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C Doucet

Publications and source records attributed to C Doucet.

At least 37 records · Page 2Linked to original sources
PubMed1998

IL-15 is produced by a subset of human melanomas, and is involved in the regulation of markers of melanoma progression through juxtacrine loops.

IL-15 is a novel cytokine active through the IL-2R/betagamma. Since several human melanoma cell lines display functional IL-2Rs, we studied the IL-15/melanoma cells interactions. Ten out of 17 melanoma cell lines express the IL-15 transcript and four of them express levels of IL-15 mRNA similar to those detected in control activated monocytes. Nine out of ten cell lines also express two transcripts for the IL-15R alpha originated by the alternative splicing of exon'3'. Two melanoma cell lines, MELP and MELREO, derived from patients with rapidly progressive primary melanomas, co-express the two IL-15 transcripts, originated by alternative splicing of exon 'A'. Intracellular IL-15 protein was only detected in these two cells lines and it is mainly retained in the Endoplasmic Reticulum (ER). However, a small amount of IL-15 is also found in the Golgi apparatus and in the early endosomes, suggesting production and intercellular trafficking of endogenous IL-15 protein. Nevertheless, no biologically active IL-15 could be detected in the supernatant of all melanoma cells. The anti IL-15 blocking mAb M111 causes the up regulation of HLA Class I in dense MELP and MELREO cultures. These data suggest that IL-15 is probably active through juxtacrine loops negatively controlling HLA Class I molecules expression. These data offer, for the first time, a likely explanation to the controversial issue of IL-15 secretion and constitute a natural model for understanding IL-15 routing. Moreover, we identify a subset of melanoma cells producing IL-15, possibly involved in tumor escape mechanisms.

Biomarkers, Tumor↗
PubMed1998

Elevated lipoprotein(a) levels and small apo(a) isoforms are compatible with longevity: evidence from a large population of French centenarians.

Epidemiological studies have shown lipoprotein(a) (Lp(a)) to be an independent risk factor for cardiovascular disease. Lp(a) is a cholesterol-rich, low-density lipoprotein (LDL)-like particle to which a large glycoprotein, apolipoprotein(a) (apo(a)) is attached. Plasma Lp(a) levels are highly genetically determined and influenced to a minor degree by environmental factors. In an effort to determine whether Lp(a) might be associated with longevity, we have evaluated Lp(a) levels and apo(a) isoform sizes in a population of French centenarians (n = 109) compared to a control group (n = 227). The mean age of centenarians was 101.5 +/- 2.4 years while the control group was 39.4 +/- 7.2 years. Plasma levels of total cholesterol and triglyceride were within the normal range in both centenarian and control subjects. Lp(a) levels were higher in centenarians (both male and female) than in the normolipidemic control group (mean Lp(a) level = 0.33 +/- 0.42 and 0.22 +/- 0.27 mg/ml, respectively, P < 0.005). The distribution of apo(a) isoforms was significantly shifted towards small isoform size in the centenarian population as compared to the controls (54.4 and 41.4% of isoforms < or = 27 kringles (kr), respectively, P = 0.04). Nonetheless, the apo(a) size distribution in centenarians did not entirely explain the high Lp(a) levels observed in this population. Factors other than apo(a) size, and which may be either genetic or environmental in nature, appear to contribute to the elevated plasma Lp(a) levels of our centenarian population. We conclude therefore that high plasma Lp(a) levels are compatible with longevity.

Adult↗
PubMed1998

IL-4 and IL-13 specifically increase adhesion molecule and inflammatory cytokine expression in human lung fibroblasts.

Subepithelial fibrosis in the bronchi of asthmatics is the result of an irreversible lung fibroblast activation, triggered by cytokines secreted by IL-4- and IL-5-activated inflammatory cells. Here, we provide evidence that human lung fibroblasts (ICIG7 cells) express a single class of high-affinity IL-4 receptor (IL-4R). This receptor is functional and composed of at least the IL-4Ralpha and IL-13Ralpha1 chains in the absence of the IL-2Rgamma chain. The IL-4Ralpha is efficiently internalized at 37 degrees C within 15 min in the presence of IL-4, whereas this process is slower with IL-13. In ICIG7 cells, IL-4 triggers the tyrosine phosphorylation of at least two proteins (110 and 180 kDa), and up-regulates the transcription of c-fos, c-jun and c-myc proto-oncogenes. In addition, the secretion of several cytokines [IL-6, granulocyte colony stimulating factor and granulocyte macrophage colony stimulating factor (GM-CSF)] as well as the expression of beta1 integrin and VCAM-1 adhesion molecules are augmented by IL-4. IL-13 displays similar biological activities, but less effectively than IL-4. On the other hand, ICIG7 cells could constitute a lung fibroblast population defined by the spontaneous release of several pro-inflammatory cytokines (IL-6, IL-11 and GM-CSF) and cell surface phenotype (CD4 and Thy-1). Through this peculiar cytokine pattern and the IL-4/IL-13-dependent activities, these cells could act as effector cells in the pathogenesis of asthma, triggering and maintaining the recruitment, homing and activation of bone marrow-derived inflammatory cells, and playing a role in the remodeling process of the airways.

Cell Adhesion Molecules↗
PubMed1998

Chimpanzee lipoprotein(A): Relationship between apolipoprotein(A) isoform size and the density profile of lipoprotein(A) in animals with different heterozygous apo(A) phenotypes.

In a previous study [C. Doucet et al., J. Lipid Res 35:263-270, 1994], we have shown that plasma lipoprotein (a) [Lp(a)] levels were significantly elevated in a population of unrelated chimpanzees as compared to those in normolipidemic human subjects. Nonetheless, the inverse correlation between Lp(a) levels and apolipoprotein (a) [apo(a)] isoforms typical of man was maintained in the chimpanzee. In the present study, we describe the density profiles of apo B- and apo A1-containing lipoproteins and of Lp(a) in chimpanzee plasmas heterozygous for apo(a) isoforms after fractionation by single spin ultracentrifugation in an isopycnic gradient. The distribution of apo(a) isoforms in the density gradient was also examined by SDS-agarose gel electrophoresis and immunoblotting using chemiluminescence detection. In all double-band phenotypes examined, the smallest isoform was present along the entire length of the density gradient. The density distribution of the second isoform varied according to the size difference between the respective isoforms. Two isoforms close in size (difference in apparent molecular mass = 60 kDa) were present together in every gradient subfraction. On the contrary, when the two isoforms displayed distinct molecular mass (maximal difference in apparent molecular mass = 340 kDa), then the largest was principally present in the densest fractions of the gradient (d > 1.1 mg/ml). These observations suggest that Lp(a) particles with small apo(a) isoforms are more susceptible to interact with other lipoproteins than are Lp(a) particles with large isoforms.

Animals↗
PubMed1998

[Attitude and behavior of hospital personnel toward smoking].

Due to a lack of data available at the national level, on smoking behaviour of hospital staff, a survey was carried out in a representative sample of hospitals. This survey, which will be repeated in the future, should enable to evaluate preventive actions at hospital and to measure the implication level of the hospital staff in tobacco prevention. The analysis of data collected among 3082 persons shows that a third of the hospital staff smokes. Among these persons a certain part smoke at their working place, this proportion is varying considerably with the working area in the hospital. The more this area is close to the patient the less is hospital staff smoking. More than one third of the interviewers (37.3%) think that trying to convince people to stop smoking is part of their role: this opinion varies considerably depending on the professional activity in the hospital. In addition a very large majority (81.4%) of the hospital staff considers that it has an exemplary role to play in front of patients. This belief is shared equally by all the different groups of staff. The smoking prevalence among hospital staff is equivalent to that of the general population; this underlines the need and the necessity to realize anti-tobacco campaigns among this health care population. Moreover since the exemplary role they have to play, is very well accepted, preventive actions could rely on a good level of participation.

Adult↗
PubMed1997

Role of interleukin (IL)-2 and IL-15 in the tumour progression of a melanoma cell line MELP, derived from an IL-2 progressor patient.

MELP is an interleukin (IL)-2 receptor (IL-2R; alpha+ beta+ gamma-) melanoma cell line that was derived, before the beginning of the immunotherapy, from a patient whose metastasis increased in size during treatment with IL-2/interferon-alpha. In these cells, continuous culture in the presence IL-2 (1000 UI/ml) causes the selection of a cell sub-line (termed MILG) expressing the gamma-chain which is tumorigenic in nude mice. Here, we further analysed the characteristics of MELP and MILG cells as well as clones selected at limiting dilution in the presence of high concentrations of IL-2 or IL-15, or those selected after transfection for the expression of a human IL-2 transgene (MELP-CL1). MELP cells, but not six other melanomas cell lines, shed two soluble immunosuppressive molecules, CD25 and intercellular adhesion molecule-1, whose levels also strongly increase in vivo during immunotherapy. In vitro MELP cells express transcripts for IL-6, transforming growth factor, basic fibroblast growth factor and vascular-endothelial growth factor. Cloning at limiting dilution was obtained in culture fed with IL-2 or IL-15. All these clones, as MILG cells, express the transcript for the IL-2R gamma chain. This could favour improved interactions with cytokines using this chain. By contrast, MELP-CL1 cells, which secrete low amounts of biologically active IL-2 (200 UI/10(6) cells) exhibit a phenotype and growth characteristics similar to those of the parental MELP cells. Indeed, a crosslinking experiment with 125I-IL-2, has showed that MELP and MELP-CL1 cells display a scant IL-2 binding ability that is strongly increased in MELP cells fed for 1 week with 1000 UI/ml IL-2. These cells, as well as MILG cells express a betagamma-complex which can also bind IL-15. IL-2 induces a rapid tyrosine phoshorylation in MILG cells, which is followed by a prolonged induction of c-fos and c-jun genes. By contrast, in MELP cells IL-2 only causes a delayed induction of c-myc gene. All MELP derivatives, but not MILG cells, express the transcripts for IL-15, which is not secreted but is present as an intracellular protein. All MELP cells express the transcript for the IL-15R alpha chain. MELP-CL1 cells are not tumorigenic in nude mice, whereas MILG cells form rapidly growing tumours in 75% of the mice. Coinjection at the same site of MILG and MELP-CL1 cells causes the rapid regression of MILG tumours in 80% of the mice, whereas their bilateral injection causes the rapid development of MILG tumours in 100% of the nude mice. Finally, treatment in nude mice of MILG cells with low amounts of IL-2 (1000 UI per mouse) and IL-15 (50 ng per mouse) induces the development of much more aggressive tumours.The expression of functional IL-2Rs in a subset of human melanomas could be responsible for tumour progression.

Adult↗
PubMed1996

Esters and amides of 6-(chloromethyl)-2-oxo-2H-1-benzopyran-3-carboxylic acid as inhibitors of alpha-chymotrypsin: significance of the "aromatic" nature of the novel ester-type coumarin for strong inhibitory activity.

A series of esters and amides of 6-(chloromethyl)-2-oxo-2H-1-benzopyran-3-carboxylic acid were synthesized and evaluated in vitro for their inhibitory activity toward bovine alpha-chymotrypsin and human leukocyte elastase. Both series behaved as time-dependent inhibitors of alpha-chymotrypsin, but ester-type coumarins were clearly more efficient than the corresponding amides in inactivating the serine proteinase. The best inactivations were observed with "aromatic" esters, in particular with meta-substituted phenyl esters such as m-chlorophenyl 6-(chloromethyl)-2-oxo-2H-1-benzopyran-3-carboxylate, which appears to be one of the most powerful inactivators of alpha-chymotrypsin yet reported (kinact/KI = 760,000 M-1 S-1 at pH 7.5 and 25 degrees C). Usually, the coumarin derivatives failed to inhibit significantly human leukocyte elastase. As a result, the reported series of aromatic coumarinic esters behaves as a new chemical family of selective alpha-chymotrypsin inhibitors.

Animals↗
PubMed1996

IL2 triggers a tumor progression process in a melanoma cell line MELP derived from a patient whose metastasis increased in size during IL2/INFalpha biotherapy.

Human melanomas may express both in vivo and in vitro functional IL-Rs and may be expected to directly respond to injected IL2. This may generate biological situations which may be favourable for the patient, but also for tumor progression. Here, we analyse the latter hypothesis. MELP is a melanoma cell line derived from a patient whose metastasis increased in size during IL2/IFN alpha biotherapy [correction of biotheraphy]. These cells have been characterized in vitro for their phenotype and for their sensitivity to IL2. In vitro MELP cells express an IL2-R alpha(+) beta(+) gamma(-) phenotype and IL2 treatment induces the acquisition of new functional characteristics represented (i) by the increased surface expression of two markers of metastatic evolution (ICAM-1 and CD44); (ii) by the stable induction of the IL2-R gamma with the appearance of functional IL2-R beta complex, which are also recognized by GM-CSF; (iii) by the inhibition of transcription of a regulatory cytokine such as IL6; (iv) by a differential effect of IL6 on CD44 surface expression in MELP cells treated or not with IL2 (MILG cells); (v) by the acquisition of faster growth rates and appearance of piling up and multilayer cellular organization; (vi) by the development of rapidly growing tumors in nude mice. IL2 induces in MELP cells a tumor progression process that could mimic the metastatic evolution observed in vivo during biotherapy. Therefore, MELP phenotype may help to define a subset of patients in which IL2 therapy may trigger unfavourable evolution.

Adult↗
PubMed1996

Synthesis and inhibition of human leucocyte elastase by functionalized N-aryl azetidin-2-ones: effect of different substituents on the aromatic ring.

N-aryl-3,3-difluoroazetidin-2-ones featured by a latent electrophilic methylene quinoniminium function have been synthesized and evaluated as inhibitors of human leucocyte elastase. To promote hydrophobic interactions with the enzyme, to increase the rates of beta-lactam ring opening and of benzylic group departure, or to induce hydrosolubility, these compounds incorporate on their aromatic ring either an alkyl moiety, a methoxy substituent or a carboxylic group. Some of these beta-lactams proved to be good inactivators of human leucocyte elastase.

Humans↗
PubMed1995

Characterization of the N-terminal and C-terminal domains of human apolipoprotein(a): relevance to fibrin binding.

The structural domains of human apolipoprotein(a) (apo(a)) in the lipoprotein(a) (Lp(a)) particle have been recently investigated by limited proteolysis [Huby, T., Doucet, C., Dieplinger, H., Chapman, J., & Thillet, J. (1994) Biochemistry 33, 3335-3341]. We have shown that apo(a) can be cleaved into two structural domains: one was of constant size (170 kDa) and corresponded to the C-terminal (Cter) domain of apo(a). This domain was linked by a disulfide bond to apo B100. By contrast, the N-terminal (Nter) domain, whose size varied according to the digested apo(a) isoform, was not linked to apo B100. We now describe the purification of these apo(a) domains and their interaction with fibrin surfaces in an in vitro binding assay. The Nter domain of apo(a) was purified as a soluble protein in a two-step procedure which involved sequential use of a heparin-Sepharose column and a lysine-Sepharose column. The Cter domain of apo(a), which remained in disulfide linkage with apo B100 of Lp(a), was isolated as a lipoprotein particle by a combination of chromatographic steps on heparin-Sepharose and Q-Sepharose columns. This particle, termed "mini-Lp(a)", appeared homogeneous in nondenaturing polyacrylamide gels and exhibited a particle size (285 A) which was intermediate between that of Lp(a) (300 A) and LDL (265 A). The cleavage site between the respective apo(a) domains was determined by N-terminal sequencing of the purified Cter domain. Such cleavage occurred between residues 3532 and 3533, which are located in the interkringle region between apo(a) kringles 4(4) and 4(5).(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1995

Non-enzymatic glycation of lipoprotein(a) in vitro and in vivo.

Nonenzymatic glycation of lipoprotein may contribute to the premature atherogenesis of patients with diabetes mellitus by diverting lipoprotein catabolism from non-atherogenic to atherogenic pathways. It has been demonstrated that the proportion of apolipoprotein B-100 (apo B-100) in glycated form is significantly higher in diabetic patients than in non-diabetic controls, and equally that plasma lipoprotein(a) Lp(a) levels may be increased in diabetic patients. Consequently, we have evaluated the glycation of Lp(a) in vitro and in vivo, by use of a combination of m-aminophenylboronate affinity chromatography and enzyme-linked immunosorbent assay (ELISA) for apo B-100 and Lp(a). In vitro studies were performed on normolipodemic plasma samples containing elevated concentrations of Lp(a). These studies establish that Lp(a) can be glycated in vitro in the presence of high concentrations of glucose, although to a lesser extent than low density lipoprotein (LDL) (15.8% +/- 4.4% and 30.2% +/- 5.4% (P = 0.0001) after a 48 h incubation at 37 degrees C, respectively). We have also shown that apo B-100 was more glycated than apo(a) in the Lp(a) particle. In vivo studies have shown that the percentage of glycated Lp(a) in diabetic patients was significantly higher than in the control population (2.8% +/- 1.07% versus 2.0% +/- 0.43%, P = 0.017). The level of glycated Lp(a) is also positively correlated with that of HbA1c in diabetic patients (r=0.6, P=0.002). Since our results show that Lp(a) is less susceptible to glycation than LDL, we speculate that glycation of LDL may be more relevant to the cardiovascular risk associated with this particle than with Lp(a).

Apolipoprotein B-100↗
PubMed1994

Structural domains of apolipoprotein(a) and its interaction with apolipoprotein B-100 in the lipoprotein(a) particle.

The structural domains of human apolipoprotein(a) [apo(a)] and its interaction with apolipoprotein B-100 (apo B-100) in the lipoprotein(a) [Lp(a)] particle were investigated by limited proteolysis with thermolysin and cathepsin D. We characterized the proteolytic products by sodium dodecyl sulfate-polyacrylamide gradient gel electrophoresis, followed by immunoblotting using different antibodies. For apo B-100 in Lp(a), the digestion patterns were found to be identical to those previously described [Chen et al. (1989) J. Biol. Chem. 264, 14369-14375; Chen et al. (1991) J. Biol. Chem. 266, 12581-12587] for apo B-100 in LDL. Thus, we compared the digestion patterns of apo B-100 in Lp(a) resolved under reducing and nonreducing migrating conditions. Using an antibody specific for a synthetic peptide of apo B-100 (residues 4004-4021), we confirmed that apo B-100 was linked to apo(a) by its C-terminal end. Various Lp(a)s isolated from several donors, and containing different isoforms, were used to study the structural domains of apo(a). Using the same procedure as for apo B-100, several common features were found for the different isoforms. (1) Apo(a) can be cleaved into two structural domains: one was of constant size (170 kDa) and was linked to apo B-100. Using an antibody specifically directed against kringle V, we demonstrated that this fragment corresponded to the C-terminal part of apo(a). (2) The other domain, whose size varied according to the digested apo(a) isoform, was not linked to apo B-100.(ABSTRACT TRUNCATED AT 250 WORDS)

Apolipoprotein B-100↗
PubMed1994

Lipoprotein[a] in the chimpanezee: relationship of apo[a] phenotype to elevated plasma Lp[a] levels.

Several studies have documented the presence of Lp[a] in nonhuman primates. However, data are lacking in great apes such as the chimpanzee. We have studied the quantitative distribution of Lp[a], as well as the frequency of apo[a] phenotypes, in a population of chimpanzees living in Gabon. Monoclonal antibody 14A12, directed against human apo[a], failed to recognize chimpanzee Lp[a]. Therefore, Lp[a] was assayed using an ELISA involving two polyclonal antibodies, an anti-human apo[a] and an anti-human apoB-100. Under these conditions, Lp[a] was detected in each of 28 animals. The plasma level of Lp[a] was found to be highly skewed toward elevated values: the mean Lp[a] level was 0.61 mg/ml (SD 0.45) as compared to 0.18 mg/ml (SD 0.16) in a normal Caucasian population (P < 0.0001). Phenotypes for apo[a] were identified by SDS-agarose-gel electrophoresis, followed by immunoblotting and detection by chemiluminescence. Seventeen different isoforms (ranging from 440 to 920 kDa) were found among all the animals as compared to 19 (540 to 960 kDa) in a human population of equivalent number. However, the distribution of apo[a] phenotypes was distinct between these populations. Thus isoforms of low molecular mass occurred with greater frequency in chimpanzee as compared to humans. In both populations, a strong inverse correlation between Lp[a] levels and apo[a] isoform sizes was found in chimpanzees (r = -0.48; P < 0.01) and in man (r = -0.68; P < 0.0002).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Elevated lipoprotein (a) levels in primary nephrotic syndrome.

Elevated plasma levels of total cholesterol and increase in the hepatic synthesis of some apo B-containing lipoproteins have been noted in the nephrotic syndrome. Apoprotein (a), the apolipoprotein distinguishing lipoprotein (a) [Lp(a)] from low-density lipoprotein, is equally of hepatic origin, and Lp(a) recently has been shown to possess both atherogenic and thrombogenic activities. However, little is known of Lp(a) levels in nephrotic patients. We measured plasma Lp(a) concentrations in 11 patients with primary nephrotic syndrome in the absence of hematuria, hypertension, and renal insufficiency. Histologic lesions were minimal-change disease in five cases, membranous glomerulopathy in four cases, and focal and segmental glomerulosclerosis in two cases. Mean levels of Lp(a) (98 +/- 92 mg/dL [mean +/- SD]) were markedly elevated in the nephrotic patients as compared with the controls (14 +/- 13 mg/dL). No correlation was noted between plasma Lp(a) and proteinuria, albuminemia, total cholesterolemia, low-density lipoprotein cholesterol, apoprotein B100, or plasminogen. Furthermore, there was no correlation between Lp(a) levels and apoprotein (a) isoform size. In four patients, the level of Lp(a) decreased approximately fourfold after remission of the nephrotic syndrome under corticosteroid treatment. Our observation that Lp(a) levels are elevated in the nephrotic syndrome is consistent with the hypothesis that these patients may be at an increased risk of cardiovascular and thrombotic complications.

Adult↗
PubMed1991

Discrepancy between anthropometry and biochemistry in the assessment of the nutritional status of the elderly.

Institutionalized aged subjects, considered free of evolutive disease and whose body weight was stable, were studied. They were divided into two groups depending on their body mass index: controls (BMI greater than or equal to 24) and depleted (BMI less than or equal to 21). The depleted group, as judged by anthropometric measurements, showed dramatically reduced body muscle and adipose masses. Usual blood parameters were normal in both groups. Biochemical markers of the protein and energy status, viz. albumin, transthyretin, transferrin, somatomedin-C, as well as serum levels of osteocalcin and apolipoproteins AI, AII, B, CII, CIII and E, were not affected in the depleted group. However, moderate iron deficiency and marked zinc deficiency were found in this group. It is concluded that in the elderly, biochemical markers of the protein and energy status are not related to the nutritional status assessed by anthropometry.

Aged↗