Foregut glands of Solenogastres (mollusca): anatomy and revised terminology

In the molluscan class Solenogastres, different types of foregut glands vary in number, structure, and location within the foregut. The present article describes their anatomy and cytology and intends to clarify their confused terminology. Pharyngeal glands, esophageal glands, and the more complex dorsal and ventrolateral foregut glands can be distinguished. The ventrolateral foregut glands (ventral foregut glandular organs, ventral salivary glands of auct.), in the literature subdivided previously into four types, are revisited here in the context of current vertebrate gland terminology. The results of recent investigations are added to earlier ones, and a classification system for these multicellular glands is proposed. This system is based on cytological characters of glandular cells (intra- or extraepithelial), characters of the associated musculature (inner or outer musculature), location of the gland relative to the pharynx epithelium (endoepithelial or exoepithelial), characters of the gland openings (paired or unpaired), morphology of the gland duct (simple or branched), and some additional features like the arrangement of glandular cells along the gland ducts. Gross morphology and anatomy of ventrolateral foregut glands constitute useful taxonomic characters in determining higher taxa (family level), and finer details of the anatomy and cytology are useful in determining lower levels (genus and species). Possible pathways for the evolution of the different gland types of Solenogastres in relation to foregut glands present in the other molluscan clades are presented. The importance of ventrolateral foregut gland characters for phylogenetic considerations within the Solenogastres is discussed.     

Warfarin resistance in a French strain of rats

A warfarin-resistant strain and a warfarin-susceptible strain of wild rats (Rattus norvegicus) maintained in enclosures of the National Veterinary School of Lyon (France) were studied to determine the mechanism of the resistance to anticoagulant rodenticides. A low vitamin K epoxide reductase (VKOR) activity has been reported for many resistant rat strains. As recently suggested, mutations in the vitamin K epoxide reductase subunit 1 (VKORC1) gene are the genetic basis of anticoagulant resistance in wild populations of rats from various locations in Europe. Here we report, for our strain, one of the seven described mutations (Tyr139Phe) for VKORC1 in rats. In addition, a low expression of mRNA encoding VKORC1 gene is observed in resistant rats, which could explain their low VKOR activity. We calculated kinetic parameters of VKOR in the warfarin-resistant and warfarin-susceptible rats. The V(max) and the K(m) of the VKOR obtained in resistant rats were lowered by 57 and 77%, respectively, compared to those obtained in susceptible rats. As a consequence, the enzymatic efficiency (V(m)/K(m)) of the VKOR was similar between resistant and susceptible rats. This result could be a good explanation to the observation that no clinical signs of vitamin K deficiency was observed in the warfarin-resistant strain, while a low VKOR activity was found. VKOR activity in warfarin-resistant rats was poorly inhibited by warfarin (K(i) for warfarin is 29 microM and 0.72 microM for resistant and susceptible rats, respectively).     

In vitro degradation of the antimicrobial human peptide HEM-gamma 130-146 in plasma analyzed by a validated quantitative LC-MS/MS procedure

In stability studies during preclinical drug development, the human antimicrobial peptide hHEM-gamma 130-146 shows progressive N-terminal degradation in plasma. To determine this effect, we developed and validated a selective and quantitative muHPLC-MS/MS procedure for this compound. Following deproteinization by precipitation, reversed-phase separation is performed with a time-saving two-column design online coupled to an ion trap mass spectrometer for electrospray ionization MS detection. Using a linear calibration curve obtained with synthetic external standards ranging nearly two orders of magnitude, we achieved good precision (repeatability and reproducibility: 5-15%), accuracy (-3 to 15%), and ruggedness with a lower limit of quantification at 0.29 microg/ml plasma (0.15 microM). Because of good linearity (r2>0.999), the recovery (84+/-3%) and ion suppression (86+/-4% remaining intensity) were calculated from specifically prepared calibration curves. The developed procedure was applied to human and animal plasma samples. Incubations in the presence and absence of proteinase inhibitors revealed at least an aminopeptidase M activity for the initial N-terminal truncation of tryptophan (W130) and a putative glutaminyl-peptide cyclotransferase activity for the resulting intermediate starting with the bared glutamine residue (Q131). The calculated periods of half-change demonstrated exceeding interspecies variations, whereas the intraspecies variations were only between 20 and 30%. The current procedure is valuable as a generic method for pharmaceutical purposes, and data give important information for further development toward a potential natural drug candidate.     

Potato homologs of Arabidopsis thaliana genes functional in defense signaling--identification, genetic mapping, and molecular cloning

Defense against pests and pathogens is a fundamental process controlled by similar molecular mechanisms in all flowering plants. Using Arabidopsis thaliana as a model, steps of the signal transduction pathways that link pathogen recognition to defense activation have been identified and corresponding genes have been characterized. Defense signaling (DS) genes are functional candidates for controlling natural quantitative variation of resistance to plant pathogens. Nineteen Arabidopsis genes operating in defense signaling cascades were selected. Solanaceae EST (expressed sequence tag) databases were employed to identify the closest homologs of potato (Solanum tuberosum). Sixteen novel DS potato homologs were positioned on the molecular maps. Five DS homologs mapped close to known quantitative resistance loci (QRL) against the oomycete Phytophthora infestans causing late blight and the bacterium Erwinia carotovora subsp. atroseptica causing blackleg of stems and tuber soft rot. The five genes are positional candidates for QRL and are highly sequence related to Arabidopsis genes AtSGT1b, AtPAD4, and AtAOS. Full-length complementary DNA and genomic sequences were obtained for potato genes StSGT1, StPAD4, and StEDS1, the latter being a putative interactor of StPAD4. Our results form the basis for further studies on the contributions of these candidate genes to natural variation of potato disease resistance.     

In vitro and ex vivo permissivity of hepatocytes for Leishmania donovani

Using models of ex vivo infection of murine, rat, and human primary hepatocytes by Leishmania donovani, we showed that hepatocytes are permissive for Leishmania at a low level. We then modeled the in vitro infection of a human hepatoma-derived cell line to examine the parasite's capability to proliferate and to cause direct damage to hepatocytes. Results showed that L. donovani can infect hepatocytes, but do not massively proliferate. This slight infection under our experimental conditions resulted in limited damage to hepatocytes. These results bring into question a possible role for hepatocytes as a parasite reservoir during latent infection.     

Trs85 (Gsg1), a component of the TRAPP complexes, is required for the organization of the preautophagosomal structure during selective autophagy via the Cvt pathway

Autophagosomes and Cvt vesicles are limited by two membrane layers. The biogenesis of these unconventional vesicles and the origin of their membranes are hardly understood. Here we identify in Saccharomyces cerevisiae Trs85, a nonessential component of the TRAPP complexes, to be required for the biogenesis of Cvt vesicles. The TRAPP complexes function in endoplasmic reticulum-to-Golgi and Golgi trafficking. Growing trs85delta cells show a defect in the organization of the preautophagosomal structure. Although proaminopeptidase I is normally recruited to the preautophagosomal structure, the recruitment of green fluorescent protein-Atg8 depends on Trs85. Autophagy proceeds in the absence of Trs85, albeit at a reduced rate. Our electron microscopic analysis demonstrated that the reduced autophagic rate of trs85delta cells does not result from a reduced size of the autophagosomes. Growing and starved cells lacking Trs85 did not show defects in vacuolar biogenesis; mature vacuolar proteinase B and carboxypeptidase Y were present. Also vacuolar acidification was normal in these cells. It is known that mutations impairing the integrity of the ER or Golgi block both autophagy and the Cvt pathway. But the phenotypes of trs85delta cells show striking differences to those seen in mutants with defects in the early secretory pathway. This suggests that Trs85 might play a direct role in the Cvt pathway and autophagy.     

Fetal loss in homozygous mutant Norrie disease mice: a new role of Norrin in reproduction

Mutations in the Norrie disease pseudoglioma gene (NDP) are known to cause X-linked recessive Norrie disease. In addition, NDP mutations have been found in other vasoproliferative retinopathies such as familial exudative vitreoretinopathy, retinopathy of prematurity, and Coats disease, suggesting a role for Norrin in vascular development. Here we report that female mice homozygous for the Norrie disease pseudoglioma homolog (Ndph) knockout allele exhibit almost complete infertility, while heterozygous females and hemizygous males are fertile. Histological examinations and RNA in situ hybridization analyses revealed defects in vascular development and decidualization in pregnant Ndph-/- females from embryonic day 7 (E7) onwards, resulting in embryonic loss. Using RT-PCR analyses we also demonstrate, for the first time, the expression of Ndph in mouse uteri and deciduae as well as the expression of NDP in human placenta. Taken together, these data provide strong evidence for Norrin playing an important role in female reproductive tissues.     

Injection of the immuno-modulatory drug alpha-galactosylceramide results in the recruitment of a large population of antigen-presenting cells into the liver of C57BL/6 mice

Injection of the immuno-modulatory drug alpha-galactosylceramide into C57BL/6 mice leads to the already known apoptosis of natural killer T (NKT) cells and to thus far undescribed large changes in the leukocyte populations of the liver. These changes are characterized by the recruitment of neutrophils and that of a population of large monocytic cells. The latter cells display the morphological and immunological features of natural suppressor cells. Their recruitment in the liver depends on the presence of NKT cells, most probably through the local release of cytokines and chemokines by activated NKT cells. We discuss the ubiquitous, long-term effects of alpha-galactosylceramide injection on immuno-pathological processes mediated through the NKT-triggered recruitment of a subset of large macrophages/monocytes.     

Design of new anti-tumor agents interrupting deregulated signaling pathways induced by tyrosine kinase proteins. Inhibition of protein-protein interaction involving Grb2

Cellular signaling pathways induced by growth-factor receptors are frequently deregulated in cancer. Anti-tumor agents that inhibit their enzymatic tyrosine kinase activity have been designed and are now used in human chemotherapy. We propose here an alternative way to interrupt over-expressed signaling by inhibiting protein-protein interactions that involve either the over-expressed proteins or proteins located downstream. The adaptor protein Grb2 over-expressed in connection with HER2/ErbB2/neu in Ras signaling pathway was chosen as a target. Peptides with very high affinity for Grb2 were rationally designed from structural data. Their capacity to interrupt the signaling pathway, their anti-proliferative activity as well as their potential anti-tumor properties are described.