2,3,7,8-Tetrachlorodibenzo-p-dioxin impairs iron homeostasis by modulating iron-related proteins expression and increasing the labile iron pool in mammalian cells

Cellular iron metabolism is essentially controlled by the binding of cytosolic iron regulatory proteins (IRP1 or IRP2) to iron-responsive elements (IREs) located on mRNAs coding for proteins involved in iron acquisition, utilization and storage. The 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is one of the most potent toxins of current interest that occurs as poisonous chemical in the environment. TCDD exposure has been reported to induce a broad spectrum of toxic and biological responses, including significant changes in gene expression for heme and iron metabolism associated with liver injury. Here, we have investigated the molecular effects of TCDD on the iron metabolism providing the first evidence that administration of the toxin TCDD to mammalian cells affects the maintenance of iron homeostasis. We found that exposure of Madin-Darby Bovine Kidney cell to TCDD caused a divergent modulation of IRP1 and IRP2 RNA-binding capacity. Interestingly, we observed a concomitant IRP1 down-regulation and IRP2 up-regulation thus determining a marked enhancement of transferrin receptor 1 (TfR-1) expression and a biphasic response in ferritin content. The changed ferritin content coupled to TfR-1 induction after TCDD exposure impairs the cellular iron homeostasis, ultimately leading to significant changes in the labile iron pool (LIP) extent. Since important iron requirement changes occur during the regulation of cell growth, it is not surprising that the dioxin-dependent iron metabolism dysregulation herein described may be linked to cell-fate decision, supporting the hypothesis of a central connection among exposure to dioxins and the regulation of critical cellular processes. This article is part of a Special Issue entitled: 11th European Symposium on Calcium.     

Palytoxin Induces Functional Changes of Anion Transport in Red Blood Cells: Metabolic Impact

Palytoxin (PTX) is classified as one of the most powerful marine biotoxins (of high molecular weight and no protein origin) because it is able to interact strongly with important cellular structures influencing their function in different biological processes. This study of the effects of PTX on red blood cells (RBC) extends the knowledge about its toxicity, which concerns not only the well-known action on Na(+)/K(+)-ATPase but also band 3 protein (B3 or AE1), the role of which is essential for anion transport and for the structure, function, and metabolic integrity of the erythrocyte. The effects of PTX on RBC can be summarized as follows: it alters the anionic flux and seriously compromises not only CO(2) transport but also the metabolic modulation centered on the oxy-deoxy cycle of hemoglobin; it stabilizes the plasma membrane by preventing lipid peroxidation; and its effect does not lead to activation of caspases 3 and 8. From what is reported in steps 2 and 3, and on the basis of the results obtained on hemolysis, methemoglobin levels, and phosphatase activity, an increase of the reducing power of the erythrocytes (RBC) in the presence of PTX clearly emerges. The results have enabled us to outline some metabolic adaptations induced in the RBC by PTX.     

Ion-exchange voltammetry and electrocatalytic sensing capabilities of cytochrome c at polyestersulfonated ionomer coated glassy carbon electrodes

The incorporation and electrochemical behaviour of cytochrome c (Cyt C) at glassy carbon electrodes modified with the polyestersulfonated ionomer Eastman AQ 55 are examined. The presence of the polyelectrolytic coating allows the preconcentration of the protein within the polymer and the observation of the direct electrochemistry of Cyt C at the modified electrode without addition of promoters or mediators in the solutions. The dependence of voltammetric signals on typical parameters such as solution pH and nature or concentration of the supporting electrolyte supports the ion-exchange nature of the incorporation process. The relevant role of the permselectivity of the polymeric modifier is highlighted also by the study of electrocatalytic processes which take place at the modified electrode loaded with Cyt C. No electrocatalytic effect is observed when the electrogenerated dication (ferricenylmethyl)trimethylammonium is present as possible oxidant. On the contrary, electrocatalytic current enhancements are observed for anionic substrates such as Fe(CN)(6)(3-) (oxidant) and ascorbate (reductant). Catalytic currents increase with the substrate concentration, with higher sensitivity for Fe(CN)(6)(3-). Due to ionic repulsion, the reaction with anions occurs at the polymer-solution interface. In the case of chemically unstable substrates, such as superoxide anion, ionic repulsion slow down the approach rate of the substrate so that spontaneous decomposition can prevail over the reaction with Cyt C incorporated in the coating.     

Gait Initiation in Children with Rett Syndrome

Rett syndrome is an X-linked neurodevelopmental condition mainly characterized by loss of spoken language and a regression of purposeful hand use, with the development of distinctive hand stereotypies, and gait abnormalities. Gait initiation is the transition from quiet stance to steady-state condition of walking. The associated motor program seems to be centrally mediated and includes preparatory adjustments prior to any apparent voluntary movement of the lower limbs. Anticipatory postural adjustments contribute to postural stability and to create the propulsive forces necessary to reach steady-state gait at a predefined velocity and may be indicative of the effectiveness of the feedforward control of gait. In this study, we examined anticipatory postural adjustments associated with gait initiation in eleven girls with Rett syndrome and ten healthy subjects. Muscle activity (tibialis anterior and soleus muscles), ground reaction forces and body kinematic were recorded. Children with Rett syndrome showed a distinctive impairment in temporal organization of all phases of the anticipatory postural adjustments. The lack of appropriate temporal scaling resulted in a diminished impulse to move forward, documented by an impairment in several parameters describing the efficiency of gait start: length and velocity of the first step, magnitude and orientation of centre of pressure-centre of mass vector at the instant of (swing-)toe off. These findings were related to an abnormal muscular activation pattern mainly characterized by a disruption of the synergistic activity of antagonistic pairs of postural muscles. This study showed that girls with Rett syndrome lack accurate tuning of feedforward control of gait.     

Developmental origins of complex radular characters in the Muricidae: the bifid rachidian edge

Muricid gastropod radulae are more complex than those of most other neogastropods, especially in the number and variety of cusps, denticles, and interlocking mechanisms. How this complexity evolved, however, is unknown. Morphological gaps between higher taxa within the Muricidae are substantial, and there are few unambiguous intermediates. Here, we use developmental data from the Patagonian trophonine muricid Trophon geversianus to investigate the evolution of an unusual condition in which there are two marginal cusps at each end of each central rachidian tooth, rather than one or none as in most muricids. Trophon geversianus begins ontogeny with one marginal cusp (the inner marginal cusp), but a second (the outer marginal cusp) appears later, arising from separation of the rachidian base edge from the radular membrane rather than through bifurcation or lateral migration of pre‐existing cusps. Truncation of development (i.e., paedomorphosis) at this second developmental phase in a trophonine ancestor provides an explanation for the lack of transitional forms between most adult trophonine muricids, which have the plesiomorphic condition of one marginal cusp, and sister group ocenebrine muricids, which have the derived condition of two marginal cusps.     

Phosphorylation of the Casein Kinase II Domain of B-50 (GAP-43) in Rat Cortical Growth Cones

Abstract: Growth-associated phosphoprotein B-50 is a neural protein kinase C (PKC) substrate enriched in nerve growth cones that has been implicated in growth cone plasticity. Here we investigated whether B-50 is a physiological substrate for casein kinase II (CKII) in purified rat cortical growth cone preparations. Using site-specific proteolysis and known modulators of PKC, in combination with immunoprecipitation, mass spectrometry, and phosphoamino acid analysis, we demonstrate that endogenous growth cone B-50 is phosphorylated at multiple sites, on both serine and threonine residues. Consistent with previous reports, stimulation of PKC activity increased the phosphorylation of only those proteolytic fragments containing Ser⁴¹. Under basal conditions, however, phosphorylation was predominantly associated with fragments not containing Ser⁴¹. Mass spectrometry of tryptic digests of B-50, which had been immunoprecipitated from untreated growth cones, revealed that in situ phosphorylation occurs within peptides B-50_181–198 and B-50_82–98. These peptides contain the major and minor in vitro CKII phosphosites, respectively. In addition, cyanogen bromide digestion of immunoprecipitated chick B-50 generated a 4-kDa C-terminal B-50 phosphopeptide, confirming that phosphorylation of the CKII domain occurs across evolutionary diverse species. We conclude that B-50 in growth cones is not only a substrate for PKC, but also for CKII.     

Increased Secretion of the Amino-Terminal Fragment of Amyloid Precursor Protein in Brains of Rats with a Constitutive Up-Regulation of Protein Kinase C

Abstract: Protein kinase C (PKC) activation stimulates release of secreted amyloid precursor protein (APP_s) in several cell lines. To ascertain the role of PKC in regulating APP metabolism in vivo, we used an animal model (methylazoxymethanol-treated rats; MAM rats) in which PKC is permanently hyperactivated in selected brain areas, i.e., cortex and hippocampus. A significant decrease in membrane-bound APP concentration was found in synaptosomes derived from cortex and hippocampus of MAM rats, where PKC is up-regulated, with a concomitant increase in APP_s production in soluble fractions of the same brain areas. In contrast, in a brain area not affected by MAM treatment (i.e., cerebellum), APP secretion is similar in control and MAM rats, indicating that altered metabolism of APP is restricted to only those areas in which the PKC system is up-regulated. In addition, phorbol esters or H-7 modulate APP_s release in hippocampal slices from both control and MAM rats, further supporting an in vivo role for this enzyme in regulating metabolism of mature APP.