Effects of Nerve Growth Factor and Phorbol Derivative on Reactivation of Herpes Simplex Virus Type 1 in Cultured Cells of Latently Infected Adult Mouse Trigeminal Ganglia

Reactivation of herpes simplex virus type 1 (HSV-1) occurred rapidly in cells of latently infected adult mouse trigeminal ganglia which were cultured in serum-free medium in the presence of sufficient nerve growth factor (NGF). However, HSV-1 reactivation was delayed significantly in ganglionic cultures in the absence of exogenous NGF or in cultures treated with 2-aminopurine in the presence of NGF. The delayed viral reactivation in ganglionic cultures without NGF was accelerated by treatment with phorbol myristate acetate or dibutyryl cyclic AMP. Culture conditions which affected HSV-1 reactivation did not affect replication of HSV-1 in normal ganglionic cultures.     

Inter-varietal differences in xylem exudate composition and growth under contrasting forms of N supply in cucumber

Variations in the inorganic and organic composition of xylem exudate, growth and N content under contrasting forms of N supply in three cucumber cultivars (Hyclos, Medusa and Victory) were studied in glasshouse conditions. The plants were grown hydroponically with two NO₃ ^-:NH₄ ⁺ ratios (100:0 and 60:40).The xylem sap of Medusa grown with both N sources displayed an increase of organic N and carboxylate concentrations and a decrease of cations, inorganic anions and carbohydrates compared with that of those grown with NO₃ ^- alone, showing a higher growth and N content in tissues and thus better utilization of N supplied as NO₃ ^- and NH₄ ⁺. Mixed N nutrition in Hyclos caused the greatest amounts of NO₃ ^- and NH₄ ⁺ in xylem sap, lower root weight and N levels in the leaves, while its root was unable to generate an adequate supply of organic N compounds. Despite the levels of cations, inorganic and organic anions were reduced by the NH₄ ⁺ supplied to Victory, the ionic balance in the xylem sap, growth and N content remained similar to that of those supplied with NO₃ ^- alone. Finally, the cucumber cultivars studied here, responded differently to the form of N supplied, it may partly be due to their ability of assimilating N in the roots and partly to the form in which the N is translocated to the shoot.     

Preliminary validation of an activation assay for ex vivo activated T cells utilized in cancer immunotherapy

An in vitro assay that measures the activation level of ex vivo activated (EVA) T cells currently being used in the adoptive immunotherapy of metastatic renal cell carcinoma has been developed. This assay is based on the ability of activated, but not resting. T cells to proliferate in response to the protein kinase C activator, phorbol myristate (PMA). To utilize this assay for in-process monitoring and control, we have begun an initial validation of the overall reproducibility of this assay. The proliferation of activated T cells in response to PMA, as measured by the mean cpm values of (3)H-thymidine incorporated, was demonstrated to have intra-assay coefficients of variation (cv's) for individual analysts that were typically less than 10% and rarely exceeded 20%. Activated T cells could be frozen and stored for at least 6 weeks with little or no deterioration in their ability to proliferate in response to PMA. Using these cells, inter-assay cv's that were typically less than 15% were obtained by individual analysts, and overall cv's of 10% to 25% were obtained for different samples assayed by different analysts at different times. This level of variability is very reasonable for a cellular assay. Furhter validation of this assay will address the issues of sensitivity, linearity and selectivity. To date, this assay has been used to analyze over 90 patient EVA cell samples and has revealed a broad range of proliferative responses to PMA. Taken together, these results suggest that this assay may be useful in defining the potency of the activated T cell used therapeutically.     

Biological destruction of CCl(4): I. Experimental design and data

A denitrifying consortium capable of transforming carbon tetrachloride (CCl(4)) was cultured from aquifer sediment from the U.S. Department of Energy's Hanford Site in southeastern Washington State. To understand the kinetics of the biological destruction of CCl(4) by these microbes, a set of experiments, the conditions of which were chosen according to a fractional factorial experimental design, were completed. This article reports on the experimental design along with the results for CCl(4), biomass, acetate, nitrate, and nitrite concentrations. These data indicate that growth is inhibited by high nitrite concentrations, whereas CCl(4) degradation is slowed by the presence of nitrate and/or nitrite. (c) 1994 John Wiley & Sons, Inc.     

Determination of ammonium and L-Glutamine in hybridoma cell cultures by sequential flow injection analysis

A flow injection anlytical system based on a gas diffusion membrane module for ammonia and an ammonium flow-through potentiometric detector has been set up for measurement of L-glutamine and ammonium ions in hybridoma cell cultures. The main feature of the system is that the same basic analytical concept and equipment is used in both measurements, the only difference being for the determination of L-glutamine, in which the sample flows through an immobilized glutaminase cartridge. The conditions to enable the performance of both analysis consecutively, avoiding potential interferences by unwanted deamination of other compounds in the samples, have been determined. Finally, the proposed system has been compared with reference analytical methods for batch hybridoma cell culture experiments.     

Effects of ammonium sulphate application on the chemistry of bulk soil,rhizosphere, fine roots and fine-root distribution in a Picea abies (L.) karst. stand

The effect of ammonium sulphate application on the bulk and rhizosphere soil chemistry, elemental concentration of living fine roots (<2 mm in diameter), amounts of living and dead fine roots, root length density and specific root length density were investigated in a 28 year old Norway spruce stand in SW Sweden. The treatments started in 1988. Core samples of the LFH layer and mineral soil layers were sampled in control (C) and ammonium sulphate (NS) treatment plots in 1988, 1989 and 1990. Soil pH and NO₃-S and SO₄-S, Al, Ca, Mg, Mn and K concentrations were measured for both the bulk soil and rhizosphere soil.The pH-values of the bulk and rhizosphere soil decreased in 1989 and 1990 in NS plots compared to control plots, while the SO₄-S concentration increased. The Ca, Mg and K concentration increased in the NS treatment in almost all layers in the bulk and the rhizosphere soil. Ammonium ions may have replaced these elements in the soil organic matter. The NS treatment reduced Mg concentration in fine roots in all layers in 1990. The Al concentrations in the rhizosphere and bulk soil were higher in NS plots in all layers, except at 0–10 cm depth, both in 1989 and 1990. The Al content of living fine roots was higher in NS plots than C plots but the differences were not significant. The NS addition did not affect the P and K contents of fine roots in any soil layer, but the S concentrations of fine roots were significantly higher in NS plots in 1989 and 1990. The fine root necromass was higher in NS than in C in 1990, in the LFH layer, indicating a gradual decrease in the vitality of the fine roots. It was suggested that the NS treatment resulted in displacement of Mg and K from exchange sites in the LFH layer leading to leaching of these cations to the mineral soil. Further application of ammonium sulphate may damage the fine roots and consequently adversely affect the water and nutrient uptake of root systems.     

Ammonium uptake in carrot cell structures is influenced by pH-dependent cell aggregation

Semicontinuously grown wild carrot ( Daucus carota L.) cells were used in an investigation of the effect of culture medium pH on ammonium uptake in suspension cultures as a first step in exploring the relationship between pH and anthocyanin biosynthesis. In contrast to published data showing decreasing uptake rates with decreasing culture medium pH, ammonium-limited, semicontinuous carrot cell cultures showed a 25% greater ammonium uptake rate at pH 4.5 than at pH 5.5. When cells that had been grown semicontinuously in medium with a pH of 4.5 or 5.5 were grown in batch cultures at pH 4.5, 5.5 or 6.5 the ammonium uptake rates were those of the semicontinuous cultures, indicating that the pH of the batch culture medium had no effect on ammonium uptake rates over 7 days. The cell culture was composed of very small aggregates when it was grown semicontinuously in medium at pH 4.5, but was composed of large aggregates when it was grown semicontinuously in medium at pH 5.5. The aggregation/disaggregation of the cells was pH dependent, as changing the pH of the semicontinuous culture medium altered the extent of the aggregation. We conclude that the change in culture medium pH caused the cells to aggregate or disaggregate which in turn decreased or increased the rate of ammonium uptake from the medium.     

Soil nitrogen transformations and nitrate utilization by Deschampsia flexuosa (L.) Trin. at two contrasting heathland sites

Two Dutch heathland sites Hoorneboeg (HB) and Ede, dominated by Deschampsia flexuosa and differing in nitrate production, were sampled for an entire growing season. A large number of soil and plant parameters were monitored in an attempt to assess the contribution of nitrate in the N supply and its assimilation by Deschampsia.Average NO₃ ^– and NH₄ ⁺ concentrations (mg kg^–1) in the top 10-cm depth were 0.03 and 2.2, respectively, for HB, and 2.1 and 6.7, respectively, for Ede. Laboratory incubations of intact cores and experiments with FH-layer suspensions showed significantly higher mineralization and nitrification rates for the Ede site during most of the season. Nitrification was largely controlled by the rate of net N-mineralization, which in turn was highly affected by soil moisture. Nitrate production was virtually zero at HB and accounted for 25% of the net N-mineralization at Ede.Shoot chemical composition showed no essential differences for the two sites, but mean in vivo (current) foliar NRA was almost 2-fold higher at Ede than at HB, indicating some utilization of nitrate at the former location. At the HB site with essentially no nitrate production, however, enzyme activities were clearly higher than basal constitutive levels in NH₄ ⁺-fed plants. Apparently, shoot NRA at the HB site became positively affected by factors other than nitrate availability and/or showed disproportional increases in response to atmospheric nitrate inputs. Root NRA displayed the same low basal level at the two sites. Nitrate fertilization (100 kg N ha^–1) yielded maximally induced foliar NRAs similar to levels found in hydroponic nitrate plants. Although no accumulation of free NO₃ ^– was observed in shoots from fertilized plots, increases in foliar concentrations of both organic N and carboxylates clearly indicated nitrate assimilation. Root NRA showed no response to nitrate addition.It is concluded that current NRA measurements in Deschampsia at heathland sites are of limited value only, especially when interpreted in isolation. A combined approach, using concurrently conducted soil and plant analyses, will allow the extent of nitrate utilization in the field to be best characterized.Publication 2013 of the Netherlands Institute of Ecology.FAX no corresponding author: +31 8306 23227