Molecular cloning, expression analysis and cellular localization of gomesin, an anti-microbial peptide from hemocytes of the spider Acanthoscurria gomesiana

Gomesin is a cationic anti-microbial peptide of 18 amino acid residues isolated from the hemocytes of unchallenged tarantula spider Acanthoscurria gomesiana. This paper reports the first study of the processing and cellular location of an anti-microbial peptide (AMP) in spiders. Gomesin cDNA sequence analysis indicated that it is processed from a precursor containing a signal peptide (23 amino acid residues) and a negative C-terminal region (43 amino acid residues). The gomesin gene was constitutively transcribed in hemocytes and the gene product localized in hemocyte granules. The constitutive production of gomesin by a spider is discussed in the context of an ancient mechanism of AMP regulation and storage.     

Bilirubin: its role in cytoprotection against oxidative stress

Bilirubin, the end product of heme catabolism in mammals, is generally regarded as a potentially cytotoxic, lipid-soluble waste product that needs to be excreted. However, in the last 10 years, in vitro and in vivo studies, have demonstrated that bilirubin exhibits potent anti-oxidant properties preventing the oxidative damage triggered by a wide range of oxidant-related stimuli. Therefore, the idea of a beneficial and physiological role for bilirubin in cytoprotection against short and long-lasting oxidant-mediated cell injury is highlighted here.     

Morphological and histochemical characterization of the pectoral fin muscle of the stripped weakfish,Cynoscion guatucupa

The fibres of superficial and deep abductor muscles of the pectoral fins of the stripped weakfish, Cynoscion guatucupa have been studied using histochemical techniques: succinic dehydrogenase (SDH) for mitochondria, periodic acid–Schiff (PAS) for glycogen, myosin‐adenosintriphosphatase (mATPase) to identify different fibre types based on the contraction speed and modified ATPase to identify capillaries. The fibre diameters were measured, and the capillaries of the main fibre types – red, pink and white— were counted. The two muscles showed both macroscopically and microscopically two well‐differentiated zones with predominant white fibres. The area of insertion of muscles into the fin rays had red, pink and white fibres. The origin zone of the muscle into the bone was composed by white fibres only. Both zones of white muscle evidenced a mosaic of small, medium and large polygonal white fibres. Red, pink and white muscles showed a wide histochemical diversity of fibre subtypes. The area per peripheral capillary increased from the red to the white muscles. Due to the predominance of white fibres, the pectoral fins of C. guatucupa were mainly involved in rapid movements to stop/discontinue and stabilize the body during swimming.     

The distinct erythropoietin functions that promote cell survival and proliferation are affected by aluminum exposure through mechanisms involving erythropoietin receptor

Erythropoietin (Epo) promotes the development of erythroid progenitors by triggering intracellular signals through the binding to its specific receptor (EpoR). Previous results related to the action of aluminum (Al) on erythropoiesis let us suggest that the metal affects Epo interaction with its target cells. In order to investigate this effect on cell activation by the Epo-EpoR complex, two human cell lines with different dependence on Epo were subjected to Al exposure. In the Epo-independent K562 cells, Al inhibited Epo antiapoptotic action and triggered a simultaneous decrease in protein and mRNA EpoR levels. On the other hand, proliferation of the strongly Epo-dependent UT-7 cells was enhanced by long-term Al treatment, in agreement with the upregulation of EpoR expression during Epo starvation. Results provide some clues to the way by which Epo supports cell survival and growth, and demonstrate that not all the intracellular factors needed to guarantee the different signaling pathways of Epo-cell activation are available or activated in cells expressing EpoR. This study then suggests that at least one of the mechanisms by which Al interfere with erythropoiesis might involve EpoR modulation.     

Oxidative stress due to aluminum exposure induces eryptosis which is prevented by erythropoietin

The widespread use of aluminum (Al) provides easy exposure of humans to the metal and its accumulation remains a potential problem. In vivo and in vitro assays have associated Al overload with anemia. To better understand the mechanisms by which Al affects human erythrocytes, morphological and biochemical changes were analyzed after long-term treatment using an in vitro model. The appearance of erythrocytes with abnormal shapes suggested metal interaction with cell surface, supported by the fact that high amounts of Al attached to cell membrane. Long-term incubation of human erythrocytes with Al induced signs of premature erythrocyte death (eryptosis), such as phosphatidylserine externalization, increased intracellular calcium, and band 3 degradation. Signs of oxidative stress, such as significant increase in reactive oxygen species in parallel with decrease in the amount of reduced glutathione, were also observed. These oxidative effects were completely prevented by the antioxidant N-acetylcysteine. Interestingly, erythrocytes were also protected from the prooxidative action of Al by the presence of erythropoietin (EPO). In conclusion, results provide evidence that chronic Al exposure may lead to biochemical and morphological alterations similar to those shown in eryptosis induced by oxidant compounds in human erythrocytes. The antieryptotic effect of EPO may contribute to enhance the knowledge of its physiological role on erythroid cells. Irrespective of the antioxidant mechanism, this property of EPO, shown in this model of Al exposure, let us suggest potential benefits by EPO treatment of patients with anemia associated to altered redox environment.     

Morphological and biochemical approaches to assess the nutritional condition of the Argentine hake Merluccius hubbsi larvae from two different nursery areas

The Argentine hake, Merluccius hubbsi, is one of the most important fishing species in the Argentine Sea due to its great abundance and high-quality meat. The study of the nutritional condition is widely used to determine the physiological state of the fish larvae and to estimate their survival possibilities. The larval nutritional condition reflects the environmental conditions to which they have been exposed and represents a useful instrument to determine favourable nursery areas. It also provides tools for the comprehensive management of a population subjected to fishing exploitation. This study aimed to determine potential differences in the nutritional condition of M. hubbsi larvae from the two fishing stocks (Northern and Southern) of the species. The authors assessed the nutritional condition of larvae captured during the 2012 main reproductive peak in the nursery areas of each population. Two different methodologies were applied: a morphometrical approach, by recording five morphometric variables, and a biochemical technique, employing the RNA/DNA index (RD_s) and its derived index of growth performance. The morphometrical indexes did not show differences in the larval condition between both stocks. Nonetheless, the RD_s index did detect differences in the nutritional condition of larvae from different stocks. The RD_s index of larvae in pre-flexion and flexion stages showed significant differences between stocks, indicating that these stages are more susceptible to starvation. The results suggest that the biochemical indexes prove to be more sensitive than the morphometric indexes to detect slight differences in hake larvae nutritional condition. The scope and limitations of these techniques for the analysis of the nutritional condition of larvae are discussed.     

Histological and histochemical study of the digestive system of the Argentine anchovy larvae (Engraulis anchoita) at different developmental stages of their ontogenetic development

In this work, histological and histochemical features of the larval digestive system of Argentine anchovy Engraulis anchoita were described. Structural changes during ontogenetic development were also characterized, and comparisons between the beginning and the end of larval development were made. Histological sections of larvae were subjected to histochemical and routine histological techniques to localize and differentiate glycoproteins (GPs). Both at an early and a late larval stage, the oesophageal goblet cells reacted more intensely than those of the rest of the digestive tract, and only the oesophagus exhibited metachromasia with toluidine blue techniques at different pHs, thus revealing diverse GPs at different concentrations. The GPs histochemical composition in the intestine varied with the developmental stage and the intestinal zone. The absence of goblet cells characterized the foregut; however, they started differentiation at an advanced stage in the midgut. These cells could be detected in the hindgut both at the beginning and at the end of development. The attached glands showed a varied glycoprotein composition. The digestive tract of E. anchoita presented a high level of complexity, related to the multiple functions of mucus in the digestive tract, such as lubrication, protection, antimicrobial function and ionic and osmotic regulation.     

TNF-alpha-induced apoptosis is prevented by erythropoietin treatment on SH-SY5Y cells

The growth factor erythropoietin (Epo) has shown neuronal protective action in addition to its well known proerythroid activity. Furthermore, Epo has dealt with cellular inflammation by inhibiting the expression of several proinflammatory cytokines, such as IL-1 and TNF-α. The action of TNF can have both apoptotic and antiapoptotic consequences due to altered balance between different cell signalling pathways. This work has focused on the apoptotic effects of this cytokine and the potential protective action of Epo. The model we used was neuroblastoma SH-SY5Y cells cultured in the presence of 25 ng/ml TNF-α or pretreated with 25 U/ml Epo for 12 h before the addition of TNF-α. Apoptosis was evaluated by differential cell count after Hoechst staining, analysis of DNA ladder pattern, and measurement of caspase activity. Despite its ability to induce NF-κB nuclear translocation, TNF-α induced cell death, which was found to be associated to upregulation of TNF Receptor 1 expression. On the other hand, cells activated by Epo became resistant to cell death. Prevention of death receptor upregulation and caspase activation may explain this antiapoptotic effect of Epo, which may be also favoured by the induction of a higher expression of protective factors, such as Bcl-2 and NF-κB, through mechanisms involving Jak/STAT and PI3K signalling pathways.     

p38 mitogen-activated protein kinase mediates the Fas-induced mitochondrial death pathway in CD8+ T cells

The p38 mitogen-activated protein kinase (MAPK) signaling pathway can be activated by a variety of stress stimuli such as UV radiation and osmotic stress. The regulation and role of this pathway in death receptor-induced apoptosis remain unclear and may depend on the specific death receptor and cell type. Here we show that binding of Fas ligand to Fas activates p38 MAPK in CD8+ T cells and that activation of this pathway is required for Fas-mediated CD8+ T-cell death. Active p38 MAPK phosphorylates Bcl-xL and Bcl-2 and prevents the accumulation of these antiapoptotic molecules within the mitochondria. Consequently, a loss of mitochondrial membrane potential and the release of cytochrome c lead to the activation of caspase 9 and, subsequently, caspase 3. Therefore, the activation of p38 MAPK is a critical link between Fas and the mitochondrial death pathway and is required for the Fas-induced apoptosis of CD8+ T cells.