Nickel oxide nanoparticles induce cell wall modifications, root anatomical changes, and nitrosative signaling in ecotypes of Ni hyperaccumulator Odontarrhena lesbiaca.

The industrial application and environmental release of nickel oxide NPs (NiO NPs) is increasing, but the details of their relationship with plants are largely unknown. In this work, the cellular, tissue, organ, and molecular level responses of three ecotypes of Ni hyperaccumulator Odontarrhena lesbiaca grown in the presence of high doses of NiO NP (250 mg/L and 500 mg/L) were studied. All three ecotypes showed a similar accumulation of Ni in the presence of nano Ni, and in the case of NiO NPs, the root-to-shoot Ni translocation was slighter compared to the bulk Ni. In all three ecotypes, the walls of the root cells effectively prevented internalization of NiO NPs, providing cellular defense against Ni overload. Exposure to NiO NP led to an increase in cortex thickness and the deposition of lignin-suberin and pectin in roots, serving as a tissue-level defense mechanism against excessive Ni. Exposure to NiO NP did not modify or cause a reduction in some biomass parameters of the Ampeliko and Loutra ecotypes, while it increased all parameters in Olympos. The free salt form of Ni exerted more negative effects on biomass production than the nanoform, and the observed effects of NiO NPs can be attributed to the release of Ni ions. Nitric oxide and peroxynitrite levels were modified by NiO NPs in an ecotype-dependent manner. The changes in the abundance and activity of S-nitrosoglutathione reductase protein triggered by NiO NPs suggest that the enzyme is regulated by NiO NPs at the post-translational level. The NiO NPs slightly intensified protein tyrosine nitration, and the slight differences between the ecotypes were correlated with their biomass production in the presence of NiO NPs. Overall, the Odontarrhena lesbiaca ecotypes exhibited tolerance to NiO NPs at the cellular, tissue, organ/organism and molecular levels, demonstrating various defense mechanisms and changes in the metabolism of reactive nitrogen species metabolism and nitrosative protein modification.

Nitro-oxidative response to internalized multi-walled carbon nanotubes in Brassica napus and Solanum lycopersicum.

In addition to their beneficial effects on plant physiology, multi-walled carbon nanotubes (MWCNTs) are harmful to plants in elevated concentrations. This study compared the effects of two doses of MWCNT (10 and 80 mg/L) in Brassica napus and Solanum lycopersicum seedlings focusing on nitro-oxidative processes. The presence of MWCNTs was detectable in the root and hypocotyl of both species. Additionally, transmission electron microscopy analysis revealed that MWCNTs are heavily transformed within the root cells forming large aggregates. The uptake of MWCNTs negatively affected root viability and root cell proliferation of both species, but more intense toxicity was observed in S. lycopersicum compared to B. napus. The presence of MWCNT triggered more intense protein carbonylation in the relative sensitive S. lycopersicum, where increased hydrogen peroxide levels were observed. Moreover, MWCNT exposure increased the level of physiological protein tyrosine nitration which was more intense in S. lycopersicum where notable peroxynitrite accumulation occurred. These suggest for the first time that MWCNT triggers secondary nitro-oxidative stress which contributes to its toxicity. Moreover, the results indicate that the extent of the nitro-oxidative processes is associated with the extent of MWCNT toxicity.

L-Aminoguanidine Induces Imbalance of ROS/RNS Homeostasis and Polyamine Catabolism of Tomato Roots after Short-Term Salt Exposure.

Polyamine (PA) catabolism mediated by amine oxidases is an important process involved in fine-tuning PA homeostasis and related mechanisms during salt stress. The significance of these amine oxidases in short-term responses to salt stress is, however, not well understood. In the present study, the effects of L-aminoguanidine (AG) on tomato roots treated with short-term salt stress induced by NaCl were studied. AG is usually used as a copper amine oxidase (CuAO or DAO) inhibitor. In our study, other alterations of PA catabolism, such as reduced polyamine oxidase (PAO), were also observed in AG-treated plants. Salt stress led to an increase in the reactive oxygen and nitrogen species in tomato root apices, evidenced by in situ fluorescent staining and an increase in free PA levels. Such alterations were alleviated by AG treatment, showing the possible antioxidant effect of AG in tomato roots exposed to salt stress. PA catabolic enzyme activities decreased, while the imbalance of hydrogen peroxide (H2O2), nitric oxide (NO), and hydrogen sulfide (H2S) concentrations displayed a dependence on stress intensity. These changes suggest that AG-mediated inhibition could dramatically rearrange PA catabolism and related reactive species backgrounds, especially the NO-related mechanisms. More studies are, however, needed to decipher the precise mode of action of AG in plants exposed to stress treatments.

Selenate triggers diverse oxidative responses in Astragalus species with diverse selenium tolerance and hyperaccumulation capacity.

Selenium (Se) hyperaccumulators are capable of uptake and tolerate high Se dosages. Excess Se-induced oxidative responses were compared in Astragalus bisulcatus and Astragalus cicer. Plants were grown on media supplemented with 0, 25 or 75 µM selenate for 14 days. Both A. bisulcatus and A. cicer accumulated >2000 µg/g dry weight Se to the shoot but the translocation factors of A. cicer were below 1 suggesting its non hyperaccumulator nature. A. cicer showed Se sensitivity indicated by reduced seedling fresh weight, root growth and root apical meristem viability, altered element homeostasis in the presence of Se. In Se-exposed A. bisulcatus, less toxic organic Se forms (mainly MetSeCys, γ-Glu-MetSeCys, and a selenosugar) dominated, while these were absent from A. cicer suggesting that the majority of the accumulated Se may be present as inorganic forms. The glutathione-dependent processes were more affected, while ascorbate levels were not notably influenced by Se in either species. Exogenous Se triggered more intense accumulation of malondialdehyde in the sensitive A. cicer compared with the tolerant A. bisulcatus. The extent of protein carbonylation in the roots of the 75 µM Se-exposed A. cicer exceeded that of A. bisulcatus indicating a correlation between selenate sensitivity and the degree of protein carbonylation. Overall, our results reveal connection between oxidative processes and Se sensitivity/tolerance/hyperaccumulation and contribute to the understanding of the molecular responses to excess Se.

Nanoforms of essential metals: from hormetic phytoeffects to agricultural potential.

Vital plant functions require at least six metals (copper, iron, molybdenum, manganese, zinc, and nickel), which function as enzyme cofactors or inducers. In recent decades, rapidly evolving nanotechnology has created nanoforms of essential metals and their compounds (e.g. nZnO, nFe2O3) with a number of favourable properties over the bulk materials. The effects of nanometals on plants are concentration-dependent (hormesis) but also depend on the properties of the nanometals, the plant species, and the treatment conditions. Here, we review studies examining plant responses to essential nanometal treatments using a (multi)omics approach and emphasize the importance of gaining a holistic view of the diverse effects. Furthermore, we discuss the beneficial effects of essential nanometals on plants, which provide the basis for their application in crop production as, for example, nanopriming or nanostimulator agents, or nanofertilizers. As lower environmental impact and increased yield can be achieved by the application of essential nanometals, they support sustainable agriculture. Recent studies have actively examined the utilization of green-synthesized metal nanoparticles, which perfectly fit into the environmentally friendly trend of future agriculture. Further knowledge is required before essential nanometals can be safely applied in agriculture, but it is a promising direction that is timely to investigate.

Nitric oxide signalling in plant nanobiology: current status and perspectives.

Plant nanobiology as a novel research field provides a scientific basis for the agricultural use of nanoparticles (NPs). Plants respond to the presence of nanomaterials by synthesizing signal molecules, such as the multifunctional gaseous nitric oxide (NO). Several reports have described the effects of different nanomaterials (primarily chitosan NPs, metal oxide NPs, and carbon nanotubes) on endogenous NO synthesis and signalling in different plant species. Other works have demonstrated the ameliorating effect of exogenous NO donor (primarily sodium nitroprusside) treatments on NP-induced stress. NO-releasing NPs are preferred alternatives to chemical NO donors, and evaluating their effects on plants has recently begun. Previous studies clearly indicate that endogenous NO production in the presence of nanomaterials or NO levels increased by exogenous treatments (NO-releasing NPs or chemical NO donors) exerts growth-promoting and stress-ameliorating effects in plants. Furthermore, an NP-based nanosensor for NO detection in plants has been developed, providing a new and excellent perspective for basic research and also for the evaluation of plants' health status in agriculture.

Dual Effect of Nanomaterials on Germination and Seedling Growth: Stimulation vs. Phytotoxicity.

Due to recent active research, a large amount of data has been accumulated regarding the effects of different nanomaterials (mainly metal oxide nanoparticles, carbon nanotubes, chitosan nanoparticles) on different plant species. Most studies have focused on seed germination and early seedling development, presumably due to the simplicity of these experimental systems. Depending mostly on size and concentration, nanomaterials can exert both positive and negative effects on germination and seedling development during normal and stress conditions, thus some research has evaluated the phytotoxic effects of nanomaterials and the physiological and molecular processes behind them, while other works have highlighted the favorable seed priming effects. This review aims to systematize and discuss research data regarding the effect of nanomaterials on germination and seedling growth in order to provide state-of-the-art knowledge about this fast developing research area.

ZnO nanoparticles induce cell wall remodeling and modify ROS/ RNS signalling in roots of Brassica seedlings.

Cell wall-associated defence against zinc oxide nanoparticles (ZnO NPs) as well as nitro-oxidative signalling and its consequences in plants are poorly examined. Therefore, this study compares the effect of chemically synthetized ZnO NPs (~45 nm, 25 or 100 mg/L) on Brassica napus and Brassica juncea seedlings. The effects on root biomass and viability suggest that B. napus is more tolerant to ZnO NP exposure relative to B. juncea. This may be due to the lack of Zn ion accumulation in the roots, which is related to the increase in the amount of lignin, suberin, pectin and in peroxidase activity in the roots of B. napus. TEM results indicate that root cell walls of 25 mg/L ZnO NP-treated B. napus may bind Zn ions. Additionally, callose accumulation possibly contribute to root shortening in both Brassica species as the effect of 100 mg/L ZnO NPs. Further results suggest that in the roots of the relatively sensitive B. juncea the levels of superoxide radical, hydrogen peroxide, hydrogen sulfide, nitric oxide, peroxinitrite and S-nitrosoglutathione increased as the effect of high ZnO NP concentration meaning that ZnO NP intensifies nitro-oxidative signalling. In B. napus; however, reactive oxygen species signalling was intensified, but reactive nitrogen species signalling wasn't activated by ZnO NPs. Collectively, these results indicate that ZnO NPs induce cell wall remodeling which may be associated with ZnO NP tolerance. Furthermore, plant tolerance against ZnO NPs is associated rather with nitrosative signalling than oxidative modifications.

Nitro-oxidative signalling induced by chemically synthetized zinc oxide nanoparticles (ZnO NPs) in Brassica species.

Due to their release into the environment, zinc oxide nanoparticles (ZnO NPs) may come in contact with plants. In elevated concentrations, ZnO NPs induce reactive oxygen species (ROS) production, but the metabolism of reactive nitrogen species (RNS) and the consequent nitro-oxidative signalling has not been examined so far. In this work, Brassica napus and Brassica juncea seedlings were treated with chemically synthetized ZnO NPs (∼8 nm, 0, 25 or 100 mg/L). At low dose (25 mg/L) ZnO NP exerted a positive effect, while at elevated concentration (100 mg/L) it was toxic to both species. Additionally, B. juncea was more tolerant to ZnO NPs than B. napus. The ZnO NPs could enter the root cells due to their small (∼8 nm) size which resulted in the release of Zn2+ and subsequently increased Zn2+ content in the plant organs. ZnO NPs disturbed superoxide radical and hydrogen peroxide homeostasis and modulated ROS metabolic enzymes (NADPH oxidase, superoxide dismutase, ascorbate peroxidase) and non-enzymatic antioxidants (ascorbate and glutathione) inducing similar changes in oxidative signalling in both Brassica species. The homeostasis of RNS (nitric oxide, peroxynitrite and S-nitrosoglutathione) was also altered by ZnO NPs; however, changes in nitrosative signalling proved to be different in the examined species. Moreover, ZnO NPs triggered changes in protein carbonylation and nitration. These results suggest that ZnO NPs induce changes in nitro-oxidative signalling which may contribute to ZnO NP toxicity. Furthermore, difference in ZnO NP tolerance of Brassica species is more likely related to nitrosative than to oxidative signalling.

Distinct redox signalling and nickel tolerance in Brassica juncea and Arabidopsis thaliana.

Despite of its essentiality, nickel (Ni) in excess is toxic for plants partly due to the overproduction of reactive oxygen species (ROS) and the consequent increase in oxidative stress signalling. However, in Ni-stressed plants little is known about the signal transduction of reactive nitrogen species (RNS) and protein tyrosine nitration as the protein-level consequence of increased RNS formation. Our experiments compared the nickel accumulation and tolerance, the redox signalling and the protein nitration in the agar-grown Arabidopsis thaliana and Brassica juncea exposed to Ni (50 µM nickel chloride). Studying GUS-tagged Arabidopsis lines (ARR5::GUS, ACS8::GUS and DR5::GUS) revealed that Ni-increased lateral root (LR) emergence, and concomitantly reduced LR initiation were accompanied by elevated levels of auxin, cytokinin, and ethylene in the LRs or in upper root parts, whereas Ni-induced primary root shortening is related to decreased auxin, and increased cytokinin and ethylene levels. These suggest the Ni-induced disturbance of hormonal balance in the root system. Results of the comparative study showed that weaker Ni tolerance of A. thaliana was coupled with a Ni-induced increase in RNS, ROS, and hydrogen sulfide levels, as well as with an increase in redox signalling and consequent increment of protein nitration. However, in relative Ni tolerant B. juncea, redox signalling (except for peroxynitrite) was not modified, and Ni-induced intensification of protein tyrosine nitration was less pronounced. Data collectively show that the better Ni tolerance of Brassica juncea may be related to the capability of preventing the induction of redox signalling and consequently to the slighter increase in protein nitration.

Zinc-induced root architectural changes of rhizotron-grown B. napus correlate with a differential nitro-oxidative response.

Roots have a noteworthy plasticity: due to different stress conditions their architecture can change to favour seedling vigour and yield stability. The development of the root system is regulated by a complex and diverse signalling network, which besides hormonal factors, includes reactive oxygen (ROS) - and nitrogen species (RNS). The delicate balance of the endogenous signal system can be affected by various environmental stimuli, such as the excess of essential heavy metals, like zinc (Zn). Zn at low concentration, is able to induce the morphological and physiological adaptation of the root system, but in excess it exerts toxic effects on plants. In this study the effect of a low, growth-inducing, and a high, growth inhibiting Zn concentrations on the early development of Brassica napus (L.) root architecture and the underlying nitro-oxidative mechanisms were studied in a soil-filled rhizotron system. The growth-inhibiting Zn treatment resulted in elevated protein tyrosine nitration due to the imbalance in ROS and RNS homeostasis, however its pattern was not changed compared to the control. This nitro-oxidative stress was accompanied by serious changes in the cell wall composition and decrease in the cell proliferation and viability, due to the high Zn uptake and disturbed microelement homeostasis in the root tips. During the positive root growth response, a tyrosine nitration-pattern reorganisation was observed; there were no substantial changes in ROS and RNS balance and the viability and proliferation of the root tips' meristematic zone decreased to a lesser extent, as a result of a lower Zn uptake. The obtained results suggest that Zn in different amounts triggers different root growth responses accompanied by distinct changes in the pattern and strength of tyrosine nitration, proposing that nitrosative processes have an important role in the stress-induced root growth responses.

The intensity of tyrosine nitration is associated with selenite and selenate toxicity in Brassica juncea L.

Selenium phytotoxicity involves processes like reactive nitrogen species overproduction and nitrosative protein modifications. This study evaluates the toxicity of two selenium forms (selenite and selenate at 0µM, 20µM, 50µM and 100µM concentrations) and its correlation with protein tyrosine nitration in the organs of hydroponically grown Indian mustard (Brassica juncea L.). Selenate treatment resulted in large selenium accumulation in both Brassica organs, while selenite showed slight root-to-shoot translocation resulting in a much lower selenium accumulation in the shoot. Shoot and root growth inhibition and cell viability loss revealed that Brassica tolerates selenate better than selenite. Results also show that relative high amounts of selenium are able to accumulate in Brassica leaves without obvious visible symptoms such as chlorosis or necrosis. The more severe phytotoxicity of selenite was accompanied by more intense protein tyrosine nitration as well as alterations in nitration pattern suggesting a correlation between the degree of Se forms-induced toxicities and nitroproteome size, composition in Brassica organs. These results imply the possibility of considering protein tyrosine nitration as novel biomarker of selenium phytotoxicity, which could help the evaluation of asymptomatic selenium stress of plants.

Selenite-induced nitro-oxidative stress processes in Arabidopsis thaliana and Brassica juncea.

Extremes of selenium (Se) exert toxic effects on plants' physiological processes; although plant species tolerate Se differently. This study focuses on the effect of Se (0, 20, 50 or 100µM sodium selenite) on secondary nitro-oxidative stress processes mainly using in situ microscopic methods in non-accumulator Arabidopsis thaliana and secondary Se accumulator Brassica juncea. Relative Se tolerance or sensitivity of the species was evaluated based on growth parameters (fresh and dry weight, root growth) and cell viability. Besides, selenite-triggered cell wall modifications (pectin, callose) and stomatal regulations were determined for the first time. In case of Arabidopsis, relative selenite sensitivity was accompanied by decreased stomatal density and induced stomatal opening, callose accumulation, pronounced oxidative stress and moderate nitrosative modifications. In contrast, the selenite-treated, relatively tolerant Brassica juncea showed larger number of more opened stomata, pectin accumulation, moderate oxidative and intense nitrosative stress. These suggest that selenite tolerance or sensitivity is rather associated with oxidative processes than secondary nitrosative modifications in higher plants.

The quality of the antioxidant defence system in term and preterm twin neonates.

OBJECTIVE: Multiple pregnancy is associated with an enhanced metabolism and demand for O2, which may lead to the overproduction of reactive oxygen species and the development of oxidative stress. The degree of oxidative damage depends on the level of the antioxidant protection system of the foetus. The objective of the study was to identify the relationship between the state of the maturity and the antioxidant status of twin neonates. Investigations of the umbilical cord blood were carried out to detect differences in the antioxidant defence system between mature and premature twin neonates. METHODS: The activities of the superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) enzymes, the levels of reduced glutathione (GSH), protein carbonyls and oxidized lipids and the total antioxidant capacity of the plasma were determined. RESULTS: The level of lipid peroxidation was significantly higher in the premature neonates. An increase in the total antioxidant capacity was accompanied by a decrease in the damaged protein concentration. Significantly elevated activities of GPx alone were observed in the premature twins, though the GSH content too tended to be increased. The activity of SOD was decreased in the premature neonates. DISCUSSION: The antioxidant status of twin neonates are mainly influenced by maturity. We suggest that the level of lipid peroxidation might be of clinical value as a marker of pre- and perinatal distress in twins.

Cadmium-induced oxidative stress and antioxidative mechanisms in germinating Indian mustard (Brassica juncea L.) seeds.

Cadmium, known as a non-essential heavy metal, can cause oxidative stress in plants. In this study we tried to find out whether oxidative changes could be measured in the early stages of ontogenesis in Indian mustard (Brassica juncea L.) seeds exposed to Cd stress. Cadmium-caused oxidative stress and antioxidative responses were investigated with respect to both time- and concentration-dependence. Parameters that were measured were follows: total antioxidant capacity (ferric reducing ability of plasma (FRAP)), glutathione (GSH) content, level of lipid peroxidation (LP), total protein content, and glutathione-S-transferase (GST, EC 2.5.1.18) activity. Seeds were germinated in vitro at 0, 50, 100 and 200mg/LCd concentrations in dark for 12, 24, 48 and 96h. Oxidative stress occurred in the seeds due to Cd treatment, the level of LP was high at the beginning of the germination at all concentrations used, but it attenuated later on. FRAP showed concentration-dependent increase during 24h, but it decreased later on. GSH content was also elevated by increasing concentrations of Cd, which referred to the activity of non-enzymatic antioxidant system. The GST activity induced with germination only after 24h at the highest Cd concentration. The results show that FRAP is a suitable parameter with which to assess the antioxidant capacity of heavy metal-stressed germinating seeds.