Comparison of rat and guinea pig as sources of the S9 fraction in the salmonella/mammalian microsome mutagenicity test

A highly significant enhancement of mutagenicity occurs with 11 polycyclic aromatic hydrocarbons when 3-methylcholanthrene-induced guinea pig liver S9 is substituted for Aroclor-induced rat liver S9 in the Ames test. The use of MC-induced guinea pig liver S9 is particularly valuable for detecting the weak mutagenicity of benz[c]acridine, which is barely positive in a standard Ames assay. However, anthracene and phenanthrene, which are generally considered not to be carcinogens, remain non-mutagenic for strain TA100. This enhancement of mutagenicity does not correlate with arylhydrocarbon hydroxylase activities of the various liver preparations and does not apply to certain other non-PAH mutagens, including β-naphthylamine, aflatoxin B₁ and 4-dimethylaminoazobenzene.     

Rhizophagus irregularis MUCL 41,833 Association with Green Cuttings of Prunus sp. Rootstocks

Journal of Plant Growth Regulation - Sour cherry ‘Latvijas Zemais’ (Prunus cerasus) is a promising dwarfing rootstock for sweet cherries in Latvia, but low growing rate of newly...     

Osteocalcin does not influence acute or chronic inflammation in human vascular cells

Some human observational studies have suggested an anti-inflammatory role of osteocalcin (OCN). An inflammatory protocol using interferon-γ and tumor necrosis factor-α (10 ng/ml) was employed to examine the acute (24 hr) and chronic (144 hr) effects of uncarboxylated OCN (ucOCN) in commercial, primary, subcultured human aortic endothelial cells (HAEC), and human smooth muscle cells (HASMCs). The inflammatory protocol increased phosphorylation of intracellular signaling proteins (CREB, JNK, p38, ERK, AKT, STAT3, STAT5) and increased secretion of adhesion markers (vascular cell adhesion molecule-1, intracellular adhesion molecule-1, monocyte chemoattractant protein-1) and proinflammatory cytokines (interleukin-6 [IL-6], IL-8). After acute inflammation, there were no additive or reductive effects of ucOCN in either cell type. Following chronic inflammation, ucOCN did not affect cell responses, nor did it appear to have any pro- or anti-inflammatory effects when administered acutely or chronically on its own in either cell type. Additionally, ucOCN did not affect lipopolysaccharide (LPS)-induced acute inflammation in HAECs or HASMCs. The findings of this study do not support a causal role for OCN within the models of vascular inflammation chosen. Further confirmatory studies are warranted.     

The effects of Eucalyptus terpenes on hepatic cytochrome P450 CYP4A, peroxisomal Acyl CoA oxidase (AOX) and peroxisome proliferator activated receptor alpha (PPARalpha) in the common brush tail possum (Trichosurus vulpecula)

Eucalyptus leaves contain a high proportion of essential oils comprising of a complex mixture of monoterpenes such as 1,8-cineole, alpha-pinene, d-limonene and p-cymene. In this study, hepatic levels of microsomal lauric acid hydroxylase and peroxisomal cyanide-intensive palmitoyl coenzyme A oxidative activities were examined in livers of possums given an artificial diet consisting of the above monoterpenes for 10 days. These values were compared with those of possums fed a control diet containing only fruits and cereals. Peroxisomal cyanide-intensive palmitoyl coenzyme A oxidative activity was significantly higher in livers of treated possums relative to that of control possums (2.96+/-0.93 vs. 0.98+/-0.88 nmol/mg protein per min, P<0.01) (mean+/-S.D., n=4). A small increase in microsomal lauric acid hydroxylase activity was observed in the treated possum in comparison with the control group (4.40+/-0.85 vs. 3.60+/-0.48 nmol/mg protein per min) (mean+/-S.D., n=4). A higher NAD/ NADP ratio was observed in treated possums as compared with control possums (4.73+/-0.65 vs. 3.51+/-0.64 nmol/mg protein per min, P<0.05) (mean+/-S.D., n=4). No other statistically significant differences in pyridine nucleotide contents were found between control and treated possums. Northern blot analysis of mRNA from rat, human, terpene treated and control possum livers, using the corresponding koala cDNA probes, detected a more intense acyl CoA oxidase (AOX) mRNA band in livers of terpene fed possums. Negligible differences in the intensity of CYP4A and PPARalpha mRNA bands were observed between the two groups. These data suggest that Eucalyptus terpenes elevate hepatic AOX expression in possums.     

Geldanamycin Enhances Retrograde Transport of Shiga Toxin in HEp-2 Cells

The heat shock protein 90 (Hsp90) inhibitor geldanamycin (GA) has been shown to alter endosomal sorting, diverting cargo destined for the recycling pathway into the lysosomal pathway. Here we investigated whether GA also affects the sorting of cargo into the retrograde pathway from endosomes to the Golgi apparatus. As a model cargo we used the bacterial toxin Shiga toxin, which exploits the retrograde pathway as an entry route to the cytosol. Indeed, GA treatment of HEp-2 cells strongly increased the Shiga toxin transport to the Golgi apparatus. The enhanced Golgi transport was not due to increased endocytic uptake of the toxin or perturbed recycling, suggesting that GA selectively enhances endosomal sorting into the retrograde pathway. Moreover, GA activated p38 and both inhibitors of p38 or its substrate MK2 partially counteracted the GA-induced increase in Shiga toxin transport. Thus, our data suggest that GA-induced p38 and MK2 activation participate in the increased Shiga toxin transport to the Golgi apparatus.     

BCL3 gene role in facial morphology

BACKGROUND: Cleft lip (CL) with or without palate (CLP) and isolated cleft palate (CP) are etiologically complex diseases with interactions among various environmental and genetic factors. The aim of the current study was to identify association with genetic markers and phenotypic craniofacial data in patients with CL/CLP/CP parents. METHODS: Posteroanterior and lateral digital radiographs of the cranium were obtained from 74 parents of patients with CL/CLP/CP. One hundred seventy‐three patients with CL/CLP/CP and 190 controls were enrolled in the study for the association test. Five genetic markers of the IRF6 gene and 14 markers of the 19q13 locus were genotyped. Linear regression analysis was performed for the relationship of cephalometric measurements with genotype data adjusted for age, gender, and cleft type. Chi‐square and transmission disequilibrium tests were performed to evaluate differences in alleles of the BCL3 gene. Positive findings were replicated in an independent sample (n = 95) of patients with CL/CLP/CP parents. RESULTS: Genetic markers of the BCL3 gene at 19q13, rs7257231, and rs1979377 in the familial association test and rs10401176 in the case‐control association test, were associated with craniofacial phenotype. Carriers of BCL3 allele rs7257231T had longer posterior cranial bases than noncarriers (p_adjusted = 0.0028), and in the familial‐based association test showed the statistically strongest relationship (p_adjusted = 0.05) to phenotype. Relation of rs7257231 to facial formation was confirmed in the replication group (p = 0.0024). CONCLUSIONS: The results indicate that BCL3, which has functions related to cell adhesion and whose downregulation can cause disruption of ectodermal development, is likely to be important in facial formation. Birth Defects Research (Part A), 2012. © 2012 Wiley Periodicals, Inc.     

Monitoring the Assembly of a Secreted Bacterial Virulence Factor Using Site-specific Crosslinking

This article describes a method to detect and analyze dynamic interactions between a protein of interest and other factors in vivo. Our method is based on the amber suppression technology that was originally developed by Peter Schultz and colleagues¹. An amber mutation is first introduced at a specific codon of the gene encoding the protein of interest. The amber mutant is then expressed in E. coli together with genes encoding an amber suppressor tRNA and an amino acyl-tRNA synthetase derived from Methanococcus jannaschii. Using this system, the photo activatable amino acid analog p-benzoylphenylalanine (Bpa) is incorporated at the amber codon. Cells are then irradiated with ultraviolet light to covalently link the Bpa residue to proteins that are located within 3-8 Å. Photocrosslinking is performed in combination with pulse-chase labeling and immunoprecipitation of the protein of interest in order to monitor changes in protein-protein interactions that occur over a time scale of seconds to minutes. We optimized the procedure to study the assembly of a bacterial virulence factor that consists of two independent domains, a domain that is integrated into the outer membrane and a domain that is translocated into the extracellular space, but the method can be used to study many different assembly processes and biological pathways in both prokaryotic and eukaryotic cells. In principle interacting factors and even specific residues of interacting factors that bind to a protein of interest can be identified by mass spectrometry.