valuable research tools for use in

Canine osteosarcoma karyotypes from an original tumor, its metastasis, and tumor cells in tissue culture

International Nuclear Information System (INIS)

Taylor, N.; Shifrine, M.; Wolf, H.G.; Trommershausen-Smith, A.

1975-01-01

Radiation-induced osteosarcoma, its metastasis, and cells grown in tissue culture were karyotyped. Both hypodiploid and hyperdiploid stem lines were observed. The hypodiploid line contained 45-55 chromosomes with 10 to 15 abnormal metacentric and submetacentric chromosomes and one subtelocentric marker. The hyperdiploid line contained 90 to 105 chromosomes with 20 to 30 abnormal metacentric and submetacentric chromosomes with two subtelocentric markers. Karyotypic analysis can be used to monitor osteosarcomas maintained in tissue culture

Radiation adaptive response for the growth of cultured glial cells

International Nuclear Information System (INIS)

Suzuki, S.; Miura, Y.; Kano, M.; Toda, T.; Urano, S.

2003-01-01

Full text: To examine the molecular mechanism of radiation adaptive response (RAR) for the growth of cultured glial cells and to investigate the influence of aging on the response, glial cells were cultured from young and aged rats (1 month and 24 months old). RAR for the growth of glial cells conditioned with a low dose of X-rays and subsequently exposed to a high dose of X-rays was examined for cell number and BrdU incorporation. Involvement of the subcellular signaling pathway factors in RAR was investigated using their inhibitors, activators and mutated glial cells. RAR was observed in cells cultured from young rats, but was not in cells from aged rats. The inhibitors of protein kinase C (PKC) and DNA-dependent protein kinase (DNA-PK) or phosphatidylinositol 3-kinase (PI3K) suppressed RAR. The activators of PKC instead of low dose irradiation also caused RAR. Moreover, glial cells cultured from severe combined immunodeficiency (scid) mice (CB-17 scid) and ataxia-telangiectasia (AT) cells from AT patients showed no RAR. These results indicated that PKC, ATM, DNAPK and/or PI3K were involved in RAR for growth and BrdU incorporation of cultured glial cells and RAR decreased with aging. Proteomics data of glial cells exposed to severe stress of H 2 O 2 or X-rays also will be presented in the conference since little or no difference has not been observed with slight stress yet

Mass production of 'Swan's Golden', Cupressus sempervirens by tissue culture and its radiation breeding. 2. Study on growth regulating substance to stimulate the elongation of shoot apex

International Nuclear Information System (INIS)

Suda, Hirokatsu

1997-01-01

Some species of 'Swan's Golden' have been propagated by means of grafting because propagation by cutting is difficult. However, not a few species of the plant are easily lodged, resulting that spreading of 'Swan's Golden' is difficult. Here, an investigation was made on growth regulating substances added into the basal medium in respects of their kinds and concentrations and also the kind of medium for successive culture. As the basal medium, SH medium supplemented with 4 g/l of gel-light and 3 g/l of carbon-powder was used. The optimum medium for the initiation of culture was the basal medium added with zeatin (Ze) and gibberellin (GA) and it was also suitable for successive culture after the 14 day. The medium added with Ze and benzyladenine (BA) or Ze alone was found to be most suitable for the culture after the 28 day. The concentration was optimum at 1 - 2.5 mg/l for either of these growth regulating substances. (M.N.)

Homeostatic pressure, tumor growth and fingering of epithelial tissues: Some generic physics arguments

Science.gov (United States)

Risler, Thomas

2011-03-01

We propose that one aspect of homeostasis is the regulation of tissues to preferred pressures, which can lead to a competition for space of purely mechanical origin and be an underlying mechanism for tumor growth. Surface and bulk contributions to pressure lead to the existence of a critical size that must be overcome by metastases to reach macroscopic sizes. This property qualitatively explains the observed size distributions of metastases, while size-independent growth rates cannot account for clinical and experimental data. It also potentially explains the observed preferential growth of metastases on tissue surfaces and membranes, suggests a mechanism underlying the seed and soil hypothesis introduced by Stephen Paget in 1889, and yields realistic values for metastatic inefficiency. Treating epithelial tissues as viscous fluids with effective cell division, we find a novel hydrodynamic instability that leads to the formation of fingering protrusions of the epithelium into the connective tissue. Arising from a combination of viscous friction effects and proliferation of the epithelial cells, this instability provides physical insight into a potential mechanism by which interfaces between epithelia and stroma undulate, and potentially by which tissue dysplasia leads to cancerous invasion. In collaboration with M. Basan, J.-F. Joanny, X. Sastre-Garau and J. Prost.

Connective tissue growth factor immunohistochemical expression is associated with gallbladder cancer progression.

Science.gov (United States)

Garcia, Patricia; Leal, Pamela; Alvarez, Hector; Brebi, Priscilla; Ili, Carmen; Tapia, Oscar; Roa, Juan C

2013-02-01

Gallbladder cancer (GBC) is an aggressive neoplasia associated with late diagnosis, unsatisfactory treatment, and poor prognosis. Molecular mechanisms involved in GBC pathogenesis remain poorly understood. Connective tissue growth factor (CTGF) is thought to play a role in the pathologic processes and is overexpressed in several human cancers, including GBC. No information is available about CTGF expression in early stages of gallbladder carcinogenesis. Objective.- To evaluate the expression level of CTGF in benign and malignant lesions of gallbladder and its correlation with clinicopathologic features and GBC prognosis. Connective tissue growth factor protein was examined by immunohistochemistry on tissue microarrays containing tissue samples of chronic cholecystitis (n = 51), dysplasia (n = 15), and GBC (n = 169). The samples were scored according to intensity of staining as low/absent and high CTGF expressers. Statistical analysis was performed using the χ(2) test or Fisher exact probability test with a significance level of P Connective tissue growth factor expression showed a progressive increase from chronic cholecystitis to dysplasia and then to early and advanced carcinoma. Immunohistochemical expression (score ≥2) was significantly higher in advanced tumors, in comparison with chronic cholecystitis (P < .001) and dysplasia (P = .03). High levels of CTGF expression correlated with better survival (P = .04). Our results suggest a role for CTGF in GBC progression and a positive association with better prognosis. In addition, they underscore the importance of considering the involvement of inflammation on GBC development.

Tissue culture of three species of Laurencia complex

Science.gov (United States)

Shen, Songdong; Wu, Xunjian; Yan, Binlun; He, Lihong

2010-05-01

To establish a micropropagation system of three Laurencia complex species ( Laurencia okamurai, Laurencia tristicha, and Chondrophycus undulatus) by tissue culture techniques, we studied the regeneration characteristics and optimal culture conditions of axenic algal fragments cultured on solid medium and in liquid medium. Regeneration structures were observed and counted regularly under a reverse microscope to investigate the regeneration process, polarity and optimal illumination, and temperature and salinity levels. The results show that in most cultures of the three species, we obtained bud regeneration on solidified medium with 0.5% agar and in liquid medium. Rhizoid-like regeneration was filamentous and developed from the lower cut surface of fragments in L. okamurai, but was discoid and developed from the apical back side of bud regeneration in L. tristicha and C. undulatus. Regeneration polarity was localized to the apical part of algal fronds in all three species, and on fragments cut from the basal part of algae buds could develop from both the upper and the lower cut surfaces. Buds could develop from both the medullary and the cortical portions in L. okamurai and C. undulatus, while in L. tristicha, buds only emerged from the cortex. The optimal culture conditions for L. okamurai were 4 500 lx, 20°C and 35 (salinity); for C. undulatus, 4 500 lx, 20°C and 30; and for L. tristicha, 4 500 lx, 25°C and 30.

Extracellular matrix production by human osteoblasts cultured on biodegradable polymers applicable for tissue engineering.

Science.gov (United States)

El-Amin, S F; Lu, H H; Khan, Y; Burems, J; Mitchell, J; Tuan, R S; Laurencin, C T

2003-03-01

The nature of the extracellular matrix (ECM) is crucial in regulating cell functions via cell-matrix interactions, cytoskeletal organization, and integrin-mediated signaling. In bone, the ECM is composed of proteins such as collagen (CO), fibronectin (FN), laminin (LM), vitronectin (VN), osteopontin (OP) and osteonectin (ON). For bone tissue engineering, the ECM should also be considered in terms of its function in mediating cell adhesion to biomaterials. This study examined ECM production, cytoskeletal organization, and adhesion of primary human osteoblastic cells on biodegradable matrices applicable for tissue engineering, namely polylactic-co-glycolic acid 50:50 (PLAGA) and polylactic acid (PLA). We hypothesized that the osteocompatible, biodegradable polymer surfaces promote the production of bone-specific ECM proteins in a manner dependent on polymer composition. We first examined whether the PLAGA and PLA matrices could support human osteoblastic cell growth by measuring cell adhesion at 3, 6 and 12h post-plating. Adhesion on PLAGA was consistently higher than on PLA throughout the duration of the experiment, and comparable to tissue culture polystyrene (TCPS). ECM components, including CO, FN, LM, ON, OP and VN, produced on the surface of the polymers were quantified by ELISA and localized by immunofluorescence staining. All of these proteins were present at significantly higher levels on PLAGA compared to PLA or TCPS surfaces. On PLAGA, OP and ON were the most abundant ECM components, followed by CO, FN, VN and LN. Immunofluorescence revealed an extracellular distribution for CO and FN, whereas OP and ON were found both intracellularly as well as extracellularly on the polymer. In addition, the actin cytoskeletal network was more extensive in osteoblasts cultured on PLAGA than on PLA or TCPS. In summary, we found that osteoblasts plated on PLAGA adhered better to the substrate, produced higher levels of ECM molecules, and showed greater cytoskeletal

Influence of ionizing radiation on synthesis and molecular heterogeneity of catalase in tissue culture of Rauwolfia serpentina; Vliyanie ioniziruyushchego izlucheniya na biosintez i molekulyarnuyu geterogennost' katalazy v kul'ture tkani Rauwolfia serpentina

Energy Technology Data Exchange (ETDEWEB)

Komov, V P; Bespalova, E V; Strelkova, M A [Sankt-Peterburgskaya Khimiko-Farmatsevticheskaya Akademiya, Kafedra Biologii, S.-Peterburg (Russian Federation)

1998-12-01

Changes in activity and molecular heterogeneity of catalase in tissue culture of Rauwolfia serpentina following irradiation in early growth period at the doses of 8 and 50 Gy has been studied. Ionizing radiation accelerate the synthesis and degradation rates of catalase and total protein. A comparative study of changes in enzyme and protein turnover during growth on irradiated and non-irradiated medium has been made.

Studies in tissue culture of some indigenous rice (Oryza glaberrima Steud.) accessions in Ghana

International Nuclear Information System (INIS)

Diawuoh, R.G.

2011-01-01

A study was conducted with the aim of developing separate protocols for callus induction and plant regeneration from different parts of three O. glaberrima accessions indigenous to Ghana. The three O. glaberrima accessions, Guame, N/4 and SARI 1 were assessed for their callus induction and plant regeneration ability from leaf segments, mature dehusked seeds and anthers on different concentrations of plant growth regulators, incorporated into Murashige and Skoog, (1962) (MS) basal medium. For leaf segments, callus was induced on MS supplemented with (0-10) mg/l 2,4-D. Callus induction frequency was significantly (p≤0.05) different among accessions, as well as among the 2,4-dichlorophenoxyacetic acid (2,4-D) levels tested. Highest callus induction frequency was exhibited at a concentration of 6 mg/l 2,4-D for all accessions tested. Callus obtained was sub-cultured on regeneration medium consisting of MS supplemented with (1:0-5) mg.l NAA:BAP. Plant regeneration was nil. Instead, prolific root formation was observed. For mature dehusked seeds, callus induction medium consisted of MS supplemented with (0-6) mg/l 2,4-D. All tested accessions exhibited highest callus frequency at 4 mg/l 2,4-D. Similarly callus induction frequency was significantly (p≤0.05) different among accessions, as well as among concentrations of 2,4-D tested. Calli obtained were sub-cultured on MS medium supplemented with (0-2.5) mg/l 6-benzylaminopurine (BAP) and exhibited the highest regeneration frequency on medium containing 2.0 mg/l BAP. However, callus induced on a concentration of 3 mg/l 2,4-D and sub-cultured on a concentration of 2 mg/l BAP gave the best response n terms of shoot proliferation, growth and root development and therefore were considered to be the optimum concentrations for callus induction and plant regeneration respectively. Plantlet regeneration was achieved only in accession N/4 while Guame and SARI 1 exhibited poor regeneration response. Among the three rice

Effect of Aflatoxin B1 on Growth of Bovine Mammary Epithelial Cells in 3D and Monolayer Culture System

Directory of Open Access Journals (Sweden)

Babak Qasemi-Panahi

2013-02-01

Full Text Available Purpose: Many studies have been showed transfer of aflatoxins, toxins produced by Aspergillus flvaus and Aspergillus parasiticus fungi, into milk. These toxins are transferred into the milk through digestive system by eating contaminated food. Due to the toxicity of these materials, it seems that it has side effects on the growth of mammary cells. Therefore, the present work aimed to investigate possible toxic effects of aflatoxin B1 (AFB1 on bovine mammary epithelial cells in monolayer and three-dimensional cultures. Methods: Specimens of the mammary tissue of bovine were sized out in size 2×2 cm in slaughterhouse. After disinfection and washing in sterile PBS, primary cell culture was performed by enzymatic digestion of tissue with collagenase. When proper numbers of cells were achieved in monolayer culture, cells were seeded in a 24-well culture plate for three-dimensional (3D culture in Matrigel matrix. After 21 days of 3D culture and reaching the required number of cells, the concentrations of 15, 25 and 35 μL of AFB1 were added to the culture in quadruplicate and incubated for 8 hours. Cellular cytotoxicity was examined using standard colorimetric assay and finally, any change in the morphology of the cells was studied by microscopic technique. Results: Microscopic investigations showed necrosis of the AFB1-exposed cells compared to the control cells. Also, bovine mammary epithelial cells were significantly affected by AFB1 in dose and time dependent manner in cell viability assays. Conclusion: According to the results, it seems that AFB1 can induce cytotoxicity and necrosis in bovine mammary epithelial cells.

Effects of basic fibroblast growth factor and insulin-like growth factor on cultured cartilage cells from skate Raja porasa

Science.gov (United States)

Fan, Tingjun; Jin, Lingyun; Wang, Xiaofeng

2003-12-01

Effects of basic fibroblast growth factor (bFGF) and insulin-like growth factor II (IGF-II) on cartilage cells from proboscis of skate, Raja porasa Günther, were investigated in this study. The cartilage cells were cultured in 20% FBS-supplemented MEM medium at 24°C. Twelve hours after culture initiation, the cartilage cells were treated with bFGF and IGF-II at different concentration combinations. It was found that 20 ng/ml of bFGF or 80 ng/ml of IGF-II was enough to have obvious stimulating effect on the growth and division of skate cartilage cells. Test of bFGF and IGF-II together, revealed that 20 ng/ml of bFGF and 80 ng/ml of IGF-II together had the best stimulating effect on the growth and division of skate cartilage cells. The cartilage cells cultured could form a monolayer at day 7.

Tissue culture of osteogenic sarcoma in rats, induced by radioactive phosphorus P-32 and the effect of the anti-cancerous agents on these tumor cells under tissue culture

International Nuclear Information System (INIS)

Osaka, Shunzo

1976-01-01

Small pieces of osteogenic sarcoma, induced into albino rats of the C.F. Wistar strain by injection of radioactive phosphorus 32 P, were cultured in mixtures of Eagle's minimum essential medium and 20% calf serum. The tumor cells cultured in this way were transplanted into the subcutaneous tissue or the intraabdominal cavity to healthy albino rats. The effect of the anticancerous agents was evaluated by the decrease of nucleic acid composition in these cultured tumor cells. As anti-cancerous agents, cyclophosphamide (CPA), mitomycin C(MMC), and 5-fluorouracil(5-FU) were put into contact with the tumor cells in cultures for two hours under the following dilutions: CPA; 10 -6 , 10 -5 , 10 -4 g/ml. MMC; 2 x 10 -8 , 2 x 10 -7 , 2 x 10 -6 g/ml. 5-FU; 2 x 10 -6 , 2 x 10 -5 , 2 x 10 -4 g/ml. The results are as follows: Three of the seven osteogenic sarcomas in rats were successfully cultured, one of them through more than eighteen generations. After about five hundred thousand cultured cells had been transplanted into the subcutaneous tissues or abdominal cavities of rats, tumors grew in all of them. The histological findings of the tumors in the second generation were quite similar to those of the original tumor. The same process was repeated three times and the tumor showed histogical findings similar to those of the original ones. The capability of nucleic acid synthesis in these cells was decreased at twenty fours after CPA contact and at forty eight hours after MMC. (J.P.N.)

Angiogenesis is not impaired in connective tissue growth factor (CTGF) knock-out mice

NARCIS (Netherlands)

Kuiper, Esther J.; Roestenberg, Peggy; Ehlken, Christoph; Lambert, Vincent; van Treslong-de Groot, Henny Bloys; Lyons, Karen M.; Agostini, Hans-Jürgen T.; Rakic, Jean-Marie; Klaassen, Ingeborg; van Noorden, Cornelis J. F.; Goldschmeding, Roel; Schlingemann, Reinier O.

2007-01-01

Connective tissue growth factor (CTGF) is a member of the CCN family of growth factors. CTGF is important in scarring, wound healing, and fibrosis. It has also been implicated to play a role in angiogenesis, in addition to vascular endothelial growth factor (VEGF). In the eye, angiogenesis and

Whole genome characterization of non-tissue culture adapted HRSV strains in severely infected children

Directory of Open Access Journals (Sweden)

Kumaria Rajni

2011-07-01

Full Text Available Abstract Background Human respiratory syncytial virus (HRSV is the most important virus causing lower respiratory infection in young children. The complete genetic characterization of RSV clinical strains is a prerequisite for understanding HRSV infection in the clinical context. Current information about the genetic structure of the HRSV genome has largely been obtained using tissue culture adapted viruses. During tissue culture adaptation genetic changes can be introduced into the virus genome, which may obscure subtle variations in the genetic structure of different RSV strains. Methods In this study we describe a novel Sanger sequencing strategy which allowed the complete genetic characterisation of 14 clinical HRSV strains. The viruses were sequenced directly in the nasal washes of severely hospitalized children, and without prior passage of the viruses in tissue culture. Results The analysis of nucleotide sequences suggested that vRNA length is a variable factor among primary strains, while the phylogenetic analysis suggests selective pressure for change. The G gene showed the greatest sequence variation (2-6.4%, while small hydrophobic protein and matrix genes were completely conserved across all clinical strains studied. A number of sequence changes in the F, L, M2-1 and M2-2 genes were observed that have not been described in laboratory isolates. The gene junction regions showed more sequence variability, and in particular the intergenic regions showed a highest level of sequence variation. Although the clinical strains grew slower than the HRSVA2 virus isolate in tissue culture, the HRSVA2 isolate and clinical strains formed similar virus structures such as virus filaments and inclusion bodies in infected cells; supporting the clinical relevance of these virus structures. Conclusion This is the first report to describe the complete genetic characterization of HRSV clinical strains that have been sequenced directly from clinical

Diagnostic utility of melanin production by fungi: Study on tissue sections and culture smears with Masson-Fontana stain

Directory of Open Access Journals (Sweden)

Challa Sundaram

2014-01-01

Full Text Available Background: Dematiaceous fungi appear brown in tissue section due to melanin in their cell walls. When the brown color is not seen on routine H and E and culture is not available, differentiation of dematiaceous fungi from other fungi is difficult on morphology alone. Aims and Objective: To study if melanin production by dematiaceous fungi can help differentiate them from other types of fungi. Materials and Methods: Fifty tissue sections of various fungal infections and 13 smears from cultures of different species of fungi were stained with Masson Fontana stain to assess melanin production. The tissue sections included biopsies from 26 culture-proven fungi and 24 biopsies of filamentous fungi diagnosed on morphology alone with no culture confirmation. Results: All culture-proven dematiaceous fungi and Zygomycetes showed strong positivity in sections and culture smears. Aspergillus sp showed variable positivity and intensity. Cryptococcus neoformans showed strong positivity in tissue sections and culture smears. Tissue sections of septate filamentous fungi (9/15, Zygomycetes (4/5, and fungi with both hyphal and yeast morphology (4/4 showed positivity for melanin. The septate filamentous fungi negative for melanin were from biopsy samples of fungal sinusitis including both allergic and invasive fungal sinusitis and colonizing fungal balls. Conclusion: Melanin is produced by both dematiaceous and non-dematiaceous fungi. Masson-Fontana stain cannot reliably differentiate dematiaceous fungi from other filamentous fungi like Aspergillus sp; however, absence of melanin in the hyphae may be used to rule out dematiaceous fungi from other filamentous fungi. In the differential diagnosis of yeast fungi, Cryptococcus sp can be differentiated from Candida sp by Masson-Fontana stain in tissue sections.

Poly(N-isopropylacrylamide) hydrogel/chitosan scaffold hybrid for three-dimensional stem cell culture and cartilage tissue engineering.

Science.gov (United States)

Mellati, Amir; Kiamahalleh, Meisam Valizadeh; Madani, S Hadi; Dai, Sheng; Bi, Jingxiu; Jin, Bo; Zhang, Hu

2016-11-01

Providing a controllable and definable three-dimensional (3D) microenvironment for chondrogenic differentiation of mesenchymal stem cells (MSCs) remains a great challenge for cartilage tissue engineering. In this work, poly(N-isopropylacrylamide) (PNIPAAm) polymers with the degrees of polymerization of 100 and 400 (NI100 and NI400) were prepared and the polymer solutions were introduced into the preprepared chitosan porous scaffolds (CS) to form hybrids (CSNI100 and CSNI400, respectively). SEM images indicated that the PNIPAAm gel partially occupied chitosan pores while the interconnected porous structure of chitosan was preserved. MSCs were incorporated within the hybrid and cell proliferation and chondrogenic differentiation were monitored. After 7-day incubation of the cell-laden constructs in a growth medium, the cell viability in CSNI100 and CSNI400 were 54 and 108% higher than that in CS alone, respectively. Glycosaminoglycan and total collagen contents increased 2.6- and 2.5-fold after 28-day culture of cell-laden CSNI400 in the chondrogenic medium. These results suggest that the hybrid structure composed of the chitosan porous scaffold and the well-defined PNIPAAm hydrogel, in particular CSNI400, is suitable for 3D stem cell culture and cartilage tissue engineering. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2764-2774, 2016. © 2016 Wiley Periodicals, Inc.

Tobacco clones derived from tissue culture with supersensitivity to ozone

International Nuclear Information System (INIS)

Sun, E.J.; Kang, H.W.

2003-01-01

New tobacco clones supersensitive to ozone were obtained from tissue culture. - At least two supersensitive tobacco somaclones were obtained from tissue culture (TC) , when this approach was used to asexually propagate Bel-W3 tobacco indicator plants. These somaclones can detect as low as 30 ppb ozone for a 4-h exposure duration both within CSTR exposure chambers and in ambient air. Comparison of the injury index and their coefficient of variance showed that the TC plantlets usually have more uniform performance in response to ozone in addition to their higher sensitivity. A quick regeneration procedure was established to preserve the supersensitive germplasm immediately when it was found. The TC plantlets will flower and produce seed similar to seed-grown tobacco. The TC approach proved to be a better propagation system for valuable indicator plant species. The mechanism that causes the variation and the possible difference in their genome from seed-grown tobacco is still unknown. Further studies are needed in the future to determine if factors in the TC system may be responsible for the sensitivity difference

Influence of the Culture Media and the Organic Matter in the Growth of Paxillus ammoniavirescens (Contu & Dessi).

Science.gov (United States)

Cagigal, Elena Fernández-Miranda; Sánchez, Abelardo Casares

2017-09-01

The genus Paxillus is characterized by the difficulty of species identification, which results in reproducibility problems, as well as the need for large quantities of fungal inoculum. In particular, studies of Paxillus ammoniavirescens have reported divergent results in the in vitro growth while little is known of its capacity to degrade organic matter. For all the above, and assuming that this variability could be due to an inappropriate culture media, the aim of this study was to analyse growth in different culture media (MMN, MS, and 1/2 MS) and in the case of MMN in presence/absence of two types of organic matter (fresh litter and senescence litter) to probe the saprophytic ability of P. ammoniavirescens . We also evaluated the effects of pH changes in the culture media. Growth kinetics was assessed by weekly quantification of the area of growth in solid culture media over 5 wk, calculating the growth curves and inflection points of each culture media. In addition, final biomass after 5 wk in the different culture media was calculated. Results showed that best culture media are MS and 1/2 MS. Moreover, an improvement in growth in culture media containing decomposing fall litter was observed, leading to confirm differences in the culture media of this species with others of the same genus. Further, we established that all growth media suffered a significant acidification after fungal growth.

Organ and plantlet regeneration of Menyanthes trifoliata through tissue culture

Directory of Open Access Journals (Sweden)

Urszula Adamczyk-Rogozińska

2014-01-01

Full Text Available The conditions for the regeneration of plants through organogenesis from callus tissues of Menyanthes trifoliata are described. The shoot multiplication rate was affected by basal culture media, the type and concentration of cytokinin and subculture number. The best response was obtained when caulogenic calli were cultured on the modified Schenk and Hildebrandt medium (SH-M containing indole-3-acetic acid (IAA 0,5 mg/l and 6-benzyladenine (BA 1 mg/l or zeatin (2 mg/l. Under these conditions ca 7 shoots (mostly 1 cm or more in length per culture in the 5th and 6th passages could be developed. In older cultures (after 11-12 passages there was a trend for more numerous but shorter shoot formation. All regenerated shoots could be rooted on the SH-M medium supplemented with 0.5 mg/l IAA within 6 weeks; 80% of in vitro rooted plantlets survived their transfer to soil.

Treatment of chronic desquamative gingivitis using tissue-engineered human cultured gingival epithelial sheets: a case report.

Science.gov (United States)

Okuda, Kazuhiro; Momose, Manabu; Murata, Masashi; Saito, Yoshinori; lnoie, Masukazu; Shinohara, Chikara; Wolff, Larry F; Yoshie, Hiromasa

2004-04-01

Human cultured gingival epithelial sheets were used as an autologous grafting material for regenerating gingival tissue in the maxillary left and mandibular right quadrants of a patient with chronic desquamative gingivitis. Six months post-surgery in both treated areas, there were gains in keratinized gingiva and no signs of gingival inflammation compared to presurgery. In the maxillary left quadrant, preoperative histopathologic findings revealed the epithelium was separated from the connective tissue and inflammatory cells were extensive. After grafting with the gingival epithelial sheets, inflammatory cells were decreased and separation between epithelium and connective tissue was not observed. The human cultured gingival epithelial sheets fabricated using tissue engineering technology showed significant promise for gingival augmentation in periodontal therapy.

Characterization of cytoskeletal and junctional proteins expressed by cells cultured from human arachnoid granulation tissue

Directory of Open Access Journals (Sweden)

Mehta Bhavya C

2005-10-01

Full Text Available Abstract Background The arachnoid granulations (AGs are projections of the arachnoid membrane into the dural venous sinuses. They function, along with the extracranial lymphatics, to circulate the cerebrospinal fluid (CSF to the systemic venous circulation. Disruption of normal CSF dynamics may result in increased intracranial pressures causing many problems including headaches and visual loss, as in idiopathic intracranial hypertension and hydrocephalus. To study the role of AGs in CSF egress, we have grown cells from human AG tissue in vitro and have characterized their expression of those cytoskeletal and junctional proteins that may function in the regulation of CSF outflow. Methods Human AG tissue was obtained at autopsy, and explanted to cell culture dishes coated with fibronectin. Typically, cells migrated from the explanted tissue after 7–10 days in vitro. Second or third passage cells were seeded onto fibronectin-coated coverslips at confluent densities and grown to confluency for 7–10 days. Arachnoidal cells were tested using immunocytochemical methods for the expression of several common cytoskeletal and junctional proteins. Second and third passage cultures were also labeled with the common endothelial markers CD-31 or VE-cadherin (CD144 and their expression was quantified using flow cytometry analysis. Results Confluent cultures of arachnoidal cells expressed the intermediate filament protein vimentin. Cytokeratin intermediate filaments were expressed variably in a subpopulation of cells. The cultures also expressed the junctional proteins connexin43, desmoplakin 1 and 2, E-cadherin, and zonula occludens-1. Flow cytometry analysis indicated that second and third passage cultures failed to express the endothelial cell markers CD31 or VE-cadherin in significant quantities, thereby showing that these cultures did not consist of endothelial cells from the venous sinus wall. Conclusion To our knowledge, this is the first report of

Growth and Plating of Cell Suspension Cultures of Datura Innoxia

DEFF Research Database (Denmark)

Engvild, Kjeld Christensen

1974-01-01

Suspension cultures of Datura innoxia Mill, were successfully grown on a modified Murashige and Skoog medium with 2,4–D, NAA or BAP as growth substances, provided the micronutrient levels were reduced to 1/10. Normal amounts of micronutrients were toxic. Attempts to identify the toxic elements did...... not succeed. Cultures grew exponentially on a shaker at 27°C in the light. Their doubling times varied from 1.1 days on 2,4–D (10–6M) or NAA (10−5M)+ 1 g/1 casein hydrolysate to 2.7 days on BAP (3 × 10−7M) and 5.1 days on supraoptimal levels of 2,4-D (10−5M). Cultures grew on NH4+-N alone (from ammonium...... malate) or on NO3−-N alone. Dry weight yield was proportional to the amount of nitrate-N added (47 mg/mg N). Filtered suspension cultures containing single cells (plating cultures) could be grown in agar in petri dishes when NAA or 2,4-D were used as growth substances. Cells grew at densities above 500...

Downregulation of connective tissue growth factor inhibits the growth and invasion of gastric cancer cells and attenuates peritoneal dissemination.

Science.gov (United States)

Jiang, Cheng-Gang; Lv, Ling; Liu, Fu-Rong; Wang, Zhen-Ning; Liu, Fu-Nan; Li, Yan-Shu; Wang, Chun-Yu; Zhang, Hong-Yan; Sun, Zhe; Xu, Hui-Mian

2011-09-28

Connective tissue growth factor (CTGF) has been shown to be implicated in tumor development and progression. However, the role of CTGF in gastric cancer remains largely unknown. In this study, we showed that CTGF was highly expressed in gastric cancer tissues compared with matched normal gastric tissues. The CTGF expression in tumor tissue was associated with histologic grade, lymph node metastasis and peritoneal dissemination (P cancer cells and decreased cyclin D1 expression. Moreover, knockdown of CTGF expression also markedly reduced the migration and invasion of gastric cancer cells and decreased the expression of matrix metalloproteinase (MMP)-2 and MMP-9. Animal studies revealed that nude mice injected with the CTGF knockdown stable cell lines featured a smaller number of peritoneal seeding nodules than the control cell lines. These data suggest that CTGF plays an important role in cell growth and invasion in human gastric cancer and it appears to be a potential prognostic marker for patients with gastric cancer.

Downregulation of connective tissue growth factor inhibits the growth and invasion of gastric cancer cells and attenuates peritoneal dissemination

Directory of Open Access Journals (Sweden)

Zhang Hong-Yan

2011-09-01

Full Text Available Abstract Background Connective tissue growth factor (CTGF has been shown to be implicated in tumor development and progression. However, the role of CTGF in gastric cancer remains largely unknown. Results In this study, we showed that CTGF was highly expressed in gastric cancer tissues compared with matched normal gastric tissues. The CTGF expression in tumor tissue was associated with histologic grade, lymph node metastasis and peritoneal dissemination (P 1 expression. Moreover, knockdown of CTGF expression also markedly reduced the migration and invasion of gastric cancer cells and decreased the expression of matrix metalloproteinase (MMP-2 and MMP-9. Animal studies revealed that nude mice injected with the CTGF knockdown stable cell lines featured a smaller number of peritoneal seeding nodules than the control cell lines. Conclusions These data suggest that CTGF plays an important role in cell growth and invasion in human gastric cancer and it appears to be a potential prognostic marker for patients with gastric cancer.

Tissue-engineered skin preserving the potential of epithelial cells to differentiate into hair after grafting.

Science.gov (United States)

Larouche, Danielle; Cuffley, Kristine; Paquet, Claudie; Germain, Lucie

2011-03-01

The aim of this study was to evaluate whether tissue-engineered skin produced in vitro was able to sustain growth of hair follicles in vitro and after grafting. Different tissues were designed. Dissociated newborn mouse keratinocytes or newborn mouse hair buds (HBs) were added onto dermal constructs consisting of a tissue-engineered cell-derived matrix elaborated from either newborn mouse or adult human fibroblasts cultured with ascorbic acid. After 7-21 days of maturation at the air-liquid interface, no hair was noticed in vitro. Epidermal differentiation was observed in all tissue-engineered skin. However, human fibroblast-derived tissue-engineered dermis (hD) promoted a thicker epidermis than mouse fibroblast-derived tissue-engineered dermis (mD). In association with mD, HBs developed epithelial cyst-like inclusions presenting outer root sheath-like attributes. In contrast, epidermoid cyst-like inclusions lined by a stratified squamous epithelium were present in tissues composed of HBs and hD. After grafting, pilo-sebaceous units formed and hair grew in skin elaborated from HBs cultured 10-26 days submerged in culture medium in association with mD. However, the number of normal hair follicles decreased with longer culture time. This hair-forming capacity after grafting was not observed in tissues composed of hD overlaid with HBs. These results demonstrate that epithelial stem cells can be kept in vitro in a permissive tissue-engineered dermal environment without losing their potential to induce hair growth after grafting.

Solid tissue culture for cytogenetic analysis: a collaborative survey for the Association of Clinical Cytogeneticists.

Science.gov (United States)

Rodgers, C S; Creasy, M R; Fitchett, M; Maliszewska, C T; Pratt, N R; Waters, J J

1996-01-01

AIMS: To survey the diagnostic service provided by UK laboratories for the culture of solid tissue samples (excluding tumours) and in particular to examine the variation in culture success rates and the problems of maternal cell overgrowth. METHODS: Twenty seven laboratories took part in a collaborative survey during 1992. Each laboratory submitted data on up to a maximum of 60 consecutive specimens (n = 1361) over a six month period. RESULTS: Skin specimens, the largest category received (n = 520), were the most problematic (51% success rate). Culture success rates were significantly lower (43%) when skin specimens (n = 140) were transported dry to the laboratory. Success rates for skin specimens also varied, depending on the origin of the specimen, from 18% for intra-uterine deaths (IUD) (n = 94) to 85% for neonatal deaths (n = 33) and 83% for live patients (n = 54). Culture of selected extra-fetal tissues from IUD, stillbirths and following elective termination of pregnancy (TOP) gave comparable success rates to those achieved for skin samples from neonatal deaths and live births. Skewed sex ratios, female > male, were identified for products of conception (POC) (n = 298) and placental biopsy specimens (n = 97). CONCLUSIONS: By appropriate selection, transport and processing of tissues, and in particular by avoiding relying solely on skin samples from IUD, stillbirths and TOP, an increase in culture success rates for solid tissue samples submitted for cytogenetic analysis could be achieved. The high risk of maternal cell contamination from POC and placental biopsy specimens was also identified in this survey. PMID:8881913

Study of the agroindustrial alterations induced by the irradiated tissue culture in sugar cane, variety NA 56-79

International Nuclear Information System (INIS)

Figueiredo Junior, O.

1991-01-01

The use of plant tissue culture and the application of gamma radiation as mutation inducing agents, in the sugar cane plant, variety NA 5679, are studied. The variation in the contents of brix, pol, fiber, purity, extraction, phosphorus, nitrogen, reducing sugars as well as the morphological characteristics are analysed. The 'callus' obtained by the tissue culture were irradiated with 20, 40, and 60 Gy doses. The statistical analysis indicated that the method of tissue culture may, eventually, increase the contents of the technological parameters and the dosages of gamma radiation were not efficient for such purpose. (M.A.C.)

Effect of induced mutagenesis in rice tissue culture

International Nuclear Information System (INIS)

Maddumage, R.

1994-01-01

The influence of chemical mutagens and ionising radiation on growth, regenerative capacity of rice callus culture and the effect o9f mutagens on frequency and spectrum of mutant regenerants, derived from calli and determination of approximate semi-lethal dose of each mutagen on rice calli was studied. Intact mature de-husked grains and pieces of primordial particles of four varieties were used as explants in the experiment. Organogenesis was induced using MS media supplemented with agar. After thirty days calluses were subjected to varying concentrations/dosage of mutagens. The effect of mutagens on growth of callus was stimulative in low concentration/doses at short exposure, but in higher concentration/doses at longer exposure it was oppressive. In x-radiation treatment all the studied doses showed only stimulative effect on growth. The effect of mutagenic treatment on regenerative capacity was negative. No specificity was found even between two chemical mutagens of their action on studied characters

Optimization of an Efficient Non-Tissue Culture Transformation Method for Brassica Juncea

International Nuclear Information System (INIS)

Naeem, I.; Munir, I.; Iqbal, A.; Ullah, F.

2016-01-01

The major hurdles in successful in vitro transformation of Brassica juncea through standard tissue culture (STC) method are: culture contamination, somaclonal variations, and lack of expertise. Moreover, the current STC method is time consuming and needs continuous electricity. In the present study, the in planta transformation method through floral dip with or without vacuum infiltration was optimized for successful transformation of B. juncea. The B. juncea CV RAYA Anmol was used for transformation through Agrobacterium tumefaciens strain GV3101 harboring the binary vector plasmid pBinGlyBar4-EADcT. Based on the resistance reaction to the herbicide Basta, 20 and 40 resistant seedlings were obtained from 2000 seed germinated from the plants transformed through floral dip and vacuum infiltration methods, respectively. The PCR analyses further confirmed the presence of transgene in 3 floral dipped plants without vacuum infiltration and 17 floral dipped plants with vacuum infiltration, giving the transformation frequencies of 1.5*10/sup -3/ and 8.5*10/sup -3/, respectively. This method, which avoids tissue culture, will reduce the somaclonal variation accompanying prolonged culture of cells in a dedifferentiated state, will facilitate functional genomics and improvement of Brassica juncea with novel desirable traits while reducing time and expense. (author)

Applications of Biomaterials in Corneal Endothelial Tissue Engineering.

Science.gov (United States)

Wang, Tsung-Jen; Wang, I-Jong; Hu, Fung-Rong; Young, Tai-Horng

2016-11-01

When corneal endothelial cells (CECs) are diseased or injured, corneal endothelium can be surgically removed and tissue from a deceased donor can replace the original endothelium. Recent major innovations in corneal endothelial transplantation include replacement of diseased corneal endothelium with a thin lamellar posterior donor comprising a tissue-engineered endothelium carried or cultured on a thin substratum with an organized monolayer of cells. Repairing CECs is challenging because they have restricted proliferative ability in vivo. CECs can be cultivated in vitro and seeded successfully onto natural tissue materials or synthetic polymeric materials as grafts for transplantation. The optimal biomaterials for substrata of CEC growth are being investigated. Establishing a CEC culture system by tissue engineering might require multiple biomaterials to create a new scaffold that overcomes the disadvantages of single biomaterials. Chitosan and polycaprolactone are biodegradable biomaterials approved by the Food and Drug Administration that have superior biological, degradable, and mechanical properties for culturing substratum. We successfully hybridized chitosan and polycaprolactone into blended membranes, and demonstrated that CECs proliferated, developed normal morphology, and maintained their physiological phenotypes. The interaction between cells and biomaterials is important in tissue engineering of CECs. We are still optimizing culture methods for the maintenance and differentiation of CECs on biomaterials.

Culture of equine bone marrow mononuclear fraction and adipose tissue-derived stromal vascular fraction cells in different media

Directory of Open Access Journals (Sweden)

Gesiane Ribeiro

2013-12-01

Full Text Available The objective of this study was to evaluate the culture of equine bone marrow mononuclear fraction and adipose tissue - derived stromal vascular fraction cells in two different cell culture media. Five adult horses were submitted to bone marrow aspiration from the sternum, and then from the adipose tissue of the gluteal region near the base of the tail. Mononuclear fraction and stromal vascular fraction were isolated from the samples and cultivated in DMEM medium supplemented with 10% fetal bovine serum or in AIM-V medium. The cultures were observed once a week with an inverted microscope, to perform a qualitative analysis of the morphology of the cells as well as the general appearance of the cell culture. Colony-forming units (CFU were counted on days 5, 15 and 25 of cell culture. During the first week of culture, differences were observed between the samples from the same source maintained in different culture media. The number of colonies was significantly higher in samples of bone marrow in relation to samples of adipose tissue.

The serum levels of connective tissue growth factor in patients with systemic lupus erythematosus and lupus nephritis.

Science.gov (United States)

Wang, F-M; Yu, F; Tan, Y; Liu, G; Zhao, M-H

2014-06-01

The expression of connective tissue growth factor mRNA in human kidneys may serve as an early marker for lupus nephritis progression. Therefore, we speculated that connective tissue growth factor may be involved in the pathogenesis of systemic lupus erythematosus and lupus nephritis. In this study, we set out to investigate the associations between serum connective tissue growth factor levels and clinicopathological features of patients with systemic lupus erythematosus and lupus nephritis. Serum samples from patients with non-renal systemic lupus erythematosus, renal biopsy-proven lupus nephritis and healthy control subjects were detected by enzyme-linked immunosorbent assay for serum connective tissue growth factor levels. The associations between connective tissue growth factor levels and clinicopathological features of the patients were further analysed. The levels of serum connective tissue growth factor in patients with non-renal systemic lupus erythematosus and lupus nephritis were both significantly higher than those in the normal control group (34.14 ± 12.17 ng/ml vs. 22.8 ± 3.0 ng/ml, plupus erythematosus and lupus nephritis group (34.14 ± 12.17 ng/ml vs. 44.1 ± 46.8 ng/ml, p = 0.183). Serum connective tissue growth factor levels were significantly higher in lupus nephritis patients with the following clinical manifestations, including anaemia (51.3 ± 51.4 ng/ml vs. 23.4 ± 9.7 ng/ml, plupus nephritis (63.3 ± 63.4 ng/ml vs. 38.3 ± 37.9 ng/ml, p = 0.035, respectively). Serum connective tissue growth factor levels were negatively associated with estimated glomerular filtration rate (r = -0.46, plupus nephritis (plupus and correlated with chronic renal interstitial injury and doubling of serum creatinine in patients with lupus nephritis. © The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

Polyploidization of glia in neural development links tissue growth to blood-brain barrier integrity.

Science.gov (United States)

Unhavaithaya, Yingdee; Orr-Weaver, Terry L

2012-01-01

Proper development requires coordination in growth of the cell types composing an organ. Many plant and animal cells are polyploid, but how these polyploid tissues contribute to organ growth is not well understood. We found the Drosophila melanogaster subperineurial glia (SPG) to be polyploid, and ploidy is coordinated with brain mass. Inhibition of SPG polyploidy caused rupture of the septate junctions necessary for the blood-brain barrier. Thus, the increased SPG cell size resulting from polyploidization is required to maintain the SPG envelope surrounding the growing brain. Polyploidization likely is a conserved strategy to coordinate tissue growth during organogenesis, with potential vertebrate examples.

Cytogenetic studies on stevia rebaudiana produced by tissue culture and affected by gamma rays and drought

International Nuclear Information System (INIS)

Awad, A.S.A

2009-01-01

The present investigation was under taken to carry out in the laboratories of the Natural Products Department, National Center for Radiation Research and Technology, Atomic Energy authority, Nasr city, Cairo, Egypt, to study the effect of gamma radiation doses, osmostress and the combined effects between them on tissue culture, some biochemical analysis and molecular genetic marker in stevia rebaudiana bertoni. The results obtained were: Tissue culture 1- micropropagation media: stevia rebaudiana plantlets cultured on MS medium hormones free for micropropagation.Hormones such as BAP and NAA with different concentrations induced callus formation and give slight growth.Study the effect of gamma radiation, osmostress and the combined effects between them : 1)The effect of gamma radiation on buds survival: Gamma radiation doses (10, 20 and 30 Gy) induced decreasing in bud survival percentage with increasing radiation dose in stevia rebaudiana. The dose 30 Gy was induced 60% mortality.2) Study the effect of gamma radiation on some biochemical analysis: Gamma radiation doses induced increase in the total carbohydrate with doses (20 and 30 Gy) but decreased with dose 10 Gy. Proline contents increased in plantlets with increasing doses . The total protein was increased with doses (10 and 20 Gy), but the dose 30 Gy induced decrease in total protein. Gamma radiation doses induced decreasing in total DNA while, the nucleic acid RNA increased.3) The effect of osmostress on buds survival: The concentrations (40000,50000,60000,70000 and 80000 ppm) from sucrose or sorbitol decreased the bud survival and shoot length in stevia plantlets with increasing sucrose or sorbitol levels. 4) The effect of osmostress on some biochemical analysis: Sucrose and sorbitol concentrations (40000,50000,60000,70000 and 80000 ppm) caused decrease in total carbohydrate.

Cell and Molecular Biology of Ataxia Telangiectasia Heterozygous Human Mammary Epithelial Cells Irradiated in Culture

Science.gov (United States)

Richmond, Robert C.

2001-01-01

Autologous isolates of cell types from obligate heterozygotes with the autosomal disorder ataxia-telangiectasia (A-T)were used to begin a tissue culture model for assessing pathways of radiation-induced cancer formation in this target tissue. This was done by establishing cultures of stromal fibroblasts and long-term growth human mammary epithelial cells (HMEC) in standard 2-dimensional tissue culture in order to establish expression of markers detailing early steps of carcinogenesis. The presumptive breast cancer susceptibility of A-T heterozygotes as a sequel to damage caused by ionizing radiation provided reason to study expression of markers in irradiated HMEC. Findings from our study with HMEC have included determination of differences in specific protein expression amongst growth phase (e.g., log vs stationary) and growth progression (e.g., pass 7 vs pass 9), as well as differences in morphologic markers within populations of irradiated HMEC (e.g., development of multinucleated cells).

Host DNA synthesis-suppressing factor in culture fluid of tissue cultures infected with measles virus

International Nuclear Information System (INIS)

Minagawa, T.; Nakaya, C.; Iida, H.

1974-01-01

Host DNA synthesis is suppressed by the culture fluid of cell cultures infected with measles virus. This activity in the culture fluid is initiated somewhat later than the growth of infectious virus. Ninety percent of host DNA synthesis in HeLa cells is inhibited by culture fluid of 3-day-old cell cultures of Vero or HeLa cells infected with measles virus. This suppressing activity is not a property of the virion, but is due to nonvirion-associated componentnent which shows none of the activities of measles virus such as hemagglutination, hemolysis, or cell fusion nor does it have the antigenicity of measles virus as tested by complement-fixation or hemagglutination-inhibiting antibody blocking tests. Neutralization of the activity of this component is not attained with the pooled sera of convalescent measles patients. This component has molecular weights of about 45,000, 20,000, and 3,000 and appears to be a heat-stable protein. The production of host DNA suppressing factor (DSF) is blocked by cycloheximide. Neither uv-inactivated nor antiserum-neutralized measles virus produce DSF. Furthermore, such activity of nonvirion-associated component is not detected in the culture fluid of cultures infected with other RNA viruses such as poliovirus, vesicular stomatitis virus, or Sindbis virus. (auth)

Fish culture in heated effluents (eastern Europe). [Growth increase in carp and rainbow trout

Energy Technology Data Exchange (ETDEWEB)

Backiel, T; Gay, B; La Croix, M J.B.; Ophel, I L [eds.

1976-01-01

Dependence of growth and reproduction of cultivated fishes on temperature are briefly reviewed. Experiments with and commercial-size operations of various methods of fish culture with the use of heated effluents are exemplified. Cage culture, earthen ponds, flowing water culture have demonstrated possibilities of growth increase in carp and rainbow trout and of controlled reproduction of the former. Problems involved in utilization of various systems and several points of view on the use of low-grade heat for fish culture are considered.

Mass production of `Swan`s Golden`, Cupressus sempervirens by tissue culture and its radiation breeding. 2. Study on growth regulating substance to stimulate the elongation of shoot apex

Energy Technology Data Exchange (ETDEWEB)

Suda, Hirokatsu [Tokyo Metropolitan Isotope Research Center (Japan)

1997-09-01

Some species of `Swan`s Golden` have been propagated by means of grafting because propagation by cutting is difficult. However, not a few species of the plant are easily lodged, resulting that spreading of `Swan`s Golden` is difficult. Here, an investigation was made on growth regulating substances added into the basal medium in respects of their kinds and concentrations and also the kind of medium for successive culture. As the basal medium, SH medium supplemented with 4 g/l of gel-light and 3 g/l of carbon-powder was used. The optimum medium for the initiation of culture was the basal medium added with zeatin (Ze) and gibberellin (GA) and it was also suitable for successive culture after the 14 day. The medium added with Ze and benzyladenine (BA) or Ze alone was found to be most suitable for the culture after the 28 day. The concentration was optimum at 1 - 2.5 mg/l for either of these growth regulating substances. (M.N.)

Effects of photoperiod, plant growth regulators and culture media on in vitro growth of seedlings of Cyrtochilum loxense (Lindl. Kraenzl. an endemic and endangered orchid from Ecuador

Directory of Open Access Journals (Sweden)

Yadira González

2014-10-01

Full Text Available Cyrtochilum loxense (Lindl. Kraenzl. is an endemic and seriously endangered orchid species endemic in the Loja Province (Southern Ecuador. The main goals of this research were to analyze how culture media, plant growth regulators and photoperiod affect the growth of C. loxense. Eight month old plants (approximate 1 – 1.5 cm in height obtained by in vitro germination, were cultivated on MS media or Knudson C; MS with three levels of naphthalene acetic acid (NAA and 6-benzylaminopurine (BAP (2/0.5; 1/0.5 y 0.5/ 0.5 mg-1L; and three photoperiodic regimes (24/0, 16/8, 8/16 h on MS with and without plant growth regulators. No significant differences of shoot induction were observed on media with or without plant growth regulators, and all tested photoperiods. The highest growth (1.2 cm was observed in plantlets cultivated on growth regulator-free media with a 16/8 photoperiod. Also the shoot and root formation was better in this species in absence of plant growth regulators. Probably this response is due to the endogenous hormone levels in the tissues or due to the kind and concentrations of PGRs used were too low to induce positive morphogenetic responses.

Discrimination and similarity evaluation of tissue-cultured and wild Dendrobium species using Fourier transform infrared spectroscopy

Science.gov (United States)

Chen, Nai-dong; Chen, Han; Li, Jun; Sang, Mang-mang; Ding, Shen; Yu, Hao

2015-04-01

The FTIR method was applied to evaluate the similarity of tissue-cultured and wild Dendrobium huoshanense C.Z. Tang et S.J. Cheng, Dendrobium officinale Kimura et Migo and Dendrobium moniliforme (Linn.) Sw and discriminate different Dendrobium species, especially D. huoshanense and its main goldbrick Dendrobium henanense J.L. Lu et L.X. Gao. Despite the general pattern of the IR spectra, different intensities, shapes and peak positions were found in the IR spectra of these samples, especially in the range of 1800-600 cm-1, which could be used to discriminate them. The methanol, aqueous extracting procedure and the second derivative transformation obviously enlarged the tiny spectral differences among these samples. The similarity evaluation based on the IR spectra and the second derivative IR spectrum revealed that the similarity of the methanol extracts between tissue-cultured and wild Dendrobiums might be lower than that between different Dendrobium species. The similarities of the powders and aqueous extracts between tissue-cultured and wild Dendrobiums were higher than those between different Dendrobium species. The further principal component analysis showed that the first three components explained 99.7%, 87.7% and 85.1% of data variance for powder, methanol extract and aqueous extract, respectively, demonstrating a good discrimination between samples. Our research suggested that the variations of secondary metabolites between different origins of the investigated Dendrobiums might be higher than what we had supposed. Tissue culture techniques were widely used in the conversation of rare and endangered medicinal amedica, however, our study suggested that the chemical constituents of tissue-cultured plants might be quite different from their wild correspondences.

Culture media profoundly affect Candida albicans and Candida tropicalis growth, adhesion and biofilm development.

Science.gov (United States)

Weerasekera, Manjula M; Wijesinghe, Gayan K; Jayarathna, Thilini A; Gunasekara, Chinthika P; Fernando, Neluka; Kottegoda, Nilwala; Samaranayake, Lakshman P

2016-11-01

As there are sparse data on the impact of growth media on the phenomenon of biofilm development for Candida we evaluated the efficacy of three culture media on growth, adhesion and biofilm formation of two pathogenic yeasts, Candida albicans and Candida tropicalis. The planktonic phase yeast growth, either as monocultures or mixed cultures, in sabouraud dextrose broth (SDB), yeast nitrogen base (YNB), and RPMI 1640 was compared, and adhesion as well as biofilm formation were monitored using MTT and crystal violet (CV) assays and scanning electron microscopy. Planktonic cells of C. albicans, C. tropicalis and their 1:1 co-culture showed maximal growth in SDB. C. albicans/C. tropicalis adhesion was significantly facilitated in RPMI 1640 although the YNB elicited the maximum growth for C. tropicalis. Similarly, the biofilm growth was uniformly higher for both species in RPMI 1640, and C. tropicalis was the slower biofilm former in all three media. Scanning electron microscopy images tended to confirm the results of MTT and CV assay. Taken together, our data indicate that researchers should pay heed to the choice of laboratory culture media when comparing relative planktonic/biofilm growth of Candida. There is also a need for standardisation of biofilm development media so as to facilitate cross comparisons between laboratories.

The effect of plant growth regulators on optimization of tissue culture ...

African Journals Online (AJOL)

Mature seeds of four upland rice cultivars namely Kusan, Lamsan, Selasi and Siam were assessed for callus induction and plant regeneration on different concentrations and combinations of plant growth regulators, incorporated into MS (Murashige and Skoog) basal medium. Callus induction frequency was significantly ...

Induction of growth and proliferation of fibroblast cells in magnetic field

Directory of Open Access Journals (Sweden)

Naghmeh Ezatti

2015-02-01

Full Text Available Background: Tissue engineering is generally defined as developing and changing the laboratory growth of molecules and cells in tissues or organs to replace and repair the damaged part of body. This study was carried out to stimulate the growth of cultured fibroblast cells by a physical electromagnetic method. Methods: First, an air-core coil was prepared and the cell culture plate was placed comfortably into the mold, then the plate containing the culture medium and human fibroblast cell along with air-core coil were placed in an incubator and then connected to the power supply. Thus, the sample underwent electromagnetic field at different times, and cell proliferation was studied by MTTassay. Results: Microscopic images indicated that the cells undergoing electromagnetic field (0.35 amps had a significant growth compared to the cells in control group in a definit range of stimulation. Conclusion: In conclusion, electromagnetic stimulation in a definite range led to cell proliferation and could be used as a positive factor in tissue engineering.

Modification of growth conditions by mm-waves of wood-decaying mushrooms cultures

International Nuclear Information System (INIS)

Avagyan, I.A.

2016-01-01

Basidial macromycetes are not only value food, but can be used as source of such biological active compounds as the genistein, β -glucans, glioxal-oxidase et al. In this work we used different frequencies of extremely high frequency of electromagnetic irradiation (EHF EMI) with the aim of obtaining mushroom cultures with increased fermentative activity by the modulation of its growth conditions during growth on the peptone media. We investigated the influence of the non-thermal extremely high frequency electromagnetic waves in the interval of 45-53 GHz on β-glucosidase activities of two species of wood-decaying mushroom. In this study we examined the most popular edible wood-decaying mushroom Pleurotus ostreatus, which is wide-spread in the forests and commercial mushroom Lentinula edodes, under influence of such an abiotic factor as the extremely high frequency waves in the interval of 45 GHz - 53 GHz during 20 and 40 min on the 7th day of mycelial culture's growth. After the treatment of cultures we continued their growth and on the 3th day we examined the influence of these waves on fermentative activity of mycelial extracts. The some conditions of such treatment led to significant rising of β-glucosidase activities in the extracts of mycelial cultures

Noninvasive Real-Time Assessment of Cell Viability in a Three-Dimensional Tissue.

Science.gov (United States)

Mahfouzi, Seyed Hossein; Amoabediny, Ghassem; Doryab, Ali; Safiabadi-Tali, Seyed Hamid; Ghanei, Mostafa

2018-04-01

Maintaining cell viability within 3D tissue engineering scaffolds is an essential step toward a functional tissue or organ. Assessment of cell viability in 3D scaffolds is necessary to control and optimize tissue culture process. Monitoring systems based on respiration activity of cells (e.g., oxygen consumption) have been used in various cell cultures. In this research, an online monitoring system based on respiration activity was developed to monitor cell viability within acellular lung scaffolds. First, acellular lung scaffolds were recellularized with human umbilical cord vein endothelial cells, and then, cell viability was monitored during a 5-day period. The real-time monitoring system generated a cell growth profile representing invaluable information on cell viability and proliferative states during the culture period. The cell growth profile obtained by the monitoring system was consistent with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide analysis and glucose consumption measurement. This system provided a means for noninvasive, real-time, and repetitive investigation of cell viability. Also, we showed the applicability of this monitoring system by introducing shaking as an operating parameter in a long-term culture.

Connective tissue growth factor (CTGF/CCN2 is negatively regulated during neuron-glioblastoma interaction.

Directory of Open Access Journals (Sweden)

Luciana F Romão

Full Text Available Connective-tissue growth factor (CTGF/CCN2 is a matricellular-secreted protein involved in complex processes such as wound healing, angiogenesis, fibrosis and metastasis, in the regulation of cell proliferation, migration and extracellular matrix remodeling. Glioblastoma (GBM is the major malignant primary brain tumor and its adaptation to the central nervous system microenvironment requires the production and remodeling of the extracellular matrix. Previously, we published an in vitro approach to test if neurons can influence the expression of the GBM extracellular matrix. We demonstrated that neurons remodeled glioma cell laminin. The present study shows that neurons are also able to modulate CTGF expression in GBM. CTGF immnoreactivity and mRNA levels in GBM cells are dramatically decreased when these cells are co-cultured with neonatal neurons. As proof of particular neuron effects, neonatal neurons co-cultured onto GBM cells also inhibit the reporter luciferase activity under control of the CTGF promoter, suggesting inhibition at the transcription level. This inhibition seems to be contact-mediated, since conditioned media from embryonic or neonatal neurons do not affect CTGF expression in GBM cells. Furthermore, the inhibition of CTGF expression in GBM/neuronal co-cultures seems to affect the two main signaling pathways related to CTGF. We observed inhibition of TGFβ luciferase reporter assay; however phopho-SMAD2 levels did not change in these co-cultures. In addition levels of phospho-p44/42 MAPK were decreased in co-cultured GBM cells. Finally, in transwell migration assay, CTGF siRNA transfected GBM cells or GBM cells co-cultured with neurons showed a decrease in the migration rate compared to controls. Previous data regarding laminin and these results demonstrating that CTGF is down-regulated in GBM cells co-cultured with neonatal neurons points out an interesting view in the understanding of the tumor and cerebral microenvironment

Co-cultures and cell sheet engineering as relevant tools to improve the outcome of bone tissue engineering strategies

OpenAIRE

Pirraco, Rogério

2011-01-01

Taking into consideration the complex biology of bone tissue it is quite clear that the understanding of the cellular interactions that regulate the homeostasis and regeneration of this remarkable tissue is essential for a successful Tissue Engineering strategy. The in vitro study of these cellular interactions relies on co-culture systems, a tremendously useful methodology where two or more cell types are cultured at the same time. Such strategy increases the complexity of typ...

Connective tissue growth factor (CTGF) and cancer progression.

Science.gov (United States)

Chu, Chia-Yu; Chang, Cheng-Chi; Prakash, Ekambaranellore; Kuo, Min-Liang

2008-11-01

Connective tissue growth factor (CTGF) is a member of the CCN family of secreted, matrix-associated proteins encoded by immediate early genes that play various roles in angiogenesis and tumor growth. CCN family proteins share uniform modular structure which mediates various cellular functions such as regulation of cell division, chemotaxis, apoptosis, adhesion, motility, angiogenesis, neoplastic transformation, and ion transport. Recently, CTGF expression has been shown to be associated with tumor development and progression. There is growing body of evidence that CTGF may regulate cancer cell migration, invasion, angiogenesis, and anoikis. In this review, we will highlight the influence of CTGF expression on the biological behavior and progression of various cancer cells, as well as its regulation on various types of protein signals and their mechanisms.

Microfluidic systems for stem cell-based neural tissue engineering.

Science.gov (United States)

Karimi, Mahdi; Bahrami, Sajad; Mirshekari, Hamed; Basri, Seyed Masoud Moosavi; Nik, Amirala Bakhshian; Aref, Amir R; Akbari, Mohsen; Hamblin, Michael R

2016-07-05

Neural tissue engineering aims at developing novel approaches for the treatment of diseases of the nervous system, by providing a permissive environment for the growth and differentiation of neural cells. Three-dimensional (3D) cell culture systems provide a closer biomimetic environment, and promote better cell differentiation and improved cell function, than could be achieved by conventional two-dimensional (2D) culture systems. With the recent advances in the discovery and introduction of different types of stem cells for tissue engineering, microfluidic platforms have provided an improved microenvironment for the 3D-culture of stem cells. Microfluidic systems can provide more precise control over the spatiotemporal distribution of chemical and physical cues at the cellular level compared to traditional systems. Various microsystems have been designed and fabricated for the purpose of neural tissue engineering. Enhanced neural migration and differentiation, and monitoring of these processes, as well as understanding the behavior of stem cells and their microenvironment have been obtained through application of different microfluidic-based stem cell culture and tissue engineering techniques. As the technology advances it may be possible to construct a "brain-on-a-chip". In this review, we describe the basics of stem cells and tissue engineering as well as microfluidics-based tissue engineering approaches. We review recent testing of various microfluidic approaches for stem cell-based neural tissue engineering.

Mammalian Cell Culture Simplified.

Science.gov (United States)

Moss, Robert; Solomon, Sondra

1991-01-01

A tissue culture experiment that does not require elaborate equipment and that can be used to teach sterile technique, the principles of animal cell line maintenance, and the concept of cell growth curves is described. The differences between cancerous and normal cells can be highlighted. The procedure is included. (KR)

Analytical and diagnostic performance of a qPCR assay for Ichthyophonus spp. compared to the tissue culture 'gold standard'.

Science.gov (United States)

Lowe, Vanessa C; Hershberger, Paul K; Friedman, Carolyn S

2018-06-04

Parasites of the genus Ichthyophonus infect many fish species and have a non-uniform distribution within host tissues. Due in part to this uneven distribution, the comparative sensitivity and accuracy of using molecular-based detection methods versus culture to estimate parasite prevalence is under debate. We evaluated the analytical and diagnostic performance of an existing qPCR assay in comparison to the 'gold standard' culture method using Pacific herring Clupea pallasii with known exposure history. We determined that the assay is suitable for use in this host, and diagnostic specificity was consistently high (>98%) in both heart and liver tissues. Diagnostic sensitivity could not be fully assessed due to low infection rates, but our results suggest that qPCR is not as sensitive as culture under all circumstances. Diagnostic sensitivity of qPCR relative to culture is likely affected by the amount of sample processed. The prevalence values estimated by the 2 methods were not significantly different when sample amounts were equal (heart tissue), but when the assayed sample amounts were unequal (liver tissue), the culture method detected a significantly higher prevalence of the parasite than qPCR. Further, culture of liver also detected significantly more Ichthyophonus infections than culture of heart, suggesting that the density and distribution of parasites in tissues also plays a role in assay sensitivity. This sensitivity issue would be most problematic for fish with light infections. Although qPCR does not detect the presence of a live organism, DNA-based pathogen detection methods provide the opportunity for alternate testing strategies when culture is not possible.

Estradiol suppresses tissue androgens and prostate cancer growth in castration resistant prostate cancer

International Nuclear Information System (INIS)

Montgomery, Bruce; Nelson, Peter S; Vessella, Robert; Kalhorn, Tom; Hess, David; Corey, Eva

2010-01-01

Estrogens suppress tumor growth in prostate cancer which progresses despite anorchid serum androgen levels, termed castration resistant prostate cancers (CRPC), although the mechanisms are unclear. We hypothesize that estrogen inhibits CRPC in anorchid animals by suppressing tumoral androgens, an effect independent of the estrogen receptor. The human CRPC xenograft LuCaP 35V was implanted into orchiectomized male SCID mice and established tumors were treated with placebo, 17β-estradiol or 17β-estradiol and estrogen receptor antagonist ICI 182,780. Effects of 17β-estradiol on tumor growth were evaluated and tissue testosterone (T) and dihydrotestosterone (DHT) evaluated by mass spectrometry. Treatment of LuCaP 35V with 17β-estradiol slowed tumor growth compared to controls (tumor volume at day 21: 785 ± 81 mm 3 vs. 1195 ± 84 mm 3 , p = 0.002). Survival was also significantly improved in animals treated with 17β-estradiol (p = 0.03). The addition of the estrogen receptor antagonist ICI 182,780 did not significantly change survival or growth. 17β-estradiol in the presence and absence of ICI 182,780 suppressed tumor testosterone (T) and dihydrotestosterone (DHT) as assayed by mass spectrometry. Tissue androgens in placebo treated LuCaP 35V xenografts were; T = 0.71 ± 0.28 pg/mg and DHT = 1.73 ± 0.36 pg/mg. In 17β-estradiol treated LuCaP35V xenografts the tissue androgens were, T = 0.20 ± 0.10 pg/mg and DHT = 0.15 ± 0.15 pg/mg, (p < 0.001 vs. controls). Levels of T and DHT in control liver tissue were < 0.2 pg/mg. CRPC in anorchid animals maintains tumoral androgen levels despite castration. 17β-estradiol significantly suppressed tumor T and DHT and inhibits growth of CRPC in an estrogen receptor independent manner. The ability to manipulate tumoral androgens will be critical in the development and testing of agents targeting CRPC through tissue steroidogenesis

Bone marrow-derived cultured mast cells and peritoneal mast cells as targets of a growth activity secreted by BALB/3T3 fibroblasts

International Nuclear Information System (INIS)

Jozaki, K.; Kuriu, A.; Hirota, S.; Onoue, H.; Ebi, Y.; Adachi, S.; Ma, J.Y.; Tarui, S.; Kitamura, Y.

1991-01-01

When fibroblast cell lines were cultured in contact with bone marrow-derived cultured mast cells (CMC), both NIH/3T3 and BALB/3T3 cell lines supported the proliferation of CMC. In contrast, when contact between fibroblasts and CMC was prohibited by Biopore membranes or soft agar, only BALB/3T3 fibroblasts supported CMC proliferation, suggesting that BALB/3T3 but not NIH/3T3 cells secreted a significant amount of a mast cell growth activity. Moreover, the BALB/3T3-derived growth activity induced the incorporation of [3H]thymidine by CMC and the clonal growth of peritoneal mast cells in methylcellulose. The mast cell growth activity appeared to be different from interleukin 3 (IL-3) and interleukin 4 (IL-4), because mRNAs for these interleukins were not detectable in BALB/3T3 fibroblasts. Although mast cells are genetically deficient in tissues of W/Wv mice, CMC did develop when bone marrow cells of W/Wv mice were cultured with pokeweed mitogen-stimulated spleen cell-conditioned medium. Because BALB/3T3 fibroblast-conditioned medium (BALB-FCM) did not induce the incorporation of [3H]thymidine by W/Wv CMC, the growth activity in BALB-FCM appeared to be a ligand for the receptor encoded by the W (c-kit) locus. Because CMC and peritoneal mast cells are obtained as homogeneous suspensions rather easily, these cells may be potentially useful as targets for the fibroblast-derived mast cell growth activity

Rotating three-dimensional dynamic culture of adult human bone marrow-derived cells for tissue engineering of hyaline cartilage.

Science.gov (United States)

Sakai, Shinsuke; Mishima, Hajime; Ishii, Tomoo; Akaogi, Hiroshi; Yoshioka, Tomokazu; Ohyabu, Yoshimi; Chang, Fei; Ochiai, Naoyuki; Uemura, Toshimasa

2009-04-01

The method of constructing cartilage tissue from bone marrow-derived cells in vitro is considered a valuable technique for hyaline cartilage regenerative medicine. Using a rotating wall vessel (RWV) bioreactor developed in a NASA space experiment, we attempted to efficiently construct hyaline cartilage tissue from human bone marrow-derived cells without using a scaffold. Bone marrow aspirates were obtained from the iliac crest of nine patients during orthopedic operation. After their proliferation in monolayer culture, the adherent cells were cultured in the RWV bioreactor with chondrogenic medium for 2 weeks. Cells from the same source were cultured in pellet culture as controls. Histological and immunohistological evaluations (collagen type I and II) and quantification of glycosaminoglycan were performed on formed tissues and compared. The engineered constructs obtained using the RWV bioreactor showed strong features of hyaline cartilage in terms of their morphology as determined by histological and immunohistological evaluations. The glycosaminoglycan contents per microg DNA of the tissues were 10.01 +/- 3.49 microg/microg DNA in the case of the RWV bioreactor and 6.27 +/- 3.41 microg/microg DNA in the case of the pellet culture, and their difference was significant. The RWV bioreactor could provide an excellent environment for three-dimensional cartilage tissue architecture that can promote the chondrogenic differentiation of adult human bone marrow-derived cells.

Influence of plant growth regulators on development and ...

African Journals Online (AJOL)

Therefore propagation of the plant material by cell cultures and the extraction of potential pharmaceutical active compounds are of great interest. Calli were established on different media from roots and shoots of seedlings and softness and colour of the tissue were compared. Optimum growth of callus cultures was ...

Bioeconomic of profit maximization of red tilapia (Oreochromis sp.) culture using polynomial growth model

Science.gov (United States)

Wijayanto, D.; Kurohman, F.; Nugroho, RA

2018-03-01

The research purpose was to develop a model bioeconomic of profit maximization that can be applied to red tilapia culture. The development of fish growth model used polynomial growth function. Profit maximization process used the first derivative of profit equation to time of culture equal to zero. This research has also developed the equations to estimate the culture time to reach the target size of the fish harvest. The research proved that this research model could be applied in the red tilapia culture. In the case of this study, red tilapia culture can achieve the maximum profit at 584 days and the profit of Rp. 28,605,731 per culture cycle. If used size target of 250 g, the culture of red tilapia need 82 days of culture time.

Growth of microbial mixed cultures under anaerobic, alkaline conditions

International Nuclear Information System (INIS)

Wenk, M.

1993-09-01

Cement and concrete are the most important engineered barrier materials in a repository for low- and intermediate-level waste and thus represent the most significant component of the total disposal inventory. Based on the chemical composition of the concrete used in the repository and the groundwater fluxes in the modelled host rock, it is to be expected that the pH in the near vicinity of the repository could exceed a value of 10.5 for more than a million years. The groundwater in the repository environment also has a limited carbon concentration. Since microorganisms will be present in a repository and can even find suitable living conditions within the waste itself, investigations were carried out in order to establish the extent to which microbial activity is possible under the extreme conditions of the repository near-field. For the investigations, alkalophilic cultures were enriched from samples from alkaline habitats and from Valanginian Marl. Anaerobic bacteria with fermentative, sulfate-reducing and methanogenic metabolism were selected. The growth and activity of the mixed cultures were studied under alkaline conditions and the dependence on pH and carbon concentration determined. All the mixed cultures investigated are alkalophilic. The optimum growth range for the cultures is between pH 9.0 and pH 10.0. The activity limit for the fermentative mixed culture is at pH 12, for the sulfate-reducers at pH 11 and for the methanogens at pH 10.5. Given the limited supply of carbon, the mixed cultures can only grow under slightly alkaline conditions. Only the fermentative cultures are capable of surviving with limited carbon supply at pH 13. (author) 24 figs., 18 tabs., 101 refs

Flavonoid Production, Growth and Differentiation of Stelechocarpus burahol (Bl.) Hook. F. and Th. Cell Suspension Culture.

Science.gov (United States)

Aini Habibah, Noor; Moeljopawiro, Sukarti; Dewi, Kumala; Indrianto, Ari

2017-01-01

Stelechocarpus burahol is a plant containing flavonoid compounds that have the potential for use as an antihyperuricemic for gout medication. This study was performed to assess flavonoid production, growth and cell differentiation of S. burahol in cell suspension culture. Mesocarp was planted in Murashige and Skoog (MS) medium supplemented with 7.5 mg L-1 picloram for the induction of callus. Non-embryonic callus obtained was used in the formation of cell suspension cultures. Growth of cells was determined by fresh and dry weights. During the culturing, the fresh weight, dry weight and flavonoid content were determined as a result of culture status. The growth of the S. burahol cell suspension was slow, the stationary phase occurred at 30 days. The production of flavonoids was not in line with the growth of cells and the maximum production occurred on the 15th day of the log phase. The globular-shaped cells dominated the cell suspension culture at all ages. Fluorescein diacetate (FDA) staining of cells derived from cell cultures aged for 36 days showed that some cells were still viable. The results show that flavonoid production, growth and cell differentiation of a S. burahol cell suspension culture differed according to the culture age.

Non-specific interference of certain components of tissue culture media with the radioimmunoassay of rat alpha-foetoprotein

International Nuclear Information System (INIS)

Dambuyant, C.; Sizaret, Ph.

1975-01-01

Interferences of 'Williams' tissue culture medium used for cultivating rat hepatocytes upon rat alpha-foetoprotein (AFP) radioimmunoassay have been investigated. They are not due to foetal calf serum proteins which are added as growth factor and can be abolished by dialysis which appears to be necessary for the distinction between AFP non-producer and low-producer cell lines. Of the three major groups of non-mineral components examined, amino acid solution played a major role. When individual amino acids were examined using the double antibody technique, arginine was found to interfere predominantly; its dose-response curve was parallel to that of rat AFP which confirmed that an immunological identity between two substances cannot be established on the basis of parallelism as the only criterion

Functional enhancement of chitosan and nanoparticles in cell culture, tissue engineering, and pharmaceutical applications

Directory of Open Access Journals (Sweden)

Wenjuan eGao

2012-08-01

Full Text Available Abstract: As a biomaterial, chitosan has been widely used in tissue engineering, wound healing, drug delivery, and other biomedical applications. It can be formulated in a variety of forms, such as powder, film, sphere, gel and fiber. These features make chitosan an almost ideal biomaterial in cell culture applications, and cell cultures arguably constitute the most practical way to evaluate biocompatibility and biotoxicity. The advantages of cell cultures are that they can be performed under totally controlled environments, allow high throughput functional screening, and are less costly, as compared to other assessment methods. Chitosan can also be modified into multilayer composite by combining with other polymers and moieties to alter the properties of chitosan for particular biomedical applications. This review briefly depicts and discusses applications of chitosan and nanoparticles in cell culture, in particular, the effects of chitosan and nanoparticles on cell adhesion, cell survival, and the underlying molecular mechanisms: both stimulatory and inhibitory influences are discussed. Our aim is to update the current status of how nanoparticles can be utilized to modify the properties of chitosan to advance the art of tissue engineering by using cell cultures.

Cholera toxin expression by El Tor Vibrio cholerae in shallow culture growth conditions.