Resveratrol inhibits protein translation in hepatic cells

Resveratrol is a plant-derived polyphenol that extends lifespan and healthspan in model organism. Despite extensive investigation, the biological processes mediating resveratrol's effects have yet to be elucidated. Because repression of translation shares many of resveratrol's beneficial effects, we hypothesized that resveratrol was a modulator of protein synthesis. We studied the effect of the drug on the H4-II-E rat hepatoma cell line. Initial studies showed that resveratrol inhibited global protein synthesis. Given the role of the mammalian Target of Rapamycin (mTOR) in regulating protein synthesis, we examined the effect of resveratrol on mTOR signaling. Resveratrol inhibited mTOR self-phosphorylation and the phosphorylation of mTOR targets S6K1 and eIF4E-BP1. It attenuated the formation of the translation initiation complex eIF4F and increased the phosphorylation of eIF2α. The latter event, also a mechanism for translation inhibition, was not recapitulated by mTOR inhibitors. The effects on mTOR signaling were independent of effects on AMP-activated kinase or AKT. We conclude that resveratrol is an inhibitor of global protein synthesis, and that this effect is mediated through modulation of mTOR-dependent and independent signaling.     

Autotaxin and LPA receptors represent potential molecular targets for the radiosensitization of murine glioma through effects on tumor vasculature

Despite wide margins and high dose irradiation, unresectable malignant glioma (MG) is less responsive to radiation and is uniformly fatal. We previously found that cytosolic phospholipase A2 (cPLA(2)) is a molecular target for radiosensitizing cancer through the vascular endothelium. Autotaxin (ATX) and lysophosphatidic acid (LPA) receptors are downstream from cPLA(2) and highly expressed in MG. Using the ATX and LPA receptor inhibitor, α-bromomethylene phosphonate LPA (BrP-LPA), we studied ATX and LPA receptors as potential molecular targets for the radiosensitization of tumor vasculature in MG. Treatment of Human Umbilical Endothelial cells (HUVEC) and mouse brain microvascular cells bEND.3 with 5 μmol/L BrP-LPA and 3 Gy irradiation showed decreased clonogenic survival, tubule formation, and migration. Exogenous addition of LPA showed radioprotection that was abrogated in the presence of BrP-LPA. In co-culture experiments using bEND.3 and mouse GL-261 glioma cells, treatment with BrP-LPA reduced Akt phosphorylation in both irradiated cell lines and decreased survival and migration of irradiated GL-261 cells. Using siRNA to knock down LPA receptors LPA1, LPA2 or LPA3 in HUVEC, we demonstrated that knockdown of LPA2 but neither LPA1 nor LPA3 led to increased viability and proliferation. However, knockdown of LPA1 and LPA3 but not LPA2 resulted in complete abrogation of tubule formation implying that LPA1 and LPA3 on endothelial cells are likely targets of BrP-LPA radiosensitizing effect. Using heterotopic tumor models of GL-261, mice treated with BrP-LPA and irradiation showed a tumor growth delay of 6.8 days compared to mice treated with irradiation alone indicating that inhibition of ATX and LPA receptors may significantly improve malignant glioma response to radiation therapy. These findings identify ATX and LPA receptors as molecular targets for the development of radiosensitizers for MG.     

Modulation of virulence and antibiotic susceptibility of enteropathogenic Escherichia coli strains by Enterococcus faecium probiotic strain culture fractions

The increasing rate of antimicrobial resistance drastically reduced the efficiency of conventional antibiotics and led to the reconsideration of the interspecies interactions in influencing bacterial virulence and response to therapy. The aim of the study was the investigation of the influence of the soluble and cellular fractions of Enterococcus (E.) faecium CMGB16 probiotic culture on the virulence and antibiotic resistance markers expression in clinical enteropathogenic Escherichia (E.) coli strains.The 7 clinical enteropathogenic E. coli strains, one standard E. coli ATCC 25,922 and one Bacillus (B.) cereus strains were cultivated in nutrient broth, aerobically at 37 °C, for 24 h. The E. faecium CMGB16 probiotic strain was cultivated in anaerobic conditions, at 37 °C in MRS (Man Rogosa Sharpe) broth, and co-cultivated with two pathogenic strains (B. cereus and E. coli O28) culture fractions (supernatant, washed sediment and heat-inactivated culture) for 6 h, at 37 °C. After co-cultivation, the soluble and cellular fractions of the probiotic strain cultivated in the presence of two pathogenic strains were separated by centrifugation (6000 rpm, 10 min), heat-inactivated (15 min, 100 °C) and co-cultivated with the clinical enteropathogenic E. coli strains in McConkey broth, for 24 h, at 37 °C, in order to investigate the influence of the probiotic fractions on the adherence capacity and antibiotic susceptibility. All tested probiotic combinations influenced the adherence pattern of E. coli tested strains. The enteropathogenic E. coli strains susceptibility to aminoglycosides, beta-lactams and quinolones was increased by all probiotic combinations and decreased for amoxicillin-clavulanic acid. This study demonstrates that the plurifactorial anti-infective action of probiotics is also due to the modulation of virulence factors and antibiotic susceptibility expression in E. coli pathogenic strains.     

Polymorphisms associated with the risk of lung cancer in a healthy Mexican Mestizo population: Application of the additive model for cancer

Lung cancer is the leading cause of cancer mortality in Mexico and worldwide. In the past decade, there has been an increase in the number of lung cancer cases in young people, which suggests an important role for genetic background in the etiology of this disease. In this study, we genetically characterized 16 polymorphisms in 12 low penetrance genes (AhR, CYP1A1, CYP2E1, EPHX1, GSTM1, GSTT1, GSTPI, XRCC1, ERCC2, MGMT, CCND1 and TP53) in 382 healthy Mexican Mestizos as the first step in elucidating the genetic structure of this population and identifying high risk individuals. All of the genotypes analyzed were in Hardy-Weinberg equilibrium, but different degrees of linkage were observed for polymorphisms in the CYP1A1 and EPHX1 genes. The genetic variability of this population was distributed in six clusters that were defined based on their genetic characteristics. The use of a polygenic model to assess the additive effect of low penetrance risk alleles identified combinations of risk genotypes that could be useful in predicting a predisposition to lung cancer. Estimation of the level of genetic susceptibility showed that the individual calculated risk value (iCRV) ranged from 1 to 16, with a higher iCRV indicating a greater genetic susceptibility to lung cancer.     

Therapeutic effect of a peptide inhibitor of TGF-β on pulmonary fibrosis

Pulmonary fibrosis encompasses several respiratory diseases characterized by epithelial cell injury, inflammation and fibrosis. Transforming growth factor (TGF)-β1 is one of the main profibrogenic cytokines involved in the pathogenesis of lung fibrosis. It induces fibroblast differentiation into myofibroblasts, which produce high levels of collagen and concomitantly loss of lung elasticity and reduction of the respiratory function. In the present study, we have investigated the effects of P17 (a TGF-β inhibitor peptide) on IMR-90 lung fibroblast differentiation in vitro, as well as on the inhibition of the development of bleomycin-induced pulmonary fibrosis in mice. It was found that in IMR-90 cells, P17 inhibited TGF-β1-induced expression of connective tissue growth factor and α-smooth muscle actin. In vivo, treatment of mice with P17 2days after bleomycin administration decreased lung fibrosis, areas of myofibroblast-like cells and lymphocyte infiltrate. P17 also reduced mRNA expression of collagen type I, fibronectin and the fibronectin splice isoform EDA in the lung, and increased the expression of IFN-γ mRNA. Finally, therapeutic treatment with P17 in mice with already established fibrosis was able to significantly attenuate the progression of lung fibrosis. These results suggest that P17 may be useful in the treatment of pulmonary fibrosis.     

Fine structure and molecular content of human chondrocytes encapsulated in alginate beads

Preservation of the chondrocytic phenotype in vitro requires a 3D (three‐dimensional) culture model. Diverse biomaterials have been tested as scaffolds for culture of animal chondrocytes; however, to date, none is considered a gold standard in regenerative medicine. Here, we studied the fine structure and the GAGs (glycosaminoglycans) content of human chondrocytes encapsulated in alginate beads by using electron microscopy and radioactive sulfate [³⁵S] incorporation, respectively. Cells were obtained from human cartilage, encapsulated in alginate beads and cultured for 28 days. [³⁵S]Na₂SO₄ was added to the culture media and later isolated for quantification of the sulfated GAGs found in three compartments: IC (intracellular), IB (intra‐bead) and EB (extra‐bead). Round cells were seen isolated or forming small groups throughout the alginate. Human chondrocytes presented the features of active cells such as euchromatic nuclei, abundant RER (rough endoplasmic reticulum) and many transport vesicles. We observed an extracellular matrix rich in collagen fibres and electrondense material adjacent to the cells. Most of the GAGs produced (74%) were found in the culture medium (EB), indicating that alginate has a limited capacity to retain the GAGs. CS (chondroitin sulfate), the major component of aggrecan, was the most prominent GAG produced by the encapsulated cells. Human chondrocytes cultured in alginate can sustain their phenotype, confirming the potential application of this biomaterial for cartilage engineering.     

Endopolyploid and proliferating trophoblast cells express different patterns of intracellular cytokeratin and glycogen localization in the rat placenta

The presence of keratin intermediate filaments is a characteristic of trophoblast differentiation. Meantime, their intracellular localization in the functionally different subtypes of placental trophoblast is poorly investigated in rodent, whereas their placentae are being broadly investigated in recent years as a model of the feto-maternal interaction. The purpose was to study the intracellular distribution of cytokeratin filaments in correlation with glycogen deposits, both being important constituents of the trophoblast cells in rat placenta. Different rat trophoblast cell populations exhibited different patterns of cytokeratin immunolocalization. The most intensive immunostaining was observed in the highly endopolyploid SGTCs (secondary giant trophoblast cells) at the border with decidua basalis. The most prominent cytokeratin-positive threads were found at the periphery of cytoplasm and in the extensive system of cytoplasmic sprouts by which the SGTC connect each other. Similar cytokeratin intensity and distribution was detected in the TSC (trabecular spongiotrophoblast cells) of the junctional zone of placenta that line the lacunae with the maternal blood. Clusters of highly proliferative pre-glycogen as well as glycogen cells showed some weaker cytokeratin signals mostly in the perinuclear and peripheral zones of cytoplasm. At the 11.5th to the 13.5th day of gestation, the interstitial and endovascular invasive endopolyploid TGTCs (tertiary giant trophoblast cells) prove the intensive cytokeratin staining throughout the cytoplasm and its sprouts. Meantime, the TGTCs were glycogen negative. By contrast, glycogen was heavily accumulated in the glycogen cells that belong both to the junctional zone of placenta and the cuff of the central arterial channel underlying the monolayer of endovascularly invading TGTCs. Thus, the TGTCs that are first to penetrate into the depth of the uterine wall do not contain glycogen but are accompanied by the glycogen-rich cells. The SGTC also contained the prominent deposits of glycogen at the periphery of cytoplasm and in the cytoplasmic sprouts. At the 16th day of gestation, an extensive interstitial invasion of the cytokeratin-positive glycogen trophoblast cells from the junctional zone was observed. The patterns of cytokeratin and glycogen intracellular localization are specific for each subtype of the rat trophoblast; that is, most probably, accounted for by the functional diversity of different trophoblast populations, i.e. patterns of invasion/phagocytosis and their involvement in a barrier at the feto-maternal interface.     

New genetic maps for globe artichoke and wild cardoon and their alignment with an SSR-based consensus map

An F1 mapping population was bred by crossing an accession of wild cardoon with a single Argentinian globe artichoke plant of the variety Estrella del Sur FCA with a view to generating new Cynara cardunculus linkage maps. Genotyping was conducting using a set of 553 SRAP, SSR, AFLP and SNP markers. The 1,465.5 cM map based on the segregation of alleles present in the wild cardoon parent comprised 214 loci distributed across 16 linkage groups (LGs), while the 910.1 cM globe artichoke-based map featured 141 loci falling into 12 LGs covering the total length. Three of the morphological traits (head spininess, leaf spininess and head color) for which the parents contrasted were inherited monogenically, and the genes conditioning them were mapped. A set of 48 co-dominant loci was used to align the LGs with those derived from a reference SSR-based consensus map of the species.     

Tree-growth responses across environmental gradients in subtropical Argentinean forests

Subtropical forests in montane ecosystems grow under a wide range of environmental conditions. However, little is known about the growth responses of subtropical trees to climate along ecological gradients. To assess how, and to what extent climate controls tree growth, we analyzed tree responses to climate for 15 chronologies from 4 different species (Schinopsis lorentzii, Juglans australis, Cedrela lilloi, Alnus acuminata) across a variety of environments in subtropical forests from northwestern Argentina (22–28°S, 64–66°W). Using correlation and principal component analysis, site and species differences in tree-growth responses to precipitation and temperature were determined along the elevation gradient from the dry-warm Chaco lowlands to the wet-cool montane Yungas. Our results show that species responses differ according to the severity in climate conditions along the elevation gradient. At sites with unfavorable conditions, mainly located at the extremes of the environmental gradient, responses of different species to climate variations are similar; in contrast, at sites with relatively mild conditions, tree growth displays a large variety of responses reflecting differences in both local environmental conditions and species physiology. Our research suggests that individualistic responses to environmental variability would determine differences in the type and timing of the responses of dominant trees to climate, which ultimately may shift species’ assemblages in montane subtropical regions of South America under future climate changes.