Inhibition of tomato (Solanum lycopersicum L.) root growth by cyanamide is not always accompanied with enhancement of ROS production

Mode of action of allelochemicals in target plants is currently widely studied. Cyanamide is one of the newly discovered allelochemical, biosynthesized in hairy vetch. Recently, it has been recognized that cyanamide is plant growth inhibitor, which affects mitosis in root tip cells and causes,e.g., disorder in phytohormonal balance. We also demonstrated that CA may act as oxidative stress agent but it strictly depends on plant species, exposure time and doses. Roots of tomato seedling treated with water solution of 1.2 mM cyanamide did not exhibit elevated reactive oxygen species concentration during the whole culture period.     

Nitric oxide and HCN reduce deep dormancy of apple seeds

Nitric oxide (NO) and cyanide (HCN) are small gaseous molecules that have been intensively studied to explain their role in plant development, metabolism and reaction to stresses. Cyanide and NO are known to be produced endogenously during early phase of seed germination or are present in the environment. Both molecules regulate breakage of seed dormancy and accelerate seed germination. Regulatory role of cyanide in breaking of dormancy seems to be understood to some extend, while the NO mode of action is much less explained. However, some similarities could be suggested. The mechanisms involved in HCN-dependent dormancy breakage in apple embryos are summarized in relation to NO-donor mediated stimulation of germination.     

The effect of phosphate deficiency on membrane phospholipid composition of bean (Phaseolus vulgaris L.) roots

Plasma membranes were isolated from roots of bean (Phaseolus vulgaris L.) plants cultured on phosphate sufficient or phosphate deficient medium. The phospholipid composition of plasma membranes was analyzed and compared with that of the microsomal fraction. Phosphate deficiency had no influence on lipid/protein ratio in microsomal as well as plasma membrane fraction. In phosphate deficient roots phospholipid content was lower in the plasma membrane, but did not change in the microsomal fraction. Phosphatidylcholine and phosphatidylethanolamine were two major phospholipids in plasmalemma and microsomal membranes (80 % of the total). After two weeks of phosphate starvation a considerable decrease (about 50 %) in phosphatidylcholine and phosphatidylethanolamine in microsomal membranes was observed. The decline in two major phospholipids was accompanied by an increase in phosphatidic acid and lysophosphatidylcholine content. The effect of alterations in plasma membrane phospholipids on membrane function e.g. nitrate uptake is discussed.     

Polyamines and Nitric Oxide Link in Regulation of Dormancy Removal and Germination of Apple (Malus domestica Borkh.) Embryos

Polyamines (PAs) belong to plant growth regulators and in complex with classical phytohormones take part in regulation of seed dormancy and germination. Although the impact of reactive oxygen (ROS) and nitrogen (RNS) species on seed germination is well described, the cross talk of PAs with ROS/RNS has never been analyzed. Due to the close connection of PAs and ethylene biosynthetic pathways to arginine (Arg)-dependent NO biosynthesis we investigated production of nitric oxide (NO), peroxynitrite (ONOO^?) and the level of O ₂ ^?? or H₂O₂ in apple embryos, germination of which was PA regulated. PAs: putrescine (Put) and spermidine (Spd) in contrast to spermine (Spm) stimulated germination of apple embryos. Among amino acids, stimulation of germination was observed in Arg and ornithine (Orn) only. Dormancy removal of embryos by PAs was associated with increased accumulation of H₂O₂ and O ₂ ^?? in embryonic axes. At the same stage of completion of sensu stricto germination the stimulatory effect of PAs (Put and Spd) and amino acids, mainly Arg and Orn, was accompanied by enhanced NO and ONOO^? production in embryonic axis. The beneficial effect of PAs (Put and Spd) and their precursors on germination of apple embryos was removed by NO scavenging, suggesting a crucial role of NO in termination of embryo germination and radicle growth. Moreover, activity of polyamine oxidase in embryo axes was greatly enhanced by embryo fumigation with NO. Our data demonstrate the interplay of RNS/ROS with PAs and point to NO action as an integrator of endogenous signals activating germination.     

Inhibition of tomato (Solanum lycopersicum L.) root growth by cyanamide is due to altered cell division, phytohormone balance and expansin gene expression

Cyanamide (CA) has been reported as a natural compound produced by hairy vetch (Vicia villosa Roth.) and it was shown also to be an allelochemical, responsible for strong allelopathic potential in this species. CA phytotoxicity has been demonstrated on various plant species, but to date little is known about its mode of action at cellular level. Treatment of tomato (Solanum lycopersicum L.) roots with CA (1.2 mM) resulted in inhibition of growth accompanied by alterations in cell division, and imbalance of plant hormone (ethylene and auxin) homeostasis. Moreover, the phytotoxic effect of CA was also manifested by modifications in expansin gene expression, especially in expansins responsible for cell wall remodeling after the cytokinesis (LeEXPA9, LeEXPA18). Based on these results the phytotoxic activity of CA on growth of roots of tomato seedlings is likely due to alterations associated with cell division.     

Nitric oxide and hydrogen cyanide as regulating factors of enzymatic antioxidant system in germinating apple embryos

Short-term (3 or 6 h) pre-treatment of apple (Malus domestica Borkh.) embryos with nitric oxide (NO) or hydrogen cyanide (HCN) induces transient accumulation of reactive oxygen species (ROS) leading to dormancy removal and germination. We demonstrated that enhanced NO emission by apple embryos during early phase of germination “sensu stricto” is required for seed transition from dormant into non-dormant state, and may be described by the model of “nitrosative door”, analogous to “oxidative window”. Cellular ROS concentration, resulting from NO or HCN embryo pre-treatment, seems to be under severe control of antioxidant system. Activity of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPX) and total peroxidases (Prxs) was determined during NO and HCN-mediated germination “sensu stricto” of embryos. CAT and SOD activity increased transiently 24 h after embryos pre-treatment, while GR and Prx activity was stimulated mainly after 96 h. The most evident alterations were detected in GPX activity, being more than threefold stimulated by NO or HCN. Based on this results, we conclude that these reactive molecules act simultaneously crossing their signaling pathways and we propose that ROS, reactive nitrogen species, HCN at accurate level are essential during seed germination as signaling factors.     

Desensitization at the first stage of IGE-mediated response as hay fever prophylaxis

The aim of our work was to assess the prophylactic aspects of desensitization at the first stage of IgE-mediated response to grass pollen antigens (GPA). Forty six patients aged 10-45 (Me = 20.45) years (group I) and 50 patients aged 11-45 (Me = 19.17) years (group II) were included in to the study. All of them: 1) suffered from allergic rhinitis due to house dust (HD), feathers (F), weed (W) or tree (T) pollen, 2) had also some other allergic diseases, 3) had positive family history of allergic disorders, 4) had reproducible, strongly positive skin reaction of type I to GPA, but 5) did not show any clinical symptoms of hypersensitivity to GPA. Specific immunotherapy with HD, F, W, and T aqueous extracts was administered to both groups for at least 3 successive years, while parallel desensitization with GPA aqueous extract was carried out in group I, only. The symptoms of grass pollinosis were searcher for and the effects of immunotherapy were evaluated in all the patients over the period of at least 5 successive years after the complete course of vaccines administration. Clinical signs of hypersensitivity to GPA became evident in 6 patients (13.04%) of group I and in 27 patients (54.00%) of group II (p less than 0.001). Thus the desensitization at the first stage of IgE-mediated response to GPA effectively prevented development of the symptomatic hay fever. However, the disease revealed itself in few cases still much later and its course was much milder than in people who were not subjected to such a preventive desensitization.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dormancy removal in apple embryos by nitric oxide or cyanide involves modifications in ethylene biosynthetic pathway

The connection between classical phytohormone-ethylene and two signaling molecules, nitric oxide (NO) and hydrogen cyanide (HCN), was investigated in dormancy removal and germination “sensu stricto” of apple (Malus domestica Borkh.) embryos. Deep dormancy of apple embryos was removed by short-term (3–6 h) pre-treatment with NO or HCN. NO- or HCN-mediated stimulation of germination was associated with enhanced emission of ethylene by the embryos, coupled with transient increase in ROS concentration in embryos. Ethylene vapors stimulated germination of dormant apple embryos and eliminated morphological anomalies characteristic for young seedlings developed from dormant embryos. Inhibitors of ethylene receptors completely impeded beneficial effect of NO and HCN on embryo germination. NO- and HCN-induced ethylene emission by apple embryo was only slightly reduced by inhibitor of 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase activity during first 4 days of germination. Short-term pre-treatment of the embryos with NO and HCN modified activity of both key enzymes of ethylene biosynthetic pathway: ACC synthase and ACC oxidase. Activity of ACC synthase declined during first 4 days of germination, while activity of ACC oxidase increased markedly at that time. Additional experiments point to non-enzymatic conversion of ACC to ethylene in the presence of ROS (H₂O₂). The results indicate that NO and HCN may alleviate dormancy of apple embryos “via” transient accumulation of ROS, leading to enhanced ethylene emission which is required to terminate germination “sensu stricto”. Therefore, ethylene seems to be a trigger factor in control of apple embryo dormancy removal and germination.     

Impact of functionalized coligands on the pharmacokinetics of 99mTcIII '4+1' mixed-ligand complexes conjugated to bombesin

Bombesins (BN) containing ^99mTc ‘4 + 1’ complexes may be useful to detect tumors expressing the gastrin-releasing peptide receptor (GRPR). Derivatives of the formula [^99mTc(NS₃R)(L2-BN_st)] were synthesized, in which Tc(III) is coordinated by an isocyanide L2-BN_st bearing the peptide (BN_st = βAla-βAla-Gln-Trp-Ala-Val-Gly-His-Cha-Nle-NH₂) and a tetradentate chelator NS₃R. NS₃R consists of 2,2′,2″-nitrilotriethanethiol (NS₃) bearing a crown ether (NS₃crown), an aliphatic amine (NS₃en) and a tricarboxylic acid (NS₃(COOH)₃). Non-radioactive Re compounds were prepared and analysed by electrospray ionization mass spectrometry. The structural similarity to the ^99mTc conjugates was demonstrated by their identical HPLC elution profiles. The lipophilicity of [^99mTc(NS₃R)(L2-BN_st)] decreased depending on the coligands NS₃crown (log D_O/W, pH = 7.4, 0.98 ± 0.11), NS₃en (− 0.49 ± 0.07) and NS₃(COOH)₃ (− 2.01 ± 0.09). Biodistribution in normal rats was characterized by an increasing kidney uptake and a decreasing uptake into the liver corresponding to the reduced lipophilicity of the conjugates. The pancreatic uptake expressed by the organ/blood ratio of standardized uptake values at 60 min p.i. in rats was 8.6 ± 1.2 for [^99mTc(NS₃en)(L2-BN_st)] and higher compared to the other conjugates. The pancreas/liver ratio of the SUV at 60 min p.i. in rats was highest for [^99mTc(NS₃(COOH)₃)(L2-BN_st)] at 8.4 ± 1.3. [^99mTc(NS₃en)(L2-BN_st)] was further studied in tumor-bearing mice and its pancreas/blood and pancreas/liver ratios were lower, however the pancreas/kidney ratios were higher in mice compared to rats. The activity uptake of [^99mTc(NS₃en)(L2-BN_st)] into the PC-3 tumor xenografts was low (%ID/g: 0.83 ± 0.18 at 60 min; SUV: 0.21 ± 0.05 at 60 min) but specific.     

Phytotoxic cyanamide affects maize (Zea mays) root growth and root tip function: From structure to gene expression

Cyanamide (CA) is a phytotoxic compound produced by four Fabaceae species: hairy vetch, bird vetch, purple vetch and black locust. Its toxicity is due to complex activity that involves the modification of both cellular structures and physiological processes. To date, CA has been investigated mainly in dicot plants. The goal of this study was to investigate the effects of CA in the restriction of the root growth of maize (Zea mays), representing the monocot species. CA (3 mM) reduced the number of border cells in the root tips of maize seedlings and degraded their protoplasts. However, CA did not induce any significant changes in the organelle structure of other root cells, apart from increased vacuolization. CA toxicity was also demonstrated by its effect on cell cycle activity, endoreduplication intensity, and modifications of cyclins CycA2, CycD2, and histone HisH3 gene expression. In contrast, the arrangement of microtubules was not altered by CA. Treatment of maize seedlings with CA did not completely arrest mitotic activity, although the frequency of dividing cells was reduced. Furthermore, prolonged CA treatment increased the proportion of endopolyploid cells in the root tip. Cytological malformations were accompanied by an induction of oxidative stress in root cells, which manifested as enhanced accumulation of H₂O₂. Exposure of maize seedlings to CA resulted in an increased concentration of auxin and stimulated ethylene emission. Taken together, these findings suggested that the inhibition of root growth by CA may be a consequence of stress-induced morphogenic responses.