Exploration of the antioxidative defense system to characterize chickpea genotypes showing differential response towards water deficit conditions

The present investigation was carried out to characterize genotypic variability in chickpea for water deficit tolerance by exploring the antioxidative defense system and seedling growth. Twenty nine chickpea genotypes including cultivars and advanced lines were grown under control and water deficit conditions induced by adding 3 % mannitol. The genotypes showed differential response in seedling growth under water deficit conditions. The activities of catalase (CAT) and superoxide dismutase (SOD) were observed to be differentially expressed in the roots of various genotypes, under control and water deficit conditions. The contents of H₂O₂, malondialdehyde (MDA) and proline were also observed to be variable in the roots of all the genotypes, under control and water deficit conditions. Stress tolerance index for the various parameters, viz, CAT and SOD activity, H₂O₂, MDA and proline content, root length, shoot length and their biomass was determined and the level of stress resistance calculated. The genotypes which showed increased activities of CAT and SOD, decreased contents of H₂O₂ and MDA together with least affected seedling growth under water deficit conditions exhibited higher stress resistance capacity. Multivariate principal component analysis for all the parameters affected under water deficit conditions, grouped the genotypes into three clusters having different (high, moderate and low) levels of stress resistance. Complete linkage clustering grouped these genotypes into two major clusters-I and II. The genotypes present in sub–sub cluster ‘A1’ and sub cluster ‘B’ of major cluster-I have been observed to possess high stress resistance levels for respective parameters. It can thus be concluded that chickpea genotypes exhibiting increased stress resistance levels in relation to SOD and CAT activities, H₂O₂ and MDA contents and seedling growth would have higher stress tolerance under water deficit conditions.     

Caffeic acid phenethyl ester reduces neurovascular inflammation and protects rat brain following transient focal cerebral ischemia

Ischemic stroke is a neurovascular disease treatable by thrombolytic therapy, but the therapy has to be initiated within 3 h of the incident. This therapeutic limitation stems from the secondary injury which results mainly from oxidative stress and inflammation. A potent antioxidant/anti-inflammatory agent, caffeic acid phenethyl ester (CAPE) has potential to mitigate stroke's secondary injury, and thereby widening the therapeutic window. We observed that CAPE protected the brain in a dose-dependent manner (1-10 mg/kg body weight) and showed a wide therapeutic window (about 18 h) in a rat model of transient focal cerebral ischemia and reperfusion. The treatment also increased nitric oxide and glutathione levels, decreased lipid peroxidation and nitrotyrosine levels, and enhanced cerebral blood flow. CAPE down-regulated inflammation by blocking nuclear factor kappa B activity. The affected mediators included adhesion molecules (intercellular adhesion molecule-1 and E-selectin), cytokines (tumor necrosis factor-alpha and interleukin-1beta) and inducible nitric oxide synthase. Anti-inflammatory action of CAPE was further documented through reduction of ED1 (marker of activated macrophage/microglia) expression. The treatment inhibited apoptotic cell death by down-regulating caspase 3 and up-regulating anti-apoptotic protein Bcl-xL. Conclusively, CAPE is a promising drug candidate for ischemic stroke treatment due to its inhibition of oxidative stress and inflammation, and its clinically relevant wide therapeutic window.     

Effect of Rice Straw Incorporation on Phytotoxicity of Isoxaflutole to an Exotic Weed Phalaris minor Retz.

Phalaris minorRetz. is a major exotic annual weed in the wheat (Triticum aestivum L.) crop. Unharvested rice (Oryza sativa L.) straw, unburned and burned, is often incorporated in the field prior to cultivating wheat. Isoxaflutole (Balance), a pre-emergent systemic soil applied herbicide, has potential to control P. minor. Glasshouse experiments were conducted to determine the phytotoxicity of isoxaflutole defined by reductions in relation to shoot length of P. minor when grown in unamended soil or soil amended with unburned or burned rice straw. A 120 g soil was amended with 0, 1, 2 and 4 g of unburned or burned rice straw, and placed in 150 mL styrofoam pots. Appropriate amount of isoxaflutole (75% active ingredient, ai) was added to pots to get final concentration of 0, 7.5, 30, 60 and 120 μg ai/pot. Unamended soil and soil amended with unburned or burned rice straw were analyzed for pH and organic matter; two important determinants of isoxaflutole activity. Results indicate a significant reduction in shoot length of P. minor when grown in soil treated with isoxaflutole at 30, 60 or 120 μg ai/pot. Inhibition in the shoot length of P. minor was observed when soil amended with unburned straw was treated with isoxaflutole at 7.5 and 30 μg ai/pot compared with unamended soil treated with similar amounts of isoxaflutole. No significant change in isoxaflutole toxicity was observed when soil amended with unburned straw was treated with isoxaflutole at 60 and 120 lg ai/pot compared with unamended soil treated with similar amounts of isoxaflutole. Isoxaflutole phytotoxicity to P. minor shoot length was eliminated when soil amended with burned straw was treated with isoxaflutole at 7.5 and 30 μg ai/pot. P. minor shoot length was greater when soil amended with burned straw was treated with isoxaflutole at 60 and 120 μg ai/pot relative to herbicide-treated unamended soils. We conclude that incorporation of burned rice straw greatly reduces the phytotoxicity of isoxaflutole toP. minor.     

Soil Microorganisms: An Important Determinant of Allelopathic Activity

Current evidence illustrates the significance of soil microbes in influencing the bioavailability of allelochemicals. This review discusses (i) the significance of soil microorganisms in influencing allelopathic expression, (ii) different ways of avoiding microbial degradation of putative allelochemicals, and (iii) the need of incorporating experiments on microbial modification of allelochemicals in laboratory bioassays for allelopathy. Several climatic and edaphic factors affect the soil microflora; therefore, allelopathy should be assessed in a range of soil types. Allelopathy can be better understood in terms of soil microbial ecology, and appropriate methodologies are needed to evaluate the roles of soil microorganisms in chemically-mediated interactions between plants.     

Plant allelochemical interference or soil chemical ecology?

While allelopathy has been defined as plant-plant chemical interference, there has been much confusion about what the concept encompasses and how important it is in nature. We distinguish between (1) direct plant-plant interference mediated by allelochemicals, and (2) the effects of secondary compounds released by plants on abiotic and biotic soil processes that affect other plants.It very difficult to demonstrate direct effects of chemicals released by a plant on nearby plants. Although soil ecology-mediated effects of secondary plant compounds do not fit the classical concept of allelopathy, we find support in the literature for the hypothesis that the most important effects of compounds released into the soil environment by plants on other plants occur through such indirect effects. The emphasis on, and skepticism of, direct plant-plant allelopathic interference has led some researchers to demand unreasonably high standards of evidence for establishing even the existence of allelopathic interactions, standards that are not demanded for other plant-plant interactions such as resource competition. While the complete elucidation of the mechanisms by which allelochemicals function in the field is many years away, such elucidation is not necessary to establish the existence of allelopathic interactions.We propose that most of the phenomena broadly referred to as allelopathic interference are better conceptualized and investigated in terms of soil chemical ecology. Even when direct plant-plant allelochemical interference occur, the levels of allelochemicals in the environment and their effects on plants are heavily influenced by abiotic and biotic components of the soil ecosystem. Putting allelopathy in the context of soil ecology can further research and reduce some of the less fruitful controversy surrounding the phenomenon.     

Experimental designs for the study of allelopathy

The primary aim of this paper is to discuss the methodological approaches that may best develop studies of allelopathy in the future. Laboratory studies on the functions of isolated chemicals, no matter how mechanistically detailed, cannot demonstrate the significance of allelopathy in communities. Evidence for allelopathy in natural plant communities should include information of concentrations and release rates such as demonstrated in field soils for (±)-catechin and Centaurea maculosaLam. Community-relevant evidence for allelopathy should include some manipulation of exudates such as performed in many experiments with activated carbon and gel filtration columns. Realistic evidence for allelopathy should include separation of resource effects from chemical effects; such as demonstrated by experiments with activated carbon additions, density-dependent responses to additions of competitors and chemicals, and resource addition treatments. Community-relevant evidence should link laboratory effects to field patterns and experiments; such as the links between the inhibitory effects of roots of Larrea tridentataCov., the highly spatially segregated root systems and regular above-ground spacing of this species, strong spatial disassociation of L. tridentata with other species, and removal experiments indicating that segregation of L. tridentata root systems via allelopathy may feed back to sequestering of resource use. Studies of allelopathy should consider chemically initiated shifts in microbial populations, and the effects of organic and inorganic soil components on the function of exudates; which has been done in a number of studies. Finally, studies of allelopathy should include large-scale manipulation of chemical effects; such as performed in field experiments in boreal forests in Sweden with Empetrum hermaphroditumHagerup and Scots pine (Pinus sylvestrisL.). Demonstrating the occurrence and importance of chemically mediated interactions among plants is not trivial. If even a small portion of the thousands of chemicals produced by different plant species have effects on their neighbours, then species-specific interactions, natural selection, community integration, and community coevolution may be quite different than predicted by conceptual models based solely on resource competition. Appropriate methodology is crucial for integrating chemically mediated interactions into ecological theory.     

Expression of antioxidant enzymes in rat kidney during ischemia-reperfusion injury

The effect of ischemia-reperfusion on activity, protein and m-RNA levels of catalase, copper-zinc and manganese containing superoxide dismutases and glutathione peroxidase, the enzymes that are involved in free radical detoxification was studied in rat kidney. Ischemia alone did not alter either the activities or protein levels of superoxide dismutase and glutathione peroxidase. However, catalase activity was found to be inhibited to 82% of control. The inhibition of catalase was due to the inactivation of the enzyme as there was no significant change in enzyme protein level. Reperfusion following ischemia, however, led to a significant decrease in both the activities as well as the protein levels of all the antioxidant enzymes. The observed overall decrease in total superoxide dismutase activity was the net effect of a decrease in copper-zinc superoxide dismutase while manganese superoxide dismutase activity was found to be increased following reperfusion. This observed increased manganese superoxide dismutase activity was the result of its increased protein level. The mRNA levels for catalase, superoxide dismutases, and glutathione peroxidase were observed to be increased (100–145% of controls) following ischemia; reperfusion of ischemic kidneys, however, resulted in a significant decrease in the levels of mRNAs coding for all the enzymes except manganese superoxide dismutase which remained high. These results suggest that in tissue, the down regulation of the antioxidant enzyme system could be responsible for the pathophysiology of ischemia-reperfusion injury.     

Postnatal Development and Isolation of Peroxisomes from Brain

We analyzed the postnatal peroxisome development in rat brain by measuring the enzyme activities of catalase and acyl-CoA oxidase and beta-oxidation of [1-14C]lignoceric acid. These enzyme activities were higher between 10 and 16 days of postnatal life and then decreased. We developed and compared two different methods for isolation of enriched peroxisomes from 10-day-old rat brain by using a combination of differential and density gradient centrifugation techniques. Peroxisomes in Percoll (self-generating gradient) banded at a density of 1.036 +/- 0.012 g/ml and in Nycodenz continuous gradient at 1.125 +/- 0.014 g/ml. Acyl-CoA oxidase, D-amino acid oxidase, L-pipecolic acid oxidase, and dihydroxyacetone phosphate acyltransferase activities and activities for the oxidation of very long chain fatty acid (lignoceric acid) were almost exclusively associated with catalase activity (a marker enzyme for peroxisomes) in the gradient. The postnatal increase in peroxisomal activity with the onset of myelination and the presence of enzyme for the biosynthesis of plasmalogens and oxidation of very long chain fatty acid (both predominant constituents of myelin) suggest that brain peroxisomes may play an important role in the assembly and turnover of myelin.     

Functional Characterization of IPSC-Derived Brain Cells as a Model for X-Linked Adrenoleukodystrophy

X-ALD is an inherited neurodegenerative disorder where mutations in the ABCD1 gene result in clinically diverse phenotypes: the fatal disorder of cerebral childhood ALD (cALD) or a milder disorder of adrenomyeloneuropathy (AMN). The various models used to study the pathobiology of X-ALD disease lack the appropriate presentation for different phenotypes of cALD vs AMN. This study demonstrates that induced pluripotent stem cells (IPSC) derived brain cells astrocytes (Ast), neurons and oligodendrocytes (OLs) express morphological and functional activities of the respective brain cell types. The excessive accumulation of saturated VLCFA, a “hallmark” of X-ALD, was observed in both AMN OLs and cALD OLs with higher levels observed in cALD OLs than AMN OLs. The levels of ELOVL1 (ELOVL Fatty Acid Elongase 1) mRNA parallel the VLCFA load in AMN and cALD OLs. Furthermore, cALD Ast expressed higher levels of proinflammatory cytokines than AMN Ast and control Ast with or without stimulation with lipopolysaccharide. These results document that IPSC-derived Ast and OLs from cALD and AMN fibroblasts mimic the respective biochemical disease phenotypes and thus provide an ideal platform to investigate the mechanism of VLCFA load in cALD OLs and VLCFA-induced inflammatory disease mechanisms of cALD Ast and thus for testing of new therapeutics for AMN and cALD disease of X-ALD.