Mineral nutrition and plant morphogenesis

Plant morphogenesis in vitro can be achieved via two pathways, somatic embryogenesis or organogenesis. Relationships between the culture medium and explant leading to morphogenesis are complex and, despite extensive study, remain poorly understood. Primarily the composition and ratio of plant growth regulators are manipulated to optimize the, quality and numbers of embryos or organs initiated. However, many species and varieties do not respond to this classical approach and require further optimization by the variation of other chemical or physical factors. Mineral nutrients form a significant component of culture media but are often overlooked as possible morphogenic elicitors. The combination of minerals for a particular plant species and developmental pathway are usually determined by the empirical manipulation of one or a combination of existing published formulations. Often only one medium type is used for the duration of culture even though this formulation may not be optimal for the different stages of explant growth and development. Furthermore, mineral studies have often focused on growth rather than morphogenesis with very little known of the relationships between mineral uptake and morphogenesis. This article examines the present knowledge of the main effects that mineral nutrients have on plant morphogenesis in vitro. In particular, the dynamics of nitrogen, phosphorus, and calcium supply during development are discussed.

The number of long-lasting functional memory CD8(+) T cells generated depends on the nature of the initial nonspecific stimulation

The mechanism of generation of memory cytotoxic T cells (CTL) following immunization remains controversial. Using tumor protection and IFN-gamma ELISPOT assays in mice to detect functional CTL, we show that the initial effector CTL burst size after immunization is not directly related to the amount of functional memory CTL formed, suggesting that memory CTL are unlikely to arise stochastically from effector CTL. Induction of MHC class II-restricted T helper cells at the time of immunization by inclusion of a T helper peptide or protein in the immunogen, is necessary to generate memory CTL, although no T helper cell induction is required to generate effector CTL to a strong MHC class I-binding peptide. Host protective T cell memory correlates with the number of CTL epitope responsive IFN-gamma-secreting memory T cells as measured in an ELISPOT assay at the time of tumor challenge. We conclude that a different antigen presenting environment is required to induce long-lasting functional memory CTL, and non-cognate stimulation of the immune system is essential to allow generation of a long-lasting host protective memory CTL response.

The identification of the transcriptional regulator CRP in Aeromonas hydrophila JMP636 and its involvement in amylase production and the 'acidic toxicity' effect

Aims: The physiological examination of amylase production by Aeromonas hydrophila JMP636 and identification of the mechanism of regulation. Methods and Results: Aeromonas hydrophila JMP636 was grown with single, then dual carbon sources; the growth cycle was followed and amylase activity throughout was monitored. The levels of cAMP, a known secondary messenger for the regulatory gene crp, were also examined. Amylase activity was regulated by catabolite repression. Physiological studies revealed that JMP636 exhibited both diauxic growth, with two carbon sources, and the 'acid toxicity' effect on glucose. The crp gene was cloned, expressed and inactivated from the JMP636 chromosome. Catabolite repression of amylase production and the 'acid toxicity' effect both require crp and were linked to cAMP levels. Conclusions: Regulation of amylase production was predicted to follow the model CRP-mediated cAMP-dependent Escherichia coli catabolite regulation system. Significance and Impact of the Study: This work provides an understanding of the physiology of the opportunistic pathogen Aer. hydrophila through identification of the mechanism of catabolite repression of amylase production and the existence of crp within this cell. It also provides a broader knowledge of global gene regulation and suggests regulatory mechanisms of other Aer. hydrophila gene/s.

Micro-mechanical approximation to the stress field around an inclined fiber of FRCC

Matrix spalling or crushing is one of the important mechanisms of fiber-matrix interaction of fiber reinforced cementitious composites (FRCC). The fiber pullout mechanisms have been extensively studied for an aligned fiber but matrix failure is rarely investigated since it is thought not to be a major affect. However, for an inclined fiber, the matrix failure should not be neglected. Due to the complex process of matrix spalling, experimental investigation and analytical study of this mechanism are rarely found in literature. In this paper, it is assumed that the load transfer is concentrated within the short length of the inclined fiber from the exit point towards anchored end and follows the exponential law. The Mindlin formulation is employed to calculate the 3D stress field. The simulation gives much information about this field. The 3D approximation of the stress state around an inclined fiber helps to qualitatively understand the mechanism of matrix failure. Finally, a spalling criterion is proposed by which matrix spalling occurs only when the stress in a certain volume, rather than the stress at a small point, exceeds the material strength. This implies some local stress redistribution after first yield. The stress redistribution results in more energy input and higher load bearing capacity of the matrix. In accordance with this hypothesis, the evolution of matrix spalling is demonstrated. The accurate prediction of matrix spalling needs the careful determination of the parameters in this model. This is the work of further study. (C) 2002 Elsevier Science Ltd. All rights reserved.

Inhibition of adventitious rooting in Backhousia citriodora F. Muell cuttings correlate with the concentration of essential oil

Backhousia citriodora is typical of the many commercially valuable woody Australian Myrtaceae species that are recalcitrant in forming adventitious roots from cuttings after maturation. A series of experiments were conducted to identify an endogenous rooting inhibitor in line with established criteria. Endogenous levels of citral were correlated with the rooting capacities of juvenile versus mature, and easy- versus difficult-to-root genotypes of B. citriodora, in both winter and summer. The biological activity of citral was confirmed in bioassays on mung beans and easy-to-root B. citriodora seedlings. Evidence of a common mechanism of root inhibition with other species in the Myrtaceae and the role of action of citral are discussed.

A review of animal models for the study of arsenic carcinogenesis

As inorganic arsenic is a proven human carcinogen, significant effort has been made in recent decades in an attempt to understand arsenic carcinogenesis using animal models, including rodents (rats and mice) and larger mammals such as beagles and monkeys. Transgenic animals were also used to test the carcinogenic effect of arsenicals, but until recently all models had failed to mimic satisfactorily the actual mechanism of arsenic carcinogenicity. However, within the past decade successful animal models have been developed using the most common strains of mice or rats. Thus dimethylarsinic acid (DMA), an organic arsenic compound which is the major metabolite of inorganic arsenicals in mammals, has been proven to be tumorigenic in such animals. Reports of successful cancer induction in animals by inorganic arsenic (arsenite and arsenate) have been rare, and most carcinogenetic studies have used organic arsenicals such as DMA combined with other tumor initiators. Although such experiments used high concentrations. of arsenicals for the promotion of tumors, animal models using doses of arsenicals species closed to the exposure level of humans in endemic areas are obviously the most significant. Almost all researchers have used drinking water or food as the pathway for the development of animal model test systems in order to mimic chronic arsenic poisoning in humans; such pathways seem more likely to achieve desirable results. (C) 2002 Elsevier Science Ireland Ltd. All rights reserved.

Phenotypic and genotypic analysis of Helicoverpa armigera nucleopolyhedrovirus serially passaged in cell culture

Rapid accumulation of few polyhedra (FP) mutants was detected during serial passaging of Helicoverpa armigera nucleopolyhedrovirus (HaSNPV) in cell culture. 100% FP infected cells were observed by passage 6. The specific yield decreased from 178 polyhedra per cell at passage 2 to two polyhedra per cell at passage 6. The polyhedra at passage 6 were not biologically active, with a 28-fold reduction in potency compared to passage 3. Electron microscopy studies revealed that very few polyhedra were produced in an FP infected cell (< 10 polyhedra per section) and in most cases these polyhedra contained no virions. A specific failure in the intranuclear nucleocapsid envelopment process in the FP infected cells, leading to the accumulation of naked nucleocapsids, was observed. Genomic restriction endonuclease digestion profiles of budded virus DNA from all passages did not indicate any large DNA insertions or deletions that are often associated with such FP phenotypes for the extensively studied Autographa californica nucleopolyhedrovirus and Gaileria mellonella nucleopolyhedrovirus. Within an HaSNPV 25K FP gene homologue, a single base-pair insertion (an adenine residue) within a region of repetitive sequences (seven adenine residues) was identified in one plaque-purified HaSNPV FP mutant. Furthermore, the sequences obtained from individual clones of the 25KFP gene PCR products of a late passage revealed point mutations or single base-pair insertions occurring throughout the gene. The mechanism of FP mutation in HaSNPV is likely similar to that seen for Lymantria dispar nucleopolyhedrovirus, involving point mutations or small insertions/deletions of the 25K FP gene.

The relationship of natural killer cell counts, perforin mRNA and CD2 expression to post-exercise natural killer cell activity in humans

The aim of this study was to further investigate the mechanism of suppression of natural killer (NK) cell cytotoxic activity In peripheral blood following strenuous exercise. Blood was collected for analysis of NK cell concentration, cytotoxic activity, CD2 surface expression and perforin gene expression from runners (RUN, n = 6) and resting controls (CONTROL, n = 4) pre-exercise, 0, 1.5, 5, and 24 h following a 60-min treadmill run at 80% of VO2 peak. Natural killer cytotoxic activity, measured using a whole blood chromium release assay, fluctuated minimally in the CONTROL group and increased by 63% and decreased by 43% 0 and 1.5 h post-exercise, respectively, in the RUN group (group x time, P < 0.001). Lytic index (cytotoxic activity per cell) did not change. Perforin mRNA, measured using quantitative real-time polymerase chain reaction (ORT-PCR) decreased from pre- to post-exercise and remained decreased through 24 h, The decrease from pre- to 0 In post-exercise was seen predominately in the RUN group and was inversely correlated r = - 0.95) to pre-exercise perform mRNA. The NK cell surface expression of CD2 (lymphocyte function-associated antigen-2) was determined using fluorescent antibodies and flow cytometry, There was no change in the proportion of NK cells expressing CD2 or CD2 density, We conclude that (1) numerical redistribution accounted for most of the change in NK cytotoxic activity following a strenuous run, (2) decrease in perforin gene expression during the run was inversely related to pre-exercise levels but did not parallel changes in cytotoxic activity, and (3) CD2 surface expression was not affected by exercise.

Columnar to equiaxed transition of eutectic in hypoeutectic aluminium-silicon alloys

Directional solidification of unmodified and strontium modified binary, high-purity aluminium-7 wt% silicon and commercial A356 alloys has been carried out to investigate the mechanism of eutectic solidification. The microstructure of the eutectic growth inter-face was investigated with optical microscopy and Electron Backscattering Diffraction (EBSD). In the commercial alloys, the eutectic solidification inter-face extends in the growth direction and creates a eutectic mushy zone. A planar eutectic growth front is observed in the high-purity alloys. The eutectic aluminium has mainly the same crystallographic orientation as the dendrites in the unmodified alloys and the strontium modified high-purity alloy. A more complex eutectic grain structure is found in the strontium modified commercial alloy. A mechanism involving constitutional undercooling and a columnar to equiaxed transition explains the differences between pure and commercial alloys. It is probably caused by the segregation of iron and magnesium and the activation of nucleants in the commercial alloy. (C) 2002 Acta Materialia Inc. Published by Elsevier Science Ltd. All rights reserved.

Adsorption of aromatic compounds onto activated carbons: Effects of the orientation of the adsorbates

Adsorption of model aromatic compounds onto two untreated activated carbons with similar physical and chemical properties is investigated. The solution pH of all experiments was lowered so that all solutes were in their molecular forms. It is shown that the difference in the maximum adsorption capacities of the solutes was mainly attributed to the difference in the sizes of the molecules. This new experimental finding is significant to gaining insight into the orientation of the adsorbed phase and hence the adsorption mechanism of aromatic compounds in aqueous solutions. It is shown that the adsorption of aromatic compounds in a stacked motif for pi-pi interactions is unlikely, and in the absence of physical restrictions such as pore width, a T-shaped motif is the preferred orientation.

Hepatic xenobiotic metabolism of cylindrospermopsin in vivo in the mouse

Cylindrospermopsin (CYN) is a hepatotoxin isolated from the blue-green alga Cylindrospermopsis raciborskii. The role of both glutathione (GSH) and the cytochrome P450 enzyme system (P450) in the mechanism of toxicity of CYN has been previously investigated in in vitro systems. We have investigated the role of GSH and P450 in vivo in mice. Mice pre-treated with buthionine sulphoximine and diethyl maleate to deplete hepatic GSH prior to dosing with 0.2 mg/kg CYN showed a seven-day survival rate of 5/13 while the control group rate was 9/14. Dosing mice with 0.2 mg/kg CYN produced a small decrease in hepatic GSH with a characteristic rebound effect at 24 h, The magnitude of this effect is however small and combined with the non-significant difference in survival rates after GSH depletion suggest depletion of GSH by CYN could not be a primary mechanism for CYN toxicity, Conversely, pro-treatment with piperonyl butoxide, a P450 inhibitor, protected mice against CYN toxicity giving a survival rate of 10/10 compared with 4/10 in the control group (p < 0.05 Chi squared) and was protective at doses up to 0.8 mg/kg, suggesting activation of CYN by P450 is of primary importance in the mechanism of action. (C) 2002 Elsevier Science Ltd. All rights reserved.

Study on diffusion and flow of benzene, n-hexane and CCl4 in activated carbon by a differential permeation method

In this paper the diffusion and flow of carbon tetrachloride, benzene and n-hexane through a commercial activated carbon is studied by a differential permeation method. The range of pressure is covered from very low pressure to a pressure range where significant capillary condensation occurs. Helium as a non-adsorbing gas is used to determine the characteristics of the porous medium. For adsorbing gases and vapors, the motion of adsorbed molecules in small pores gives rise to a sharp increase in permeability at very low pressures. The interplay between a decreasing behavior in permeability due to the saturation of small pores with adsorbed molecules and an increasing behavior due to viscous flow in larger pores with pressure could lead to a minimum in the plot of total permeability versus pressure. This phenomenon is observed for n-hexane at 30degreesC. At relative pressure of 0.1-0.8 where the gaseous viscous flow dominates, the permeability is a linear function of pressure. Since activated carbon has a wide pore size distribution, the mobility mechanism of these adsorbed molecules is different from pore to pore. In very small pores where adsorbate molecules fill the pore the permeability decreases with an increase in pressure, while in intermediate pores the permeability of such transport increases with pressure due to the increasing build-up of layers of adsorbed molecules. For even larger pores, the transport is mostly due to diffusion and flow of free molecules, which gives rise to linear permeability with respect to pressure. (C) 2002 Elsevier Science Ltd. All rights reserved.

Effects of surface chemistry on aromatic compound adsorption from dilute aqueous solutions by activated carbon

Adsorption of one nondissociating and four dissociating aromatic compounds onto three untreated activated carbons from dilute aqueous solutions were investigated. All adsorption experiments were preformed in pH-controlled solutions. The experimental isotherms were analyzed using the homogeneous Langmuir model. The surface chemical properties of the activated carbons were characterized using a combination of water adsorption, X-ray photoemission spectroscopy, and mass titration. These data give rise to a new insight into the adsorption mechanism of aromatic solutes, in their molecular and ionic forms, onto untreated activated carbons. It was found that, for the hydrophilic activated carbons, the dominant adsorption forces were observed to be dipolar interactions when the solutes were in their molecular form whereas dispersive forces, such as pi-pi interactions, were most likely dominant in the case of the basic hydrophobic carbons. However, when the solutes were in their ionic form adsorption occurs in all cases through dispersive forces.

Secondary structure of the third extracellular loop responsible for ligand selectivity of a mammalian gonadotropin-releasing hormone receptor

The extracellular loop 3 (ECL3) of the mammalian gonadotropin-releasing hormone receptor (GnRH-R) contains an acidic amino acid (Glu(301) in the mouse GnRH-R,) that confers agonist selectivity for Are in mammalian GnRH. It is proposed that a specific conformation of ECL3 is necessary to orientate the carboxyl side chain of the acidic residue for interaction with Arg(8) of GnRH, which is supported by decreased affinity for Arg(8) GnRH but not Gln(8) GnRH when an adjacent Pro is mutated to Ala. To probe the structural contribution of the loop domain to the proposed presentation of the carboxyl side chain, we synthesized a model peptide (CGPEMLNRVSEPGC) representing residues 293-302 of mouse ECL3, where Cys and Gly residues are added symmetrically at the N and C termini, respectively, allowing the introduction of a disulfide bridge to simulate the distances at which the ECL3 is tethered to the transmembrane domains 6 and 7 of the receptor. The ability of the ECL3 peptide to bind GnRH with low affinity was demonstrated by its inhibition of GnRH stimulation of inositol phosphate production in cells expressing the GnRH-R. The CD bands of the ECL3 peptides exhibited a superposition of predominantly unordered structure and partial contributions from beta-sheet structure. Likewise, the analysis of the amide I and amide III bands from micro-Raman and FT Raman experiments revealed mainly unordered conformations of the cyclic and of the linear peptide. NMR data demonstrated the presence of a beta-hairpin among an ensemble of largely disordered structures in the cyclic peptide. The location of the turn linking the two strands of the hairpin was assigned to the three central residues L-296, N-297, and R-298. A small population of structured species among an ensemble of predominantly random coil conformation suggests that the unliganded receptor represents a variety of structural conformers, some of which have the potential to make contacts with the ligand. We propose a mechanism of receptor activation whereby binding of the agonist to the inactive receptor state induces and stabilizes a particular structural state of the loop domain, leading to further conformational rearrangements across the transmembrane domain and signal propagating interaction with G proteins. Interaction of the Glu(301) of the receptor with Arg(8) of GnRH induces a folded configuration of the ligand. Our proposal thus suggests that conformational changes of both ligand and receptor result from this interaction.

Inhibition of rat platelet aggregation by the diazeniumdiolate nitric oxide donor MAHMA NONOate

1 Inhibition of rat platelet aggregation by the nitric oxide (NO) donor MAHMA NONOate (Z-1-{N-methyl-N-[6-(N-methylammoniohexyl)amino]}diazen-l-ium-1,2-diolate) was investigated. The aims were to compare its anti-aggregatory effect with vasorelaxation, to determine the effects of the soluble guanylate cyclase inhibitor, ODQ (1H-[1,2,4]oxadiazolo[4,3-ajquinoxalin-1-one), and to investigate the possible role of activation of sarco-encloplasmic reticulum calcium-ATPase (SERCA), independent of soluble guanylate cyclase, using thapsigargin. 2 MAHMA NONOate concentration-dependently inhibited sub-maximal aggregation responses to collagen (2 - 10 mug ml(-1)) and adenosine diphosphate (ADP; 2 mum) in platelet rich plasma. It was (i) more effective at inhibiting aggregation induced by collagen than by ADP, and (ii) less potent at inhibiting platelet aggregation than relaxing rat pulmonary artery. 3 ODQ (10 mum) caused only a small shift (approximately half a log unit) in the concentration-response curve to MAHMA NONOate irrespective of the aggregating agent. 4 The NO-independent activator of soluble guanylate cyclase, YC-1 (3-(5'-hydroxymethyl-2'-furyl)-1-benzy] indazole; 1 - 100 mum), did not inhibit aggregation. The cGMP analogue, 8-pCPT-cGMP (8-(4-chlorophenylthio)guanosine 3'5' cyclic monophosphate; 0.1 - 1 mm), caused minimal inhibition. 5 On collagen-aggregated platelets responses to MAHMA NONOate (ODQ 10 PM present) were abolished by thapsigargin (200 nm). On ADP-aggregated platelets thapsigargin caused partial inhibition. 6 Results with S-nitrosoglutathione (GSNO) resembled those with MAHMA NONOate. Glyceryl trinitrate and sodium nitroprusside were poor inhibitors of aggregation. 7 Thus inhibition of rat platelet aggregation by MAHMA NONOate (like GSNO) is largely ODQ-resistant and, by implication, independent of soluble guanylate cyclase. A likely mechanism of inhibition is activation of SERCA.