Effect of Triphala on oxidative stress and on cell-mediated immune response against noise stress in rats

Stress is one of the basic factors in the etiology of number of diseases. The present study was aimed to investigate the effect of Triphala (Terminalia chebula, Terminalia belerica and Emblica officinalis) on noise-stress induced alterations in the antioxidant status and on the cell-mediated immune response in Wistar strain male albino rats. Noise-stress employed in this study was 100 dB for 4 h/d/15 days and Triphala was used at a dose of 1 g/kg/b.w/48 days. Eight different groups of rats namely, non-immunized: control, Triphala, noise-stress, Triphala with noise-stress, and corresponding immunized groups were used. Sheep red blood cells (5×10⁹ cells/ml) were used to immunize the animals. Biochemical indicators of oxidative stress namely lipid peroxidation, antioxidants superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), ascorbic acid in plasma and tissues (thymus and spleen) and SOD, GPx and corticosterone level in plasma were estimated. Cell-mediated immune response namely foot pad thickness (FPT) and leukocyte migration inhibition (LMI) test were performed only in immunized groups. Results showed that noise-stress significantly increased the lipid peroxidation and corticosterone level with concomitant depletion of antioxidants in plasma and tissues of both non-immunized and immunized rats. Noise-stress significantly suppressed the cell-mediated immune response by decreased FPT with an enhanced LMI test. The supplementation with Triphala prevents the noise-stress induced changes in the antioxidant as well as cell-mediated immune response in rats. This study concludes that Triphala restores the noise-stress induced changes may be due to its antioxidant properties.     

Light-regulated overexpression of an Arabidopsis phytochrome A gene in rice alters plant architecture and increases grain yield

The phytochromes are a family of red/far-red light absorbing photoreceptors that control plant developmental and metabolic processes in response to changes in the light environment. We report here the overexpression of Arabidopsis thaliana PHYTOCHROME A (PHYA) gene in a commercially important indica rice variety (Oryza sativa L. Pusa Basmati-1). The expression of the transgene was driven by the light-regulated and tissue-specific rice rbcS promoter. Several independent homozygous sixth generation (T₅) transgenic lines were characterized and shown to accumulate relatively high levels of PHYA protein in the light. Under both far-red and red light, PHYA-overexpressing lines showed inhibition of the coleoptile extension in comparison to non-transgenic seedlings. Furthermore, compared with non-transgenic rice plants, mature transgenic plants showed significant reduction in plant height, internode length and internode diameter (including differences in cell size and number), and produced an increased number of panicles per plant. Under greenhouse conditions, rice grain yield was 6–21% higher in three PHYA-overexpressing lines than in non-transgenic plants. These results demonstrate the potential of manipulating light signal-transduction pathways to minimize the problems of lodging in basmati/aromatic rice and to enhance grain productivity.     

Model of a putative pore: the pentameric alpha-helical bundle of SARS coronavirus E protein in lipid bilayers

The coronavirus responsible for the severe acute respiratory syndrome contains a small envelope protein, E, with putative involvement in host apoptosis and virus morphogenesis. To perform these functions, it has been suggested that protein E can form a membrane destabilizing transmembrane (TM) hairpin, or homooligomerize to form a TM pore. Indeed, in a recent study we reported that the alpha-helical putative transmembrane domain of E protein (ETM) forms several SDS-resistant TM interactions: a dimer, a trimer, and two pentameric forms. Further, these interactions were found to be evolutionarily conserved. Herein, we have studied multiple isotopically labeled ETM peptides reconstituted in model lipid bilayers, using the orientational parameters derived from infrared dichroic data. We show that the topology of ETM is consistent with a regular TM alpha-helix. Further, the orientational parameters obtained unequivocally correspond to a homopentameric model, by comparison with previous predictions. We have independently confirmed that the full polypeptide of E protein can also aggregate as pentamers after expression in Escherichia coli. This interaction must be stabilized, at least partially, at the TM domain. The model we report for this pentameric alpha-helical bundle may explain some of the permabilizing properties of protein E, and should be the basis of mutagenesis efforts in future functional studies.     

Induction of brain natriuretic peptide and monocyte chemotactic protein-1 gene expression by oxidized low-density lipoprotein: relevance to ischemic heart failure

Brain natriuretic peptide (BNP) and monocyte chemotactic protein-1 (MCP-1) are biomarkers of heart failure (HF). The aim of the present study was to determine the role of oxidized low-density lipoprotein (Ox-LDL) in the induction of these biomarkers and the signaling pathways involved in vitro. Incubation of HL-1 cardiomyocytes and human myocytes with Ox-LDL induced the expression of BNP and MCP-1 genes, while native LDL had no effect. When peroxides associated with Ox-LDL were reduced to hydroxides, the ability to induce BNP and MCP-1 gene expression was abolished. Furthermore, exposure of HL-1 cells to ischemic conditions alone had no effect on BNP gene expression, while ischemia followed by reperfusion resulted in increased expression of BNP gene. Inhibitors of ERK and JNK inhibited the induction of BNP. Signaling array results suggested that the induction of both MAPK and NF-κB pathways is involved in the induction of BNP by Ox-LDL. These results suggest that Ox-LDL or peroxidized lipids formed in oxidatively stressed myocytes during ischemia-reperfusion injury may play a role in the induction of BNP and MCP-1.     

NALP-3 inflammasome silencing attenuates ceramide-induced transepithelial permeability

The hallmark of acute lung injury (ALI) is the influx of proinflammatory cytokines into lung tissue and alveolar permeability that ultimately leads to pulmonary edema. However, the mechanisms involved in inflammatory cytokine production and alveolar permeability are unclear. Recent studies suggest that excessive production of ceramide has clinical relevance as a mediator of pulmonary edema and ALI. Our earlier studies indicate that the activation of inflammasome promotes the processing and secretion of proinflammatory cytokines and causes alveolar permeability in ALI. However, the role of ceramide in inflammasome activation and the underlying mechanism in relation to alveolar permeability is not known. We hypothesized that ceramide activates the inflammasome and causes inflammatory cytokine production and alveolar epithelial permeability. To test this hypothesis, we analyzed the lung ceramide levels during hyperoxic ALI in mice. The effect of ceramide on activation of inflammasome and production of inflammatory cytokine was assessed in primary mouse alveolar macrophages and THP-1 cells. Alveolar transepithelial permeability was determined in alveolar epithelial type-II cells (AT-II) and THP-1 co-cultures. Our results reveal that ceramide causes inflammasome activation, induction of caspase-1, IL-1β cleavage, and release of proinflammatory cytokines. In addition, ceramide further induces alveolar epithelial permeability. Short-hairpin RNA silencing of inflammasome components abrogated ceramide-induced secretion of proinflammatory cytokines in vitro. Inflammasome silencing abolishes ceramide-induced alveolar epithelial permeability in AT-II. Collectively, our results demonstrate for the first time that ceramide-induced secretion of proinflammatory cytokines and alveolar epithelial permeability occurs though inflammasome activation.     

Expression and purification of coronavirus envelope proteins using a modified β-barrel construct

Coronavirus envelope (E) proteins are short (～100 residues) polypeptides that contain at least one transmembrane (TM) domain and a cluster of 2-3 juxtamembrane cysteines. These proteins are involved in viral morphogenesis and tropism, and their absence leads in some cases to aberrant virions, or to viral attenuation. In common to other viroporins, coronavirus envelope proteins increase membrane permeability to ions. Although an NMR-based model for the TM domain of the E protein in the severe acute respiratory syndrome virus (SARS-CoV E) has been reported, structural data and biophysical studies of full length E proteins are not available because efficient expression and purification methods for these proteins are lacking. Herein we have used a novel fusion protein consisting of a modified β-barrel to purify both wild type and cysteine-less mutants of two representatives of coronavirus E proteins: the shortest (76 residues), from SARS-CoV E, and one of the longest (109 residues), from the infectious bronchitis virus (IBV E). The fusion construct was subsequently cleaved with cyanogen bromide and all polypeptides were obtained with high purity. This is an approach that can be used in other difficult hydrophobic peptides.     

Mir-206 Regulates Pulmonary Artery Smooth Muscle Cell Proliferation and Differentiation

Pulmonary Arterial Hypertension (PAH) is a progressive devastating disease characterized by excessive proliferation of the Pulmonary Arterial Smooth Muscle Cells (PASMCs). Studies suggest that PAH and cancers share an apoptosis-resistant state featuring excessive cell proliferation. MicroRNA-206 (miR-206) is known to regulate proliferation and is implicated in various types of cancers. However, the role of miR-206 in PAH has not been studied. In this study, it is hypothesized that miR-206 could play a role in the proliferation of PASMCs. In the present study, the expression patterns of miR-206 were investigated in normal and hypertensive mouse PASMCs. The effects of miR-206 in modulating cell proliferation, apoptosis and smooth muscle cell markers in human pulmonary artery smooth muscle cells (hPASMCs) were investigated in vitro. miR-206 expression in mouse PASMCs was correlated with an increase in right ventricular systolic pressure. Reduction of miR-206 levels in hPASMCs causes increased proliferation and reduced apoptosis and these effects were reversed by the overexpression of miR-206. miR-206 over expression also increased the levels of smooth muscle cell differentiation markers α-smooth muscle actin and calponin implicating its importance in the differentiation of SMCs. miR-206 overexpression down regulated Notch-3 expression, which is key a factor in PAH development. These results suggest that miR-206 is a potential regulator of proliferation, apoptosis and differentiation of PASMCs, and that it could be used as a novel treatment strategy in PAH.     

Phenotyping of tianma-stimulated differentiated rat neuronal b104 cells by quantitative proteomics

Gastrodia elata blume (tianma) is a traditional Chinese herb often used in the treatment of convulsions, headaches, and hypertension. Although interest in neuronal-related actions of tianma is increasing, minimal studies have been conducted to determine its specific effects on neuronal cells. This study was designed to examine the effects of tianma on the metabolism in differentiated neuroblastoma cells using the isobaric tag for relative and absolute quantitation (iTRAQ) technology. Stimulation of these cells with tianma caused changes in the expression of 38 proteins that were subsequently classified according to their physiological functions and association with neurodegenerative diseases. We identified six proteins with altered functional activities in neurodegenerative disease states that were modulated by tianma: triosephosphate isomerase (Tpi1), peptidyl-prolyl cis-trans isomerase A (Ppia), neural cell adhesion molecule 1 (Ncam1), ubiquitin carboxyl-terminal hydrolase isozyme L1 (Uchl1), septin-2 (Sept2) and heat shock protein 90 (Hsp90aa1). We postulate that tianma mediates its neuroprotective effects via upregulation of Ncam1, Hsp90aa1, Tpi1 and Ppia while downregulating Sept2 and Uchl1. These changes in protein expression aid in the restoration of the intracellular environment to a metabolically balanced state, promoting cell survival. Based on these observed data, we conclude that tianma has therapeutic potential, especially for neurodegenerative diseases.