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Biomedical subjects

C Philippe

Publications and source records attributed to C Philippe.

At least 37 records · Page 2Linked to original sources
PubMed1997

Eleven X chromosome breakpoints associated with premature ovarian failure (POF) map to a 15-Mb YAC contig spanning Xq21.

Eleven balanced X-autosome translocations associated with premature ovarian failure (POF) were mapped to a YAC contig spanning most of Xq21 and constructed between the DXS223 and DXS1171 loci. The contig corresponds to a genomic region of about 15 Mb and contains the whole X-Y homologous region. The most proximal and most distal breakpoints associated with POF were mapped 15 Mb apart. The remaining breakpoints were localized along this large region, in the X-specific and in the X-Y homologous region. Four of the YACs contained two breakpoints in the same or in flanking STS intervals. Our results confirm the cytological findings and suggest that a minimum number of eight different genes in Xq21 may be involved with ovary development. Interruption of such loci could be the cause of POF.

Carrier Proteins↗
PubMed1997

Pharmacological and biochemical evidence for the simultaneous expression of CCKB/gastrin and CCKA receptors in the pig pancreas.

1. In the pig, the secretory response of the pancreas is not inhibited by the antagonist MK329 suggesting that cholecystokininA (CCKA) receptors are not involved. 2. Membranes were isolated from the pancreas of 6 Large White pigs to characterize their CCK receptors. 3. The binding of [125I]-BH-[Thr, Nle]CCK-9 was dependent on pH, maximal after a 90 min incubation period, saturable and reversible. Saturation analysis of the binding demonstrated a single class of high affinity sites (Kd = 0.22 +/- 0.02 nM) and a binding capacity, Bmax = 110.64 +/- 12.50 fmol mg-1 protein. 4. Competition binding by agonists and antagonists of CCKA and CCKB/gastrin receptors demonstrated the presence of two distinct binding components, sites presenting a high affinity for [Thr, Nle]CCK-9, gastrin, PD 135158, L-365, 260 and a low affinity for MK329, SR 27897, and sites presenting a high affinity for [Thr, Nle]CCK-9, MK329, SR 27897 and a low affinity for gastrin, PD 135158, L-365,260. 5. These pharmacological data demonstrate the presence of both CCKA and CCKB/gastrin receptors in the pig pancreas, the latter being predominant. 6. Two distinct membrane proteins (50 and 85-100 kDa, respectively) display pharmacological features of CCKB/gastrin and CCKA receptors. 7. In pigs, as in calves and humans, CCKB/gastrin receptors are predominant in the pancreas.

Affinity Labels↗
PubMed1997

Mutation screening of the EXT1 and EXT2 genes in patients with hereditary multiple exostoses.

Hereditary multiple exostoses (HME), the most frequent of all skeletal dysplasias, is an autosomal dominant disorder characterized by the presence of multiple exostoses localized mainly at the end of long bones. HME is genetically heterogeneous, with at least three loci, on 8q24.1 (EXT1), 11p11-p13 (EXT2), and 19p (EXT3). Both the EXT1 and EXT2 genes have been cloned recently and define a new family of potential tumor suppressor genes. This is the first study in which mutation screening has been performed for both the EXT1 and EXT2 genes prior to any linkage analysis. We have screened 17 probands with the HME phenotype, for alterations in all translated exons and flanking intronic sequences, in the EXT1 and EXT2 genes, by conformation-sensitive gel electrophoresis. We found the disease-causing mutation in 12 families (70%), 7 (41%) of which have EXT1 mutations and 5 (29%) EXT2 mutations. Together with the previously described 1-bp deletion in exon 6, which is present in 2 of our families, we report five new mutations in EXT1. Two are missense mutations in exon 2 (G339D and R340C), and the other three alterations (a nonsense mutation, a frameshift, and a splicing mutation) are likely to result in truncated nonfunctional proteins. Four new mutations are described in EXT2. A missense mutation (D227N) was found in 2 different families; the other three alterations (two nonsense mutations and one frameshift mutation) lead directly or indirectly to premature stop codons. The missense mutations in EXT1 and EXT2 may pinpoint crucial domains in both proteins and therefore give clues for the understanding of the pathophysiology of this skeletal disorder.

Electrophoresis, Polyacrylamide Gel↗
PubMed1997

Insulin-like growth factors and their binding proteins in pleural fluid.

We investigated the expression and potential regulatory role of insulin-like growth factors (IGFs) and their specific binding proteins (BPs) in tuberculous and nontuberculous pleuritis. By using a radioimmunoassay after acid gel filtration chromatography, we found that mean concentrations of IGF-I were 211.9 +/- 20.2 microg/l and 203.2 +/- 31.1 microg/l in pleural fluid of 14 patients with tuberculous pleuritis and 9 patients with malignant pleuritis respectively. These values were near those in serum of the same patients (221.3 +/- 19.5 microg/l and 204.6 +/- 21.0 microg/l respectively). By using a specific protein-binding assay, we found that mean concentrations of IGF-II were 345.3 +/- 61.0 microg/l and 167.6 +/- 22.7 microg/l in tuberculous and malignant pleural effusions respectively. These values were significantly lower than those in serum of the same patients (628.3 +/- 79.0 microg/l, P<0.025 and 532.0 +/- 85.9 microg/l, P<0.025 respectively). Because bioavailability and bioactivity of IGFs may be regulated by their binding to IGFBPs, we studied IGFBP patterns in the pleural fluid of 6 patients with tuberculous pleuritis. As assessed by Western ligand blotting the levels of IGFBP-1 and IGFBP-2 were increased whereas those of IGFBP-3 were decreased in pleural fluid in comparison with serum. The decrease in IGFPB-3 levels reflected increased proteolysis, as assessed by Western immunoblotting. In spite of this presence of IGFBPs, IGFs could be responsible for the local biosynthesis of 1.25-dihydroxyvitamin D (1,25-(OH)2D) since pleural fluid levels of both IGF-I and IGF-II significantly correlated with those of 1,25-(OH)2D. These results indicate that IGFs are detectable in pleural fluid and may contribute to control the activity of 25-hydroxyvitamin D-1alpha hydroxylase in tuberculous pleuritis.

Adult↗
PubMed1997

[Low extracellular pH has a role in the induction of NO synthase type 2 in macrophages].

Stimulation of macrophages with endotoxin and/or cytokines is responsible for the expression of the inducible isoform of nitric oxide synthase (iNOS). Because macrophages are exposed to low pH within the microenvironment of inflammatory lesions, the potential role of low pH as an additional regulator of iNOS was investigated. Substitution of the culture medium of rat peritoneal macrophages at pH 7.4 with medium at pH 7.0 upregulated iNOS activity, as reflected by a 2.5-fold increase in nitrite accumulation. The increase in iNOS activity was associated with a similar increase in iNOS mRNA expression. Low environmental pH-induced iNOS gene expression involved the activation of nuclear factor-kappa B (NF-kappa B) transcription factor since [1] exposure of macrophages to low environmental pH increased NF-kappa B binding activity in the nucleus, and [2] treatment of macrophages with pyrrolidine dithiocarbamate or n-acetyl-leucinyl-norleucinal, two drugs preventing NF-kappa B translocation to the nucleus, canceled low pH-induced nitrite accumulation. The overall mechanism required the synthesis of tumor necrosis factor-alpha (TNF-alpha). Indeed, [1] elevated TNF-alpha bioactivity was observed in the medium of macrophages exposed to pH 7.0, and [2] incubation of macrophages with a neutralizing anti-TNF-alpha antibody impaired both NF-kappa B activation and nitrite accumulation in response to acid challenge. In summary, exposure of macrophages to acidic microenvironment in inflammatory lesions leads to the upregulation of iNOS activity through the activation of NF-kappa B.

Animals↗
PubMed1996

Regulation of interleukin-10 production by beta-adrenergic agonists.

Catecholamines have been shown to inhibit some aspects of macrophage activation through a beta receptor-dependent mechanism. This study was undertaken to analyze the effects of isoproterenol, a specific beta-adrenergic agonist, on the synthesis of interleukin-10 (IL-10), a major macrophage-deactivating factor. Isoproterenol increased IL-10 release from lipopolysaccharide-(LPS)-activated mouse peritoneal macrophages in a dose-dependent manner. A significant effect was already observed with 1 microM isoproterenol, while a 4.5-fold increase was achieved with 10 microM. This increase was observed only if macrophages were exposed to isoproterenol for at least 2 h before LPS challenge. It was apparent within 0.5 h and persisted through 24 h at all the LPS concentrations used. A similar increase was observed at the IL-10 mRNA level, as judged by enzyme-linked immunosorbent assay-polymerase chain reaction. The macrophage response to isoproterenol that led to cyclic AMP accumulation was markedly inhibited by H-89, a potent inhibitor of protein kinase A. These data suggest the involvement of cyclic AMP in the regulation of IL-10 synthesis by isoproterenol. IL-10 was in turn partly responsible for a reduction in tumor necrosis factor-alpha synthesis. In vivo, the administration of oxprenolol, a beta-receptor antagonist, significantly reduced serum IL-10 levels 90 min after LPS challenge. Thus, the present study provides the first evidence that endogenous catecholamines are of critical importance in determining the magnitude of the IL-10 response in experimental endotoxemia.

Adjuvants, Immunologic↗
PubMed1996

Immunochemical study of galanin in the cat digestive tract and autonomic ganglia.

The presence of galanin was examined in the cat gut and related autonomic nervous structures using radioimmunoassay (RIA) and high performance liquid chromatography (HPLC). In the gut wall, the concentration of galanin-like immunoreactivity (GAL-Lt) was assayed separately in the muscular layers with the nervous plexuses and in the mucosa and ranged from 0.35 to 0.55 pmol/g wet tissue. In the autonomic nervous structures, GAL-L1 concentrations ranged from 0.22 (thoracic spinal ganglia) to 0.81 (inferior mesenteric ganglion) pmol/g wet tissue. The presence of galanin was checked by HPLC in the antrum, intestine, and colon. HPLC of extractable material revealed a major peak coeluting with the synthetic porcine peptide and minor earlier peaks representing likely different molecular forms of galanin. Our study strengthens the notion that galanin acts in nervous control of the cat gut functions.

Animals↗
PubMed1996

Pseudoknot and translational control in the expression of the S15 ribosomal protein.

Translational autocontrol of the expression of the ribosomal protein S15 proceeds through the transitory formation of a pseudoknot. A synopsis of the known data is used to propose a molecular model of the mechanism involved and for the role of the pseudoknot. This latter structure is able to recruit 30S ribosomal subunits to initiate translation, but also to bind S15 and to stop translation by trapping the ribosome on its loading site. Information on the S15 protein recognition of the messenger RNA site was deduced from mutational analyses and chemical probing. A comparison of this messenger site with the S15 ribosomal binding site was conducted by analysing hydroxyl radical footprintings of these two sites. The existence of two subsites in 16S RNA suggests that the ribosomal protein S15 might present either two different binding sites or at least one common subsite. Clues for the presence of a common site between the messenger and 16S RNA are given which cannot rule out that recognition specificity is linked to a few other determinants. Whether these determinants are different or not remains an open question.

Base Sequence↗
PubMed1996

Cloning and characterization of DXS6673E, a candidate gene for X-linked mental retardation in Xq13.1.

In several families with non-specific X-linked mental retardation (XLMR) linkage analyses have assigned the underlying gene defect to the pericentromeric region of the X chromosome, but none of these genes have been isolated so far. Here, we report on the cloning and characterization of a novel gene, DXS6673E, that maps to Xq13.1, is subject to X-inactivation and is disrupted in the 5' untranslated region by a balanced X;13 translocation in a mentally retarded female. The DXS6673E gene is highly conserved among vertebrates and its expression is most abundant in brain. It encodes a hydrophilic protein of 1358 amino acids (aa) that does not show sequence homology to other known proteins. A segment of this protein consisting of neutral and hydrophobic aa with a proline residue in every second position may represent a transmembrane domain. Almost complete sequence identity was found between the 3' end of the DXS6673E gene and two expressed sequence tags (ESTs) and between the 5' end of the DXS6673E gene and a third EST. Moreover, weaker sequence similarity was observed between coding regions and two other ESTs.

Amino Acid Sequence↗
PubMed1996

Diffuse panbronchiolitis in an Asian immigrant.

Diffuse panbronchiolitis (DPB) is a disease with chronic inflammation exclusively located in the region of the respiratory bronchiole. It is largely restricted geographically to the Far East, and cases in Western countries are exceptional, even among Asian immigrants. A patient of Asian origin with DPB who had been living in France for 10 years is described. Only re-examination of the initial open lung biopsy specimen after an eight year period allowed this rare disease to be diagnosed correctly. The known efficacy of low dose erythromycin in DPB was again confirmed after failure of long term high dose corticosteroid therapy administered before an accurate diagnosis had been made.

Adult↗
PubMed1995

Insulin activates nuclear factor kappa B in mammalian cells through a Raf-1-mediated pathway.

We examined the effect of insulin on nuclear factor kappa B (NF-kappa B) activity in Chinese ovary (CHO) cells overexpressing wild-type (CHO-R cells) or -defective insulin receptors mutated at Tyr1162 and Tyr1163 autophosphorylation sites (CHO-Y2 cells). In CHO-R cells, insulin caused a specific, time-, and concentration-dependent activation of NF-kappa B. The insulin-induced DNA-binding complex was identified as the p50/p65 heterodimer. Insulin activation of NF-kappa B: 1) was related to insulin receptor number and tyrosine kinase activity since it was markedly reduced in parental CHO cells which proved to respond to insulin growth factor-1 and phorbol 12-myristate 13-acetate (PMA) activation, and was dramatically decreased in CHO-Y2 cells; 2) persisted in the presence of cycloheximide and was blocked by pyrrolidine dithiocarbamate, aspirin and sodium salicylate, three compounds interfering with I kappa B degradation and/or NF-kappa B.I kappa B complex dissociation; 3) was independent of both PMA-sensitive and atypical (zeta) protein kinases C; and 4) was dependent on Raf-1 kinase activity since insulin-stimulated NF-kappa B DNA binding activity was inhibited by 8-bromo-cAMP, a Raf-1 kinase inhibitor. Moreover, insulin activation of NF-kappa B-driven luciferase reporter gene expression was blocked in CHO-R cells expressing a Raf-1 dominant negative mutant. This is the first evidence that insulin activates NF-kappa B in mammalian cells through a post-translational mechanism requiring both insulin receptor tyrosine kinase and Raf-1 kinase activities.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1995

Transforming growth factor-beta stimulates arginase activity in macrophages. Implications for the regulation of macrophage cytotoxicity.

Macrophage arginine metabolism via nitric oxide (NO) synthase and arginase pathways reduces and enhances tumor cell proliferation, respectively. Transforming growth factor-beta (TGF-beta) has been shown to down-regulate the NO synthase pathway. The present study describes the effect of TGF-beta on the arginase pathway. TGF-beta up-regulated arginase activity in rat peritoneal macrophages as assessed by measuring the generation of [14C]urea from [14C]-L-arginine in the presence of NG-monomethyl-L-arginine (L-NMMA). The stimulation, which reached fivefold after a 48-h exposure of macrophages to 10 ng/ml TGF-beta, was due to reduction in Km value of arginase. TGF-beta-induced up-regulation of arginase activity led to the release of more polyamines, mainly putrescine. The role of this up-regulation on macrophage cytotoxicity toward L-929 tumor cells was analyzed in coculture experiments. Macrophages blunted DNA synthesis by L-929 cells as assessed by measuring the incorporation of [3H]TdR into the cells and the proportion of cells in the G2 phase. Addition of TGF-beta in the presence of L-NMMA permitted L-929 cells cocultured with macrophages to resume DNA synthesis. The mechanism responsible for this restoration was the up-regulation of arginase activity rather than the down-regulation of NO synthase activity since TGF-beta in the presence of L-NMMA failed to further reduce NO synthase activity whereas it still enhanced arginase activity; synthetic putrescine (1-10 microM) also blunted macrophage cytotoxicity toward L-929 cells. This is the first evidence that TGF-beta up-regulates arginase activity in macrophages and, hence, limits macrophage-dependent cytostasis.

Animals↗
PubMed1995

A high-resolution interval map of the q21 region of the human X chromosome.

In a previous study, we have developed a panel of chromosomal rearrangements for the physical mapping of the q13-q21 region of the human X chromosome (Philippe et al., Genomics 17: 147-152, 1993). Here, we report the physical localization of 36 additional polymorphic markers by polymerase chain reaction analysis. The high density of chromosomal breakpoints in Xq21 allows us to map 58 DNA loci in 22 intervals. As a result, this segment of the X chromosome is saturated with approximately three sequence tagged sites per megabase of DNA, which will facilitate the construction of a YAC contig of this region.

Chromosome Mapping↗
PubMed1995

Molecular dissection of the pseudoknot governing the translational regulation of Escherichia coli ribosomal protein S15.

The ribosomal protein S15 controls its own translation by binding to a mRNA region overlapping the ribosome binding site. That region of the mRNA can fold in two mutually exclusive conformations that are in dynamic equilibrium: a structure with two hairpins and a pseudoknot. A mutational analysis provided evidence for the existence and requirement of the pseudoknot for translational control in vivo and S15 recognition in vitro. In this study, we used chemical probing to analyze the structural consequences of mutations and their effect on the stem-loop/pseudoknot equilibrium. Interactions between S15 and the pseudoknot structure were further investigated by footprinting experiments. These data, combined with computer modelling and the previously published data on S15 binding and in vivo control, provide important clues on pseudoknot formation and S15 recognition. An unexpected result is that the relevant control element, here the pseudoknot form, can exist in a variety of topologically equivalent structures recognizable and shapable by S15. S15 sits on the deep groove of the co-axial stack and makes contacts with both stems, shielding the bridging adenine. The only specific sequence determinants are found in the helix common to the pseudoknot and the hairpin structures.

Base Sequence↗
PubMed1995

Role of CCK in the regulation of secretion and adaptation in the pig pancreas.

The effect of cholecystokinin (CCK) on the pancreas was investigated in the pig in two experiments. Fifteen pigs were fed a diet containing 17 or 48% protein with or without MK329 (4.5 mg per meal). MK329 enhanced the postprandial peak of plasma CCK during the first 30 min, but pancreas adaptation to high protein was not affected. Sixteen pigs were divided into two groups: 12 pigs were infused with CCK-8 + secretin for 1 h and four pigs received a standard meal. In both groups, pancreatic secretion tests were performed under infusion of the vehicle alone or with MK329. After CCK + secretin, MK329 (65-500 micrograms/kg/h) did not alter CCK plasma levels and reduced the early pancreatic protein response by about 30%. Enzyme outputs in pancreatic juice were modestly affected by MK329. After the meal, MK329 (500 micrograms/kg/h) doubled the postprandial peak of plasma CCK and lowered the pancreatic protein output by 35-40% for the first 30 min. We suggest that (a) pancreatic adaptation to high dietary protein is not mediated via CCK-A receptors and (b) the stimulation of pancreatic protein secretion by a meal or by exogenous CCK-8 is mediated partly by CCK-A receptors.

Adaptation, Physiological↗
PubMed1995

Monosomy 21q: two cases of del(21q) and review of the literature.

We report on two cases of partial monosomy 21 and review cases with a partial or an apparently full monosomy 21. In situ hybridization and/or molecular studies appear to be necessary tools to study imbalance in such a small chromosome and to perform further genotype-phenotype correlations. The segregation mode in cases with a translocation is adjacent 1, adjacent 2, and 3:1 in about 1/4, 1/4 and 1/2 of the cases, respectively.

Child, Preschool↗
PubMed1995

Mesangial cell-derived interleukin-10 modulates mesangial cell response to lipopolysaccharide.

Interleukin (IL)-10 is a novel cytokine produced by a variety of cells, including monocytes/macrophages, upon exposure to lipopolysaccharide (LPS). Recent observations indicate that, in turn, IL-10 exerts suppressive effects on macrophage response to LPS. Because mesangial cells are also a target for LPS, we have examined the potential role of IL-10 in the regulation of mesangial cell response to LPS. To this aim, we have studied the synthesis and the autocrine/paracrine function of IL-10 in cultured mouse mesangial cells. IL-10 mRNA expression and IL-10 protein secretion were determined by a reverse transcription polymerase chain reaction technique and a specific enzyme-linked immunosorbent assay, respectively. No IL-10 mRNA expression was detectable in unactivated cells. LPS induced IL-10 mRNA expression in a dose-dependent fashion (1 to 100 micrograms/ml). In addition, LPS induced IL-10 protein release that was both dose dependent (1 to 100 micrograms/ml) and time dependent (24 to 72 hours). We have also studied the effect of IL-10 on the production of inflammatory mediators by LPS-activated mouse mesangial cells. Whereas recombinant IL-10 inhibited the generation of tumor necrosis factor-alpha (TNF-alpha) and IL-1 beta by 90 and 60%, respectively, it did not affect the formation of nitric oxide-derived nitrite (NO2-) and nitrate (NO3-). As shown by the use of anti-IL-10 monoclonal antibody, endogenously produced IL-10 affected the generation of TNF-alpha but neither that of IL-1 beta nor that of NO2- and NO3-. Finally, we have examined whether conditions known to also reduce the generation of TNF-alpha modified the expression of IL-10. Of all the conditions tested, only the addition of desferrioxamine and transforming growth factor-beta were found to increase IL-10 release. Together, these data demonstrate that mesangial cell-derived IL-10 has important regulatory effects on the inflammatory response of these cells to LPS because of its capacity to blunt TNF-alpha generation.

Animals↗