3-Amino derivative of beta-cyclodextrin: thermodynamics of copper(II) complexes and exploitation of its enantioselectivity in the separation of amino acid racemates by ligand exchange capillary electrophoresis

The systems that the 3-amino derivative of beta-cyclodextrin (CD3NH2) forms with the proton, the copper(II) ion and each of the enantiomers of certain amino acids (alanine, phenylalanine, tyrosine and tryptophan) were investigated. The enantioselectivity shown by the potentiometric measurements carried out on the phenylalanine ternary systems was exploited in capillary electrophoresis by ligand exchange capillary electrophoresis (LECE) to obtain the separation of phenylalanine racemate. The tyrosine racemate was also separated by LECE. The comparison between thermodynamic and capillary electrophoresis (CE) results is discussed, in order to get a better insight into the separation mechanism.     

PI3 kinase is important for Ras,MEK and Erk activation of Epo-stimulated human erythroid progenitors

Background  

Erythropoietin is a multifunctional cytokine which regulates the number of erythrocytes circulating in mammalian blood. This is crucial in order to maintain an appropriate oxygen supply throughout the body. Stimulation of primary human erythroid progenitors (PEPs) with erythropoietin (Epo) leads to the activation of the mitogenic kinases (MEKs and Erks). How this is accomplished mechanistically remained unclear.     

Solution chemistry and cytotoxic properties of novel organogold(III) compounds

The solution behaviour of some novel organogold(III) compounds was investigated, and their cytotoxic properties evaluated against a few human tumour cell lines (A2780/S, A2780/R, MCF7, HT29 and A549). Specifically, the following compounds were considered: [Au(bipy(dmb)-H)(2,6-xylidine-H)][PF(6)] (AuXyl) and [Au(bipy(dmb)-H)(p-toluidine-H)][PF(6)] (AuTol) (in which bipy(dmb)=6-(1,1-dimethylbenzyl)-2,2'-bipyridine), [Au(py(dmb)-H)(AcO)(2)] (AuPyAcO) (in which py(dmb)=2-(1,1-dimethylbenzyl)-pyridine) and [Au(pz(Ph)-H)Cl(3)]K (AuPzCl) (in which pz(Ph)=1-phenylpyrazole). The solution chemistry of these compounds, under physiological-like conditions, was investigated through UV-vis absorption and (1)H NMR spectroscopies. Significant cytotoxic effects in vitro were observed in selected cases.     

Protection by herpes simplex virus glycoprotein D against Fas-mediated apoptosis: role of nuclear factor kappaB

Signals involved in protection against apoptosis by herpes simplex virus 1 (HSV-1) were investigated. Using U937 monocytoid cells as an experimental model, we have demonstrated that HSV-1 rendered these cells resistant to Fas-induced apoptosis promptly after infection. UV-inactivated virus as well as the envelope glycoprotein D (gD) of HSV-1, by itself, exerted a protective effect on Fas-induced apoptosis. NF-kappaB was activated by gD, and protection against Fas-mediated apoptosis by gD was abolished in cells stably transfected with a dominant negative mutant I-kappaBalpha, indicating that NF-kappaB activation plays a role in the antiapoptotic activity of gD in our experimental model. Moreover, NF-kappaB-dependent protection against Fas-mediated apoptosis was associated with decreased levels of caspase-8 activity and with the up-regulation of intracellular antiapoptotic proteins.     

Essential role of A-kinase anchor protein 121 for cAMP signaling to mitochondria

A-Kinase anchor proteins (AKAPs) immobilize and concentrate protein kinase A (PKA) isoforms at specific subcellular compartments. Intracellular targeting of PKA holoenzyme elicits rapid and efficient phosphorylation of target proteins, thereby increasing sensitivity of downstream effectors to cAMP action. AKAP121 targets PKA to the cytoplasmic surface of mitochondria. Here we show that conditional expression of AKAP121 in PC12 cells selectively enhances cAMP.PKA signaling to mitochondria. AKAP121 induction stimulates PKA-dependent phosphorylation of the proapoptotic protein BAD at Ser(155), inhibits release of cytochrome c from mitochondria, and protects cells from apoptosis. An AKAP121 derivative mutant that localizes on mitochondria but does not bind PKA down-regulates PKA signaling to the mitochondria and promotes apoptosis. These findings indicate that PKA anchored by AKAP121 transduces cAMP signals to the mitochondria, and it may play an important role in mitochondrial physiology.     

Single-sample preparation for simultaneous cellular redox and energy state determination

A simple and reliable method for the preparation of biological samples for the evaluation of biochemical parameters representative of the redox and energy states, such as glutathione (GSH), oxidized glutathione (GSSG), oxidized nicotinamide adenine dinucleotide (NAD+), reduced nicotinamide adenine dinucleotide (NADH), oxidized nicotinamide adenine dinucleotide phosphate (NADP+), reduced nicotinamide adenine dinucleotide phosphate (NADPH), coenzyme A (CoASH), oxidized CoASH, ascorbate, malondialdehyde, oxypurines, nucleosides, and energy metabolites, is presented. Fast deproteinization under nonoxidizing conditions is obtained by tissue homogenization in ice-cold, nitrogen-saturated CH3CN + 10 mM KH2PO4 (3:1; v:v), pH 7.40. After sample centrifugation to pellet precipitated proteins, organic solvent removal is performed on clear supernatants by three washings with large volumes of high-performance liquid chromatography (HPLC)-grade chloroform. The remaining aqueous phase, free of solvent and any lipid-soluble substances that may interfere with the further metabolite analysis, is used for the simultaneous ion-pairing HPLC determination of 39 compounds by means of a Kromasil C-18, 250 x 4.6-mm, 5-microm-particle-size column with tetrabutylammonium hydroxide as the pairing reagent. Results obtained by using the present method to prepare different rat tissue extracts demonstrate that it is possible to perform a single tissue preparation only for monitoring, in the same sample, compounds representative of the redox state (through the direct determination of GSH, GSSG, NAD+, NADH, NADP+, NADPH, CoASH, and oxidized CoASH) and of the cell energy state (by the analysis of oxypurines, nucleosides, and energy metabolites). Applicability of this sample processing procedure to quantify variations of the aforementioned compounds under pathological conditions was effected in rats subjected to moderate closed-head trauma.     

3-hydroxykynurenine as a substrate/activator for mushroom tyrosinase

3-Hydroxykynurenine is a tryptophan metabolite with an o-aminophenol structure. It is both a tyrosinase activator and a substrate, reducing the lag phase, stimulating the monophenolase activity, and being oxidized to xanthommatin. In the early stage of monophenol hydroxylation, catechol accumulation takes place, whereas 3-hydroxykynurenine is substantially unchanged and no significant amounts of the o-quinone are produced. These results suggest an activating action of 3-hydroxykynurenine toward o-hydroxylation of monophenols. 3-Hydroxykynurenine could therefore well act as a physiological device to control phenolics metabolism to catechols and quinonoids.