Cleavage of the plasma membrane Na+/Ca2+ exchanger in excitotoxicity

In brain ischemia, gating of postsynaptic glutamate receptors and other membrane channels triggers intracellular Ca2+ overload and cell death. In excitotoxic settings, the initial Ca2+ influx through glutamate receptors is followed by a second uncontrolled Ca2+ increase that leads to neuronal demise. Here we report that the major plasma membrane Ca2+ extruding system, the Na+/Ca2+ exchanger (NCX), is cleaved during brain ischemia and in neurons undergoing excitotoxicity. Inhibition of Ca2+-activated proteases (calpains) by overexpressing their endogenous inhibitor protein, calpastatin or the expression of an NCX isoform not cleaved by calpains, prevented Ca2+ overload and rescued neurons from excitotoxic death. Conversely, down-regulation of NCX by siRNA compromised neuronal Ca2+ handling, transforming the Ca2+ transient elicited by non-excitotoxic glutamate concentrations into a lethal Ca2+overload. Thus, proteolytic inactivation of NCX-driven neuronal Ca2+ extrusion is responsible for the delayed excitotoxic Ca2+ deregulation and neuronal death.     

Tumor necrosis factor, cancer and anticancer therapy

Tumor necrosis factor (TNF) is being utilized as an antineoplastic agent for the treatment of patients with locally advanced solid tumors. However, its role in cancer therapy is debated. Although a large body of evidence supports TNF's antineoplastic activity, the cascade of molecular events underlying TNF-mediated tumor regression observed in vivo is still incompletely elucidated. Intriguingly, some pre-clinical findings suggest that TNF may promote cancer development and progression, which has led to propose anti-TNF therapy as a novel approach to malignancies. In the present work, we summarize the molecular biology of TNF with particular regard to its tumor-related properties, and review the experimental and clinical evidence currently available describing the complex and sometime conflicting relationship between this cytokine, cancer and antitumor therapy. Recent insights that might pave the way to further exploitation of the antineoplastic potential of TNF are also discussed.     

Relationships between macroalgal biomass and microbiological quality of water in a phytotreatment pond

The microbiological quality of wastewater in phytotreatment ponds with foliose macroalgae can be influenced by biofilm formation on thallus surface. This hypothesis was tested with an in situ experiment which was carried out in a pond with Ulva spp. receiving wastewater from a land based fish farm at Piombino (Italy). The total bacterial load (TBL) was determined in the inflowing and outflowing waters and a multifactorial design was employed to investigate the effect of different macroalgal biomass. Microbiological analysis revealed a high TBL in the water column (18.4 ± 7.4 × 10⁸ cells ml⁻¹). TBL of inlet water was significantly correlated with quantity and quality of particulate organic matter (POM) of inflowing water, whereas no correlation was found between TBL and POM in the outlet water. A significant decrease in the POM concentration was detected within macroalgal ponds, due to the mechanical action of thalli which favoured POM sedimentation. Nevertheless, great TBLs were found in the outlet water. These findings suggest that TBL probably depended upon macroalgae. Indeed high bacterial density was found on macroalgal thallus surface (~10⁸ cells cm²). Furthermore, high plate counts of faecal bacteria (faecal enterococci) were determined on thallus surface (~ 40 CFU cm²) and outlet water (11 886 ± 3984 CFU 100 ml⁻¹) supporting the evidence that macroalgae negatively affect the microbiological quality of treated water. Bacterial activities in terms of exoenzymatic rates and secondary production were two folds higher in the water within macroalgal beds, than in the open water. These preliminary results suggest that high macroalgal biomass represents a ‚hot spot’ of bacterial density and activity that may affect microbiological quality of the treated water. Bacterial control of inlet water and management of macroalgal biomass through periodic removal are essential for a more efficient treatment of wastewater in phytotreatment ponds.     

New statistical software for intralaboratory and interlaboratory quality control in clinical cytology. Validation in a simulation study on clinical samples

OBJECTIVE: To design a statistical software package to provide automated calculations of normal and weighted and 3 indices. STUDY DESIGN: Prompted by the lack of commonly available software to compute weighted kappa and the nonproportionate workload needed to calculate our 3 variability indices manually, the new statistical software package was designed. To demonstrate the performance of the new CONQUISTADOR software, a simulation study (both intralaboratory and interlaboratory) was designed using 5,000 clinical samples randomly selected from a data file of > or = 200,000 conventional Pap smears and programmed to become "analyzed" by 12 cytologists in 5 imaginary laboratories. RESULTS: A representative set of both complete and partial outputs provided by the software, in Excel format (Microsoft, Redmond, Washington, U.S.A.) are shown to illustrate the different functions of the program. In the interlaboratory mode, the software calculates accuracy indicators (sensitivity, specificity, positive and negative predictive value, and their 95% CI), which are not common features of regular statistical packages; kappa and weighted kappa; and their 95% CI (comparison of single laboratories to all laboratories and pairwise comparisons between single laboratories). The 3 diagnostic variability indices can be computed separately for all samples or for only the positive samples. In the intralaboratory mode, the software calculates the same indices for individual cytologists. CONCLUSION: The CONQUISTADOR statistical package has properties that are useful in monitoring cytologic laboratory quality in both intralaboratory and interlaboratory settings. The software will be distributed by the National Institute of Health, Rome, for the delivery costs only.     

Characterization of the Prion Protein in Human Urine

The presence of the prion protein (PrP) in normal human urine is controversial and currently inconclusive. This issue has taken a special relevance because prion infectivity has been demonstrated in urine of animals carrying experimental or naturally occurring prion diseases, but the actual presence and tissue origin of the infectious prion have not been determined. We used immunoprecipitation, one- and two-dimensional electrophoresis, and mass spectrometry to prove definitely the presence of PrP in human urine and its post-translational modifications. We show that urinary PrP (uPrP) is truncated mainly at residue 112 but also at other residues up to 122. This truncation makes uPrP undetectable with some commonly used antibodies to PrP. uPrP is glycosylated and carries an anchor which, at variance with that of cellular PrP, lacks the inositol-associated phospholipid moiety, indicating that uPrP is probably shed from the cell surface. The detailed characterization of uPrP reported here definitely proves the presence of PrP in human urine and will help determine the origin of prion infectivity in urine.     

Biochemical Origin and Refractory Properties of Humic Acid Extracted from the Maize Plant

Humic acids (HA) contribute to soil fertility because of their chemical, physical, and biological properties. The origin of HAs in soils has puzzled scientists for decades, and what HAs are and what their origin is remain unclear. The isolation of HAs in plants, which have characteristics close to soil HAs, suggests the probable origin of soil-HA is the preservation of plant tissue, indicating biochemical origin. In this paper HA from maize plant at different stages of maturity is isolated, from which it was found that the evolution of this fraction depends on and is derived from cell wall formation. Evidence was also found that HA was above all composed of lignin and cutin residues, and was characterized by low surface area. After 8 months of incubation in both mineral-artificial and natural soils, humic acid isolated form maize plant could be recovered intact.     

Polyphasic screening, homopolysaccharide composition, and viscoelastic behavior of wheat Sourdough from a Leuconostoc lactis and Lactobacillus curvatus exopolysaccharide-producing starter culture

After isolation from different doughs and sourdoughs, 177 strains of lactic acid bacteria were screened at the phenotypic level for exopolysaccharide production on media containing different carbohydrate sources. Two exopolysaccharide-producing lactic acid bacteria (Lactobacillus curvatus 69B2 and Leuconostoc lactis 95A) were selected through quantitative analysis on solid media containing sucrose and yeast extract. The PCR detection of homopolysaccharide (gtf and lev) and heteropolysaccharide (epsA, epsB, epsD and epsE, and epsEFG) genes showed different distributions within species and strains of the lactic acid bacteria studied. Moreover, in some strains both homopolysaccharide and heteropolysaccharide genes were detected. Proton nuclear magnetic resonance spectra suggest that Lactobacillus curvatus 69B2 and Leuconostoc lactis 95A produced the same exopolysaccharide, which was constituted by a single repeating glucopyranosyl unit linked by an α-(1→6) glycosidic bond in a dextran-type carbohydrate. Microbial growth, acidification, and viscoelastic properties of sourdoughs obtained by exopolysaccharide-producing and nonproducing lactic acid bacterial strains were evaluated. Sourdough obtained after 15 h at 30°C with exopolysaccharide-producing lactic acid bacteria reached higher total titratable acidity as well as elastic and dissipative modulus curves with respect to the starter not producing exopolysaccharide, but they showed similar levels of pH and microbial growth. On increasing the fermentation time, no difference in the viscoelastic properties of exopolysaccharide-producing and nonproducing samples was observed. This study suggests that dextran-producing Leuconostoc lactis 95A and Lactobacillus curvatus 69B2 can be employed to prepare sourdough, and this would be particularly useful to improve the quality of baked goods while avoiding the use of commercially available hydrocolloids as texturizing additives.     

Mirna expression profiles identify drivers in colorectal and pancreatic cancers

Background and Aim

Altered expression of microRNAs (miRNAs) hallmarks many cancer types. The study of the associations of miRNA expression profile and cancer phenotype could help identify the links between deregulation of miRNA expression and oncogenic pathways.

Methods

Expression profiling of 866 human miRNAs in 19 colorectal and 17 pancreatic cancers and in matched adjacent normal tissues was investigated. Classical paired t-test and random forest analyses were applied to identify miRNAs associated with tissue-specific tumors. Network analysis based on a computational approach to mine associations between cancer types and miRNAs was performed.

Results

The merge between the two statistical methods used to intersect the miRNAs differentially expressed in colon and pancreatic cancers allowed the identification of cancer-specific miRNA alterations. By miRNA-network analysis, tissue-specific patterns of miRNA deregulation were traced: the driving miRNAs were miR-195, miR-1280, miR-140-3p and miR-1246 in colorectal tumors, and miR-103, miR-23a and miR-15b in pancreatic cancers.

Conclusion

MiRNA expression profiles may identify cancer-specific signatures and potentially useful biomarkers for the diagnosis of tissue specific cancers. miRNA-network analysis help identify altered miRNA regulatory networks that could play a role in tumor pathogenesis.