Induction of Cell Death through Alteration of Oxidants and Antioxidants in Lung Epithelial Cells Exposed to High Energy Protons

Radiation affects several cellular and molecular processes, including double strand breakage and modifications of sugar moieties and bases. In outer space, protons are the primary radiation source that poses a range of potential health risks to astronauts. On the other hand, the use of proton irradiation for tumor radiation therapy is increasing, as it largely spares healthy tissues while killing tumor tissues. Although radiation-related research has been conducted extensively, the molecular toxicology and cellular mechanisms affected by proton irradiation remain poorly understood. Therefore, in this study, we irradiated rat lung epithelial cells with different doses of protons and investigated their effects on cell proliferation and death. Our data show an inhibition of cell proliferation in proton-irradiated cells with a significant dose-dependent activation and repression of reactive oxygen species and antioxidants glutathione and superoxide dismutase, respectively, compared with control cells. In addition, the activities of apoptosis-related genes such as caspase-3 and -8 were induced in a dose-dependent manner with corresponding increased levels of DNA fragmentation in proton-irradiated cells compared with control cells. Together, our results show that proton irradiation alters oxidant and antioxidant levels in cells to activate the apoptotic pathway for cell death.     

Subjective Duration Distortions Mirror Neural Repetition Suppression

Background

Subjective duration is strongly influenced by repetition and novelty, such that an oddball stimulus in a stream of repeated stimuli appears to last longer in duration in comparison. We hypothesize that this duration illusion, called the temporal oddball effect, is a result of the difference in expectation between the oddball and the repeated stimuli. Specifically, we conjecture that the repeated stimuli contract in duration as a result of increased predictability; these duration contractions, we suggest, result from decreased neural response amplitude with repetition, known as repetition suppression.

Methodology/Principal Findings

Participants viewed trials consisting of lines presented at a particular orientation (standard stimuli) followed by a line presented at a different orientation (oddball stimulus). We found that the size of the oddball effect correlates with the number of repetitions of the standard stimulus as well as the amount of deviance from the oddball stimulus; both of these results are consistent with a repetition suppression hypothesis. Further, we find that the temporal oddball effect is sensitive to experimental context – that is, the size of the oddball effect for a particular experimental trial is influenced by the range of duration distortions seen in preceding trials.

Conclusions/Significance

Our data suggest that the repetition-related duration contractions causing the oddball effect are a result of neural repetition suppression. More generally, subjective duration may reflect the prediction error associated with a stimulus and, consequently, the efficiency of encoding that stimulus. Additionally, we emphasize that experimental context effects need to be taken into consideration when designing duration-related tasks.     

Cultivable Bacterial Diversity of Alkaline Lonar Lake, India

Aerobic, alkaliphilic bacteria were isolated and characterized from water and sediment samples collected in the winter season, January 2002 from alkaline Lonar lake, India, having pH 10.5. The total number of microorganisms in the sediment and water samples was found to be 10²–10⁶ cfu g⁻¹ and 10²–10⁴ cfu ml⁻¹, respectively. One hundred and ninety-six strains were isolated using different enrichment media. To study the bacterial diversity of Lonar lake and to select the bacterial strains for further characterization, screening was done on the basis of pH and salt tolerance of the isolates. Sixty-four isolates were subjected to phenotypic, biochemical characterization and 16S rRNA sequencing. Out of 64, 31 bacterial isolates were selected on the basis of their enzyme profile and further subjected to phylogenetic analysis. Phylogenetic analysis indicated that most of the Lonar lake isolates were related to the phylum Firmicutes, containing Low G+C, Gram-positive bacteria, with different genera: Bacillus, Paenibacillus, Alkalibacillus, Exiguobacterium, Planococcus, Enterococcus and Vagococcus. Seven strains constituted a Gram-negative bacterial group, with different genera: Halomonas, Stenotrophomonas and Providencia affiliated to γ-Proteobacteria, Alcaligenes to β-Proteobacteria and Paracoccus to α-Proteobacteria. Only five isolates were High G+C, Gram-positive bacteria associated with phylum Actinobacteria, with various genera: Cellulosimicrobium, Dietzia, Arthrobacter and Micrococcus. Despite the alkaline pH of the Lonar lake, most of the strains were alkalitolerant and only two strains were obligate alkaliphilic. Most of the isolates produced biotechnologically important enzymes at alkaline pH, while only two isolates (ARI 351 and ARI 341) showed the presence of polyhydroxyalkcanoate (PHA) and exopolysaccharide (EPS), respectively.     

Glutathione-S-transferase, superoxide dismutase, xanthine oxidase, catalase, glutathione peroxidase and lipid peroxidation in the liver of exercised rats

Glutathione-S-transferase (GST), superoxide dismutase (SOD), Xanthine oxidase, selenium-dependent glutathione peroxidase (GPxI), catalase activities and malondialdehyde (MDA) content were determined in liver of three groups of exercised rats (E) viz., one day (E1), 10 days (E10) and 60 days (E60). GST, SOD and xanthine oxidase activities increased significantly with the increase in exercise period. Lipid peroxidation, expressed in terms of MDA formation, also increased in the liver of all the three groups. But catalase activity decreased significantly during exercise. Further, GPxI did not show any significant change in its activity in response to exercise. Our findings indicate that: 1) The significant increase in GST activity suggests their induction aimed at counteracting the oxidant stress induced during exercise; 2) The significant increase in xanthine oxidase and SOD activities indicates the generation of more superoxide anion radicals and their removal, respectively. 3) The significant reduction in catalase activity denotes the decreased formation of hydrogenperoxides during exercise; and 4) The pattern of changes in the activity level of GPxI indicate its least participation during exercise. However, in another way it is giving a scope for the involvement of GPxII associated with GST in the reduction of organic hydroperoxides. Further more, the relative increase in MDA is considered as the indicator of the rate of lipid peroxidation in the wake of exhaustive exercise.     

Computational characterization of substrate and product specificities,and functionality of S‐adenosylmethionine binding pocket in histone lysine methyltransferases from Arabidopsis,rice and maize

Histone lysine methylation by histone lysine methyltransferases (HKMTs) has been implicated in regulation of gene expression. While significant progress has been made to understand the roles and mechanisms of animal HKMT functions, only a few plant HKMTs are functionally characterized. To unravel histone substrate specificity, degree of methylation and catalytic activity, we analyzed Arabidopsis Trithorax‐like protein (ATX), Su (var)3‐9 h omologs protein (SUVH), Su(var)3‐9 related protein (SUVR), ATXR5, ATXR6, and E(Z) HKMTs of Arabidopsis, maize and rice through sequence and structure comparison. We show that ATXs may exhibit methyltransferase specificity toward histone 3 lysine 4 (H3K4) and might catalyse the trimethylation. Our analyses also indicate that most SUVH proteins of Arabidopsis may bind histone H3 lysine 9 (H3K9). We also predict that SUVH7, SUVH8, SUVR1, SUVR3, ZmSET20 and ZmSET22 catalyse monomethylation or dimethylation of H3K9. Except for SDG728, which may trimethylate H3K9, all SUVH paralogs in rice may catalyse monomethylation or dimethylation. ZmSET11, ZmSET31, SDG713, SDG715, and SDG726 proteins are predicted to be catalytically inactive because of an incomplete S‐adenosylmethionine (SAM) binding pocket and a post‐SET domain. E(Z) homologs can trimethylate H3K27 substrate, which is similar to the Enhancer of Zeste homolog 2 of humans. Our comparative sequence analyses reveal that ATXR5 and ATXR6 lack motifs/domains required for protein‐protein interaction and polycomb repressive complex 2 complex formation. We propose that subtle variations of key residues at substrate or SAM binding pocket, around the catalytic pocket, or presence of pre‐SET and post‐SET domains in HKMTs of the aforementioned plant species lead to variations in class‐specific HKMT functions and further determine their substrate specificity, the degree of methylation and catalytic activity.     

Structure-function correlation during the etioplast-chloroplast transition in cucumber cotyledonary leaves

We studied the development of chloroplasts from etioplasts in the cotyledonary leaves of 4-d-old dark-grown cucumber (Cucumis sativus) seedlings after irradiation (20 μmol m^-2 s^-1). Upon irradiation, the triggering of chlorophyll (Chl) synthesis and accumulation showed a relatively short lag phase. The irradiation of etiolated seedlings initiated the synthesis of apoproteins of pigment-protein complexes. While Chl-protein 2 (CP2) was detected at 6 h after irradiation, CP1 only after 29 h. The appearance and accumulation of some of the apoproteins were monitored by Western-blotting. LHC2 apoprotein was detected after a 6 h-irradiation. The amounts of D1 protein of photosystem (PS) 2 and PsaA/B protein of PS1 were quantitated by ELISA. Further, the thylakoid membrane function during this time period in terms of PS1- and PS2-mediated electron transfer activity and intersystem electron pool size were analyzed. While PS1 activity was detected after 4 h, PS2-mediated O₂ evolution was detected only after a 17 h-irradiation. F_v/F_m value of Chl a fluorescence measurements indicated that the photochemical efficiency of these leaves reached its maximum after 29 h of irradiation. The intersystem pool size of cotyledonary leaves was equivalent to that of the control cotyledonary leaves grown for 25 h under continuous irradiation. Thus etioplasts develop into fully functional chloroplasts after approximately 25 h when 4 d-dark grown cucumber seedlings are continuously moderately irradiated. The development of photosynthetic electron transport chain seems to be limited in time at the level of PS2, possibly at the donor side. This revised version was published online in September 2006 with corrections to the Cover Date.     

Effect of bacoside A on brain antioxidant status in cigarette smoke exposed rats

Free radicals mediated oxidative stress has been implicated in the pathogenesis of smoking-related diseases and antioxidant nutrients are reported to prevent the oxidative damage induced by smoking. Therefore, the present study was conducted to evaluate the antioxidant role of bacoside A (triterpenoid saponin isolated from Bacopa monniera) against chronic cigarette smoking induced oxidative damage in rat brain. Adult male albino rats were exposed to cigarette smoke for a period of 12 weeks and simultaneously administered with bacoside A (10 mg/kg b.w./day, p.o.). Antioxidant status of the brain was assessed from the levels of reduced glutathione, vitamin C, vitamin E, and vitamin A and the activities of superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase. The levels of copper, iron, zinc and selenium in brain and serum ceruloplasmin activity were also measured. Oxidative stress was evident from the diminished levels of both enzymatic and non-enzymatic antioxidants. Alterations in the levels of trace elements with accumulation of copper and iron, and depletion of zinc and selenium were also observed. Bacoside A administration improved the antioxidant status and maintained the levels of trace elements. These results suggest that chronic cigarette smoke exposure enhances oxidative stress, thereby disturbing the tissue defense system and bacoside A protects the brain from the oxidative damage through its antioxidant potential.     

Biological and conformational evaluation of angiotensin II lactam bridge containing analogues

Angiotensin II (AII) is the active octapeptide product of the renin enzymatic cascade, which is responsible for sustaining blood pressure. In an attempt to establish the AII-receptor-bound conformation of this octapeptide, we designed conformationally constrained analogues by scanning the entire AII sequence with an i-(i+2) and i-(i+3) lactam bridge consisting of an Asp-(Xaa)(n)-Lys scaffold. Most analogues presented low agonistic activity when compared to AII in the different bioassays tested. The exceptions are cyclo(0-1a) [Asp(0), endo-(Lys(1a))]-AII (1) and [Asp(0), endo-(Lys(1a))]-AII (2), both of which showed activity similar to AII. Based on peptide 1 and the analogue cyclo(3-5)[Sar(1), Asp(3), Lys(5)]-AII characterized by Matsoukas et al., we analyzed the agonistic and antagonistic activities, respectively, through a new monocyclic peptide series synthesized by using the following combinations of residues as bridgehead elements for the lactam bond formation: D- or L-Asp combined with D- or L-Lys or L-Glu combined with L-Orn. Six analogues showed an approximately 20% increase in biological activity when compared with peptide (1) and were equipotent to AII. In contrast, six analogues presented antagonistic activity. These results suggest that the position of the lactam bridge is more important than the bridge length or chirality for recognition of and binding to the angiotensin II AT1-receptor.