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P Roussel

Publications and source records attributed to P Roussel.

At least 73 records · Page 4Linked to original sources
PubMed1994

Quantification of Ag-NOR proteins using Ag-NOR staining on western blots.

Ribosomal genes are associated with a set of silver-stained nucleolar proteins, the Ag-NOR proteins, whose amount is directly related to the duration of the cell cycle. Quantification of Ag-NOR proteins by image analysis is presently used to evaluate the rate of proliferation of cancer cells and nucleolar activity. Our objective was to establish a procedure to quantify independently each major Ag-NOR protein in cell extracts. Computerized densitometry established that the specific silver staining of Ag-NOR proteins (Ag-NOR staining) performed on Western blots makes it possible to quantify Ag-NOR proteins. Using purified Ag-NOR proteins, nucleolin, and protein B23, we observed that the intensity of Ag-NOR staining is proportional to the amount of protein. A linear relationship exists between the intensity of Ag-NOR staining and the amount of nucleolin, in the range of 0.2-1.6 micrograms. Using total nuclear extracts prepared from mammalian cells, the proportionality was maintained for total Ag-NOR-stained proteins or for a particular protein. We also determined the levels of nuclear proteins suitable for quantitative analysis. Individual Ag-NOR proteins can be quantified by computerized densitometry in nuclear extracts after Ag-NOR staining on Western blots. This procedure can be applied to establish the contribution of each Ag-NOR protein in general staining, estimate the variability of each Ag-NOR protein in normal and pathological conditions, and quantify each Ag-NOR protein contained per cell.

Animals↗
PubMed1993

[Nociceptive flexion reflex of the leg. Use after surgical treatment of herniated disk].

In patients suffering from acute sciatica due to herniated disk Lasegue's manoeuvre inhibits the leg flexion nociceptive reflex (RIII reflex) in either the ipsilateral or the contralateral limb. This is thought to be due what is called "diffuse noxious inhibitor control" modulating the activity of a group of convergent spinal neurons. The purpose of this study was to determine whether surgical resection of the herniated disk in patients suffering from sciatica might relieve this so-called "counter-irritation". Among 10 patients whose sciatica resulted from herniated disk, the decrease in amplitude of the RIII response observed when Lasegue's manoeuvre is applied disappeared in 7 patients, 6 to 8 days after surgical resection of the herniated disk. These results suggest that this test might be a useful tool for postoperative prediction of radicular compression.

Adult↗
PubMed1993

Isolation and structural characterization of novel sialylated oligosaccharide-alditols from respiratory-mucus glycoproteins of a patient suffering from bronchiectasis.

The carbohydrate chains of the respiratory-mucus glycoproteins of a patient (blood group O) suffering from bronchiectasis due to Kartagener's syndrome, were released by alkaline borohydride treatment of a pronase digest. The structures of 82 neutral and low-molecular-mass sialylated oligosaccharides have been described previously [van Kuik A., de Waard P., Vliegenthart J. F. G., Klein A., Carnoy C., Lamblin G. Roussel P. (1991) Eur. J. Biochem. 198, 169-182]. In the present work, medium-size sialylated oligosaccharides were obtained after ion-exchange chromatography and were subsequently separated into 36 fractions utilizing gel filtration, HPLC on normal-phase alkylamine-bonded silica and reverse-phase HPLC. From these fractions, the following six sialylated hepta- and octa-saccharide-alditols have been characterized by employing 500-MHz 1H-NMR spectroscopy, in conjunction with fast-atom-bombardment mass spectroscopy and methylation analysis. [formula: see text]

Bronchiectasis↗
PubMed1993

Altered carbohydrate composition of salivary mucins from patients with cystic fibrosis and the adhesion of Pseudomonas aeruginosa.

We compared the chemical composition of salivary mucin glycopeptides from cystic fibrosis (CF) and from non-CF subjects and the adhesion of Pseudomonas aeruginosa to these different salivary glycopeptides. Three pools of CF saliva, four pools of non-CF saliva, one individual CF saliva, and one individual non-CF saliva were studied. The soluble fraction of the saliva was treated with pronase, and gel filtration was performed to obtain high and low molecular mass salivary mucin glycopeptides. The yield of total glycopeptides was significantly higher from CF than from non-CF saliva. Furthermore, the chemical composition revealed a significantly higher sialic acid content in CF than in non-CF mucin glycopeptides, and higher sulfate and fucose content in CF than in non-CF high molecular mass glycopeptides. We studied the adhesion of a nonmucoid strain of P. aeruginosa (1244), its nonpiliated isogenic derivative, and a mucoid strain (M35) to salivary mucin glycopeptides from patients with CF and from non-CF subjects. The three strains bound significantly more to the CF salivary glycopeptides than to the corresponding non-CF salivary glycopeptides. The nonpiliated isogenic mutant of P. aeruginosa 1244 also bound to CF salivary glycopeptides, suggesting that the adhesion of P. aeruginosa could involve nonpilus adhesions. Furthermore, neuraminidase treatment of CF glycopeptides decreased the adhesion of P. aeruginosa 1244. Altogether these results suggested that differences in mucins may in part explain the specificity of P. aeruginosa for CF.

Bacterial Adhesion↗
PubMed1993

Localization of the RNA polymerase I transcription factor hUBF during the cell cycle.

Autoantibodies directed against nucleoli that recognized a doublet of 97-94 kDa in HeLa nuclear protein extracts were identified. The two polypeptides bound equal amounts of antibody, and each was recognized by antibodies affinity purified using the other polypeptide. These antigens were localized in the secondary constriction of PtK1 cells, i.e. the nucleolar organizer regions (NORs) where ribosomal genes accumulate. They were observed in human cells in the same sites as the NOR-silver-stained proteins. The molecular mass of the antigens, their characteristics in Western blotting and their localization in nucleoli and NORs during mitosis are consistent with them being RNA polymerase I transcriptional factor, UBF. This identification was confirmed on Western blotted proteins by their identical labelling patterns, using these autoantibodies and an anti-mUBF antibody that had been previously described. We obtained definitive evidence that these autoantibodies recognize UBF by the strong positive labelling of purified hUBF (1 to 4 ng). During interphase, these autoantibodies directed against UBF labelled in a folded filament pattern as small beads that may correspond to individual transcriptional units. In electron microscopy, the antibodies were observed in the dense fibrillar component (DFC) of the nucleoli and at the periphery of the fibrillar centers (FCs). At the end of G2 phase, transcription inactivation was concomitant with the gathering of UBF at mitotic NORs. UBF was not equally distributed between NORs in human cells: some NORs scored negative (2 to 4) and the intensity of labelling of positive NORs (6 to 8) differed. In confocal microscopy, 3-dimensional analysis of mitosis indicated that UBF remained associated with NORs during all mitotic stages and that there was equal partition of UBF between the daughter cells. The relationship between proteins associated with the NORs and ribosomal gene transcription is discussed.

Antibodies, Antinuclear↗
PubMed1993

[Specificity of bronchopulmonary infection in cystic fibrosis].

Chronic lung colonization in CF is specific and due mainly to Pseudomonas aeruginosa. Since no phagocytic or immunological abnormalities seem to be responsible for this specificity, it is possible that post-translational alterations of respiratory mucins biosynthesis occur and modify the affinity of mucins for Pseudomonas aeruginosa.

Bacterial Infections↗
PubMed1992

Separation of mucin oligosaccharide-alditols by high performance liquid chromatography on alkylamine-bonded silica columns. Effects of structural parameters.

Forty five oligosaccharide-alditols, purified after reductive beta-elimination of human bronchial mucins, were analyzed by HPLC on alkylamine-bonded silica column (Lichrosorb-NH2). The comparison of their structural features and retention times permitted the extension of some previous findings. The chromatographic behavior of the oligosaccharide depends more on the accessibility of oligosaccharide hydroxyl groups than on the sugar composition. The use of this type of fractionation is very efficient for low-molecular-mass oligosaccharide-alditols but needs to be completed by a second chromatographic step for higher molecular-mass oligosaccharide-alditols.

Alkylation↗
PubMed1992

The broad diversity of neutral and sialylated oligosaccharides derived from human salivary mucins.

Mucin glycopeptides were prepared from the salivary mucins of 20 healthy donors with blood group O. The carbohydrate chains of the high-molecular-weight mucins were released by alkaline borohydride treatment. Neutral and monosialylated oligosaccharide-alditols were purified by ion-exchange chromatography, gel filtration, and HPLC. The structures of the oligosaccharide-alditols were determined by high-resolution 1H-NMR spectroscopy in combination with fast atom bombardment mass spectrometry and methylation analysis. Thirty-seven oligosaccharide-alditols were characterized and illustrate the extreme diversity of the salivary mucins carbohydrate chains. This diversity might represent a mosaic of bacterial adhesion sites and be involved in the early events of the nonimmune defense of the oral cavity. Among these 37 oligosaccharide-alditols, 31 have not been previously described in human saliva and five of these are novel structures: [formula: see text]

Carbohydrate Conformation↗
PubMed1992

Nucleolin is an Ag-NOR protein; this property is determined by its amino-terminal domain independently of its phosphorylation state.

The Ag-NOR proteins are defined as markers of "active" ribosomal genes. They correspond to a set of proteins specifically located in the nucleolar organizer regions (NORs), but have not yet been clearly identified. We adapted the specific detection method of the Ag-NOR proteins to Western blots in order to identify these proteins. Using a purified protein, Western blots, and immunological characterization, the present study brings the first direct evidence leading to the identity of one Ag-NOR protein. We found that nucleolin is specifically revealed by Ag-NOR staining. Using different nucleolin fragments generated by CNBr cleavage and by overexpression in Escherichia coli, we demonstrate that the amino-terminal domain of nucleolin and not the carboxy-part of the protein is involved in silver staining. Moreover, as the pattern of staining does not vary using casein kinase II- and cdc2-phosphorylated nucleolin or dephosphorylated nucleolin, we conclude that the reduction of the silver ions is not linked to the phosphorylation state of the molecule. We propose that the concentration of acidic amino acids in the amino-terminal domain of nucleolin is responsible for Ag-NOR staining. This hypothesis is also supported by the finding that poly L-glutamic acid peptides are silver stained. These results provide data that can be used to explain the specificity of Ag-NOR staining. Furthermore, we clearly establish that proteolysis of the amino-terminal Ag-NOR-sensitive part of nucleolin occurs in vitro, leading to the accumulation of the carboxy-terminal Ag-NOR-negative part of the protein. We argue that this cleavage occurs in vivo as already proposed, bearing in mind that nucleolin is present in the fibrillar and in the granular component of the nucleolus, whereas no Ag-NOR staining is observed in the latter nucleolar component.

Amino Acid Sequence↗
PubMed1992

A comparative study of controlled-release morphine (CRM) suspension and CRM tablets in chronic cancer pain.

This study compared the efficacy and the adverse effects of controlled-release morphine (CRM) suspension (SAR 213) and CRM tablets (Moscontin) in the treatment of cancer pain. This multicenter, randomized, double-blind, double-dummy, crossover study was carried out on 52 patients. Each patient received both study treatments given at an equivalent dosage of morphine during each of two 7-day periods. The primary outcome variable was the severity of pain assessed three times daily by means of a visual analogue scale. Secondary criteria of efficacy were the severity of pain assessed by verbal rating scale, the need for "rescue" doses of immediate-release morphine, treatment preference, and indices of quality of life (activity, mood, sleep). There were no statistically significant differences in the parameters assessed when comparing the two groups. This study shows that, when prescribed at the same doses, CRM suspension and CRM tablets have similar efficacy and adverse effects, as well as the same duration of action. The results of this first clinical study carried out on CRM suspension are especially relevant for patients with cancer pain who have difficulty swallowing.

Delayed-Action Preparations↗
PubMed1992

The binding of amikacin to macromolecules from the sputum of patients suffering from respiratory diseases.

Antibiotics bind to whole cystic fibrosis sputum, however, the composition of sputum varies from one patient to another, making the interpretation of binding studies difficult. This problem has been examined by standardizing the macromolecule concentration of sputum from 13 patients suffering from cystic fibrosis, chronic bronchitis or bronchiectasis and adding amikacin to the sputum components. Also binding to fractions of sputum was studied before and after treatment with DNase or chondroitinase. These studies indicated that the degree of amikacin binding is dependent on the DNA concentration and the presence of acidic mucins.

Amikacin↗
PubMed1992

Procedures for specific detection of silver-stained nucleolar proteins on western blots.

Nucleolar organizer region (NOR)-silver staining of the chromosomes and nucleoli is a method that enables the detection of proteins associated with the ribosomal genes. We adapted the most commonly used cytochemical NOR-silver staining techniques to Western-blotted proteins of HeLa cells, mimicking the silver staining of cells in situ, and testing several parameters that may influence the in situ reaction. Two of these techniques, both one-step methods with colloidal developers, were standardized to obtain reproducible results. The specificity of NOR staining is documented by: (a) only a few bands are revealed among the many proteins detected by total proteins staining on gels or blots; two major groups of bands are found around 100 KD and 40 KD that could correspond at least in part to nucleolin and B23 nucleolar proteins; (b) the silver staining of bands was not the result of the high relative protein concentrations; and (c) the same number of NOR-silver-stained bands was observed across a large range of protein concentrations. The reaction appeared to be specific for a subset of nucleolar proteins, because the same bands were observed with the use of nucleolar, nuclear, or total cell protein extracts, and the silver grains observed in electron microscopy were clearly confined to the nucleolar fibrillar centers and dense fibrillar component. The efficiency of the reaction was not modified by any of the tested fixative pre-treatments except that involving methanol. The presented standardization of NOR-silver staining on Western blots allows the characterization of the Ag-NOR proteins and their specific regions responsible for silver staining of the nucleolus.

Blotting, Western↗
PubMed1992

Human respiratory mucins.

Human respiratory mucins are secreted by goblet cells and mucous glands of the respiratory mucosa. They consist of a broad family of complex glycoproteins with different peptides, or apomucins, corresponding to several genes located on at least three different chromosomes. Glycosylation, the major posttranslational phenomenon, is responsible for about 70-80% of the weight of mucins: it produces an extraordinary diversity of O-glycosidically linked carbohydrate chains which are expressed as several hundreds of different chains in the mucins of a single individual. The variety of mucin peptides and the diversity of carbohydrate chains probably allows many interactions, especially with microorganisms: this may be an essential factor in the defence of the underlying respiratory mucosa.

Animals↗