Structural and dynamical properties of KTS‐disintegrins: A comparison between Obtustatin and Lebestatin

Obtustatin and Lebestatin are lysine‐threonine‐serine (KTS)‐disintegrins, which are a family of low molecular weight polypeptides present in many viperidae venoms and are potent and specific inhibitors of collagen‐binding integrins. The integrin binding loop, harboring the ²¹KTS²³ motif, and the C‐terminal tail are known to be responsible for the selective binding to the α1β1 integrin. Despite a very high sequence homology (only two mutations are present in Lebestatin relative to Obtustatin, namely R24L and S38L), Lebestatin exhibits a higher inhibitory effect than Obtustatin on cell adhesion and cell migration to collagens I and IV. Here we show, by means of molecular dynamics simulations of the two polypeptides in aqueous solution, that Lebestatin possesses a higher flexibility of the C‐terminal tail and a greater solvent accessibility of the integrin binding loop than Obtustatin. It may be hypothesized that these properties may contribute to the higher binding‐affinity of Lebestatin to its biological partner. © 2012 Wiley Periodicals, Inc.     

Impact of DNA methyltransferases on the epigenetic regulation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor expression in malignant melanoma

Aberrant promoter methylation and resultant silencing of TRAIL decoy receptors were reported in a variety of cancers, but to date little is known about the relevance of this epigenetic modification in melanoma. In this study, we examined the methylation and the expression status of TRAIL receptor genes in cutaneous and uveal melanoma cell lines and specimens and their interaction with DNA methyltransferases (DNMTs) DNMT1, DNMT3a, and DNMT3b. DR4 and DR5 methylation was not frequent in cutaneous melanoma but on the contrary it was very frequent in uveal melanoma. No correlation between methylation status of DR4 and DR5 and gene expression was found. DcR1 and DcR2 were hypermethylated with very high frequency in both cutaneous and uveal melanoma. The concordance between methylation and loss of gene expression ranged from 91% to 97%. Here we showed that DNMT1 was crucial for DcR2 hypermethylation and that DNMT1 and DNMT3a coregulate the methylation status of DcR1. Our work also revealed the critical relevance of DcR1 and DcR2 expression in cell growth and apoptosis either in cutaneous or uveal melanoma. In conclusion, the results presented here claim for a relevant impact of aberrant methylation of decoy receptors in melanoma and allow to understand how the silencing of DcR1 and DcR2 is related to melanomagenesis.     

Fluorescent ligands for adenosine receptors

Interest is increasing in developing fluorescent ligands for characterization of adenosine receptors (ARs), which hold a promise of usefulness in the drug discovery process. The size of a strategically labeled AR ligand can be greatly increased after the attachment of a fluorophore. The choice of dye moiety (e.g. Alexa Fluor 488), attachment point and linker length can alter the selectivity and potency of the parent molecule. Fluorescent derivatives of adenosine agonists and antagonists (e.g. XAC and other heterocyclic antagonist scaffolds) have been synthesized and characterized pharmacologically. Some are useful AR probes for flow cytometry, fluorescence correlation spectroscopy, fluorescence microscopy, fluorescence polarization, fluorescence resonance energy transfer, and scanning confocal microscopy. Thus, the approach of fluorescent labeled GPCR ligands, including those for ARs, is a growing dynamic research field.     

Freshwater algae of Southern Africa

Observations are reported on akinete formation, sheath formation and the breaking away of the rest of the trichome after akinete formation in Gloeotrichia ghosei R. N. Singh.     

Small molecules interacting with α-synuclein: antiaggregating and cytoprotective properties

Curcumin, a dietary polyphenol, has shown a potential to act on the symptoms of neurodegenerative disorders, including Alzheimer’s and Parkinson’s diseases, as a consequence of its antioxidant, anti-inflammatory and anti-protein aggregation properties. Unfortunately, curcumin undergoes rapid degradation at physiological pH into ferulic acid, vanillin and dehydrozingerone, making it an unlikely drug candidate. Here, we evaluated the ability of some curcumin by-products: dehydrozingerone (1), its O-methyl derivative (2), zingerone (3), and their biphenyl analogues (4–6) to interact with α-synuclein (AS), using CD and fluorescence spectroscopy. In addition, the antioxidant properties and the cytoprotective effects in rat pheochromocytoma (PC12) cells prior to intoxication with H₂O₂, MPP+ and MnCl₂ were examined while the Congo red assay was used to evaluate the ability of these compounds to prevent aggregation of AS. We found that the biphenyl zingerone analogue (6) interacts with high affinity with AS and also displays the best antioxidant properties while the biphenyl analogues of dehydrozingerone (4) and of O-methyl-dehydrozingerone (5) are able to partially inhibit the aggregation process of AS, suggesting the potential role of a hydroxylated biphenyl scaffold in the design of AS aggregation inhibitors.     

Negative modulation of mitochondrial oxidative phosphorylation by epigallocatechin-3 gallate leads to growth arrest and apoptosis in human malignant pleural mesothelioma cells

Increasing evidence reveals a large dependency of epithelial cancer cells on oxidative phosphorylation (OXPHOS) for energy production. In this study we tested the potential of epigallocatechin-3-gallate (EGCG), a natural polyphenol known to target mitochondria, in inducing OXPHOS impairment and cell energy deficit in human epitheliod (REN cells) and biphasic (MSTO-211H cells) malignant pleural mesothelioma (MMe), a rare but highly aggressive tumor with high unmet need for treatment. Due to EGCG instability that causes H₂O₂ formation in culture medium, the drug was added to MMe cells in the presence of exogenous superoxide dismutase and catalase, already proved to stabilize the EGCG molecule and prevent EGCG-dependent reactive oxygen species formation. We show that under these experimental conditions, EGCG causes the selective arrest of MMe cell growth with respect to normal mesothelial cells and the induction of mitochondria-mediated apoptosis, as revealed by early mitochondrial ultrastructure modification, swelling and cytochrome c release. We disclose a novel mechanism by which EGCG induces apoptosis through the impairment of mitochondrial respiratory chain complexes, particularly of complex I, II and ATP synthase. This induces a strong reduction in ATP production by OXPHOS, that is not adequately counterbalanced by glycolytic shift, resulting in cell energy deficit, cell cycle arrest and apoptosis. The EGCG-dependent negative modulation of mitochondrial energy metabolism, selective for cancer cells, gives an important input for the development of novel pharmacological strategies for MMe.     

Evidence of Cross-Reactive Immunity to 2009 Pandemic Influenza A Virus in Workers Seropositive to Swine H1N1 Influenza Viruses Circulating in Italy

Background

Pigs play a key epidemiologic role in the ecology of influenza A viruses (IAVs) emerging from animal hosts and transmitted to humans. Between 2008 and 2010, we investigated the health risk of occupational exposure to swine influenza viruses (SIVs) in Italy, during the emergence and spread of the 2009 H1N1 pandemic (H1N1pdm) virus.

Methodology/Principal Findings

Serum samples from 123 swine workers (SWs) and 379 control subjects (Cs), not exposed to pig herds, were tested by haemagglutination inhibition (HI) assay against selected SIVs belonging to H1N1 (swH1N1), H1N2 (swH1N2) and H3N2 (swH3N2) subtypes circulating in the study area. Potential cross-reactivity between swine and human IAVs was evaluated by testing sera against recent, pandemic and seasonal, human influenza viruses (H1N1 and H3N2 antigenic subtypes). Samples tested against swH1N1 and H1N1pdm viruses were categorized into sera collected before (n. 84 SWs; n. 234 Cs) and after (n. 39 SWs; n. 145 Cs) the pandemic peak. HI-antibody titers ≥10 were considered positive. In both pre-pandemic and post-pandemic peak subperiods, SWs showed significantly higher swH1N1 seroprevalences when compared with Cs (52.4% vs. 4.7% and 59% vs. 9.7%, respectively). Comparable HI results were obtained against H1N1pdm antigen (58.3% vs. 7.7% and 59% vs. 31.7%, respectively). No differences were found between HI seroreactivity detected in SWs and Cs against swH1N2 (33.3% vs. 40.4%) and swH3N2 (51.2 vs. 55.4%) viruses. These findings indicate the occurrence of swH1N1 transmission from pigs to Italian SWs.

Conclusion/Significance

A significant increase of H1N1pdm seroprevalences occurred in the post-pandemic peak subperiod in the Cs (p<0.001) whereas SWs showed no differences between the two subperiods, suggesting a possible occurrence of cross-protective immunity related to previous swH1N1 infections. These data underline the importance of risk assessment and occupational health surveillance activities aimed at early detection and control of SIVs with pandemic potential in humans.     

A Combination of Mitochondrial Oxidative Stress and Excess Fat/Calorie Intake Accelerates Steatohepatitis by Enhancing Hepatic CC Chemokine Production in Mice

Mitochondrial oxidative stress is considered as a key accelerator of fibrosis in various organs including the liver. However, the production of oxidative stress and progression of liver fibrosis may merely represent the independent consequences of hepatocellular injury caused by the primary disease. Because of a lack of appropriate experimental models to evaluate the sole effects of oxidative stress, it is virtually unknown whether this stress is causatively linked to the progression of liver fibrosis. Here, we examined the direct effects of mitochondrial reactive oxygen species (ROS) on the progression of high fat/calorie diet-induced steatohepatitis using Tet-mev-1 mice, in which a mutated succinate dehydrogenase transgene impairs the mitochondrial electron transport and generates an excess amount of ROS in response to doxycycline administration. Wild type and Tet-mev-1 mice that had been continuously given doxycycline-containing water were subsequently fed either normal chow or a cholesterol-free high-fat/high-sucrose diet for 4 months at approximately 1 or 2 years of age. Histopathological examinations indicated that neither the mitochondrial ROS induced in Tet-mev-1 mice nor the feeding of wild type animals with high-fat/high-sucrose diet alone caused significant liver fibrosis. Only when the Tet-mev-1 mice were fed a high-fat/high-sucrose diet, it induced lipid peroxidation in hepatocytes and enhanced hepatic CC chemokine expression. These events were accompanied by increased infiltration of CCR5-positive cells and activation of myofibroblasts, resulting in extensive liver fibrosis. Interestingly, this combinatorial effect of mitochondrial ROS and excess fat/calorie intake on liver fibrosis was observed only in 2-year-old Tet-mev-1 mice, not in the 1-year-old animals. Collectively, these results indicate that mitochondrial ROS in combination with excess fat/calorie intake accelerates liver fibrosis by enhancing CC chemokine production in aged animals. We have provided a good experimental model to explore how high fat/calorie intake increases the susceptibility to nonalcoholic steatohepatitis in aged individuals who have impaired mitochondrial adaptation.