Interspecies comparison of cellular localization of the cyanide metabolizing enzyme rhodanese within olfactory mucosa

The observation of high levels of xenobiotic metabolizing enzyme activity in the olfactory mucosa has produced speculation on the functional significance of these enzymes in the nose. Hypothesized roles include protection of the nasal epithelium, lung, and other downstream tissues, and termination or modification of olfactory responses. The enzyme rhodanese metabolizes cyanide, which is a commonly inhaled toxicant and an odorant and therefore of interest to both toxicologists and olfactory neurobiologists. The cellular localization of this enzyme within the olfactory mucosa will have important consequences for its ability to protect specific cells, as well as its ability to alter the concentration of inhaled cyanide at receptors, and therefore could provide clues as to its function in this tissue. We have compared the distribution of this enzyme in two species, the rat and the cow, using immunohistochemical localization techniques employing species-specific polyclonal antisera raised in our laboratory. In the rat, rhodanese-like immunoreactivity was greatest within the apical portion of the sustentacular cells, the basal cells, and the duct cells of Bowman's glands. Very little to no reaction was observed in the acinar cells of Bowman's glands. In the cow, however, the acinar cells and duct cells of Bowman's glands showed intense immunoreactivity with little to no reaction observed in the sustentacular or basal cells. The differences in localization of rhodanese in these two species may have important implications for cell types at risk during inhalation of cyanide or organonitrile compounds metabolized to cyanide within the nasal mucosa. In addition, the difference in distribution in the two species emphasizes the importance of considering enzyme activity and localization in the determination of an appropriate animal model for study of both nasal toxicology and olfactory responsiveness in humans.     

Distribution of the exfoliative toxin D gene in clinical Staphylococcus aureus isolates in France

Exfoliative toxin D (ETD) was identified recently as a new exfoliative toxin serotype. Like other exfoliative toxins, ETD induces intra-epidermal cleavage through the granular layer of the epidermis of neonatal mice. The distribution of ETD production was investigated in Staphylococcus aureus isolates from infected and colonised patients in France. The etd gene was found in 55 (10.5%) of 522 isolates tested. Isolates responsible for bullous impetigo and generalised staphylococcal scalded skin syndrome did not harbour etd, but etd was significantly more frequent in isolates causing cutaneous abscesses and furuncles. Most etd- and Panton-Valentine leukocidin-positive strains belonged to the clone of community-acquired methicillin-resistant S. aureus spreading currently throughout France.     

Assessing Alzheimer severity with a global clinical scale.

Diagnosis of dementia needs to be complemented by precise determination of disease severity across the broad spectrum of disease progression. The Mini-Mental State Exam (MMS), the Activities-of-Daily-Living assessment (ADL) and the Clinical Dementia Rating scale (CDR) were modified for direct comparability and administered to 112 outpatients and 45 nursing home residents with a range of dementia severity from mild to profound. The scales showed the highest correlations for the probable Alzheimer's disease patient group (62) (Global Assessment of Dementia; GAD vs. ADL: r = 0.91; Extended Mini-Mental Assessment; EMA vs. GAD: r = 0.91; ADL vs. EMA: r = 0.86). For these patients, scores on the individual scales tended to be similar. Disparity among the three scores for individual cases was associated with the presence of comorbidities. The high correlations and correspondence among these scales demonstrate their reliability, validity, and utility in the assessment of dementia severity. The use of an average of these measures, with their increased precision, may give a more accurate indication of dementia severity over a broader range of impairment.     

Indomethacin-Dipalmitoylphosphatidylcholine Interaction. A Calorimetric Study of Drug Release from Poly(Lactide-co-glycolide) Microspheres into Multilamellar Vesicles

A comparative study of indomethacin controlled release from poly(lactide-co-glycolide) (50:50, molecular weight 3000) (PLGA) microspheres loaded with two different amounts of drug (10.9 ± 1%, and 34.1 ± 1% w/w) and pure free indomethacin, considering the effects exerted by the drug on the thermotropic behavior of dipalmitoylphosphatidylcholine multilamellar vesicles, was carried out by differential scanning calorimetry (DSC). The release was monitored by comparing the effect exerted by the free indomethacin on lipid thermotropic behavior with that of the drug released by the microspheres and relating these effects to a lipid aqueous dispersion containing the molar ratio of drug able to cause it. By DSC measurements, the pure free indomethacin was found to be able to have a fluidifying effect on the model membrane, causing a shift toward lower values of the transitional temperature (T_m), characteristic of phospholipid liposomes, without variations in the enthalpic changes (ΔH). This shift was found to be modulated by the drug molar fraction with respect to the lipid concentration in the aqueous dispersion. Successively, calorimetric measurements were performed on suspensions of blank liposomes added to weighed amounts of unloaded and indometha-cin-loaded microspheres as well as free powdered indomethacin, and the T_m shifts of the lipid bilayer caused by the drug released from the polymeric system, as well as by the free drug, were compared with that caused by free drug increasing molar fractions dispersed directly on the membrane, employed as a calibration curve to obtain the fraction of drug released. This drug release model could be employed to determine the different kinetics involved in the drug transfer from the microspheres to a membrane. This in vitro study suggests that the kinetic process involved in drug release is influenced by the amount of drug loaded in the microspheres. This calorimetric study shows that the PLGA microspheres are a good delivery system able to sustain drug release. Moreover, the DSC technique applied to the drug interaction with biomembranes constitutes a good tool for determining the drug release representing an innovative alternative in vitro model.     

Early embryonic expression patterns of the mouse Flamingo and Prickle orthologues.

The Drosophila melanogaster proteins Flamingo and Prickle act in the planar cell polarity (PCP) pathway, which is required for acquisition of epithelial polarity in the wing, eye, and epidermis. In mammals, PCP signaling has been shown to regulate cell movements and polarity in a variety of tissues. Here, we show that the murine Flamingo orthologues Celsr1-3 and the Prickle orthologues Prickle1, Prickle2, and Testin have dynamic patterns of expression during pregastrulation and gastrulation stages. Celsr1 is expressed in the anterior visceral endoderm and nascent mesoderm, Celsr2 and Celsr3 mark the prospective neuroectoderm, Prickle1 is expressed in the primitive streak and mesoderm, Prickle2 in the node, and Testin in the anterior visceral endoderm, the extraembryonic ectoderm, primitive streak, and mesoderm. Analysis of a gene-trap mutation in Testin indicates that this gene is not required for embryogenesis; therefore, other Prickle homologues may compensate for its function during development.     

Multicontrast MRI of remyelination in the central nervous system

Although magnetic resonance imaging (MRI) represents the most sensitive tool for the detection of white matter abnormalities in patients with multiple sclerosis (MS), the heterogeneity of MS placques severely hampers the elucidation of specific pathophysiological processes. In order to identify putative MRI markers for de- and remyelination, we employed the cuprizone mouse model which leads to a selective and reversible demyelination of the corpus callosum with little or no axonal damage. Apart from histopathology, animals were studied with high-resolution three-dimensional MRI in vivo using multiple contrasts. While individual MRI findings significantly correlated with electron microscopy, the differentiation of regions with normal, demyelinated or remyelinated white matter by one contrast alone was less specific than by histology or electron microscopy. However, an accurate MRI prediction of the in vivo myelin status was achieved by a discriminant function analysis using a combination of T1, T2 and magnetization transfer contrast. With a correct assignment of 95% of all animals examined, the procedure will allow for the survey of new therapeutic approaches aiming at improved remyelination.     

Frequent carriage of Panton-Valentine leucocidin genes by Staphylococcus aureus isolates from surgically drained abscesses

Between 1 February and 15 April 2002, 95 patients were admitted to Gaston Bourret Territorial Hospital (New Caledonia, France) for drainage of community-acquired soft tissue abscesses. Staphylococcus aureus was detected in 68 cases (72%). Two-thirds of the patients with S. aureus infection had furuncles, which were located on the limbs in 82% of cases. The median interval between symptom onset and hospital admission was 5.7 days. Three-quarters of the patients were Melanesians living in tribes. Fifty-four S. aureus isolates were screened for toxin genes. Panton-Valentine leucocidin (PVL) genes were detected in 48 isolates (89%), the exfoliative toxin A gene was detected in 1 isolate, and no toxin genes were detected in 4 isolates. S. aureus nasal carriage was detected in 39.7% of patients with S. aureus infections. Two infecting S. aureus strains and two nasal carriage strains were resistant to methicillin. Comparative pulsed-field gel electrophoresis, performed in 16 cases, showed that five of six patients with PVL-positive nasal carriage strains were infected by the same strains. In contrast, 8 of 10 patients with PVL-negative nasal carriage strains were infected by PVL-positive strains. PVL genes thus appear to be a major virulence factor in both primary and secondary S. aureus skin infections.     

Stimulation and proliferation of CD4+ peripheral blood T lymphocytes induced by an anti-CD4 monoclonal antibody

There is experimental evidence that the CD4 molecule participates in the antigen-driven activation of T cells expressing this surface glycoprotein. Whether CD4, a member of the immunoglobulin supergene family, acts as a ligand-binding molecule and/or is directly involved in the activation pathway has yet to be established. In this study, we show that human CD4⁺ lymphocytes can be activated by exposure to the anti-CD4 monoclonal antibody (mAb) B66. Normal peripheral blood CD4⁺ cells were induced to proliferate and to synthesize interleukin 2 (IL2) by the antibody. The specificity of the antibody stimulatory activity was tested by using IL2-producing clones bearing either CD4 or CD8 on their surface. IL2 production was induced by mAb B66 in CD4+, but not CD8⁺, clones, whereas both types of clones responded to stimulation by the anti-CD3 mAb Leu-4. Despite its unique stimulatory activity, mAb B66 shared with other anti-CD4 antibodies the ability to inhibit the specific cytolytic activity of CD4⁺ effector cells. These results clearly indicate that cross-linking of surface CD4 molecules with appropriate antibodies can fully activate CD4⁺ lymphocytes. Whether the natural ligand for CD4 can trigger this activation pathway remains to be defined.     

Regional mapping of the gene encoding dihydroorotate dehydrogenase,an enzyme involved in UMP synthesis,electron transport,and superoxide generation,to human chromosome region 16q22

De novo UMP synthesis is a critical metabolic pathway for nucleic acid synthesis and for a variety of metabolic pathways. The pathway is a target for many widely used cancer chemotherapy agents, several of which are pyrimidine analogs. Humans and cattle have been described with mutations in UMP synthesis that lead to serious inborn errors of metabolism. Dihydroorotate dehydrogenase (EC 1.3.3.1) (DHODH) carries out the fourth committed step in the pathway and may also be important for mitochondrial electron transport and oxygen radical metabolism. We report here that the gene encoding this enzyme in humans is located in the chromosomal region 16q22. With the mapping of DHODH, the mapping of all the steps of UMP synthesis is complete. All three genes involved map to different human chromosomes. This information is important in consideration of regulation of UMP synthesis in mammals, including humans.