An improved polyurethane support system for monitoring growth in plant cell cultures

An improved culture system for plant cells that employs filter paper resting on polyurethane saturated with liquid medium is described. It combines a simplified version of the system outlined by Weber and Lark [1979, Theor Appl Genet 55: 81–86] with the method of growth estimation described by Horsch et al. [1980, Can J Bot 58: 2402–2406]. The growth of plated cells or callus can be conveniently monitored through repeated non-destructive fresh weight measurements of the filter paper and adhering cells, thereby allowing the construction of a complete growth curve over the course of an experiment. Experiments with 3 Nicotiana genotypes (N. plumbaginifolia Viv., N. tabacum L. ‘SC 58’ and N. tabacum ‘WI 38’) and 3 Vitis vinifera L. genotypes (‘Chenin Blanc’, ‘Dogridge’ and ‘White Riesling’) clearly demonstrate higher growth rates of plated cells on polyurethane supports compared with agar. Further experiments with N. plumbaginifolia illustrate the use of polyurethane supports for culturing cells at low pH (4.0) and the recovery of spent medium for monitoring changes in pH. These features will greatly facilitate quantitative studies of mineral nutrition and metal toxicity in cultured cells. Polyurethane supports also allow the incorporation of conditioned medium or feeder cells to support the growth of cells at low densities and facilitate the rapid recovery of variant cells.     

Endotoxin-induced behavioural fever in the Madagascar cockroach, Gromphadorhina portentosa

Injections of a suspension of E. coli into the haemocoel of Gromphadorhina portentosa induced dose-dependent behavioural fevers. Cockroaches injected with bacteria and placed in a thermal gradient preferred temperatures significantly elevated over those chosen by control cockroaches. The fever response could also be induced by the injection of bacterial endotoxin, lipopolysaccharide-W. This is the first indication that insects are capable of febrogenesis.     

Expression and refolding of recombinant human fibroblast procathepsin D

Procathepsin D is a precursor of the human lysosomal protease cathepsin D. Due to its short half-life, procathepsin D is difficult to obtain in quantities sufficient to allow structural and enzymatic studies. To obtain large quantities of this precursor, procathepsin D was expressed using the T7 promoter vector pET3a in bacteria that carry a chromosomal copy of the T7 RNA polymerase gene under the control of the lac promoter. At high cell density in rich medium, basal levels of T7 RNA polymerase were sufficient to express recombinant procathepsin D without addition of an exogenous inducer of the lac promoter. The recombinant protein, constituting almost half of the total cell protein, accumulated in intracytoplasmic inclusion bodies and was isolated from the insoluble fraction of lysed cells. Antibodies prepared against the purified recombinant protein were shown to crossreact with native human placental and porcine spleen cathepsin D. Recombinant procathepsin D was solubilized in denaturants and was refolded. After extended preincubation of the denatured protein at acid pH to allow folding and activation of the zymogen, pepstatin inhibitable catalytic proteolysis was detected. These data demonstrated that the glycosylated aspartic protease, procathepsin D can be refolded and activated in an unglycosylated form and thus provides a system for the study of procathepsin D structure and function.     

Influence of hurricanes on coastal ecosystems along the northern Gulf of Mexico

Available literature indicates that hurricanes do not generally produce long-term detrimental impacts to unmodified coastal systems and that they often provide net benefits along the U.S. Gulf Coast. While there is normally initial erosion from hurricanes, they also often result in a large influx of inorganic sediments, creating new wetlands and contributing to the maintenance of existing wetlands. The formation of washover deposits is disastrous where cultural development has occurred, but in natural areas these deposits are part of the natural cycle of shoreline development and contribute to habitat diversity and productivity. Abundant rainfall typically associated with hurricanes often results in large increases of sediment and nutrient inputs into coastal estuaries, leading to both short-term and long-term increases in productivity. Rainfall during tropical disturbances accounts for a significant part of total precipitation along the northern gulf. The immediate impact of hurricanes may be to reduce populations of some species but these populations generally recover rapidly. Overall, productivity in natural systems seems to be increased by periodic hurricanes. Hurricane impacts are often severe and long lasting in wetlands that have been modified by human impacts such as semi- or complete impoundments.     

Orthophosphate flux across the membrane of Tetrahymena pyriformis W

Tetrahymena pyriformis W suspended in a buffered glucose solution accumulated orthophosphate [³²P] from the external solution at a measurable rate. The uptake of orthophosphate by the organisms was linear with respect to time when corrections were made to account for a constant efflux observed during the one hour time course of the experiments. Such corrections were based on the measured lowering of the relative specific activity of the suspension medium and led to the derivation of the expressions for the influx and efflux coefficients. The derived expressions for the coefficients are based solely upon the isotopic measurements and by means of these equations it is possible to describe the observed net inward flux of orthophosphate in quantitative terms. The dependence of the uptake of orthophosphate on the external concentration of orthophosphate followed Michaelis-Menten kinetics, and the temperature coefficient (18–28°) of 1.7 for the process fell into the range normally associated with a chemical reaction. The kinetic pattern, per se, does not distinguish a membrane transport mechanism from metabolic incorporation of P_i. Deviations from the expected pattern of uptake were observed at low temperatures.     

Effect of intravenous vitamin C on cytokine activation and oxidative stress in end-stage renal disease patients receiving intravenous iron sucrose

Reticuloendothelial blockade in hemodialysis patients prevents optimal intravenous (IV) iron utilization. Vitamin C has emerged as a potential therapy to improve anemia treatment by enhancing iron mobilization. However, Vitamin C can act as a pro-oxidant in the presence of iron. This was a prospective, open-label, crossover study. Thirteen patients with end-stage renal disease on hemodialysis and four healthy controls were assigned to receive 100?mg of IV iron sucrose (IS) or 100?mg of IV IS co-administered with 300?mg of IV Vitamin C (IS?+?C) in random sequence. Serum samples for IL-1, IL-6, TNF-α and IL-10 and non-transferrin bound iron were obtained at baseline, 45?min and 105?min post study medication administration. Peripheral blood mononuclear cells were isolated at the same time points and stained with fluorescent probes to identify intracellular reactive oxygen species and mitochondrial membrane potential (Δψm) by flow cytometry. Lipid peroxidation was assessed by plasma F2-isoprosatane concentration. Both IS and IS?+?C were associated with increased plasma F2-isoprostanes concentrations post-infusion. Maximal plasma F2-isoprostane concentrations after IS?+?C were significantly elevated from baseline (234?±?0.04 vs. 0.198?±?0.028?ng/mL, p?=?0.02). After IS?+?C, IL-1, IL-6, IL-10, and TNF-alpha were significantly elevated compared to baseline. After IS alone only IL-6 was noted to be elevated. Intracellular production of H(2)O(2) and loss of mitochondrial membrane potential (Δψm) was observed after IS while IS?+?C was associated with increased O (2) (·-) production. Both IS and IS?+?C induced serum cytokine activation accompanied by lipid peroxidation, however, IS?+?C induced higher plasma concentrations of F2-isoprostanes, IL-1, IL-10, and TNF-α post-infusion. Long-term safety studies of IV iron co-administered with Vitamin C are warranted.