Cancer genes hypermethylated in human embryonic stem cells

Developmental genes are silenced in embryonic stem cells by a bivalent histone-based chromatin mark. It has been proposed that this mark also confers a predisposition to aberrant DNA promoter hypermethylation of tumor suppressor genes (TSGs) in cancer. We report here that silencing of a significant proportion of these TSGs in human embryonic and adult stem cells is associated with promoter DNA hypermethylation. Our results indicate a role for DNA methylation in the control of gene expression in human stem cells and suggest that, for genes repressed by promoter hypermethylation in stem cells in vivo, the aberrant process in cancer could be understood as a defect in establishing an unmethylated promoter during differentiation, rather than as an anomalous process of de novo hypermethylation.     

Transgenic expression of a fungal endo-polygalacturonase increases plant resistance to pathogens and reduces auxin sensitivity

Polygalacturonases (PGs), enzymes that hydrolyze the homogalacturonan of the plant cell wall, are virulence factors of several phytopathogenic fungi and bacteria. On the other hand, PGs may activate defense responses by releasing oligogalacturonides (OGs) perceived by the plant cell as host-associated molecular patterns. Tobacco (Nicotiana tabacum) and Arabidopsis (Arabidopsis thaliana) plants expressing a fungal PG (PG plants) have a reduced content of homogalacturonan. Here, we show that PG plants are more resistant to microbial pathogens and have constitutively activated defense responses. Interestingly, either in tobacco PG or wild-type plants treated with OGs, resistance to fungal infection is suppressed by exogenous auxin, whereas sensitivity to auxin of PG plants is reduced in different bioassays. The altered plant defense responses and auxin sensitivity in PG plants may reflect an increased accumulation of OGs and subsequent antagonism of auxin action. Alternatively, it may be a consequence of perturbations of cellular physiology and elevated defense status as a result of altered cell wall architecture.     

Novel imidazoline compounds as partial or full agonists of D2-like dopamine receptors inspired by I2-imidazoline binding sites ligand 2-BFI

Based on the well known biological versatility of the imidazoline nucleus, we prepared the novel derivatives 3a–k inspired by 2-BFI scaffold to assess imidazoline molecules as D₂-like dopamine receptor ligands. Conservative chemical modifications of the lead structure, such as the introduction of an hydroxy group in the aromatic ring alone or associated with N-benzyl substitution, provided partial (3f) or nearly full (3e and 3h) agonists, all endowed with D₂-like potency comparable to that of dopamine.     

Thyroid hormone regulation of cell migration and oxidative metabolism in polymorphonuclear leukocytes: clinical evidence in thyroidectomized subjects on thyroxine replacement therapy

Migration and superoxide anion (O2-) generation were studied in polymorphonuclear leukocytes (PMNs) from 14 athyreotic patients, previously treated by total thyroidectomy and radioiodine therapy for differentiated thyroid carcinoma, and from age- and sex-matched euthyroid healthy controls. Patients were studied twice: in hypothyroidism (visit 1) and after TSH-suppressive L-T4 replacement therapy (visit 2). Random migration and N-formyl-Met-Leu-Phe (fMLP) 0.1-microM induced chemotaxis were similar in cells from patients at both visit 1 and visit 2 and from healthy controls. On the contrary, resting O2- generation in cells from patients was significantly lower than control values, both at visit 1 and 2. At visit 1, fMLP 0.1 muM-induced O2- generation was significantly lower than control values, while phorbol-myristate acetate (PMA) 100-ng/ml induced O2- generation was similar in cells from patients and from controls. At visit 2 both responses increased, resulting in fMLP-induced O2- generation superimposable to control values and PMA-induced O2- generation significantly higher with respect to both visit 1 and cells from controls. In vitro exposure of PMNs from healthy subjects to L-T4 did not affect O2- generation in resting cells, and significantly increased that induced by fMLP or PMA only at high, supra-physiological concentrations. Neither TSH nor T3 had significant effects at any of the concentrations tested. The present results document the existence of a correlation between thyroid status and oxidative metabolism of human PMNs, which is however unlikely to depend upon a direct action of thyroid hormones on these cells.     

Carborane-conjugated 2-quinolinecarboxamide ligands of the translocator protein for boron neutron capture therapy

Potential boron neutron capture therapy (BNCT) agents have been designed on the basis of the evidence about translocator protein (TSPO) overexpression on the outer mitochondrial membrane of tumor cells. The structure of the first TSPO ligand bearing a carborane cage (compound 2d) has been modified in order to find a suitable candidate for in vivo studies. The designed compounds were synthesized and evaluated for their potential interaction with TSPO and tumor cells. In vitro biological evaluation showed in the case of fluoromethyl derivative 4b a nanomolar TSPO affinity very similar to that of 2d, a significantly lower cytotoxicity, and a slightly superior performance as boron carrier toward breast cancer cells. Moreover, compound 4b could be used as a 1?F magnetic resonance imaging (MRI) agent as well as labeled with 11C or 1?F to obtain positron emission tomography (PET) radiotracers in order to apply the "see and treat" strategy in BNCT.     

Differential effects of quercetin and resveratrol on Band 3 tyrosine phosphorylation signalling of red blood cells

The protective effects of eating fruits and vegetables in the prevention of several degenerative pathologies have been attributed at least in part to the antioxidant and anti-inflammatory properties of polyphenols. In this study, we investigated the effects of two polyphenols, quercetin and resveratrol, on red blood cell Band 3 tyrosine phosphorylation signalling activated by peroxynitrite. Peroxynitrite is a physiological oxidant scavenged largely by the erythrocyte and formed by the reaction between nitrogen monoxide and superoxide anion. Quercetin and its structurally analogous (+)-catechin inhibited the peroxynitrite-dependent upregulation of Band 3 tyrosine phosphorylation. Quercetin was found to downregulate the activity of syk, which is upstream in the Band 3 tyrosine phosphorylation cascade, and partially prevented peroxynitrite-mediated phosphotyrosine phosphatase inhibition. Resveratrol and hydroxytyrosol, unexpectedly, amplified peroxynitrite-dependent upregulation of Band 3 tyrosine phosphorylation through the activation of lyn, a kinase of the src family. The present results clearly indicate that polyphenols may activate cell transduction pathways in different and sometimes opposite ways.     

Increased acetylcholinesterase activities in specimens of Sparus auratus exposed to sublethal copper concentrations

The present study looks at possible changes in the activity of acetylcholinesterase (AChE) in tissues (brain and white muscle) of the Mediterranean bony fish Sparus auratus after a 20 days exposure to sublethal concentrations (0.1 or 0.5 ppm) of copper in the marine water and on control untreated animals. The trials also included measurements of Cu concentration in the tissues to evaluate possible metal accumulation. Moreover, sedimentation analysis as well as V(max) and K(m) determination were carried out in tissue extracts of Cu-exposed or control animals. V(max) and K(m) were also determined with or without addition of Cu(2+) in the assay. No Cu accumulation occurred in brain and muscle after Cu exposure. AChE showed in both tissues a molecular polymorphism with putative globular (G) and asymmetric (A) forms. Cu exposition led to an increased specific activity and improved catalytic efficiency of AChE in brain and muscle, seemingly regarding G forms. The increase in catalytic efficiency also resulted from the in vitro assay with tissue extracts and Cu(2+) addition. The higher AChE activity and catalytic efficiency in both tissues after Cu exposition and without metal accumulation, suggests an increase of free Cu aliquot into the cells, likely due to mechanisms of metal homeostasis.     

Atrial natriuretic peptide and endothelin-3 target renal sodium-glucose cotransporter

Atrial natriuretic peptide (ANP) and endothelin (ET) are endogenous vasoactive factors that exert potent diuretic and natriuretic actions. We have previously shown that ANP and ET-3 act through an NO pathway to inhibit the sodium-glucose cotransporter (SGLT) in the intestine [Gonzalez Bosc LV, Elustondo PA, Ortiz MC, Vidal NA. Effect of atrial natriuretic peptide on sodium-glucose cotransport in the rat small intestine. Peptides 1997; 18: 1491-5; Gonzalez Bosc LV, Majowicz MP, Ortiz MC, Vidal NA. Effects of endothelin-3 on intestinal ion transport. Peptides 2001; 22: 2069-75.]. Here we address the role of ANP and ET-3 on SGLT activity in renal proximal tubules. In rat renal cortical brush border membranes (BBV), fluorescein isothiocianate (FITC) labeling revealed a specific 72-kD peptide that exhibits increased FITC labeling in the presence of Na+ and D-glucose. Using alpha-14C-methylglucose active uptake, rat BBV were shown to possess SGLT activity with an affinity constant (K(0.5) approximately 2.4 mM) that is consistent with the expression of the low-affinity, high-capacity SGLT2 isoform. SGLT2 activity in these preparations is dramatically inhibited by ANP and ET-3. This inhibition is independent of changes in membrane lipids and is mimicked by the cGMP analogue, 8-Br-cGMP, suggesting the involvement of cGMP/PKG pathways. These results are the first demonstration that both ANP and ET-3 inhibit rat cortical renal SGLT2 activity, and suggest a novel mechanism by which these vasoactive substances modulate hydro-saline balance at the proximal tubular nephron level.