Culture of the terrestrial cyanobacterium, Nostoc flagelliforme (Cyanophyceae), under aquatic conditions

Both colonies and free-living cells of the terrestrial cyanobacterium, Nostoc flagelliforme (Berk. & Curtis) Bornet & Flahault, were cultured under aquatic conditions to develop the techniques for the cultivation and restoration of this endangered resource. The colonial filaments disintegrated with their sheaths ruptured in about 2 days without any desiccating treatments. Periodic desiccation played an important role in preventing the alga from decomposing, with greater delays to sheath rupture with a higher frequency of exposure to air. The bacterial numbers in the culture treated with seven periods of desiccation per day were about 50% less compared with the cultures without the desiccation treatment. When bacteria in the culture were controlled, the colonial filaments did not disintegrate and maintained the integrity of their sheath for about 20 days even without the desiccation treatments, indicating the importance of desiccation for N. flagelliforme to prevent them from being disintegrated by bacteria. On the other hand, when free-living cells obtained from crushed colonial filaments were cultured in liquid medium, they developed into single filaments with sheaths, within which multiple filaments were formed later on as a colony. Such colonial filaments were developed at 15, 25, and 30degreesC at either 20 or 60 mumol photons.m(-2).s(-1); colonies did not develop at 180 mumol photons.m(-2).s(-1), though this light level resulted in the most rapid growth of the cells. Conditions of 60 mumol photons.m(-2).s(-1) and 25degrees C appeared to result in the best colonial development and faster growth of the sheath-held colonies of N. flagelliforme when cultured indoor under aquatic conditions.

Kinetics of alkaline phosphatase in lake sediment associated with cage culture of Oreochromis niloticus

Variations in kinetics of alkaline phosphatase occurring in different sites of sediment associated with cage culture of Oreochromis niloticus in a shallow Chinese freshwater lake (Lake Donghu) were described. In addition, the kinetic parameters of each 2.5-cm stratum in the sediment from the surface down to 37.5 cm were analyzed. Horizontally, the V-max values of alkaline phosphatase in surface sediments increased markedly at sites immediately under and adjacent to the cage that would be subjected to the deposition of fish feces. Peak V-max values in the top 5 cm of the sediment under the cage were also observed relative to their deeper control. After a treatment where the fish feces were added over 12 days, the sediment in deeper layer exhibited a significantly higher V-max value, thereby corroborating the relationship between V-max values of alkaline phosphatase and fish feces in sediments. The fish feces exhibited a remarkable alkaline phosphatase activity (APA). Thus, it is indeed a source of the enzyme. Effects of the fish feces were dose- and time-dependent. The V-max values in sediments were always stimulated, but the K-m values showed much more variability. (C) 2001 Elsevier Science B.V. All rights reserved.

In vitro culture of embryonic hearts from guppy fish (Poecilia reticulata)

Forty embryonic hearts were taken out by anatomical needle from denuded embryos of the ovoviviparity guppy fish that were dechorioned by mechanic method or by trypsin digestion, and were in vitro cultured. In the cultured hearts, 80% have maintained beating in vitro for 4 weeks, and the longest record for beating was 142 d. Owing to fish embryo transparency, beating frequency and blood color changes are easily viewed from the embryonic hearts under a dissecting microscope. The current study established the in vitro culture method of embryonic hearts in guppy fish, which can be used as a model for the study of heart and cardiovascular system in vertebrates.

Suspension culture of gametophytes of transgenic kelp in a photobioreactor

Transgenic Laminaria japonica gametophytes producing a recombinant tissue-type plasminogen activator (rtPA) protein, which is an effective third-generation thrombolytic agent for acute myocardial infarction (AMI), were cultured in an illuminated bubble column bioreactor. A maximum final dry cell weight of 1120 mg l(-1) was obtained in batch culture with an initial dry cell weight of 126 mg l(-1) and with aeration rate of 1.2 l air min(-1) l(-1) culture, nitrate at 1.5 mM and phosphate at 0.17 mM. The yield of rtPA was 56 mu g g(-1) dry cell wt. This is the first report regarding cultivation of a transgenic macroalga in a bioreactor.

Mass culture of Undaria gametophyte clones and their use in sporeling culture

Undaria pinnatifida (Harv.) Sur. is one of the three main seaweed species under commercial cultivation in China. In the mid-1990s the annual production was about 20000 tons dry. The supply of healthy sporelings is key to the success of commercial cultivation of Undaria. Previous studies demonstrated that instead of the zoospore collection method, sporelings can be cultured through the use of gametophyte clones. This paper reports the experimental results on mass culture of clones and sporeling raising in commercial scale. Light had an obvious effect on growth of gametophyte clones. Under an irradiance of 80 mumol m(-2) s(-1) and favorable temperature of 22-25degreesC, mean daily growth rate may reach as high as 37%. Several celled gametophyte fragments were sprayed onto the palm rope frame. Gametogenesis occurred after 4-6 days. Juvenile sporeling growth experiments showed that nitrate and phosphate concentrations of 2.9 10(-4) mol 1(-1) and 1.7 10(-5) mol 1(-1) were sufficient to enable the sporelings to maintain a high daily growth rate. Sporelings can reach a length of 1 cm in a month. Since 1997, extension of the clone technique has been carried out in Shandong Province. Large-scale production of sporelings for commercial cultivation of 14 and 31 hectares in 1997 and 1998 had been conducted successfully.

Enhanced anthocyanin production by repeated-batch culture of strawberry cells with medium shift

Repeated-batch cultures of strawberry cells (Fragaria ananassa cv. Shikinari) subjected to four medium-shift procedures (constant LS medium, constant B5 medium, alternation between LS and B5 starting from LS and alternation between LS and B5 starting from B5) were investigated for the enhanced anthocyanin productivity. To determine the optimum period for repeated batch cultures, two medium-shift periods of 9 and 14 days were studied, which represent the end of the exponential growth phase and the stationary phase. By comparison with the corresponding batch cultures, higher anthocyanin productivity was achieved for all the repeated-batch cultures at a 9-day medium-shift period. The average anthocyanin productivity was enhanced 1.7-and 1.76-fold by repeated-batch cultures in constant LS and constant B5 medium at a 9-day shift period for 45 days, respectively. No further improvement was observed when the medium was alternated between LS (the growth medium) and B5 (the production medium). Anthocyanin production was unstable at a 14-day shift period regardless of the medium-shift procedures. The results show that it is feasible to improve anthocyanin production by a repeated-batch culture of strawberry cells.

Formulation of a basal medium for primary cell culture of the marine sponge Hymeniacidon perleve

Marine sponge cell culture is a potential route for the sustainable production of sponge-derived bioproducts. Development of a basal culture medium is a prerequisite for the attachment, spreading, and growth of sponge cells in vitro. With the limited knowledge available on nutrient requirements for sponge cells, a series of statistical experimental designs has been employed to screen and optimize the critical nutrient components including inorganic salts (ferric ion, zinc ion, silicate, and NaCl), amino acids (glycine, glutamine, and aspartic acid), sugars (glucose, sorbitol, and sodium pyruvate), vitamin C, and mammalian cell medium (DMEM and RPMI 1640) using MTT assay in 96-well plates. The marine sponge Hymeniacidon perleve was used as a model system. Plackett-Burman design was used for the initial screening, which identified the significant factors of ferric ion, NaCl, and vitamin C. These three factors were selected for further optimization by Uniform Design and Response Surface Methodology (RSM), respectively. A basal medium was finally established, which supported an over 100% increase in viability of sponge cells.

The ways in which new public management ideas impact upon the administrative culture of 'street-level' bureaucrats and professionals working within Irish social policy

The conventional meaning of culture is ‘widely shared and strongly held values’ of a particular group or society (Bradley and Parker, 2006: 89). Culture is not a rigid concept; it can be influenced or altered by new ideas or forces. This research examines the ways in which one set of ideas in particular, that is, those associated with New Public Management, have impacted upon the administrative culture of 'street-level' bureaucrats and professionals within Irish social policy. Lipsky (1980: 3) defined 'street-level' bureaucrats as ‘public service workers who interact directly with citizens in the course of their jobs, and who have substantial discretion in the execution of their work’. Utilising the Competing Values Framework (CVF) in the analysis of eighty three semi-structured interviews with 'street-level' bureaucrats and professionals, an evaluation is made as to the impact of NPM ideas on both visible and invisible aspects of administrative culture. Overall, the influence of NPM is confined to superficial aspects of administrative culture such as; increased flexibility in working hours and to some degree job contracts; increased time commitment; and a customer service focus. However, the extent of these changes varies depending on policy sector and occupational group. Aspects of consensual and hierarchical cultures remain firmly in place. These coincide with features of developmental and market cultures. Contrary to the view that members of hierarchical and consensual culture would pose resistance to change, this research clearly illustrates that a very large appetite for change exists in the attitudes of 'street-level' bureaucrats and professionals within Irish social policy, with many of them suggesting changes that correspond to NPM ideas. This study demonstrates the relevance of employing the CVF model as it is clear that administrative culture is very much a dynamic system of competing and co-existing cultures.

Ex vivo culture of patient tissue and examination of gene delivery

Gene therapy has emerged as a realistic prospect for the treatment of cancer due to its potential for selective tumour cell targeting. The greatest challenge gene delivery vectors face is the ability to safely and efficiently deliver genes into target cells. The overall objectives of this thesis are to evaluate the efficacy of various gene delivery methods in a clinically relevant tumour model and to also investigate potential strategies for tumour selective delivery. We began with the development of a tumour slice model system using patient waste tissue. This model involves the use of fresh human tumour tissue, cut into thin slices and maintained ex vivo and is universally applicable to gene delivery methods, using a real-time luminescence detection method to assess gene delivery. The nature of the ex vivo culture system permitted examination of specific physiological variables, the influence of intratumoural factors and tissue specific effects on vector expression. Adenoviral vectors under the control of the human CXCR4 promoter demonstrated a 'tumour on' and 'normal off' expression profile when compared with the ubiquitously active CMV promoter when tested in patient tumour tissue. In addition, we developed an ex vivo system of changing oxygenation using the hypoxia inducer, cobalt, to mimic the transient hypoxic conditions found in solid tumours. We found that Adenoviral transgene expression was robust in the cycling hypoxic conditions relevant to solid tumours and re-oxygenation of chronically hypoxic tissue enhanced transgene expression. Finally, we demonstrated an AAV-based tumour targeting strategy using a tumour-selective promoter allowing for the efficient targeting of AAV vectors to cancer cells and the sparing of normal tissue in both murine metastatic liver tumours models and patient tissue. The thesis highlights the importance of indepth preclinical assessment of novel therapeutics and may serve as a platform for further testing of novel gene delivery approaches.

Impact of various endocrine and paracrine factors on in vitro culture of preantral follicles in rodents

Folliculogenesis is a complex process regulated by various paracrine and autocrine factors. In vitro growth systems of primordial and preantral follicles have been developed for future use of immature oocytes, as sources of fertilizable oocytes and for studying follicular growth and oocyte maturation mechanisms. Rodents were often chosen for in vitro follicular culture research and a lot of factors implicated in folliculogenesis have been identified using this model. To date, the mouse is the only species in which the whole process of follicular growth, oocyte maturation, fertilization and embryo transfer into recipient females was successfully performed. However, the efficiency of in vitro culture systems must still be considerably improved. Within the follicle, numerous events affect cell proliferation and the acquisition of oocyte developmental competency in vitro, including interactions between the follicular cells and the oocyte, and the composition of the culture medium. Effects of the acting factors depend on the stage of follicle development, the culture system used and the species. This paper reviews the action of endocrine, paracrine factors and other components of culture medium on in vitro growth of preantral follicles in rodents.

Chondrogenesis and mineralization during in vitro culture of human mesenchymal stem cells on three-dimensional woven scaffolds.

Human mesenchymal stem cells (hMSCs) and three-dimensional (3D) woven poly(ɛ-caprolactone) (PCL) scaffolds are promising tools for skeletal tissue engineering. We hypothesized that in vitro culture duration and medium additives can individually and interactively influence the structure, composition, mechanical, and molecular properties of engineered tissues based on hMSCs and 3D poly(ɛ-caprolactone). Bone marrow hMSCs were suspended in collagen gel, seeded on scaffolds, and cultured for 1, 21, or 45 days under chondrogenic and/or osteogenic conditions. Structure, composition, biomechanics, and gene expression were analyzed. In chondrogenic medium, cartilaginous tissue formed by day 21, and hypertrophic mineralization was observed in the newly formed extracellular matrix at the interface with underlying scaffold by day 45. Glycosaminoglycan, hydroxyproline, and calcium contents, and alkaline phosphatase activity depended on culture duration and medium additives, with significant interactive effects (all p < 0.0001). The 45-day constructs exhibited mechanical properties on the order of magnitude of native articular cartilage (aggregate, Young's, and shear moduli of 0.15, 0.12, and 0.033 MPa, respectively). Gene expression was characteristic of chondrogenesis and endochondral bone formation, with sequential regulation of Sox-9, collagen type II, aggrecan, core binding factor alpha 1 (Cbfα1)/Runx2, bone sialoprotein, bone morphogenetic protein-2, and osteocalcin. In contrast, osteogenic medium produced limited osteogenesis. Long-term culture of hMSC on 3D scaffolds resulted in chondrogenesis and regional mineralization at the interface between soft, newly formed engineered cartilage, and stiffer underlying scaffold. These findings merit consideration when developing grafts for osteochondral defect repair.

[In Backpages 18.1] The culture of new writing

The influence of writing workshops on the content of plays is discussed.