Phenolic compounds isolated from Dioscorea zingiberensis protect against pancreatic acinar cells necrosis induced by sodium taurocholate

One new bibenzyl (1) and one new phenanthrene (2), together with two known bibenzyls (3–4) and four known diarylheptanoids (5–8) were isolated from the rhizomes of Dioscorea zingiberensis. The structures of 1–2 were elucidated by spectroscopic methods including 1D and 2D NMR. Phenols 1–8 were evaluated for their anti-pancreatitic activities on sodium taurocholate (NaT)-induced pancreatic acinars necrosis. Notably, 0.5 mM of compound 6 exhibited comparable inhibitory effect with 5 mM of caffeine. Furthermore, compound 6 prevented the ATP depletion and excessive ROS production which could be also involved in mitochondria-mediated injuries in acute pancreatitis. As a result, compound 6 has been demonstrated to be a potential candidate for mediating mitochondrial dysfunction to prevent pancreatic necrosis. This study is also the first report on the isolation of bibenzyls and diarylheptanoids from this plant.     

In vivo protein tyrosine nitration in S. cerevisiae: Identification of tyrosine-nitrated proteins in mitochondria

Protein tyrosine nitration (PTN) is a selective post-translational modification often associated with pathophysiological conditions. Although yeast cells lack of mammalian nitric oxide synthase (NOS) orthologues, still it has been shown that they are capable of producing nitric oxide (NO). Our studies showed that NO or reactive nitrogen species (RNS) produced in flavohemoglobin mutant (Δyhb1) strain along with the wild type strain (Y190) of Saccharomyces cerevisiae can be visualized using specific probe 4,5-diaminofluorescein diacetate (DAF-2DA). Δyhb1 strain of S. cerevisiae showed bright fluorescence under confocal microscope that proves NO or RNS accumulation is more in absence of flavohemoglobin. We further investigated PTN profile of both cytosol and mitochondria of Y190 and Δyhb1 cells of S. cerevisiae using two-dimensional (2D) gel electrophoresis followed by western blot analysis. Surprisingly, we observed many immunopositive spots both in cytosol and in mitochondria from Y190 and Δyhb1 using monoclonal anti-3-nitrotyrosine antibody indicating a basal level of NO or nitrite or peroxynitrite is produced in yeast system. To identify proteins nitrated in vivo we analyzed mitochondrial proteins from Y190 strains of S. cerevisiae. Among the eight identified proteins, two target mitochondrial proteins are aconitase and isocitrate dehydrogenase that are involved directly in the citric acid cycle. This investigation is the first comprehensive study to identify mitochondrial proteins nitrated in vivo.     

Regulatory volume decrease in cardiomyocytes is modulated by calcium influx and reactive oxygen species

We investigated the role of Ca²⁺ in generating reactive oxygen species (ROS) induced by hyposmotic stress (Hypo) and its relationship to regulatory volume decrease (RVD) in cardiomyocytes. Hypo-induced increases in cytoplasmic and mitochondrial Ca²⁺. Nifedipine (Nife) inhibited both Hypo-induced Ca²⁺ and ROS increases. Overexpression of catalase (CAT) induced RVD and a decrease in Hypo-induced blebs. Nife prevented CAT-dependent RVD activation. These results show a dual role of Hypo-induced Ca²⁺ influx in the control of cardiomyocyte viability. Hypo-induced an intracellular Ca²⁺ increase which activated RVD and inhibited necrotic blebbing thus favoring cell survival, while simultaneously increasing ROS generation, which in turn inhibited RVD and induced necrosis.     

Surface disinfection properties of the combination of an antimicrobial peptide,ranalexin, with an endopeptidase,lysostaphin, against methicillin‐resistant Staphylococcus aureus (MRSA)

Aims: To characterize the antibacterial synergy of the antimicrobial peptide, ranalexin, used in combination with the anti‐staphylococcal endopeptidase, lysostaphin, against methicillin‐resistant Staphylococcus aureus (MRSA), and to assess the combination’s potential as a topical disinfectant or decolonizing agent for MRSA. MRSA causes potentially lethal infections, and pre‐operative patients colonized with MRSA are often treated with chlorhexidine digluconate and mupirocin cream to eradicate carriage. However, chlorhexidine is unsuitable for some patients, and mupirocin resistance is increasingly encountered, indicating new agents are required. Methods and Results: Using an ex vivo assay, ranalexin and lysostaphin tested in combination reduced viable MRSA on human skin to a greater extent than either compound individually. The combination killed bacteria within 5 min and remained effective and synergistic even in high salt and low pH conditions. Conclusions: The combination is active against MRSA on human skin and under conditions that may be encountered in sweat. Significance and Impact of the Study: Although the exact mechanism of activity remains unresolved, considering its specific spectrum of activity, fast killing kinetics and low likelihood of resistance arising, the combination of ranalexin with lysostaphin warrants consideration as a new agent to eradicate nasal and skin carriage of Staph. aureus, including MRSA.     

Functional characterization of a synthetic abscisic acid analog with anti-inflammatory activity on human granulocytes and monocytes

The phytohormone abscisic acid (ABA), in addition to regulating several important physiological functions in plants, is also produced and released by human granulocytes and monocytes where it stimulates cell activities involved in the innate immune response.Here we describe the properties of an ABA synthetic analog that competes with the hormone for binding to human granulocyte membranes and to purified recombinant LANCL2 (the human ABA receptor) and inhibits several ABA-triggered inflammatory functions of granulocytes and monocytes in vitro: chemotaxis, phagocytosis, reactive oxygen species production and release of prostaglandin E₂ (PGE₂) by human granulocytes, release of PGE₂ and of monocyte chemoattractant protein-1 by human monocytes. This observation provides a proof of principle that ABA antagonists may represent a new class of anti-inflammatory agents.     

Modulation of Bax mitochondrial insertion and induced cell death in yeast by mammalian protein kinase Cα

Protein kinase Cα (PKCα) is a classical PKC isoform whose involvement in cell death is not completely understood. Bax, a major member of the Bcl-2 family, is required for apoptotic cell death and regulation of Bax translocation and insertion into the outer mitochondrial membrane is crucial for regulation of the apoptotic process. Here we show that PKCα increases the translocation and insertion of Bax c-myc (an active form of Bax) into the outer membrane of yeast mitochondria. This is associated with an increase in cytochrome c (cyt c) release, reactive oxygen species production (ROS), mitochondrial network fragmentation and cell death. This cell death process is regulated, since it correlates with an increase in autophagy but not with plasma membrane permeabilization. The observed increase in Bax c-myc translocation and insertion by PKCα is not due to Bax c-myc phosphorylation, and the higher cell death observed is independent of the PKCα kinase activity. PKCα may therefore have functions other than its kinase activity that aid in Bax c-myc translocation and insertion into mitochondria. Together, these results give a mechanistic insight on apoptosis regulation by PKCα through regulation of Bax insertion into mitochondria.     

Hydogen peroxide-dependent photocytotoxicity by phloxine B,a xanthene-type food colorant

Background

Phloxine B (PhB; 2′,4′,5′,7′-tetrabromo-4,5,6,7-tetrachloro-fluorescein), an artificial xanthene colorant, has been used as a red coloring agent in drugs and cosmetics as well as foods in some countries. However, little effort has been devoted to the study of this colorant as a potentially useful medicinal agent.

Methods

We investigated the daily light-induced photocytotoxicity of PhB in two human leukemia cells, HL-60 and Jurkat, and its underlying mechanisms by in vitro experiments using antioxidants.

Reuslts and conclusions

PhB inhibited cell proliferation more preferentially to HL-60 cells than to Jurkat cells. Co-treatment of catalase completely blocked the photocytotoxicity by PhB in HL-60 cells, whereas the effect of histidine was only partial, suggesting that hydrogen peroxide (H₂O₂), rather than singlet oxygen, might be a prerequisite for the PhB-induced HL-60 cell death. Actually, PhB produced a significant amount of H₂O₂ in the media as well as in the cells in concentration- and light-dependent manners. Furthermore, methionine, a hypochlorous acid (HOCl) scavenger, also significantly attenuated the cytotoxicity in HL-60 cells, but not in Jurkat cells, indicating the involvement of myeloperoxidase (MPO)-dependent hypohalous acid formation during the photocytotoxicity. In vitro experiments revealed that halogenated tyrosine was generated from the reaction of bovine serum albumin with PhB and HL-60 cell lysate. The present findings suggested that PhB induced a differential photodynamic action in the MPO-containing leukemia cells through an H₂O₂-dependent mechanism.

General significance

Our findings provide new insights into the molecular mechanisms underlying the PhB-induced apoptosis and also evaluated PhB as a promising PDT agent.     

The emerging roles of nitric oxide (NO) in plant mitochondria

In recent years nitric oxide (NO) has been recognized as an important signal molecule in plants. Both, reductive and oxidative pathways and different subcellular compartments appear involved in NO production. The reductive pathway uses nitrite as substrate, which is exclusively generated by cytosolic nitrate reductase (NR) and can be converted to NO by the same enzyme. The mitochondrial electron transport chain is another site for nitrite to NO reduction, operating specifically when the normal electron acceptor, O₂, is low or absent. Under these conditions, the mitochondrial NO production contributes to hypoxic survival by maintaining a minimal ATP formation. In contrast, excessive NO production and concomitant nitrosative stress may be prevented by the operation of NO-scavenging mechanisms in mitochondria and cytosol. During pathogen attacks, mitochondrial NO serves as a nitrosylating agent promoting cell death; whereas in symbiotic interactions as in root nodules, the turnover of mitochondrial NO helps in improving the energy status similarly as under hypoxia/anoxia. The contribution of NO turnover during pathogen defense, symbiosis and hypoxic stress is discussed in detail.     

Methods of nitric oxide detection in plants: A commentary

Over the last decade nitric oxide (NO) has been shown to influence a range of processes in plants. However, when, where and even if NO production occurs is controversial in several physiological scenarios in plants. This arises from a series of causes: (a) doubts have arisen over the specificity of widely used 4,5-diaminofluorescein diacetate (DAF-2DA)/4-amino-5-methylamino-2,7-difluorofluorescein (DAF-FM) dyes for NO, (b) no plant nitric oxide synthase (NOS) has been cloned, so that the validity of using mammalian NOS inhibitors to demonstrate that NO is being measured is debatable, (c) the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-l-oxyl-3-oxide (cPTIO) needs to be used with caution, and (d) some discrepancies between assays for in planta measurements and another based on sampling NO from the gas phase have been reported. This review will outline some commonly used methods to determine NO, attempt to reconcile differing results obtained by different laboratories and suggest appropriate approaches to unequivocally demonstrate the production of NO.     

Roles of intracellular hydrogen peroxide accumulation in abscisic acid signaling in Arabidopsis guard cells

Reactive oxygen species (ROS), including hydrogen peroxide (H₂O₂), are among the important second messengers in abscisic acid (ABA) signaling in guard cells. In this study, to investigate specific roles of H₂O₂ in ABA signaling in guard cells, we examined the effects of mutations in the guard cell-expressed catalase (CAT) genes, CAT1 and CAT3, and of the CAT inhibitor 3-aminotriazole (AT) on stomatal movement. The cat3 and cat1 cat3 mutations significantly reduced CAT activities, leading to higher basal level of H₂O₂ in guard cells, when assessed by 2′,7′-dichlorodihydrofluorescein, whereas they did not affect stomatal aperture size under non-stressed condition. In addition, AT-treatment at concentrations that abolish CAT activities, showed trivial affect on stomatal aperture size, while basal H₂O₂ level increased extensively. In contrast, cat mutations and AT-treatment potentiated ABA-induced stomatal closure. Inducible ROS production triggered by ABA was observed in these mutants and wild type as well as in AT-treated guard cells. These results suggest that ABA-inducible cytosolic H₂O₂ elevation functions in ABA-induced stomatal closure, while constitutive increase of H₂O₂ do not cause stomatal closure.