Insight into the apoptosis-inducing action of alpha-bisabolol towards malignant tumor cells: involvement of lipid rafts and Bid.

In a precedent report we showed that alpha-bisabolol, a sesquiterpene present widely in the plant kingdom, exerts a rapid and efficient apoptosis-inducing action selectively towards human and murine malignant glioblastoma cell lines through mitochondrial damage. The present study extends these data demonstrating the apoptosis-inducing action of alpha-bisabolol towards highly malignant human pancreatic carcinoma cell lines without affecting human fibroblast viability. The present study further shows the preferential incorporation of alpha-bisabolol to transformed cells through lipid rafts on plasma membranes and, thereafter, direct interaction between alpha-bisabolol and Bid protein, one of pro-apoptotic Bcl-2 family proteins, analyzed either by Surface Plasmon Resonance method or by intrinsic fluorescence measurement. Notions that lipid rafts are rich in plasma membranes of transformed cells and that Bid, richly present in lipid rafts, is deeply involved in lipid transport make highly credible the hypothesis that the molecular mechanism of alpha-bisabolol action may include its capacity to interact with Bid protein.

Regulation of serine racemase activity by D-serine and nitric oxide in human glioblastoma cells.

In the human glioblastoma cell line U87, the activity of serine racemase (SR), catalyzing the isomerisation of serine, was inversely regulated by D-serine and nitric oxide (NO), a neuromodulator and a neurotransmitter, respectively. SR activity was dose-dependently enhanced up to five times in cells treated with 10 mM D-serine, whereas it was inhibited by NO. Furthermore, D-serine was found to induce the denitrosylation of SR purified from mouse brain. These results suggest that serine racemase activity in astrocyte is regulated inversely by d-serine and NO. SR should be inhibited through nitrosylation by NO and activated through denitrosylation elicited by D-serine.

Mutual regulation between serine and nitric oxide metabolism in human glioblastoma cells.

D-Serine indirectly caused dose- and time-dependent inhibition of neuronal nitric oxide synthase (nNOS) without affecting endothelial nitric oxide synthase (eNOS) in human glioblastoma cell line U87. Activity of D-amino acid oxidase (DAAO), catalyzing the oxidative deamination of d-amino acid, was enhanced by NO in a dose-dependent manner. Recently, we have reported that serine racemase (SR) is inhibited by NO and activated by D-serine through nitrosylation and denitrosylation, respectively [K. Shoji, S. Mariotto, A.R. Ciampa, H. Suzuki, Regulation of serine racemase activity by D-serine and nitric oxide in human glioblastoma cells, Neurosci. Lett., in press]. Thus, the metabolism of both d-serine and NO in U87 cells is functionally correlated in a complex manner. Suppression of NO production by d-serine in U87 cells contrasts its known action in enhancing nNOS in neurons.

Corrosion by bacteria of concrete in sewerage systems and inhibitory effects of formates on their growth.

Not only sulfur-oxidizing bacteria but also an acidophilic iron-oxidizing bacterium (or bacteria) were found in the corroded concrete from several sewerage systems in Japan. The surface pH of concrete test piece exposed to an atmosphere containing hydrogen sulfide of the concentrations more than 600 ppm in the systems was usually below 2 after a month. This was attributable to ability of the sulfur-oxidizing bacteria to grow in the thin water layer which contained hydrogen sulfide and covered the piece even when the surface pH of concrete was 12-13. When the sulfuroxidizing bacteria grew in the surface of concrete and produced sulfuric acid, the pH of the inner parts of concrete was lowered where the bacteria were hardly found. Probably, sulfuric acid formed by the bacteria in the surface parts penetrated into the inner parts. The different species of sulfur-oxidizing bacteria were found in different sewerage systems. The growth of the sulfur-oxidizing and acidophilic iron-oxidizing bacteria was completely inhibited by formates, especially by calcium formate of concentrations more than 50 mM. Calcium formate can protect concrete in sewerage systems from bacterial corrosion.

Direct interaction of natural and synthetic catechins with signal transducer activator of transcription 1 affects both its phosphorylation and activity.

Our previous studies showed that (-)-epigallocatechin-3-gallate (EGCG) inhibits signal transducer activator of transcription 1 (STAT1) activation. Since EGCG may be a promising lead compound for new anti-STAT1 drug design, 15 synthetic catechins, characterized by the (-)-gallocatechin-3-gallate stereochemistry, were studied in the human mammary MDA-MB-231 cell line to identify the minimal structural features that preserve the anti-STAT1 activity. We demonstrate that the presence of three hydroxyl groups of B ring and one hydroxyl group in D ring is essential to preserve their inhibitory action. Moreover, a possible molecular target of these compounds in the STAT1 pathway was investigated. Our results demonstrate a direct interaction between STAT1 protein and catechins displaying anti-STAT1 activity. In particular, surface plasmon resonance (SPR) analysis and molecular modeling indicate the presence of two putative binding sites (a and b) with different affinity. Based on docking data, site-directed mutagenesis was performed, and interaction of the most active catechins with STAT1 was studied with SPR to test whether Gln518 on site a and His568 on site b could be important for the catechin-STAT1 interaction. Data indicate that site b has higher affinity for catechins than site a as the highest affinity constant disappears in the H568A-STAT1 mutant. Furthermore, Janus kinase 2 (JAK2) kinase assay data suggest that the contemporary presence in vitro of STAT1 and catechins inhibits JAK2-elicited STAT1 phosphorylation. The very tight catechin-STAT1 interaction prevents STAT1 phosphorylation and represents a novel, specific and efficient molecular mechanism for the inhibition of STAT1 activation.

Two naturally occurring terpenes, dehydrocostuslactone and costunolide, decrease intracellular GSH content and inhibit STAT3 activation.

The main purpose of the present study is to envisage the molecular mechanism of inhibitory action of dehydrocostuslactone (DCE) and costunolide (CS), two naturally occurring sesquiterpene lactones, towards the activation of signal transducer and activator of transcription 3 (STAT3). We report that, in human THP-1 cell line, they inhibit IL-6-elicited tyrosine phosphorylation of STAT3 and its DNA binding activity with EC(50) of 10 µM with concomitant down-regulation of the phosphorylation of the tyrosine Janus kinases JAK1, JAK2 and Tyk2. Furthermore, these compounds that contain an α-ß-unsaturated carbonyl moiety and function as potent Michael reaction acceptor, induce a rapid drop in intracellular glutathione (GSH) concentration by direct interaction with it, thereby triggering S-glutathionylation of STAT3. Dehydrocostunolide (HCS), the reduced form of CS lacking only the α-ß-unsaturated carbonyl group, fails to exert any inhibitory action. Finally, the glutathione ethylene ester (GEE), the cell permeable GSH form, reverts the inhibitory action of DCE and CS on STAT3 tyrosine phosphorylation. We conclude that these two sesquiterpene lactones are able to induce redox-dependent post-translational modification of cysteine residues of STAT3 protein in order to regulate its function.

Dual cross-talk between nitric oxide and D-serine in astrocytes and neurons in the brain.

The present review describes the role of the putative cross-talk between two neurotransmitters, nitric oxide (NO) and D-serine, in the brain. Under physiological conditions NO homeostasis guarantees the correct function of NO in a number of events in the brain such as neurotransmission and vascular tone regulation. D-serine, produced in astrocytes, acts synergistically with glutamate at NMDA receptors on postsynaptic neurons. Neuronal and endothelial NO synthase (nNOS and eNOS) in astrocytes cross-talk with serine racemase (SR) and D-amino acid oxydase (DAAO), catalyzing the synthesis and degradation of D-serine, respectively. SR is inhibited by NO which activates DAAO. D-serine inhibits nNOS but not eNOS and activates SR. Astrocytes and neurons also cross-talk through NO/D-serine system. D-serine released from astrocytes induces a rapid increase in NO contents in postsynaptic neurons. Overall, D-serine production in astrocytes is negatively regulated by NO. Under inflammatory conditions, pro-inflammatory cytokines or Abeta induce, first, a drop in NO contents and an increase in the amounts of D-serine in astrocytes. Together with enhanced glutamate release from presynaptic neurons, D-serine induces an increase in Ca(2+) up-take into presynaptic neurons. In astrocytes an initial drop in NO contents triggers NF-kappaB activation followed by inducible NOS (iNOS) expression. iNOS-derived massive amounts of NO may potentially be toxic. Under schizophrenic conditions, D-serine production is down-regulated. Together with reduced glutamate release, this situation leads to the decreased NO production in postsynaptic neurons. In astrocytes induction of iNOS expression becomes predominant. Initial drop in nNOS-derived NO is potentially toxic in this scenario.

Targeting STAT1 by myricetin and delphinidin provides efficient protection of the heart from ischemia/reperfusion-induced injury.

Flavonoids exhibit a variety of beneficial effects in cardiovascular diseases. Although their therapeutic properties have been attributed mainly to their antioxidant action, they have additional protective mechanisms such as inhibition of signal transducer and activator of transcription 1 (STAT1) activation. Here, we have investigated the cardioprotective mechanisms of strong antioxidant flavonoids such as quercetin, myricetin and delphinidin. Although all of them protect the heart from ischemia/reperfusion-injury, myricetin and delphinidin exert a more pronounced protective action than quercetin by their capacity to inhibit STAT1 activation. Biochemical and computer modeling analysis indicated the direct interaction between STAT1 and flavonoids with anti-STAT1 activity.

Protective effect of epigallocatechin-3-gallate on ischemia/reperfusion-induced injuries in the heart: STAT1 silencing flavonoid.

The beneficial effect of naturally occurring flavonoids in health is believed to be due to their strong antioxidant activity. However, recent laboratory evidence indicates the involvement of a more specific action. Here, we present evidence that, among a number of catechins present in green tea extract, only epigallocatechin-3-gallate (EGCG) exerts a strong inhibitory action on interferon-gamma-elicited activation of signal transducer and activator of transcription 1 (STAT1). Protective action of EGCG in ischemia/reperfusion injury in the heart and the molecular mechanism of action, which has nothing to do with its anti-oxidant capacity are described.