The regeneration potential of wheat shoot meristems in the presence and absence of 2,4-dichlorophenoxyacetic acid

Summary Tissue cultures capable of plant regeneration were successfully initiated from extremely immature shoot meristems of 21 randomly selected genotypes of wheat on nutrient media containing 2,4-dichlorophenoxyacetic acid (2,4-D). By means of scanning electron microscopy it was demonstrated that cultures consisted of teratomatous primordia, which were kept in a proliferating budding state by the 2,4-D. These are characteristic of cereal tissue cultures. Release of the primordia and outgrowth of normal shoots and roots occurred when the cultures were no longer exposed to 2,4-D. Shoot primordia which were clearly identifiable were always associated with root primordia in a quasi-bipolar fashion. Sometimes regions assumed the shape of zygotic embryos, but the transition from apparently normal embryos with scutellum to abnormal configurations with shoot and root regions was gradual. The differences between genotypes in shoot regeneration potential was minimal compared to cultures derived from explants which were taken from regions temporally and spatially more distant from the shoot apex. It is concluded that the ability to give rise to cultures capable of shoot regeneration was lost within a fraction of a millimeter distance from the apical meristem in many genotypes. The proliferating tissues were subcultured at regular intervals over a period of one year and the regeneration potential was monitored. Areas capable of shoot regeneration tended to deteriorate more or less rapidly and were overgrown by root-type tissue in a number of genotypes. The results are discussed in the context of the frequently observed, but largely unexplained, variability in the regeneration potential of cereal tissue cultures.     

Se和环境中富里酸对小麦种子发芽的影响及其生理特性

利用种子发芽实验研究了亚硒酸钠（ＮＶ２ＳｅＯ３）和从土壤中提取的富里酸（Ｆｕｌｖｉｃ ａｃｉｄ, ＦＡ）对小麦种子发芽和生长的影响,以及 ＦＡ对亚硒酸钠毒性的抑制作用．结果表 明．ＦＡ和低浓度Ｎａ２ＳｅＯ３对种子发芽及发芽后的生长有促进作用,高浓度 Ｎａ２ＳｅＯ３明显 降低种子发芽率、种子活力、α－淀粉酶活性及幼苗的生长,ＦＡ对亚硒酸钠的毒性有一定的 抑制作用．     

Protective effects of selenium against cadmium induced hematological disturbances,immunosuppressive, oxidative stress and hepatorenal damage in rats

Cadmium is a non-essential toxic metal used in industrial process, causes severe risk to human health. Selenium (Se) is an essential trace mineral of fundamental importance for human health. Selenium has antioxidant enzymes roles and is needed for the proper function of the immune system. In this study, the protective effects of selenium against cadmium intoxication in rats have been investigated by monitoring some selective cytokines (IL-1β, TNF α, IL-6, IL-10 and IFN-γ), antioxidant enzymes reduced glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and lipid peroxidation malondialdehyde (MDA) as well as some selective biochemical markers of liver and kidney functions. Thirty-two rats were divided into four equal groups; the first group was used as a control. Groups 2–4 were treated with selenium (Se; 0.1 mg/kg BW), cadmium (Cd; 40 mg/L drinking water) and selenium plus cadmium, respectively. Rats were orally administered their relevant doses daily for 30 days. Blood samples were collected from heart puncture at the end of the experiment (30 days) for complete blood picture (CBC) and serum was separated to evaluate the different immunological parameters and biochemical parameters, as well as liver specimens for Cd and Se estimation. Rats in the Cd treated group have a significantly higher hepatic concentration of Cd than in other treated groups. Results revealed that cadmium significantly increased IL-1β, TNF α, IL-6 and IL-10, beside peripheral neutrophils count, while the IFN-γ and lymphocytes were decreased in rat sera. In addition, GSH level, CAT, SOD and GPx activities were significantly decreased while lipid peroxidation (MDA) was increased. Regarding, liver and renal markers, they were significantly increased in the activities of aminotransferases (AST, ALT), urea and creatinine, while total plasma proteins and albumin were significantly decreased. On the other hand, selenium treated group, showed significantly increased IFN-γ, GSH level, CAT, and GPx activities, as well as lymphocyte count while IL-10 was decreased. Selenium in combination with cadmium, significantly improved the elevation of serum IL-1β, IL-6, TNF α, IL-10 and malondialdehyde in addition to enhancing the antioxidant enzyme activities of GSH, CAT, GPx and SOD. Moreover, selenium has ameliorated the cadmium-induced liver and kidney damage by improving hepatic and renal markers. The results of this investigation demonstrated that selenium has the potential to countermeasure the immunosuppressive as well as hepatic and renal oxidative damage induced by cadmium in rats; selenium has shown promising effects against Cd toxicity.     

Modulatory effects of lipoic acid and selenium against cadmium-induced biochemical alterations in testicular steroidogenesis

Exposure to toxic metals including cadmium has become an increasingly recognized source of illness worldwide. Cadmium (Cd(2+) ) is one of the environmental pollutants affecting various tissues and organs including testis. The protective effect of lipoic acid and selenium on Cd(2+) -induced testicular damage was investigated. Accordingly, male Wistar rats were allocated into four groups (n = 8; each). Gp I: (control), whereas the other 3 groups received CdCl(2) (2 mg/kg, i.p. for 28 days) alone or in combination with either (i) lipoic acid (35 mg/kg, p.o) or (ii) selenium (0.35 mg/kg, p.o) throughout the experiment. Serum testosterone, luteinizing hormone and follicle-stimulating hormone levels significantly decreased in the Cd(2+) -exposed rats. The activities of testicular key androgenic enzymes, 3β-hydroxysteroid dehydrogenase and 17 β-HSD significantly decreased in Cd(2) exposed rats compared to the control counterparts. In addition, the activities of testicular marker enzymes were significantly altered in cadmium-treated animals. Significant reductions in body and testicular weight as well as antioxidant status were also observed in Cd(2+) -exposed rats. Moreover, some testicular metal levels were altered. Lipoic acid and selenium significantly increased serum testosterone level and restored testicular activity of 3β-HSD and 17 β-HSD and were effective in modulation of most of the measured biochemical parameters. The biochemical parameters were further confirmed with histopathological findings. In conclusion, the present study demonstrated the beneficial influences of lipoic acid and selenium in reducing harmful effects of Cd(2+) in rats' testes.     

Electrophoretic variation in glutamate dehydrogenase and other isozymes in wild carrot cells cultured in the presence and absence of 2,4-dichlorophenoxyacetic acid

Summary Electrophoretic analysis of isozymal differences was performed with extracts of wild carrot (Daucus carota L.) cells, grown in the presence and absence of 2,4-dichlorophenoxyacetic acid (2,4-D). There were no differences in the patterns of malate dehydrogenase, acid phosphatase, aspartate aminotransferase, and γ-glutamyl transferase. Quantitative differences in peroxidase isozymes were detected, the plus 2,4-D cultures having lower activities. Esterase patterns were similar, but there were differences in individual isozyme activities and an additional form present in the minus 2,4-D cells. the greatest differences were in patterns of glutamate dehydrogenase with the minus 2,4-D cultures containing only the slowly migrating isozymes. The changes in glutamate dehydrogenase, as revealed by isozyme changes, together with the requirement for ammonia in embryogenesis, suggests that this enzyme may be associated with differentiation in wild carrot cells.     

Changes in membrane properties of rat red blood cells induced by cadmium accumulating in the membrane fraction

When rat red blood cells were incubated in a cadmium (Cd)-free medium following 1-h pretreatment with 0.5 mM CdCl2, incorporated Cd was retained in the cell during 14-h incubation and progressively accumulated in the membrane fraction, especially in the cytoskeleton fraction. In parallel to this accumulation, red cell filterability decreased and echinocytic cells increased, although intracellular ATP was maintained at the control level. The echinocytic shape was maintained in ghosts and cytoskeletons prepared from the Cd-loaded cells. In addition, the association of bands 2.1, 3, 4.2, and 4.5 with cytoskeletons increased and dissociation of cytoskeletal networks at low ionic strength decreased as the incubation time increased. Pretreatment of red blood cells with Cd also induced a release of small vesicles. These vesicles contained hemoglobin but were depleted of spectrin and actin, showing a phospholipid composition similar to that of red cell ghosts. These results suggest that the organization of cell membranes, especially cytoskeletal networks, is altered by Cd accumulation in the cytoskeleton fraction, which results in acceleration of age-related changes of red blood cells such as shape change and decreased filterability.     

Involvement of non-enzymatic antioxidant defenses in the protective effect of diphenyl diselenide on testicular damage induced by cadmium in mice

The involvement of non-enzymatic antioxidant defenses in the protective effect of diphenyl diselenide (PhSe)₂ on testicular damage caused by cadmium in mice was investigated. Mice received a single dose of CdCl₂ (5 mg/kg, intraperitoneally). Thirty minutes after the CdCl₂ injection, they received a single oral dose of (PhSe)₂ (400 μmol/kg). Twenty-four hours after CdCl₂ administration, blood samples were collected and mice were killed and had their testes dissected. Parameters in plasma (aspartate (AST) and alanine (ALT) aminotransferases and lactato dehydrogenase (LDH) activities as well as creatinine levels) were determined. The activity of δ-aminolevulinate dehydratase (δ-ALA-D), the levels of thiobarbituric acid-reactive substances (TBARS), ascorbic acid and nonprotein thiols (NPSH) and histological analysis were determined in collected samples. Results demonstrated that (PhSe)₂ protected against toxicity induced by CdCl₂ on δ-ALA-D activity, ascorbic acid and NPSH levels. (PhSe)₂ protected against the increase in plasma AST, ALT and LDH activities caused by CdCl₂. Testes of mice exposed to CdCl₂ showed marked histopathological alterations that were ameliorated by administration of (PhSe)₂. (PhSe)₂ protected against toxicity induced by CdCl₂ in testes of mice. Ascorbic acid and NPSH, non-enzymatic antioxidant defenses, are involved in the protective effect of (PhSe)₂ against testicular damage caused by CdCl₂ in mice.     

Changes of 6-benzylaminopurine and 2,4-dichlorophenoxyacetic acid concentrations in plant tissue culture media in the presence of activated charcoal

Somatic embryos and embryogenic callus were initiated from immature zygotic embryos of ginseng (Panax ginseng C.A. Meyer). These somatic embryos were multiplied by adventitious (secondary and tertiary) embryogenesis and their growth and development were dependent on growth hormones in the medium. Auxins, 2,4-d, NAA, and IAA at 1.0 mg l^-1 were effective in inducing secondary and tertiary somatic embryos, which proliferated directly from the apical or cotyledonary portions of the primary somatic embryos. Single somatic embryos or clusters or embryos developed from the explanted primary embryos. Cytokinin (Kn, BA) inhibited adventitious embryogenesis. Secondary somatic embryos developed to maturation and later regenerated into plantlets in two stage process; firstly elongation of the shoot axes on MS +1.0 mg l^-1 Kn, secondly formation of root on 1.0 mg l^-1 Kn+1.0 mg^-1 GA₃ medium.Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid - NAA naphthaleneacetic acid - IAA in-doleacetic acid - Kn kinetin - BA benzylaminopurine - PSE primary somatic embryo - SSE secondary somatic embryo - TSE tertiary somatic embryo     

Effect of 2,4-dichlorophenoxyacetic acid on callus induction and plant regeneration in anther culture of wheat (Triticum aestivum L.)

 Anthers from a doubled-haploid line of spring wheat (Triticum aestivum L.) cv. Pavon 76 were plated in liquid P-4 medium supplemented with 2,4-dichlorophenoxyacetic acid (2,4-D) at four concentrations (0.5, 1.0, 2.0, 4.0 mg/l) for 5, 10, 15, and 25 days before being transferred to another medium with the same or reduced 2,4-D concentrations for the remainder of the induction phase for a total of 45 days. Incubation with 0.5 mg/l 2,4-D for 45 days produced lower callus yield and plant regeneration, indicative of insufficient auxin for callus induction. Callus yield and regeneration frequencies were higher with 1.0 mg/l 2,4-D. With 2.0 or 4.0 mg/l 2,4-D, an induction period of 10 or 15 days was sufficient for initiation of callus development. The extended presence of 2–4 mg/l 2,4-D in the medium beyond the initiation phase was detrimental to plant regeneration. Thus optimal callus induction and plant regeneration could be obtained through manipulating the 2,4-D concentration and the duration of its presence in the induction medium. Received: 1 December 1997 / Revision received: 15 February 1999 / Accepted: 26 February 1999     

The uptake of cadmium,lead and selenium by barley and cabbage grown on soils amended with refuse incinerator fly ash

Summary Barley and cabbage plants grown in the greenhouse on soils amended with refuse incinerator fly ash contained significantly elevated levels of Cd, Pb and Se, with Cd uptake being greatest in both plant species. Cabbage grown on 20% ash amended soil contained 146 times more Cd than controls. Cadmium and Se appeared to be less available in a successive barley crop after overwintering the pots of soil outside, but elemental concentrations still remained elevated. Comparisons with data from other studies indicated that Cd availability was greater from refuse fly ash than from sewage sludge.     

Alterations of wheat root plasma membrane lipid composition induced by copper stress result in changed physicochemical properties of plasma membrane lipid vesicles

A response when wheat is grown in excess copper is an altered lipid composition of the root plasma membrane (PM). With detailed characterisation of the root PM lipid composition of the copper-treated plants as a basis, in the present study, model systems were used to gain a wider understanding about membrane behaviour, and the impact of a changed lipid composition.PMs from root cells of plants grown in excess copper (50 μM Cu²⁺) and control (0.3 μM Cu²⁺) were isolated using the two-phase partitioning method. Membrane vesicles were prepared of total lipids extracts from the isolated PMs, and also reference vesicles of phosphatidylcholine (PC). In a series of tests, the vesicle permeability for glucose and for protons was analysed. The vesicles show that copper stress reduced the permeability for glucose of the lipid bilayer barrier. When vesicles from stressed plants were modified by addition of lipids to resemble vesicles from control plants, the permeability for glucose was very similar to that of vesicles from control plants. The permeability for protons did not change upon stress.Electron paramagnetic resonance (EPR) of the lipid vesicles spin probed with n-doxylstearic acid (nDSA) was used to explore the lipid rotational freedom at different depth of the bilayer. The EPR measurements supported the permeability data, indicating that the copper stress resulted in more tightly packed bilayers of the PMs with reduced acyl chain motion.     

Events surrounding the early development of Euglena chloroplasts. 15. Origin of plastid thylakoid polypeptides in wild-type and mutant cells

Techniques are described for the isolation of plastid thylakoid membranes from light-grown and dark-grown cells of Euglena gracilis var. bacillaris, and from mutants affecting plastid development. These membranes, which have minimal contamination with other cell fractions, are localized in sucrose gradients by using the thylakoid membrane sulfolipid as a specific marker. The plastid thylakoid membrane polypeptides isolated from these membranes were separated on SDS polyacrylamide gels and yielded patterns containing 30–40 polypeptides. Light-grown strain Z gave patterns identical with bacillaris. Since the plastid thylakoid polypeptide patterns obtained from dark-grown wild-type cells and from a bleached mutant W₃BUL in which plastid DNA is undetectable are identical, it appears that the proplastid thylakoid polypeptides of wild-type cannot be coded in plastid DNA and are probably coded in nuclear DNA. The plastid thylakoid polypeptide patterns obtained from various dark-grown mutants are identical to those obtained from dark-grown wild-type cells. Light-grown mutants, making large but abnormal chloroplasts, show a correlation between the amount of chlorophyll formed and the amount of a plastid thylakoid polypeptide thought to be associated with one of the pigment-protein light-harvesting complexes. Treatment with SAN 9789 (4-chloro-5-(methyl-amino)-2-(α,α,α,-trifluoro-m-tolyl)-3-(2H(pyridazinone) known to block carotenoid synthesis at the level of phytoene, causes a progressive loss of all plastid thylakoid polypeptides during growth in darkness and results in the establishment of a new, lower steady-state level of sulfolipid. At least ten of the plastid thylakoid polypeptides become labeled when isolated chloroplasts are supplied with radioactive amino acids; of these six are undectable in W₃BUL and are, therefore, candidates for coding by plastid DNA.     

Fluorescence characterization of the interaction of Al3+ and Pd2+ with Suwannee River fulvic acid in the absence and presence of the herbicide 2,4-dichlorophenoxyacetic acid

In an effort to understand the role of environmental metal ions in the interaction of charged pesticides with humic substances, a fluorescence study of the interaction of the widely-used herbicide 2,4-dichlorophenoxyacetic acid (DCPAA) with Al(3+) and Pd(2+) and Suwannee River fulvic acid (SRFA) was undertaken. Initial fluorescence experiments on binary solutions clearly indicated that both Al(3+) and Pd(2+) strongly interact with both SRFA and DCPAA when alone in solution with the metal ion. Titrations of SRFA with Al(3+) at pH values of 4.0, 3.0 and 2.0 revealed decreased degrees of fluorescence emission enhancement (at lambda(emission, max)=424 nm) with decreasing pH, consistent with the expected loss of rigidity in the SRFA-Al(3+) complexes formed as pH is lowered. In contrast, titrations of SRFA with Pd(2+) at all of these pH values resulted in significant fluorescence quenching. Al(3+) additions to solutions of DCPAA at pH values above the pK(a) (2.64) of DCPAA resulted primarily in significant changes in the wavelength of maximum emission (without significant quenching or enhancement of emission intensity), while Pd(2+) additions to DCPAA solutions resulted primarily in very significant fluorescence quenching. The DCPAA fluorescence results strongly support the formation of an Al(3+)-DCPAA complex at pH values above the pK(a) of DCPAA. The fluorescence results obtained for solutions of Pd(2+) and DCPAA are best explained by a collisional quenching mechanism, that is, energy transfer from excited DCPAA molecules to Pd(2+) following the collision of these two species in solution. Excitation-emission matrix plots obtained on ternary solutions (at environmentally-relevant pH 4.0) containing SRFA, DCPAA and metal ions (i.e., either Al(3+) or Pd(2+)) provides evidence (especially for systems containing Al(3+)) for the existence of ternary complexes between fulvic acid species, the herbicide DCPAA and metal ion, suggesting (at least at pH 4.0, where the predominant DCPAA species is negatively-charged) that metal ions may function to "bridge" negatively-charged fulvic acids to negatively-charged pesticides.     

Glutathione reductase activity and isoforms in leaves and roots of wheat plants subjected to cadmium stress

The behavior of glutathione reductase (GR, EC 1.6.4.2) activity and isoforms were analyzed in wheat (Triticum aestivum L.) leaves and roots exposed to a chronic treatment with a toxic cadmium (Cd) concentration. A significant growth inhibition (up to 55%) was found in leaves at 7, 14 and 21 days, whereas roots were affected (51%) only after three weeks. Wheat plants grown in the presence of 100microM Cd showed a time-dependent accumulation of this metal, with Cd concentration being 10-fold higher in roots than in leaves. Nevertheless, lipid peroxidation was augmented in leaves in all experiments, but not in roots until 21 days. Cadmium treatment altered neither the GR activity nor the isoform pattern in the leaves. However, GR activity increased 111% and 200% in roots at 7 and 14 days, respectively, returning to control levels after 21 days. Three GR isoforms were found in roots of control and treated plants, two of which were enhanced by Cd treatment at 7 and 14 days, as assessed by activity staining on native gels. The changes in the isoform pattern modified the global kinetic properties of GR, thereby decreasing significantly (2.5-fold) the Michaelis constant (K(m)) value for oxidized glutathione. Isozyme induction was not associated with an enhancement of GR mRNA and protein expression, indicating that post-translational modification could occur. Our data demonstrated that up-regulation of GR activity by the induction of distinctive isoforms occurs as a defense mechanism against Cd-generated oxidative stress in roots.     

Rates of uptake and metabolism of indole-3-acetic acid and 2,4-dichlorophenoxyacetic acid by cultured leaf segments at different stages of development in wheat

Immature leaf tissue of Triticum timopheevi Zukh. responded to supplied auxin and showed cell division in culture. The rates of uptake and of metabolism of indole-3-acetic acid and 2,4-dichlorophenoxyacetic acid by such tissues were measured and compared with those of mature auxin-unresponsive tissue. The purpose of these experiments was to determine whether or not the concentration of auxin in cultured mature tissue was a factor limiting the cell division response to auxin. The data indicate that neither alterations in rates of uptake nor alterations in rates of metabolism could explain the loss of responsiveness to auxin which apparently occurs during cell differentiation. The results are discussed in the context of the view that changes in cell sensitivity to growth substances and not only the concentration of these compounds, play an important role in plant growth and development.     

Changes in the nucleotide pools in leaves and buds of tobacco plants upon treatment with 2,4-dichlorophenoxyacetic acid

Tobacco ( Nicotiana tabacum L. cv. Samsun) plants were treated once with 2,4-dichlorophenoxyacetic acid (2,4-D) at the 8-leaf stage. The effect of the herbicide on leaf metabolism was followed over 7 days by determination of the ribonucleotide pools, including NAD⁺, NADP⁺ and UDP-sugars, by high-preformance liquid chromatography. 2,4-D treatment resulted in large changes in the nucleotide concentrations, the magnitude and sign of which were dependent upon the leafage. The nucleotide pools decreased in the apical tissue, but increased strongly in the mature leaves with the highest relative increase in the oldest leaf tested. The time course of the changes revealed a maximum on day 5 after 2,4-D treatment. The increase in the adenine nucleotide pools, energy charge and the NADVNADP⁺ ratio are interpreted to indicate a stress situation. The different responses of young, mature and senescent tissue to the synthetic auxin could reflect their different inherent sensitivity due to the natural auxin gradient.     

Effects of selenium content in green parts of plants on the amount of ATP and ascorbate-glutathione cycle enzyme activity at various growth stages of wheat and oilseed rape

The aim of this experiment, conducted under greenhouse conditions, was to assess the influence of various H(2)SeO(3) concentrations added to soil (0.05, 0.15, and 0.45mMkg(-1)) on selenium and adenosine triphosphate (ATP) content, and on the activity of the ascorbate-glutathione cycle enzymes in green parts of wheat and oilseed rape. Selenium uptake by the test plants was found to vary, with content increasing from one developmental stage to the next over four stages of the developmental cycle. At the lowest H(2)SeO(3) dose (0.05mMkg(-1)), the wheat plants took up much more selenium than did the oilseed rape plants, while the amount of selenium taken up at higher doses (0.15 and 0.45mMkg(-1)) was markedly higher in rape. The increasing Se content in the wheat to about 10mgkg(-1) (in the dark) and to about 16mgkg(-1) (in the light) was accompanied by a concurrent increase in the ATP content, which remained unchanged in the light-exposed plants, while clearly decreasing in those kept in the dark. On the other hand, the ATP content of the light-exposed oilseed rape was maintained at a stable level to about 10mg Sekg(-1), following which ATP content was observed to decrease. In contrast, the tendency for the ATP content to decrease appeared immediately in the dark. The increasing plant selenium concentration was accompanied by decreased APX activity in wheat, increased activity in oilseed rape, no major change in the dehydroascorbate reductase (DHAR) activity in oilseed rape and a slight increase in wheat to about 8mg Sekg(-1), followed by a reduction. The glutathione reductase (GR) activity in wheat differed from the activity of DHAR; an increase in the selenium content to about 8mgkg(-1) was accompanied by a distinct reduction, while a significant increase was observed at higher selenium contents; in oilseed rape, the activity was observed to increase slightly within a narrow range of selenium contents (up to 5mgkg(-1)), and to decrease thereafter.     

Physical distribution of translocation breakpoints in homoeologous recombinants induced by the absence of the Ph1 gene in wheat and triticale

The physical distribution of translocation breakpoints was analyzed in homoeologous recombinants involving chromosomes 1A, 1B, 1D of wheat and 1R of rye, and the long arms of chromosome 7S of Aegilops speltoides and 7A of wheat. Recombination between homoeologues was induced by removal of the Ph1 gene. In all instances, translocation breakpoints were concentrated in the distal ends of the chromosome arms and were absent in the proximal halves of the arms. The relationship between the relative distance from the centromere and the relative homoeologous recombination frequency was best explained by the function f(x)=0.0091e^0.0592x. The pattern of recombination in homoeologous chromosomes was essentially the same as in homologues except that there were practically no double exchanges. Among 313 recombinant chromosomes, only one resulted from a double crossing-over. The distribution of translocation breakpoints in translocated arms indicated that positive chiasma interference operated in homoeologous recombination. This implies that the reduction of the length of alien chromosome segments present in translocations with wheat chromosomes may be more difficult than the production of the original recombinants.