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C Aubert

Publications and source records attributed to C Aubert.

At least 55 records · Page 3Linked to original sources
PubMed1993

Comparison of the sensitivities of ribonucleic acid and oligonucleotide probes for in situ detection of Theiler's virus mRNA.

To evaluate the sensitivity of in situ detection of the Theiler's virus genome, we hybridized BHK-21-infected cells with antisense ribo- and oligonucleotide 35S-labeled probes. The sensitivity achieved with the anti-sense 280-nucleotide riboprobe was similar to that obtained with a 93-mer oligonucleotide probe. However, more reproducible and accurate results were obtained with the riboprobe. With long exposure times, the background was higher with the oligonucleotide probe than with the RNA probe. The background was improved by using freshly labeled oligonucleotide probe.

Animals↗
PubMed1993

In vitro hepatic biotransformation of moclobemide (Ro 11-1163) in man and rat.

1. Moclobemide, an inhibitor of monoamine oxidase, shows mixed MAO A/B inhibition in rat, but pure MAO A inhibition in man. This is attributed to a primary amine metabolite which inhibits MAO B in vitro, but which is not detected in human plasma in vivo. A secondary amine metabolite, also present in rat but not human plasma, inhibitors MAO B in vivo but not in vitro. 2. We have studied the biotransformation of moclobemide in vitro, to investigate whether hepatocytes and hepatic subcellular fractions can reproduce the in vivo interspecies differences. 3. Moclobemide was more extensively metabolized by rat liver preparations, compared with man. For example, of an initial 100 nmol, 78 and 25 nmol were metabolized within 24 h by rat and human hepatocytes in primary culture, respectively. 4. Substantial amounts of secondary amine (12.5 nmol) were found with the rat preparation, compared with low amounts (1.5 nmol) from human hepatocytes. Similarly, for the primary amine, 1.5 nmol were formed by the rat hepatocytes compared with trace amount in the human preparations. 5. Identities of the two amines were confirmed by h.p.l.c. cochromatography and negative CI GC-MS. 6. In conclusion, all the in vitro models, but particularly hepatocytes, reflected the metabolism of moclobemide in vivo. Consequently, liver preparations can be used prospectively to screen the selectivity of related development compounds.

Animals↗
PubMed1992

Interleukin-2 in association with increasing doses of interferon-gamma in patients with advanced cancer.

Twenty-seven patients with evaluable metastatic cancer were treated with recombinant interleukin-2 (rIL-2) and escalating doses of interferon gamma (IFN-gamma). rIL-2 was infused over a 15 min period at a constant dose (8 x 10(6) IU/m2/8 h x 5 days first cycle, and 8 x 10(6) IU/m2/12 h x 5 days second and third cycles, with 9 days rest between each cycle). IFN-gamma was started 4 days before each cycle of rIL-2 and was given every other day at a dosage of 1 x 10(6) U/m2 x 3/cycle (four patients), 5 x 10(6) U/m2 x 3/cycle (four patients), 5 x 10(6) U/m2 x 5/cycle (four patients) and 10 x 10(6) U/m2 x 5/cycle (15 patients). Common side effects were fluid retention and hepatic toxicity (27 and 15% grade greater than or equal to 2); one ischemic chest pains and one acute respiratory distress occurred. Toxicities were not greater than those described with high dose rIL-2 alone and were similar in each dose level of IFN-gamma. No patient died from the procedure. Four patients responded, one complete response and three partial responses; all were treated with 25 or 50 x 10(6) U/m2/cycle of IFN-gamma (melanoma, two patients; renal cell carcinoma, one patient; lymphoma, one patient). Further phase II studies at these dosages are justified to precisely define the antitumoral efficacy of this association.

Adolescent↗
PubMed1992

Differentiation of new metastatic variants of B16 melanoma under different culture conditions.

The purpose of this study was to examine the differentiation of variant tumors of the B16 metastatic melanoma when tumors were grown serially under different culture conditions and transplanted into C57BL/6J black mice, lethal yellow Ay/a, albino c/c, and C+/c mutant mice. Morphological and biochemical markers of melanogenesis were examined in cells in culture and in the corresponding tumors. Cellular pigmentation was assessed in terms of the levels of DOPA and 5-S-CD and in terms of tyrosinase activity in the various cell lines and tumors. The observed change from high to low metastatic capacity, which was dependent on culture conditions, appeared to be unrelated to melanogenesis even though changes were observed in the biochemical melanotic phenotype. Overall, tumor cells from spontaneous pulmonary metastases appear to differentiate in ways that are unrelated to the instability of experimental metastatic capacity. The melanotic phenotype in albino c/c and C+/c mice was dependent on the phenotype of the parental tumors. A marked difference was observed between two pigmentation compartments, one of which was stable in the B16 control, while the other was unstable in YB16 and MB16 variant cells and in the tumors derived from them. It appears, therefore, that the metastatic capacity of B16 metastatic variants is changeable and is independent of the unstable melanogenic behavior. The production of metastases and the differentiation of tumors in the present experiments appeared to be related to the genetic background of the mice and the epigenetic metabolic environment of tumors and cells.

Animals↗
PubMed1992

Theiler's virus replication in brain macrophages cultured in vitro.

Infection of the mouse with Theiler's virus is one of the best animal models for the study of multiple sclerosis, a chronic demyelinating disease of the human central nervous system. The identification of the virus target cell(s) is fundamental to an understanding of the viral persistence as well as the inflammation and demyelination observed in the chronic phase of the disease. This paper reports that a small fraction of brain macrophages grown in vitro can be efficiently infected with Theiler's virus without significant cytolytic effect. Viral replication as well as continuous production of infectivity were observed in these cultures.

Animals↗
PubMed1991

Determinants of persistence and demyelination of the DA strain of Theiler's virus are found only in the VP1 gene.

The DA strain of Theiler's virus persists in the central nervous systems of mice and causes chronic inflammation and demyelination. The GDVII strain, on the other hand, causes an acute encephalitis that kills the host in a matter of days. We constructed a series of recombinants between two infectious cDNA clones of the genomes of DA and GDVII viruses. Analysis of the phenotypes of the recombinant viruses yielded the following results. (i) Determinants of persistence and demyelination are found only in the VP1 capsid protein of DA virus. (ii) Whereas the VP1 capsid protein of DA virus is able to fully attenuate the neurovirulence of GDVII virus and to allow the chimeric virus to persist and demyelinate, the VP1 capsid protein of GDVII virus is unable to render DA virus neurovirulent. (iii) The mere attenuation of the neurovirulence of GDVII virus does not allow it to persist and demyelinate.

Animals↗
PubMed1991

Role of the humoral immune response in resistance to Theiler's virus infection.

Theiler's virus, a murine picornavirus, persists in the central nervous system of susceptible strains of mice, causing chronic inflammation and demyelination in the white matter of the spinal cord. Resistant strains, however, clear the virus and do not develop late disease. In this study, we compared the characteristics of T and B lymphocytes in C57BL/6 (resistant) and SJL/J (susceptible) mice 1 week after intracerebral infection. We detected a marked increase of the number of immunoglobulin M (IgM)-secreting cells in the spleens of C57BL/6 detected a marked increase of the number of immunoglobulin M (IgM)-secreting cells in the spleens of C57BL/6 mice (but not in those of SJL/J mice), which correlated with higher levels of serum IgM antiviral antibodies. The role of the humoral response in virus clearance and resistance was demonstrated by a marked decrease in the number of infected spinal cord cells in SJL/J mice after passive transfer of serum from infected C57BL/6 donors. The B-cell response was found to be partly T cell independent. These results suggest an important role of the early humoral immune response in resistance to Theiler's virus-induced disease.

Animals↗
PubMed1991

In situ analysis of proteolipid protein gene transcripts during persistent Theiler's virus infection.

SJL/J mice inoculated intracranially with the DA strain of Theiler's virus exhibit a persistent demyelinating disease of the central nervous system. To investigate the effect of persistent infection of oligodendrocytes on the expression of myelin genes, we analyzed the level of PLP mRNA in infected as well as uninfected oligodendrocytes. This study was performed at the single-cell level using the simultaneous detection of viral antigens by immunocytochemistry and PLP mRNAs by in situ hybridization with 35S-labeled oligonucleotide probes. Our data indicate that viral infection of oligodendrocytes reduces the level of PLP mRNA by about 80%.

Animals↗
PubMed1990

New variants of the B16 melanoma: tumorigenicity and metastatic properties under different culture conditions.

Variants of the B16 melanoma exhibiting markedly different tumorigenic and metastatic potential in Ay/a and a/a C57BL/6J syngeneic mice were investigated and compared, with respect to their relative growth and metastatic potential after culture under different conditions. Over 2 to 6 months of growth, in vitro cells demonstrated a rapid and significant decrease in their ability to form spontaneous lung colonies. Such a decrease depended on the conditions of culture. We suggest that the in vitro environment influences the phenotype of cells with respect to their capacity for spontaneous metastasis. This possibility must not be ignored, when conclusions obtained from studies of established cell lines are extended to cancers.

Animals↗
PubMed1990

Splitting cell adhesiveness into independent measurable parameters by comparing ten human melanoma cell lines.

The concept of cell adhesiveness was analyzed by looking for correlations between the adhesive behavior and measurable biological properties of different cell populations. Ten established lines of melanoma cells were assayed for passive deformability (by micropipet aspiration), active spreading (by measuring the height/diameter ratio after incubation on different surfaces), density and mobility of concanavalin A binding sites (by quantitative analysis of fluorescence microscopic images), spontaneous and concanavalin A-mediated agglutination (by measuring the number of cell conjugates resisting calibrated shearing forces), and binding to glass capillary tubes (with a quantitative assay of binding strength). Forty-four different parameters were thus measured, and each set of determinations was repeated 2 or 3 t at different days on each cell line. Analysis of variance was performed to assess the capacity of each parameter to discriminate between different lines. Correlations between different parameters were studied in order to understand a possible influence of cell intrinsic properties on the behavior of individual cells. The following conclusions were suggested by experimental data 1. Cell spreading ability, resistance to slow deformation within a micropipette and ability to form shear-resistant bonds, are independent properties. It is therefore suggested that different mechanisms rule the cell deformations on time scales of several minutes, tens of seconds, and fractions of a second. 2. Cell spreading ability may effectively influence binding strength only when adhesive stimuli are low, since in this case, cell stiffness is likely to impair the formation of extensive contact areas. 3. Individual cells may display marked heterogeneity within a given population, that emphasizes the danger of using averaged parameters to predict rare events (such as metastasis formation). 4. The most useful parameters to discriminate between different cell lines were, spreading ability and shear-resistant lectin agglutination, and substrate adhesion. It is concluded that cell adhesion is influenced by several measurable cellular properties that may display independent variations. The importance of a given parameter depends on the conditions of bond formation and rupture.

Cell Adhesion↗
PubMed1990

Isolation of a specific cellular mRNA by subtractive hybridization in Theiler's virus persistent infection.

Viruses change the mRNA repertoire of the tissues they infect. They add viral mRNAs and they specifically alter the expression of some host genes. These events can play important parts in pathogenesis. In principle, it should be possible to isolate viral mRNAs and to identify changes in host gene expression using subtractive hybridization. We tested this approach in the persistent infection of mouse central nervous system by Theiler's virus. A cDNA library was constructed with poly A+ RNA from infected mouse spinal cords. The library was screened with a subtracted probe. We identified one mitochondrial gene, coding for subunit 1 of cytochrome oxidase, which is overexpressed in infected tissues whereas another mitochondrial gene, URF 2, is not. Subtractive hybridization should prove to be invaluable in studying the pathogenesis of chronic human central nervous system diseases of unknown etiology.

Animals↗
PubMed1990

Genetic mapping of the ability of Theiler's virus to persist and demyelinate.

Theiler's virus, a murine picornavirus, is responsible for two different types of disease: strains DA, BeAn, and WW persist for more than a year in the white matter of the central nervous system and cause primary demyelination; strains GDVII and FA, on the other hand, cause an acute encephalitis that kills the host in a matter of days. To map the regions of the viral genome responsible for persistence and demyelination, cDNA clones of the entire genomes of the DA and GDVII strains were constructed and cloned into Bluescript plasmid (A. McAllister, F. Tangy, C. Aubert, and M. Brahic, Microb. Pathogen. 7:381-388, 1989; F. Tangy, A. McAllister, and M. Brahic, J. Virol. 63:1101-1106, 1989). We constructed chimeric viruses obtained by exchanging regions between the cDNA clones. Analysis of the disease phenotypes produced by the chimeric viruses allowed us to map persistence and demyelination to a genome segment coding for the VP1 capsid protein and 27 amino acids of protein 2A.

Animals↗
PubMed1990

Simultaneous in situ detection of two mRNAs in the same cell using riboprobes labeled with biotin and 35S.

We used 35S-labeled and biotinylated cRNAs (riboprobes) to detect simultaneously two different mRNAs by in situ hybridization. In a first step we established the conditions under which each type of probe achieved the same high level of sensitivity. We then used these conditions to hybridize BHK cells infected with Theiler's virus, a murine picornavirus, with a mixture of a virus-specific biotinylated riboprobe and a 35S-labeled riboprobe specific for beta-actin mRNA. Both mRNAs could be detected in the same cell, although the sensitivity achieved by the radiolabeled probe was reduced by about 40% by the simultaneous hybridization with the biotinylated probe.

Actins↗
PubMed1990

Calmodulin content and distribution in six human melanoma cell lines.

Calmodulin content and distribution between soluble and particulate fractions were determined by radioimmunoassay in six human melanoma cell lines exhibiting differences in tumor origin (primary or metastatic), degree of tumorigenicity and of pigmentation (amelanotic or melanotic). The results indicate that a) total, soluble and particulate calmodulin levels expressed as ng/10(6) cells or ng/micrograms of proteins remained constant for five out of six cell lines when cells grew from subconfluency to confluency. For IGR 37 line, derived from metastatic melanoma, the calmodulin content decreases from 2.39 to 1.27 ng/micrograms protein for total calmodulin, from 2.17 to 1.52 ng/micrograms protein for soluble calmodulin and from 2.61 to 1.02 ng/micrograms protein for particulate calmodulin, b) total, soluble and particulate calmodulin levels expressed as ng/microgram proteins were twofold (at confluency) to fourfold (at subconfluency) higher in the two cell lines from metastatic origin, IGR 37 and IPC 167. As for example, for total calmodulin, values in IGR 37 and IPC 167 cell lines, were, respectively at subconfluency, 2.39 and 2.31 ng/micrograms protein as compared with the four other cell lines: 0.76 to 0.96 ng/micrograms protein and at confluency: 1.27 and 1.98 ng/micrograms protein as compared with the four other cell lines: 0.76 to 0.90 ng/micrograms protein, c) ratio of calmodulin between soluble and particulate fractions was about 1 for the two autologous cell lines IGR 37 and IGR 39 and varies from 2 to 3 for the four other cell lines.

Calmodulin↗
PubMed1989

Capillary gas chromatographic-mass spectrometric method for the identification and quantification of some benzodiazepines and their unconjugated metabolites in plasma.

A gas chromatographic-mass spectrometric method for the identification and/or quantification of diazepam, clobazam, flunitrazepam, triazolam, midazolam, oxazepam and lorazepam and some of their desmethylated and hydroxylated metabolites in plasma is described. Benzodiazepines were extracted from plasma with butyl acetate at pH 9; the hydroxylated compounds were then silylated with N,O-bis (trimethylsilyltrifluoroacetamide). Analysis was performed using a compact mass-selective detector operating in the electron-impact mode. Depending on the concentration, identification was performed either by direct comparison of the observed mass spectra with reference spectra or by the relative intensities of the most intense and characteristic ions in the selected-ion monitoring (SIM) mode. Quantification was performed in the SIM mode using the most intense ion. The intra-assay precision and accuracy were better than 5-6%; linearity was satisfactory up to 1-2 micrograms/ml. The detection limit was 1-5 ng/ml for most of the benzodiazepines. This method can be easily used in clinical situations when a safe and rapid response is essential for patient treatment.

Benzodiazepines↗
PubMed1989

Direct liquid inlet liquid chromatographic/mass spectrometric identification and high-performance liquid chromatographic analysis of a benzodiazepine glucuronide.

The glucuronide of N-1-hydroxy-ethyl flurazepam has been analysed by a direct liquid inlet liquid chromatographic/mass spectrometric system using MeOH/H2O (70:30 v/v) as mobile phase, at a flow rate of 0.7 ml min-1. Urine samples were purified by amberlite XAD-2 chromatography; the glucuronide was quantified by high-performance liquid chromatography using a counterion (tetrabutyl ammonium nitrate in methanol). Chromatographic results were validated by an enzymatic method: treatment of the samples with beta-glucuronidase and extraction of the parent drug with ethyl ether at pH 9. The biological application of this method was demonstrated by determination of this glucuronide in the urine of healthy human volunteers following a single intravenous administration of 50 mg of N-1-hydroxy-ethyl flurazepam.

Chromatography, High Pressure Liquid↗