Reproducibility of histologic classification of gastric cancer

A panel review of histologic specimens was carried out as part of a multi-centre case-control study of gastric cancer (GC) and diet. Comparisons of diagnoses of 100 GCs by six pathologists revealed agreement in histologic classification for about 70-80% of the cancers. Concordance was somewhat higher when using the Lauren rather than the Ming or World Health Organization classification systems. Histologic types from reading biopsy tissue agreed with those derived from surgical specimens for 65-75% of the 100 tumours. Intra-observer agreement in histologic classification, assessed by repeat readings up to 3 years apart by one pathologist, was 95%. The findings indicate that, although overall concordance was good, it is important to standardise diagnoses in multi-centre epidemiologic studies of GC by histologic type

Perturbation with Intrabodies Reveals That Calpain Cleavage Is Required for Degradation of Huntingtin Exon 1

Background: Proteolytic processing of mutant huntingtin (mHtt), the protein that causes Huntington's disease (HD), is critical for mHtt toxicity and disease progression. mHtt contains several caspase and calpain cleavage sites that generate N-terminal fragments that are more toxic than full-length mHtt. Further processing is then required for the degradation of these fragments, which in turn, reduces toxicity. This unknown, secondary degradative process represents a promising therapeutic target for HD. Methodology/Principal Findings: We have used intrabodies, intracellularly expressed antibody fragments, to gain insight into the mechanism of mutant huntingtin exon 1 (mHDx-1) clearance. Happ1, an intrabody recognizing the proline-rich region of mHDx-1, reduces the level of soluble mHDx-1 by increasing clearance. While proteasome and macroautophagy inhibitors reduce turnover of mHDx-1, Happ1 is still able to reduce mHDx-1 under these conditions, indicating Happ1-accelerated mHDx-1 clearance does not rely on these processes. In contrast, a calpain inhibitor or an inhibitor of lysosomal pH block Happ1-mediated acceleration of mHDx-1 clearance. These results suggest that mHDx-1 is cleaved by calpain, likely followed by lysosomal degradation and this process regulates the turnover rate of mHDx-1. Sequence analysis identifies amino acid (AA) 15 as a potential calpain cleavage site. Calpain cleavage of recombinant mHDx-1 in vitro yields fragments of sizes corresponding to this prediction. Moreover, when the site is blocked by binding of another intrabody, V_L12.3, turnover of soluble mHDx-1 in living cells is blocked. Conclusions/Significance: These results indicate that calpain-mediated removal of the 15 N-terminal AAs is required for the degradation of mHDx-1, a finding that may have therapeutic implications

C2-phytoceramide perturbs lipid rafts and cell integrity in Saccharomyces cerevisiae in a sterol-dependent manner

Specific ceramides are key regulators of cell fate, and extensive studies aimed to develop therapies based on ceramide-induced cell death. However, the mechanisms regulating ceramide cytotoxicity are not yet fully elucidated. Since ceramides also regulate growth and stress responses in yeast, we studied how different exogenous ceramides affect yeast cells. C2-phytoceramide, a soluble form of phytoceramides, the yeast counterparts of mammalian ceramides, greatly reduced clonogenic survival, particularly in the G2/M phase, but did not induce autophagy nor increase apoptotic markers. Rather, the loss of clonogenic survival was associated with PI positive staining, disorganization of lipid rafts and cell wall weakening. Sensitivity to C2-phytoceramide was exacerbated in mutants lacking Hog1p, the MAP kinase homolog of human p38 kinase. Decreasing sterol membrane content reduced sensitivity to C2-phytoceramide, suggesting sterols are the targets of this compound. This study identified a new function of C2-phytoceramide through disorganization of lipid rafts and induction of a necrotic cell death under hypo-osmotic conditions. Since lipid rafts are important in mammalian cell signaling and adhesion, our findings further support pursuing the exploitation of yeast to understand the basis of synthetic ceramides' cytotoxicity to provide novel strategies for therapeutic intervention in cancer and other diseases.This work was supported by Fundacao para a Ciencia e Tecnologia through projects PTDC/BIA-BCM/69448/2006 and PEst-C/BIA/UI4050/2011, and fellowships to A. P. (SFRH/BPD/65003) and F. A. (SFRH/BD/80934/2011), as well as by FEDER through POFC - COMPETE. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Concurrent weed growth suppression with essential oils and species-specific response to fractionated coconut oil

Plant-derived oils are phytotoxic and thus are potential non-synthetic herbicides. The present study investigated the effectiveness of three essential oils (red thyme, clove bud, cinnamon bark) and a vegetable oil, fractionated coconut oil (FCO), used alone or in 2-way essential/FCO mixes for controlling some troublesome weeds at seedling stage. The tested weeds were mugwort (Artemisia vulgaris L.), crown daisy (Glebionis coronaria (L.) Spach), milk thistle (Sylibum marianum (L.) Gaertn.), dense-flowered mullein (Verbascum densiflorum Bertol.), goosegrass (Eleusine indica L. (Gaertn.)), entireleaf morningglory (Ipomoea hederacea (L.) Jacq.), small-flower morningglory (Jacquemontia tamnifolia (L.) Griseb), large crabgrass (Digitaria sanguinalis (L.) Scop.), hemp sesbania (Sesbania exaltata (Raf.) Rydb. ex A.W. Hill). Greenhouse trials included a negative control (non-treated) and a positive control (glyphosate) and were conducted in Sassari (IT) and Auburn (USA). In both locations, all the essential oils used severely injured weeds by 5 days after treatment (DAT), with least or no recovery by plants, with no harvestable plant biomass 20 DAT. FCO provoked more diverse species-specific responses, reducing biomass of crown daisy, large crabgrass, entireleaf morningglory and hemp sesbania compared to the non-treated control while it stimulated the growth of mugwort, dense-flowered mullein, goosegrass, and small-flower morningglory. Milk thistle was the only plant not influenced by FCO compared to the non-treated control. The essential oils mixed with FCO confirmed high phytotoxic effects with a significant improved action when red thyme oil + FCO was applied to goosegrass. Our research confirms the potential herbicidal effect of some phytotoxic essential oils and the potential beneficial growth effect of FCO to some species