Search PubMed⌕ Search

Biomedical subjects

S Chevalier

Publications and source records attributed to S Chevalier.

At least 91 records · Page 5Linked to original sources
PubMed1994

Involvement of gp130/interleukin-6 receptor transducing component in interleukin-11 receptor.

The recently cloned interleukin (IL)-11 displays many biological properties in common with those reported for IL-6. In order to analyze the nature and the functionality of the IL-11 receptor we developed a proliferative assay using the human multifactor-dependent cell line TF1. We showed that a blocking monoclonal antibody GPX7 raised against the gp130/IL-6 receptor transducing subunit was also able to inhibit the IL-11-triggered TF1 line proliferation. In addition, involvement of gp130 in IL-11 signaling was demonstrated by an induction of the transducing protein phosphorylation in response to IL-11, as observed for IL-6. In contrast, the blocking monoclonal antibody B-R6, which recognized the gp80/IL-6 binding subunit failed to interfere with the IL-11 proliferative signal in the TF1 cell line. Similarly, we did not observe any competition between IL-6 and IL-11 for a putative common binding site on the cell surface. These results suggest that the IL-11 binding component is different from the gp80/IL-6 receptor. In conclusion, IL-11, along with IL-6, leukemia inhibitory factor, oncostatin M and ciliary neurotrophic factor, belongs to the same family of cytokines, using gp130 as a transducing protein.

Antibodies, Monoclonal↗
PubMed1994

Detection of phosphotyrosine in glutaraldehyde-crosslinked and alkali-treated phosphoproteins following their partial acid hydrolysis in gels.

Soluble fractions and particulate extracts from human prostate, and extracts from rat-liver membranes were used as a source of kinases to phosphorylate endogenous proteins in the presence of gamma- 32P-labeled ATP. Histone was also added as a substrate in order to compare the direct partial acid hydrolysis of phosphoproteins in gels to an indirect procedure involving partial acid hydrolysis after extraction in sodium dodecyl sulfate followed by precipitation with acetone. These procedures led to recoveries of 32P-labeled material of 90% and 40%, respectively, with a similar proportion of radiolabeled phosphoamino acids. Several 32P-labeled phosphoproteins separated in gels were therefore directly HCl-hydrolyzed and their phosphoamino acids were quantitated either prior to, or after glutaraldehyde crosslinking, with and without alkali treatment. By preventing protein losses occurring in hot alkali, glutaraldehyde crosslinking increased by an average factor of 6.5 the 32P-labeled material available for phosphoamino-acid analyses. For eight phosphoproteins analyzed, the overall effect of combined glutaraldehyde and alkali treatments was a relative decrease in phosphoserine (up to 8-fold), with concomitant relative increases in phosphotyrosine and phosphothreonine (up to 62- and 6-fold, respectively). This method will especially be useful for the detection of pTyr, a less abundant phosphoamino acid, in proteins which suffer from poor transfer efficiency in Western blot, are weakly antigenic towards anti-phosphotyrosine antibodies, can hardly be extracted from a gel and for identification of protein tyrosine kinases renatured in gels.

Acrylic Resins↗
PubMed1994

[Long-term follow-up of patients with complete congenital auriculo-ventricular block treated with a cardiac stimulator].

The congenital complete heart block (CCHB) is a rare affection. In our two centers, in a period of 10 years, 1405 pacemakers were placed and only 10 (0.71%) for CCHB. We review the clinical history, the EKG's, echocardiography and Holter monitoring of these 10 patients. All of them were symptomatic. After pacemaker implantation, symptoms were improved in 90%. No death or other morbidity were observed during the long-term follow-up. So, the clinical outcome of patients with CCHB who underwent prophylactic pacemaker implantation seems to be associated with an excellent prognosis.

Adult↗
PubMed1993

Dietary restriction reduces the incidence of NMU-induced mammary tumors and alters retinoid tissue concentrations in rats.

Previous studies suggested a relationship between dietary restriction (DR) effects on mammary carcinogenesis and DR effects on liver retinoids. Therefore, in this study, retinoid concentrations were measured by high-performance liquid chromatography in the plasma, liver, and peripheral organs of DR rats with chemically induced carcinogenesis. Rats were injected with N-methyl-N-nitrosourea (MNU) and maintained on graded levels of DR (reduction of 10-40% from energy ingested by control animals with free access to food). Mammary tumor incidence and multiplicity induced by MNU were reduced in relation to the degree of DR, with virtual prevention occurring at 30% and 40% DR. Total hepatic retinoid concentrations (retinol + retinyl esters) were significantly greater in rats given MNU and subjected to DR, but liver total retinoid content was comparable between the groups. However, plasma retinol concentrations were significantly lower in DR rats than in controls given the carcinogen without DR. Retinoid concentrations were also elevated in adipose tissue, lungs, and intestine of DR rats, while renal concentrations remained unaltered. Retinoid concentrations in mammary glands and mammary tumors were similar in all groups. Thus, in DR rats, vitamin A concentrations in liver and other target tissues are maintained or increased despite decreases in plasma. It remains to be investigated whether these alterations in retinoid content have any relationship to the cancer-preventive effect of DR.

Adipose Tissue↗
PubMed1993

Protection of islets of Langerhans from antibodies by microencapsulation with alginate-poly-L-lysine membranes.

Microencapsulation of islets has been proposed to prevent their immune destruction following transplantation. An indirect immunofluorescence technique has been developed and used to study the permeability of the alginate-poly-L-lysine microcapsules to antibodies. Wistar rat islets were incubated with the R2D6 monoclonal mouse IgM antibody against rat islets, microencapsulated, and incubated with fluorescein-labeled goat IgG antibodies against mouse IgG and IgM. For the negative controls, the first antibody was omitted or both antibodies were omitted. The positive controls included islets incubated with both antibodies before they were encapsulated. Our study demonstrated that the alginate-poly-L-lysine membranes are not permeable to IgG when poly-L-lysine of molecular weights ranging from 21,000 to 390,000 are used. This simple immunofluorescence technique demonstrated the nonpermeability of the microcapsules to IgG, and could be useful for the initial evaluation of new types of membranes.

Alginates↗
PubMed1993

High circulating leukaemia inhibitory factor (LIF) in patients with giant cell arteritis: independent regulation of LIF and IL-6 under corticosteroid therapy.

Leukaemia inhibitory factor (LIF) is a cytokine which possesses a wide range of biological activities including, like IL-6, the capacity to stimulate acute phase protein (APP) synthesis. We have developed a sensitive and specific ELISA for human LIF, and tested the circulating cytokine levels in various disease states, some of which are associated with inflammation. LIF was detected in 11/20 sera from patients with giant cell arteritis (GCA), a vasculitis syndrome affecting particularly the temporal artery, characterized by panarteritis with inflammatory cell infiltration. LIF levels were considerably elevated in some patients who also displayed elevated levels of IL-6 and C-reactive protein (CRP); however, no correlation was observed between the levels of circulating LIF and levels of IL-6 or CRP. Furthermore, LIF levels were not affected by corticosteroid therapy, whereas IL-6 and CRP decreased rapidly, as clinical symptoms resolved. A putative role for LIF in the persistence of histological lesions is discussed. This is the first report of the presence of circulating LIF in sera. These results are in agreement with the complexity of induced inflammatory cytokines and corticoid regulation of APP synthesis observed in vitro and in vivo.

Administration, Oral↗
PubMed1992

Identification of cytosolic protein tyrosine kinases of human prostate by renaturation after SDS/PAGE.

The identification of protein tyrosine kinases (PTKs) was successfully achieved by renaturation in gels after SDS/PAGE. To this effect, samples were mixed with a PTK substrate, namely the polydispersed co-polymer of glutamic acid and tyrosine [poly(Glu, Tyr), M(r) from 30,000 to 94,000], and were simultaneously submitted to electrophoresis. Following guanidine hydrochloride denaturation, renaturation and phosphorylation with [gamma-32P]ATP, kinase activity was detected by autoradiography. When applied to cytosol from human hyperplastic prostate, eleven protein kinases were detected, among which one major (M(r) 50,000) and two minor proteins (M(r) 40,000 and 38,000) were identified as PTKs by the presence of phosphotyrosine. Incubation of the gel in hot alkali after glutaraldehyde cross-linking almost completely eliminated the detection of non-PTK enzymes. On the other hand, in the absence of poly(Glu,Tyr), no PTK activity was detected. Partial purification of cytosolic PTKs indicates that the native M(r) of the major phosphotransferase was 44,000, as estimated by gel filtration following ammonium sulphate precipitation and anion-exchange chromatography. Upon renaturation after electrophoresis, this fraction showed only one major band active on poly(Glu,Tyr) which was associated with the polypeptide of M(r) 50,000. This enzyme was also identified following two-dimensional electrophoresis and renaturation in the presence of poly(Glu,Tyr), allowing the determination of a pI in the range 7.5-7.8. Thus PTKs can be easily renatured following electrophoresis and rapidly identified on the basis of their M(r) and pI in both crude or partially purified preparations. With the crucial role played by PTKs in the activation of cell function and carcinogenesis, this procedure could be useful in the identification of such enzymes and in distinguishing them from their substrates in gels.

Amino Acids↗
PubMed1992

The zona pellucida binds the mature form of an oviductal glycoprotein (oviductin).

The use of a monoclonal antibody (MAb) specific for the oviductal zona pellucida (ZP) of the hamster has demonstrated that a new antigen (oviductin) is acquired by the ZP during transit of the oocyte in the oviduct. The epitope that is recognized by the MAb bears a terminal N-acetyl-D-galactosamine residue. We conducted a study in order to determine whether this immunoreactivity of the oviductal ZP results from the addition of the terminal sugar residue to a preformed ZP protein or from the transfer of the mature glycoprotein produced by oviductal secretory cells. We measured the incorporation of [35S]methionine into proteins using four different incubation systems: cumulus oophorus (CO) alone, CO in the presence of oviductal fluid, CO co-incubated with empty oviducts, and CO within intact oviducts. At the end of the incubation period, the ZP, vitelli, dispersed cumulus without oocyte, oviducts, and culture medium were isolated and analyzed for their protein content by sodiumdodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE), autoradiography, and immunodetection. The cumulus cells synthesized several proteins, independently of the oviductal environment; however, none of these proteins corresponded to oviductin. The ZP and the vitelli of cumulus oophorus that were incubated either alone or in the presence of oviductal fluid did not contain radioactive oviductin. When the oviduct (empty or intact) was present in the incubation system, radiolabeled oviductin was synthesized and secreted into the incubation medium. The ZP picked up a detectable amount of radioactive antigen only in the system in which intact oviducts were incubated.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

The major form of protein tyrosine kinase in the dog prostate is expressed by a 50 kDa polypeptide.

We have already reported that the protein tyrosine kinase (PTK) activity in the dog prostate is distributed in cytosolic (75%) and particulate (Triton X-100-solubilized) fractions and that upon gel filtration, both PTKs migrate as entities of Mr 44,000 [(1991) Biochem. Cell. Biol. 69, 146-153]. Herein we demonstrate by immunoprecipitation with anti-phosphotyrosine antibodies that the soluble PTK has the ability to undergo self-phosphorylation. In addition, the polypeptide responsible for that enzymatic activity has been identified by 2 approaches: (1) a two-dimensional electrophoresis, in which the first dimension performed in non-denaturing conditions allowed the localization of the native enzyme, while the second dimension (SDS-PAGE) permitted the analysis of alkali-resistant phosphoproteins corresponding to the activity; (2) protein renaturation after SDS-PAGE followed by in situ phosphorylation (with [gamma-32P]ATP) of polyGT electrophoresed together with the enzyme preparation; the exclusive presence of the radiolabeled phosphotyrosine in the renatured protein confirmed its enzymatic nature. Using these methods, the major form of PTK in the dog prostate was shown to be expressed by a 50 kDa polypeptide which possesses autophosphorylation sites and which is present in the cytosol as an active monomer.

Animals↗
PubMed1992

Identification of RNA-binding proteins specific to Xenopus Eg maternal mRNAs: association with the portion of Eg2 mRNA that promotes deadenylation in embryos.

Maternal Xenopus Eg mRNAs have been previously identified as transcripts that are specifically deadenylated after fertilization and degraded after the mid blastula transition. Destabilizing cis sequences were previously localised in the 3' untranslated region of Eg2 mRNA. In order to characterize possible trans-acting factors which are involved in the post-transcriptional regulation of Eg mRNAs, gel-shift and u.v. cross-linking experiments were performed, which allowed the identification of a p53-p55 RNA-binding protein doublet specific for the 3' untranslated regions of Eg mRNAs. These p53-p55 proteins do not bind to the 3' untranslated regions of either ornithine decarboxylase or phosphatase 2Ac mRNAs, which remain polyadenylated in embryos. These novel RNA-binding proteins are distinct from the cytoplasmic polyadenylation element-binding protein that controls the polyadenylation of maternal mRNAs in maturing Xenopus oocytes, and from previously identified thermoresistant RNA-binding proteins present in oocyte mRNP storage particles. The p53-p55 bind a portion of the Eg2 mRNA 3' untranslated region, distinct from the previously identified destabilizing region, that is able to confer the postfertilization deadenylation of CAT-coding chimeric mRNAs. This suggests that the p53-p55 RNA-binding proteins are good candidates for trans-acting factors involved in the deadenylation of Eg mRNAs in Xenopus embryos.

Animals↗
PubMed1991

Radiation-inactivation size of transformed and non-transformed androgen receptors.

The nucleic acid sequence of the androgen receptor (AR) gene predicts that the protein structure possesses DNA- and steroid-binding domains that show high degrees of sequence similarity with those of other steroid receptors. Since the steroid-binding domain of the AR corresponds to a 30 kDa portion of the protein, and the AR structure may be monomeric or hetero-oligomeric depending on its transformation state, we have herein determined the AR radiation-inactivation size (RIS) in relation to the molecular structure whose binding activity toward methyltrienolone (R1881) is abolished by a radiation 'hit'. Soluble fractions from whole canine prostatic tissue were used as a source of non-transformed AR. The AR transformation was induced by the addition of 0.6 M-KCl, and these preparations were used together with high-salt nuclear extracts as a source of transformed AR. To maximize the binding activity, molybdate and dithiothreitol were included during AR extraction. Receptor transformation was verified by modifications of both the sedimentation coefficients (from 7.5 S to 4.1 S on sucrose gradients) and molecular masses (from 260 kDa to 115 kDa by gel filtration). The RIS values of the non-transformed and transformed ARs were not statistically different: 92 +/- 19 kDa and 110 +/- 25 kDa respectively. In addition, the inactivation of AR binding activity by radiation was attributed to a loss of binding sites, with no significant change in the Kd. When benzoic acid, a free-electron scavenger, was added together with dithiothreitol before and after irradiation, no change in the RIS value was observed. Thus, in the canine prostate, the RIS value of the AR represents the monomeric protein, independently of its association with other proteins, and this value corresponds to that predicted by cloning studies and photoaffinity-labelling of AR.

Animals↗
PubMed1991

Prostatic epithelial cells in culture: phosphorylation of protein tyrosyl residues and tyrosine protein kinase activity.

The ability of dividing canine prostatic epithelial cells in primary monolayers to phosphorylate protein tyrosyl residues was evaluated by metabolic studies performed through incorporation of [32P]-phosphate into alkali-resistant phosphoproteins and by the assay of their tyrosine protein kinase activity. The presence of sodium orthovanadate during cell incubation with [32P]-phosphate greatly enhanced the relative labelling intensity of a 44 kDa alkali-resistant phosphoprotein and the total cellular content of phosphotyrosine in proteins; in this respect, growth factors such as epidermal growth factor, insulin, and insulin-like growth factor I, and the steroids dihydrotestosterone and estradiol were inactive. When the cells were solubilized, sodium orthovanadate stimulated their tyrosine protein kinase activity and inhibited their phosphotyrosine phosphatase activity. To characterize the tyrosine protein kinase of these cultured cells, conditions for optimal activity were established using the substrate poly [Glu80Na, Tyr20]. The subcellular localization of the enzyme was determined upon cell fractionation: 88% of the kinase activity was associated with the particulate fraction and 30% of this activity was partially solubilized with 0.5% Triton X-100; this solubilization was improved to 83% in the presence of 0.25 M KCI. The enzyme directly solubilized from prostatic cells with Triton X-100 (38% of activity) mainly catalyzed the alkali-resistant phosphorylation of pp63, pp59, and pp44, which contained phosphotyrosine. These proteins were also phosphorylated by the major peak of kinase activity which was eluted at an apparent molecular weight of 300-350 kDa upon gel filtration.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Involvement of vitronectin and of a natural extracellular matrix in prostatic cell to cell contact and interaction with the substratum in culture.

The kinetics and regulation of attachment, the spreading and initial growth of canine prostatic epithelial cells in primary monolayers, as well as the ability of the cells to produce a natural extracellular matrix (nECM) have been studied. Within 1 day of culture, attachment without spreading occurred in uncoated dishes while spreading was observed in nECM-coated dishes. The attachment was stimulated by dialyzed fetal bovine serum (dFBS) and/or by the nECM and was not affected by the presence of cycloheximide at 10(-6) M. This effect of dFBS on attachment was maximal at a concentration of 10% and increased with cell density and was mainly attributed to vitronectin as demonstrated by gel filtration, selective adsorption of serum proteins to the substratum, and indirect immunofluorescence of prostatic epithelial cells incubated with a rabbit antiserum against bovine vitronectin. Dog serum reduced attachment while bovine serum albumin, phenol red, and steroids were inactive. The maximal effect of the nECM on attachment was observed with matrices produced by canine prostatic epithelial cells cultured in the presence of dog serum for more than 10 days. As revealed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, several proteins of relative mass (Mr) of 210, 113, 61, 56, 52, 49, 45, and 42 kDa remained in the dish following nECM denudation and could be responsible for its effect on attachment. Their deposition as well as intercell adhesion and cell-to-substratum interactions required an active protein synthesis plus non-steroidal serum components. Once attached and spread in primary monolayers, prostatic epithelial cells divided by the 3rd to 4th day; their growth, stimulated by dFBS at an optimal concentration of 20%, was inhibited by cycloheximide while the nECM was inactive. Thus, vitronectin in serum and the nECM synthesized by canine prostatic epithelial cells are responsible for the attachment and spreading of homologous cells to an in vitro substratum; nECM proteins also contribute to cell-to-cell contact and cell-to-substratum interaction in the monolayers.

Animals↗
PubMed1991

Serum and prostatic growth-promoting factors for steroid-independent epithelial cells of adult dog prostate.

A growth factor-like effect has been observed on canine prostatic epithelial cells when cultured in the presence of their homologous serum and prostatic extracts; the mitogenic activities of both preparations were dose-dependent and not altered by charcoal treatment. The effect of dog serum decreased when the density of the epithelial cell cultures increased and was minimal on canine prostatic fibroblasts. Trace amounts of intracellular sex steroids did not contribute to epithelial cell proliferation since the presence of sex steroid action inhibitors did not alter growth rate; in those conditions, cycloheximide completely prevented cell division. When various hormones and known mitogenic agents were tested alone or in combination with steroids, none elicited an increase in the number of epithelial cells cultured in serum-free medium or altered the proliferative effect of dog serum observed in parallel cultures. On gel filtration, dog serum or tissue cytosol showed a major mitogenic activity at an apparent molecular mass of 150 kDa and a minor one of 1.5 kDa as evaluated by gel filtration of dog serum ultrafiltrate. Acidic extraction of prostatic tissue followed by chromatography on a hydrophobic C-18 column and subsequent gel filtration also led to the detection of the low Mr component. Thus, humoral and/or tissular factors present in vivo and different from known mitogens may be of importance as direct modulators of the basal epithelial cell growth in the adult canine prostate.

Animals↗
PubMed1990

Protein tyrosine kinases in human breast cancer: kinetic properties and evidence for the presence of two forms of native enzyme.

The protein tyrosine kinase (PTK) of human breast tumors classified as positive (TM+) or negative (TM-) according to their estrogen and progestin receptor levels was partially characterized with regard to its distribution, kinetic parameters, molecular size, and ability to phosphorylate endogenous mammary proteins. For both types of tumors, PTK activity depended upon the presence of Mn++ (2-5 mM) and/or Mg++ (10-20 mM). The activities, total (per g of tissue) and specific (per mg of protein), were similar for both types of tumors, and an average of 60% of activity was located in cytosolic fractions. The autoradiographic detection of alkali-resistant phosphoproteins after SDS-PAGE showed very similar patterns between corresponding fractions from both types of tumors. Upon gel filtration, two peaks of activity of apparent Mr 245 kDa (peak I) and 47 kDa (peak II) were observed. Peak II was found in both cytosols and extracts from particulate fractions, while peak I was present only in the latter fraction for both TM+ and TM- tumors. The apparent Km's for ATP ranged from 4.1 to 6.6 microM, and from 11 to 34 micrograms/ml for the synthetic substrate poly [Glu80, Tyr20], at an optimal pH of 6.5-7.5. When endogenous alkali-resistant phosphorylation of peaks I and II was determined by autoradiography after SDS-PAGE, two major mammary proteins of Mr 60 and 45 kDa were phosphorylated by peak II and three, Mr 145, 74, and 62 kDa, by peak I.(ABSTRACT TRUNCATED AT 250 WORDS)

Breast Neoplasms↗