Glyoxalase Research Articles

Modulation of growth performance and coordinated induction of ascorbate-glutathione and methylglyoxal detoxification systems by salicylic acid mitigates salt toxicity in choysum (Brassica parachinensis L.)

Salinity represents a serious environmental threat to crop production and by extension, to world food supply, social and economic prosperity of the developing world. Salicylic acid (SA) is an endogenous plant signal molecule involved in regulating various plant responses to stress. In the present study, we characterized the regulatory role of exogenous SA for their ability to ameliorate deleterious effects of salt stress (0, 100, 150, 200 mM NaCl) in choysum plants through coordinated induction of antioxidants, ascorbate glutathione (AsA-GSH) cycle, and the glyoxalase enzymes. An increase in salt stress dramatically declined root and shoot growth, leaf chlorophyll and relative water content (RWC), subsequently increased electrolyte leakage (EL) and osmolytes accumulation in choysum plants. Salt stress disrupted the antioxidant and glyoxalase defense systems which persuaded oxidative damages and carbonyl toxicity, indicated by increased H2O2 generation, lipid peroxidation, and methylglyoxal (MG) content. However, application of SA had an additive effect on the growth of salt-affected choysum plants, which enhanced root length, plant biomass, chlorophyll contents, leaf area, and RWC. Moreover, SA application effectively eliminated the oxidative and carbonyl stress by improving AsA and GSH pool, upregulating the activities of antioxidant enzymes and the enzymes associated with AsA-GSH cycle and glyoxalase system. Overall, SA application completely counteracted the salinity-induced deleterious effects of 100 and 150 mM NaCl and partially mediated that of 200 mM NaCl stress. Therefore, we concluded that SA application induced tolerance to salinity stress in choysum plants due to the synchronized increase in activities of enzymatic and non-enzymatic antioxidants, enhanced efficiency of AsA-GSH cycle and the MG detoxification systems.

Spermine ameliorates prolonged fluoride toxicity in soil-grown rice seedlings by activating the antioxidant machinery and glyoxalase system.

The current manuscript presents the first report on the ameliorative roles of exogenous spermine (Spm) during prolonged fluoride-induced toxicity and oxidative damages in the susceptible rice cultivar, IR-64. The application of Spm increased the overall growth in the stressed seedlings by significantly restricting fluoride bioaccumulation within the shoots and roots. The Spm-treated stressed seedlings exhibited low chlorosis and induced activity of pyruvate dehydrogenase and nitrate reductase due to reduced accumulation and localization of reactive oxygen species (ROS) in the shoot and root. Spm-supplementation during stress reduced the levels of molecular damages by lowering malondialdehyde, electrolyte leakage and protein carbonylation, and lipoxygenase and protease activity due to effective detoxification of ROS by the antioxidants like proline, glycine-betaine, anthocyanin, flavonoids, phenolics and higher polyamines like Spm and spermidine. Excessive accumulation of the toxic methylglyoxal was reversed due to the activation of the glyoxalase system (comprising of glyoxalase I and II) and the ascorbate-glutathione cycle. Exogenous Spm also triggered the activity of superoxide dismutase, guaiacol peroxidase, glutathione peroxidase and phenylalanine ammonia lyase, which efficiently scavenged ROS in the stressed seedlings. Overall, Spm treatment mitigated the fluoride-induced injuries in IR-64 by reducing fluoride bioaccumulation and elaborately refining the various defence machineries.

Selenium mitigates the chromium toxicity in Brassicca napus L. by ameliorating nutrients uptake, amino acids metabolism and antioxidant defense system

The phytotoxicity of chromium (Cr) makes it obligatory for the researchers to develop strategies that seek to hinder its accumulation in food chains. While, protective role of selenium (Se) has not been discussed in detail under adverse conditions in oilseed rape. Here, our aim was to investigate the potential use of Se (0, 5 and 10 μM) in alleviating the Cr toxicity (0, 100 and 200 μM) in Brassica napus L. Results delineated that Se-supplementation notably recovered the Cr-phytotoxicity by reducing the Cr accumulation in plant tissues and boosted the inhibition in plant growth and biomass. Under Cr stress, the exogenously applied Se significantly recovered the impairment in photosynthesis related parameters (chlorophyll a, chlorophyll b, carotenoids, net photosynthetic rate, stomatal conductance, and photochemical efficiency of photosystem II), and counteracted the reduction in nutrients uptake and improved the essential amino acids (EAAs) levels. In addition, Se activated the antioxidants enzymes included in AsA-GSH cycle (SOD, CAT, APX, GR, DHAR, MDHAR, GSH, and AsA) and glyoxalase (Gly) system (Gly I and Gly II) and minimized the excessive generation of reactive oxygen species (ROS) and methylglyoxal (MG) contents in response to Cr stress. In a nutshell, Se (more effective at 5 μM) alleviated the Cr and MG induced phytotoxicity and oxidative damages by minimizing their (Cr and MG) accumulation and enhanced the plant growth, nutrients element level, nutrition quality by improving EAAs, antioxidant and Gly system. By considering the above-mentioned biomarkers, the addition of exogenous Se in Cr polluted soils might be effective approach to decrease the Cr uptake and its linked phytotoxicity in B. napus.

24-Epibrassinolide (EBR) Confers Tolerance against NaCl Stress in Soybean Plants by Up-Regulating Antioxidant System, Ascorbate-Glutathione Cycle, and Glyoxalase System.

The present research was performed to assess the effect of 24-epibrassinolide (EBR) on salt-stressed soybean plants. Salt stress suppressed growth, biomass yield, gas exchange parameters, pigment content, and chlorophyll fluorescence, but all these parameters were up-regulated by EBR supply. Moreover, salt stress increased hydrogen peroxide, malondialdehyde, and electrolyte leakage. EBR supplementation reduced the accumulation of oxidative stress biomarkers. The activities of superoxide dismutase and catalase, and the accumulation of proline, glycinebetaine, total phenols, and total flavonoids increased with NaCl stress, but these attributes further increased with EBR supplementation. The activities of enzymes and the levels of non-enzymatic antioxidants involved in the Asc-Glu cycle also increased with NaCl stress, and further enhancement in these attributes was recorded by EBR supplementation. Salinity elevated the methylglyoxal content, but it was decreased by the EBR supplementation accompanying with up-regulation of the glyoxalase cycle (GlyI and GlyII). Salinity enhanced the Na+ uptake in root and shoot coupled with a decrease in uptake of Ca2+, K+, and P. However, EBR supplementation declined Na+ accumulation and promoted the uptake of the aforementioned nutrients. Overall, EBR supplementation regulated the salt tolerance mechanism in soybean plants by modulating osmolytes, activities of key enzymes, and the levels of non-enzymatic antioxidants.

GLYI4 Plays A Role in Methylglyoxal Detoxification and Jasmonate-Mediated Stress Responses in Arabidopsis thaliana

Plant hormones play a central role in various physiological functions and in mediating defense responses against (a)biotic stresses. In response to primary metabolism alteration, plants can produce also small molecules such as methylglyoxal (MG), a cytotoxic aldehyde. MG is mostly detoxified by the combined actions of the enzymes glyoxalase I (GLYI) and glyoxalase II (GLYII) that make up the glyoxalase system. Recently, by a genome-wide association study performed in Arabidopsis, we identified GLYI4 as a novel player in the crosstalk between jasmonate (JA) and salicylic acid (SA) hormone pathways. Here, we investigated the impact of GLYI4 knock-down on MG scavenging and on JA pathway. In glyI4 mutant plants, we observed a general stress phenotype, characterized by compromised MG scavenging, accumulation of reactive oxygen species (ROS), stomatal closure, and reduced fitness. Accumulation of MG in glyI4 plants led to lower efficiency of the JA pathway, as highlighted by the increased susceptibility of the plants to the pathogenic fungus Plectospherella cucumerina. Moreover, MG accumulation brought about a localization of GLYI4 to the plasma membrane, while MeJA stimulus induced a translocation of the protein into the cytoplasmic compartment. Collectively, the results are consistent with the hypothesis that GLYI4 is a hub in the MG and JA pathways.

Methylglyoxal, a Highly Reactive Dicarbonyl Compound, in Diabetes, Its Vascular Complications, and Other Age-Related Diseases.

The formation and accumulation of methylglyoxal (MGO), a highly reactive dicarbonyl compound, has been implicated in the pathogenesis of type 2 diabetes, vascular complications of diabetes, and several other age-related chronic inflammatory diseases such as cardiovascular disease, cancer, and disorders of the central nervous system. MGO is mainly formed as a byproduct of glycolysis and, under physiological circumstances, detoxified by the glyoxalase system. MGO is the major precursor of nonenzymatic glycation of proteins and DNA, subsequently leading to the formation of advanced glycation end products (AGEs). MGO and MGO-derived AGEs can impact on organs and tissues affecting their functions and structure. In this review we summarize the formation of MGO, the detoxification of MGO by the glyoxalase system, and the biochemical pathways through which MGO is linked to the development of diabetes, vascular complications of diabetes, and other age-related diseases. Although interventions to treat MGO-associated complications are not yet available in the clinical setting, several strategies to lower MGO have been developed over the years. We will summarize several new directions to target MGO stress including glyoxalase inducers and MGO scavengers. Targeting MGO burden may provide new therapeutic applications to mitigate diseases in which MGO plays a crucial role.

Brain Senescence Caused by Elevated Levels of Reactive Metabolite Methylglyoxal on D-Galactose-Induced Aging Mice.

Aging is a complex natural phenomenon that is manifested by degenerative changes in the structure and function of cells and tissues. D-Galactose-induced aging mice are an artificial accelerated aging model that causes memory and learning impairment, oxidative stress, and neuroinflammation. In this study, we examined the underlying mechanism of an aging mouse model induced by D-galactose. Our behavioral Morris water maze results revealed that D-galactose administration for 2 months significantly induced memory and learning impairment in C57BL/6J mice. High performance liquid chromatography (HPLC) results showed elevated levels of the metabolite methylglyoxal (MG) in D-galactose-induced aging mice. Whether and how D-galactose induces senescence by elevated levels of reactive metabolite MG remain unclear. In our study, MG mainly accumulated through the following two aspects: to increase its source, namely, the triose phosphate produced by the glycolysis pathway, and to reduce its detoxification system, namely, the glyoxalase system. Therefore, elevated MG levels may be one of the causes of brain senescence in D-galactose-induced mice. However, the molecular mechanism of the increased level of the reaction metabolite methylglyoxal requires further exploration.

Comparative Physiological and Biochemical Changes in Tomato (Solanum lycopersicum L.) Under Salt Stress and Recovery: Role of Antioxidant Defense and Glyoxalase Systems.

Salinity toxicity and the post-stress restorative process were examined to identify the salt tolerance mechanism in tomato, with a focus on the antioxidant defense and glyoxalase systems. Hydroponically grown 15 day-old tomato plants (Solanum lycopersicum L. cv. Pusa Ruby) were treated with 150 and 250 mM NaCl for 4 days and subsequently grown in nutrient solution for a further 2 days to observe the post-stress responses. Under saline conditions, plants showed osmotic stress responses that included low leaf relative water content and high proline content. Salinity induced oxidative stress by the over-accumulation of reactive oxygen species (H2O2 and O2•−) and methylglyoxal. Salinity also impaired the non-enzymatic and enzymatic components of the antioxidant defense system. On the other hand, excessive Na+ uptake induced ionic stress which resulted in a lower content of other minerals (K+, Ca2+, and Mg2+), and a reduction in photosynthetic pigment synthesis and plant growth. After 2 days in the normal nutrient solution, the plants showed improvements in antioxidant and glyoxalase system activities, followed by improvements in plant growth, water balance, and chlorophyll synthesis. The antioxidant and glyoxalase systems worked in concert to scavenge toxic reactive oxygen species (ROS), thereby reducing lipid peroxidation and membrane damage. Taken together, these findings indicate that tomato plants can tolerate salinity and show rapid post-stress recovery by enhancement of their antioxidant defense and glyoxalase systems.

Glyoxalase 1 expression is downregulated in preimplantation blastocysts of diabetic rabbits.

In a diabetic pregnancy, an altered maternal metabolism led to increased formation of reactive α-dicarbonyls such as glyoxal (GO) and methylglyoxal (MGO) in the reproductive organs and embryos. The enzyme glyoxalase (GLO) 1 detoxifies reactive α-dicarbonyls thus protecting cells against malfunction or modifications of proteins by advanced glycated end products (AGEs). The aim of this study was to analyse the influence of a maternal insulin-dependent diabetes mellitus (IDD) on GLO1 expression and activity in preimplantation embryos in vivo and human trophoblast cells (Ac-1M88) in vitro. Maternal diabetes was induced in female rabbits by alloxan before conception and maintained during the preimplantation period. GLO1 expression and activity were investigated in 6-day-old blastocysts from healthy and diabetic rabbits. Furthermore, blastocysts and human trophoblast cells were exposed in vitro to hyperglycaemia, GO and MGO and analysed for GLO1 expression and activity. During gastrulation, GLO1 was expressed in all compartments of the rabbit blastocyst. Maternal diabetes decreased embryonic GLO1 protein amount by approx. 30 per cent whereas the enzymatic activity remained unchanged, indicating that the specific GLO1 activity increases along with metabolic changes. In in vitro cultured embryos, neither hyperglycaemia nor MGO and GO had an effect on GLO1 protein amount. In human trophoblast cells, a stimulating effect on the GLO1 expression was shown in the highest GO concentration, only. Our data show that maternal diabetes mellitus affects the specific activity of GLO1, indicating that GLO1 was post-translationally modified due to changes in metabolic processes in the preimplantation embryos.

Discovery of a nanomolar inhibitor of the human glyoxalase-I enzyme using structure-based poly-pharmacophore modelling and molecular docking.

The glyoxalase-I (GLO-I) enzyme, which is the initial enzyme of the glyoxalase system that is responsible for the detoxification of cytotoxic α-ketoaldehydes, such as methylglyoxal, has been approved as a valid target in cancer therapy. Overexpression of GLO-I has been observed in several types of carcinomas, including breast, colorectal, prostate, and bladder cancer. In this work we aimed to identify potential GLO-I inhibitors via employing different structure-based drug design techniques including structure-based poly-pharmacophore modelling, virtual screening, and molecular docking. Poly-pharmacophore modelling was applied in this study in order to thoroughly explore the binding site of the target enzyme, thereby, revealing hits that could bind in a nonconventional way which can pave the way for designing more potent and selective ligands with novel chemotypes. The modelling phase has resulted in the selection of 31 compounds that were biologically evaluated against human GLO-I enzyme. Among the tested set, seven compounds showed excellent inhibitory activities with IC50 values ranging from 0.34 to 30.57μM. The most active compound (ST018515) showed an IC50 of 0.34 ± 0.03μM, which, compared to reported GLO-I inhibitors, can be considered a potent inhibitor, making it a good candidate for further optimization towards designing more potent GLO-I inhibitors.

Preliminary Characterization of a Ni2+-Activated and Mycothiol-Dependent Glyoxalase I Enzyme from Streptomyces coelicolor

The glyoxalase system consists of two enzymes, glyoxalase I (Glo1) and glyoxalase II (Glo2), and converts a hemithioacetal substrate formed between a cytotoxic alpha-ketoaldehyde, such as methylglyoxal (MG), and an intracellular thiol, such as glutathione, to a non-toxic alpha-hydroxy acid, such as d-lactate, and the regenerated thiol. Two classes of Glo1 have been identified. The first is a Zn2+-activated class and is exemplified by the Homo sapiens Glo1. The second class is a Ni2+-activated enzyme and is exemplified by the Escherichia coli Glo1. Glutathione is the intracellular thiol employed by Glo1 from both these sources. However, many organisms employ other intracellular thiols. These include trypanothione, bacillithiol, and mycothiol. The trypanothione-dependent Glo1 from Leishmania major has been shown to be Ni2+-activated. Genetic studies on Bacillus subtilis and Corynebacterium glutamicum focused on MG resistance have indicated the likely existence of Glo1 enzymes employing bacillithiol or mycothiol respectively, although no protein characterizations have been reported. The current investigation provides a preliminary characterization of an isolated mycothiol-dependent Glo1 from Streptomyces coelicolor. The enzyme has been determined to display a Ni2+-activation profile and indicates that Ni2+-activated Glo1 are indeed widespread in nature regardless of the intracellular thiol employed by an organism.

Reduction of glyoxalase 1 (GLO1) aggravates cerebrovascular remodeling via promoting the proliferation of basilar smooth muscle cells in hypertension

Uncontrollable vascular smooth cell proliferation is responsible for vascular remodeling during hypertension development. Glyoxalase 1 (GLO1), the major enzyme detoxifying methylglyoxal, has a critical role in regulating proliferation of several cell types. However, little is known whether GLO1 is involved in cerebrovascular remodeling and basilar smooth muscle cell (BASMC) proliferation during hypertension. Here we explored the role of GLO1 in angiotensin II (Ang II)-induced cerebrovascular remodeling and proliferation of BASMCs and the underlying mechanisms. The protein expression of GLO1 in basilar arteries from hypertensive mice was decreased, and GLO1 expression was negatively correlated with medial cross-sectional area and blood pressure in basilar arteries during hypertension. Knockdown of GLO1 promoted while overexpression of GLO1 prevented Ang II-induced cell proliferation and cell cycle transition in BASMCs. These results were related to the inhibitory effects of GLO1 on PI3K/AKT/CDK2 cascade activation upon Ang II treatment. In addition, in vivo study, GLO1 overexpression with adeno-associated virus harboring GLO1 cDNA improved cerebrovascular remodeling in basilar artery tissue during Ang II-induced hypertension development. These data indicate that GLO1 reduction mediates cerebrovascular modeling via PI3K/AKT/CDK2 cascade-dependent BASMC proliferation. GLO1 acts as a negative regulator of hypertension-induced cerebrovascular remodeling and targeting GLO1 may be a novel therapeutic strategy to prevent hypertension-associated cardiovascular complications such as stroke.

Quercetin Mediated Salt Tolerance in Tomato through the Enhancement of Plant Antioxidant Defense and Glyoxalase Systems.

Quercetin (Qu) is a strong antioxidant among the phenolic compounds having physiological and biochemical roles in plants. Hence, we have studied the Qu evolved protection against salinity in tomato (Solanum lycopersicum L.). Salinity caused ionic toxicity by increasing Na+ content in seedlings along with nutritional starvation of K+, Ca2+, and Mg2+. While osmotic stress was detected by higher free proline (Pro) content and lower leaf relative water content (LRWC) in salt-stressed seedlings. Salt toxicity also induced higher H2O2 generation, malondialdehyde (MDA) content and lipoxygenase (LOX) activity as a sign of oxidative stress. Tomato seedlings suffered from methylglyoxal (MG) toxicity, degradation of chlorophyll, along with lower biomass accumulation and growth due to salt exposure. However, Qu application under salinity resulted in lower Na+/K+ due to reduced Na+ content, higher LRWC, increased Pro, and reduction of H2O2 and MDA content, and LOX activity, which indicated alleviation of ionic, osmotic, and oxidative stress respectively. Quercetin caused oxidative stress, lessening through the strengthening of both enzymatic and non-enzymatic antioxidants. In addition, Qu increased glutathione S-transferase activity in salt-invaded seedlings, which might be stimulated reactive oxygen species (ROS) scavenging along with higher GSH content. As a result, toxic MG was detoxified in Qu supplemented salt-stressed seedlings by increasing both Gly I and Gly II activities. Moreover, Qu insisted on better plant growth and photosynthetic pigments synthesis in saline or without saline media. Therefore, exogenous applied Qu may become an important actor to minimize salt-induced toxicity in crops.

Oleuropein-Induced Apoptosis Is Mediated by Mitochondrial Glyoxalase 2 in NSCLC A549 Cells: A Mechanistic Inside and a Possible Novel Nonenzymatic Role for an Ancient Enzyme.

Oleuropein (OP) is a bioactive compound derived from plants of the genus Oleaceae exhibiting antitumor properties in several human cancers, including non-small-cell lung cancer (NSCLC). Recent evidence suggests that OP has proapoptotic effects on NSCLC cells via the mitochondrial apoptotic pathway. However, the exact molecular mechanisms behind the apoptogenic action of OP in NSCLC are still largely unknown. Glyoxalase 2 (Glo2) is an ancient enzyme belonging to the glyoxalase system involved in the detoxification of glycolysis-derived methylglyoxal. However, emerging evidence suggests that Glo2 may have also nonenzymatic roles in some malignant cells. In the present study, we evaluated whether and how Glo2 participated in the proapoptotic effects of OP in NSCLC A549 cells. Our results indicate that OP is able to induce apoptosis in A549 cells through the upregulation of mitochondrial Glo2 (mGlo2), mediated by the superoxide anion and Akt signaling pathway. Moreover, our data shows that the proapoptotic role of mGlo2, observed following OP exposure, occurs via the interaction of mGlo2 with the proapoptotic Bax protein. Conversely, OP does not alter the behavior of nonmalignant human BEAS-2B cells or mGlo2 expression, thus suggesting a specific anticancer role for this bioactive compound in NSCLC. Our data identify a novel pathway through which OP exerts a proapoptotic effect in NSCLC and suggest, for the first time, a novel, nonenzymatic antiapoptotic role for this ancient enzyme in NSCLC.

Regulation of Reactive Oxygen Species Metabolism and Glyoxalase Systems by Exogenous Osmolytes Confers Thermotolerance in Brassica napus

In the current study, the beneficial role of proline (Pro) and glycinebetaine (GB) in alleviating high temperature (HT) stress was investigated in Brassica napus L. (rapeseed) seedlings. Ten-d-old rapeseed seedlings were treated with and/or without 2 mM Pro and 2 mM GB and exposed to 38/28 °C day/night (16 and 8 h) temperature for 24 and 48 h. Heat stress induced high amount of hydrogen peroxide (H2O2), lipoxygenase (LOX) and malondialdehyde (MDA) level. Exogenous Pro and GB addition in HT-affected plants increased the levels of ascorbate (AsA) and reduced glutathione (GSH) and their redox pool. Exogenous Pro and GB increased functions of AsA-GSH pathways enzymes along with catalase (CAT), glutathione S-transferase (GST) and glutathione peroxidase (GPX) in HT exposed plants. The enhanced antioxidant defense system by supplementation of Pro and GB helped to reduce oxidative stress and photosynthetic pigments damage caused by HT-induced stress. High methylglyoxal (MG) content decreased upon exogenous Pro and GB application along with enhanced activities of glyoxalase I (Gly I) and Gly II enzymes in HT-stressed plants. Applied Pro and GB under HT stress enhanced endogenous Pro level further to prevent excess water loss and also improved relative water content. Thus, Pro- and/or GB-induced regulatory interactions between ROS and MG detoxification systems may be a useful approach for the reversal of HT-induced oxidative stress.

EDTA reduces cadmium toxicity in mustard (Brassica juncea L.) by enhancing metal chelation, antioxidant defense and glyoxalase systems

To investigate the possible role of EDTA in mitigating cadmium (Cd) toxicity, we treated mustard (<em>Brassica juncea</em> L.) seedlings with CdCl<sub>2</sub> (0.5 mM and 1.0 mM, 3 days) alone and in combination with 0.5 mM EDTA in a semihydroponic medium. In the absence of EDTA, mustard seedlings accumulated Cd in their roots and shoots in a concentration dependent manner. Overaccumulation of Cd boosted generation of hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) and superoxide anions (O<sub>2</sub><sup>•−</sup>), increased lipoxygenase (LOX) activity, lipid peroxidation, and cytotoxic methylglyoxal (MG) content. It also disturbed components of the antioxidant defense and glyoxalase systems. Furthermore, Cd stress decreased growth, leaf relative water content (RWC) and chlorophyll (chl) content but augmented the proline (Pro) content. On the other hand, EDTA supplemented Cd-stressed seedlings improved the constituents of the AsA-GSH cycle with the upregulated activities of ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), glutathione reductase (GR), glutathione peroxidase (GPX), superoxide dismutase (SOD), and catalase (CAT). Moreover, addition of EDTA to the Cd-stressed seedlings notably enhanced Gly I activity in contrast to the stress treatment. Ethylenediaminetetraacetic acid decreased Cd accumulation in the both shoots and roots, as well as increased other nonprotein thiols (NPTs) in leaves, including the phytochelatin (PC) content. It also decreased H<sub>2</sub>O<sub>2</sub> and O<sub>2</sub><sup>•−</sup> generation, lipid peroxidation and MG content but enhanced RWC, chl and Pro contents in the leaves, which confirmed the improved growth of seedlings. The findings of the study suggest that exogenous application of EDTA to the Cd-treated seedlings reduces Cd-induced oxidative injuries by restricting Cd uptake, increasing NPTs concentration and upregulating most of the components of their antioxidant defense and glyoxalase systems.

Exogenous application of gibberellic acid mitigates drought-induced damage in spring wheat

Drought stress is a major problem in wheat production but it could be managed by using various exogenous protectants such as gibberellic acid (GA). Although GA is a plant growth hormone, it shows a potential to protect the plant in stress conditions. To investigate the possible role of GA in mitigating drought stress, we treated wheat (<em>Triticum aestivum</em> ‘BARI Gom-21’) seedlings with a GA spray under semihydroponic conditions. In the experiment, the combined effect of GA and drought stress (induced by 12% polyethylene glycol) was studied after 48 h and 72 h. In the absence of exogenous GA, drought-stressed wheat seedlings showed various physiological and biochemical changes in a time-dependent manner. Malondialdehyde (MDA), hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) and free proline (Pro) concentrations were increased, whereas catalase (CAT) and ascorbate peroxidase (APX) activities were reduced under drought stress. Gibberellic acid played a role in restoring the ascorbate (AsA) level, decreased the reduced/oxidized glutathione (GSH/GSSG) ratio and reduced monodehydroascorbate reductase (MDHAR) and dehydroascorbate reductase (DHAR) activities. Gibberellic acid significantly affected the glyoxalase system. Under drought stress, the methylglyoxal (MG) concentration was increased but GA application stimulated glyoxalase I (Gly I) and glyoxalase II (Gly II) activities to protect the wheat seedlings against stress. The study concluded that the severity of drought stress in wheat depends on the growth stage and it increases with an increase in the duration of stress, whereas exogenous GA helped the seedlings to survive by upregulating antioxidant defense mechanisms and the glyoxalase system.

Oxidative Damage and Antioxidant Defense in Sesamum indicum after Different Waterlogging Durations.

The present study was designed to investigate the duration-dependent changes in the biochemical attributes of sesame in response to waterlogging stress. Sesame plants (Sesamum indicum L. cv. BARI Til-4) were subjected to waterlogging for 2, 4, 6, and 8 days during the vegetative stage and data were measured following waterlogging treatment. The present study proves that waterlogging causes severe damage to different attributes of the sesame plant. The plants showed an increasing trend in lipid peroxidation as well as hydrogen peroxide (H2O2) and methylglyoxal contents that corresponded to increased stress duration. A prolonged period of waterlogging decreased leaf relative water content and proline content. Photosynthetic pigments, like chlorophyll (chl) a, b, and chl (a+b) and carotenoid contents, also decreased over time in stressed plants. Glutathione (GSH) and oxidized glutathione (GSSG) contents increased under waterlogging, while the GSH/GSSG ratio and ascorbate content decreased, indicating the disruption of redox balance in the cell. Ascorbate peroxidase, monodehydroascorbate reductase, and glutathione peroxidase activity increased under waterlogging, while dehydroascorbate reductase, glutathione reductase, and catalase activity mostly decreased. Waterlogging modulated the glyoxalase system mostly by enhancing glyoxalase II activity, with a slight increase in glyoxalase I activity. The present study also demonstrates the induction of oxidative stress via waterlogging in sesame plants and that stress levels increase with increased waterlogging duration.

2-Iodo-4'-Methoxychalcone Attenuates Methylglyoxal-Induced Neurotoxicity by Activation of GLP-1 Receptor and Enhancement of Neurotrophic Signal, Antioxidant Defense and Glyoxalase Pathway.

Methylglyoxal (MG) acts as a reactive precursor of advanced glycation end products (AGEs). This compound is often connected with pathologies such as diabetes, neurodegenerative processes and diseases of aging. 2-iodo-4′-methoxychalcone (CHA79), a synthetic halogen-containing chalcone derivative, has been reported its anti-diabetic activity. This study aims to investigate the potential protective capability of CHA79 against MG-mediated neurotoxicity in SH-SY5Y cells. Results indicated CHA79 increased viability of cells and attenuated the rate of apoptosis in MG-exposed SH-SY5Y. CHA79 up-regulated expression of anti-apoptotic protein (Bcl-2) and down-regulated apoptotic proteins (Bax, cytochrome c, caspase-3, caspase-9). Moreover, CHA79 significantly up-regulated expression of neurotrophic factors, including glucagon-like peptide-1 receptor (GLP-1R), brain derived neurotrophic factor (BDNF), p75NTR, p-TrkB, p-Akt, p-GK-3β and p-CREB. CHA79 attenuated MG-induced ROS production and enhanced the antioxidant defense including nuclear factor erythroid 2-related factor 2 (Nrf2), HO-1, SOD and GSH. Furthermore, CHA79 attenuated MG-induced reduction of glyoxalase-1 (GLO-1), a vital enzyme on removing AGE precursors. In conclusion, CHA79 is the first novel synthetic chalcone possessing the GLP-1R and GLO-1 activating properties. CHA 79 also exhibits neuroprotective effects against MG toxicity by enhancing neurotrophic signal, antioxidant defense and anti-apoptosis pathway.

406-P: Effect of Metformin on Dicarbonyl Stress in Patients with Type 2 Diabetes and Chronic Heart Failure

Background and Aims: Treatment with metformin (MET) in diabetic patients with chronic heart failure (HF) is associated with better cardiovascular outcomes. The endothelial dysfunction, atherosclerosis and macrovascular diabetic complications are associated with excessive generation of reactive dicarbonyls like methylglyoxal (MG). MET has been proposed as a scavenger of methylglyoxal. The aim of the study was to evaluate the effects of MET on substrate metabolism and dicarbonyl stress in patients with DM and HF. Materials and Methods: A randomized, double-blind, placebo-controlled, crossover study in a total time of six months testing the effect of 3- month usage of MET (2 g/day) vs. placebo. A group of 40 treatment naive patients with type 2 DM and HF were included. At the beginning and at the end of each intervention period various metabolic tests were done including hyperinsulinemic-isoglycemic clamp, parameters of dicarbonyl stress like MG, Glutathione (GSH), glutathione disulfide (GSSG), and the activity of Glyoxalase1 (GLO-1). Results: MET significantly reduced HbA1c compared to placebo (p˂0.01). Insulin sensitivity, measured as a metabolic clearance rate of glucose (MCR) during the clamp, improved significantly with MET (p˂0.001) but also, although less significantly, with placebo (p˂0.05), difference between MET and placebo was not significant. Compared to placebo, GHS levels increased after the intervention (baseline vs. MET vs. PL: 866 ± 231 vs. 994 ± 342 vs. 878 ± 266µmol/l, p<0.05). GSH/GSSG ratio was not affected. The levels of MG decreased after MET (p<0.05), decrease after placebo was not significant. The activity of GLO-1 was not affected. Conclusions: Our results on a unique group of patients with heart failure and diabetes support the evidence that one of the cardioprotective pathways of metformin is through reducing dicarbonyl stress. As there was no change in the GLO-1 activity, it might be due to binding and deactivation of reactive dicarbonyls by MET. Disclosure J. Kopecky: None. E. Hošková: None. J. Veleba: None. H. Malinska: None. K. Velebova: None. V. Melenovsky: None. T. Pelikanova: None. Funding Agentura pro zdravotnický výzkum Ceské Republiky (AZV16-27496A)