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P Poncet

Publications and source records attributed to P Poncet.

At least 37 records · Page 2Linked to original sources
PubMed1995

Mercuric chloride-induced programmed cell death of a murine T cell hybridoma. I. Effect of the proto-oncogene Bcl-2.

Mercuric chloride (HgCl2) as well as several drugs can induce T cell activation leading to systemic immune-mediated diseases in genetically susceptible individuals or rodents. T cell hybridomas represent a well-characterized model system for in vivo mechanisms of various stimuli-induced cell death. The cellular response to HgCl2 was examined using various T cell lines and particularly the murine T cell hybridoma 2B4.11. Exposure to HgCl2 induced both necrosis and apoptosis in a dose- and time-dependent way as demonstrated by DNA fragmentation analysis, flow cytometry of the whole cells and of isolated nuclei, and morphological examination. HgCl2-induced cell death was partly inhibited by cycloheximide. The expression of human Bcl-2 in 2B4.11 cells after transfection significantly prevented HgCl2-induced cell death but did not affect the susceptibility to apoptosis induced by an anti-CD3 epsilon mAb. Subcytotoxic doses of HgCl2 enhanced metabolic activity of Bcl-2 transfectants in contrast with mock-transfected cell line. Thus, we conclude that apoptosis is part of the cell death process induced by HgCl2 and that the ability of Bcl-2 to prevent the death of one particular cell line is stimulus-dependent suggesting the existence of different pathways leading to cell death.

Animals↗
PubMed1995

Mercuric chloride-induced programmed cell death of a murine T cell hybridoma. II. Opposite effect of interleukin-2 and interleukin-4.

In susceptible animals evidence is accumulating for a primary role for Th2 cells in the course of HgCl2-induced autoimmunity, and for a contribution of Th1 cells in the self-regulated phase of this disease. We have reported that incubation of 2B4.11 T cell hybridoma with HgCl2 induced programmed cell death. This paper shows that recombinant IL-2 significantly diminished HgCl2-induced 2B4.11 cell death. Although no effect was observed upon incubation with exogenous IL-4, we observed a significant protection by adding an anti-IL-4 monoclonal antibody to the culture. Accordingly, by RT-PCR we found the presence of IL-2 receptor-encoding mRNA, and by cytofluorometry, the expression of the protein was detected only after exposure to HgCl2. Moreover, upon HgCl2 treatment, 2B4.11 cells were induced to produce IL-4. Altogether these findings showed that cytokine environment, IL-2, IL-4 otherwise defining the Th1/Th2 dichotomy, in conjunction with a chemical may differentially influence the fate of cell populations, death or survival.

Animals↗
PubMed1993

Anti-histone autoantibodies react specifically with the B cell surface.

In an attempt to induce an immune response against Mls-1a antigens by immunizing C57B1/6 mouse (Mls-1b) with purified B cells from DBA/2 mouse (Mls-1a), we generated a panel of monoclonal antibodies from which the 5B9.6 mAb, taken as a representative antibody, was thoroughly investigated. This antibody specifically reacts with B cells from all mouse strains studied including C57Bl/6 mice as shown by FACS analysis of double-antibody labelled spleen cells. Using enzyme immunoassays and immunoprecipitation techniques, 5B9.6 mAb was found to be specific for histones. Amino acid sequence analysis of a peptide derived from a 5B9.6-immunoprecipitated polypeptide from DBA/2 B cells showed a 100% homology with a sequence within H2B histones. Furthermore, 5B9.6 mAb was able to interact with the cell surface of 7OZ/3 cell line, known as a typical pre-B cell line. The presence of histones can be modulated on the surface of 7OZ/3 cells since this antigen was upregulated after exposure of these cells to a cocktail of IL-1 and cAMP. Finally, 5B9.6 mAb was shown to interact with freshly isolated B cells from human peripheral blood.

Amino Acid Sequence↗
PubMed1991

Biased VH gene expression in murine CD5 B cells results from age-dependent cellular selection.

Flow cytometry-purified, peritoneal and splenic CD5+ and CD5- B cells from neonatal and adult C57BL/6 mice were studied for expression of VH and Vx gene families in RNA colony blot assays, and for frequencies of clones secreting antibodies to bromelain-treated mouse red blood cells (BrMRBC), single-stranded DNA, trimethyl ammonium and bovine gamma-globulin, by limiting dilution. The results show few overall differences between the two B cell subsets, which both manifest ontogenic D-proximal VH preferences that are lost with age. Biased VH11 expression in CD5 B cells is high in adult peritoneum and spleen but absent in newborns. It only partly correlates with the selection of anti-BrMRBC reactivity, which is considerably higher in peritoneum than in spleen. No particular Vx bias was observed in any of the populations studied with the possible exception of Vx22 in peritoneal CD5+ B cells. We conclude that the antibody repertoire expressed by peritoneal CD5+ B cells of adult mice is not the result of a genetic program, but rather the consequence of local, age-dependent cellular selection mechanisms.

Aging↗
PubMed1990

All VH11 genes expressed in peritoneal lymphocytes encode anti-bromelain-treated mouse red blood cell autoantibodies but other VH gene families contribute to this specificity.

We have studied the relationship between B cell reactivity to bromelain-treated autologous mouse erythrocytes (BrMRBC) and expression of the VH11 gene family in splenic, peritoneal and pleuropericardial cell populations from normal C57BL/6 mice. B lymphocytes producing antibodies to BrMRBC were selectively enriched or depleted from normal populations by rosette formation with BrMRBC, followed by centrifugation over density gradients. This selection method, based on the presence of functional receptors (membrane IgM), is harmless for the cells and allowed subsequent cloning in agar (colony-forming unit-B). The utilization of the 10 VH gene families was then scored in mRNA colony blot assays. The analysis of greater than 650 anti-BrMRBC clones and greater than 350 VH11-expressing colonies indicates that about half of those antibody reactivities are encoded by VH11 genes. Furthermore, it appears that all VH11-expressing B cells in the peritoneal cavity produce anti-BrMRBC antibodies.

Animals↗
PubMed1990

Cerebrovascular accidents in infective endocarditis: role of anticoagulation.

Anticoagulation is still a matter of debate in infective endocarditis, since it can increase the risk of complications, mostly neurological. In our series of 269 patients with native valve endocarditis studied between 1970 and 1982, 35 were anticoagulated. We observed 14 patients with brain infarcts, of whom five died, and 12 patients with cerebromeningeal or brain haemorrhage of whom six died. In a similar series of 63 patients with prosthetic valve endocarditis, all of whom were on anticoagulation and were studied between 1972 and 1987, we observed five patients with brain infarcts, three of whom died, and two patients with brain haemorrhage, one of whom died. The frequency of cerebrovascular accident (CVA) was similar for both groups (11.1% in prosthetic endocarditis vs 11.5% in native valve endocarditis, P = ns), as was mortality rate (57% vs 48.4%, P = ns). CVA are significantly more frequent among anticoagulated patients (19/94 vs 19/238: P less than 0.01), but the mortality rate in CVA is similar for anticoagulated and non-anticoagulated patients (11/19 vs 8/19: P = ns). The indications for anticoagulation in infective endocarditis remain similar to those in valvular heart disease. In patients with infective endocarditis, anticoagulation with heparin should be maintained whenever a brain infarct is present, unless it is large and/or haemorrhagic.

Adolescent↗
PubMed1989

Immunoglobulin VH gene expression in Ly-1+ and conventional B lymphocytes.

Lymphocyte populations in which Ly-1 B cells are differentially represented were studied for the expression of ten VH gene families, either by an RNA colony blot assay or by in situ hybridization of single cells, in BALB/c and C57BL/6 mice. The comparisons of cells from lymph nodes, Peyer's patches and adult spleen (poor in Ly-1 B cells) with cells from peritoneal cavity and neonatal spleen (rich in Ly-1 B cells) were confirmed by the analysis of adult peritoneal Ly-1- and Ly-1+ B cells sorted on the fluorescence-activated cell sorter. The results indicate that the peritoneal Ly-1+ B subset uses the whole spectrum of known VH gene families, and shows a preferential utilization of CP12 VH genes, most likely as a result of a selective process during life.

Age Factors↗
PubMed1989

Ontogenic development of autoantibody repertoires in spleen and peritoneal cavity of normal mice: examples of T cell-dependent and -independent reactivities.

The ontogenic development of B cell clonal precursors (BCP) reactive to bromelain-treated, syngeneic erythrocytes (BrMRC) and to single-stranded DNA has been studied by limiting dilution of both spleen and peritoneal cells. It was found that the frequency of anti-BrMRC BCP in the spleen is very low up to 4 weeks of age and slowly increases thereafter, to reach adult levels by 6-10 weeks. In the peritoneal cavity, no such BCP can be found before 2 weeks, but they occur at a very high frequency already by 3 weeks of age. Injection of adult, normal syngeneic T cells at birth has no apparent effect on the representation of anti-BrMRC BCP in the peritoneal cavity, but brings these to adult levels or even higher in the spleen already at 3 weeks of age. Accordingly, adult athymic (nude) mice contain normal frequencies of BrMRC-specific BCP in the peritoneal cavity but are devoid of such clones in the spleen. In contrast, the frequency of anti-DNA BCP is very high throughout postnatal development in both spleen and peritoneal cavity, of normal and athymic mice, in both resting and naturally activated splenic B cell compartments, and it is independent of T cell transfers into nude animals. These results indicate the role of T cells in the establishment of some clonal specificities in the adult, splenic autoreactive B cell repertoire.

Age Factors↗
PubMed1988

Immunochemical studies of polyspecific natural autoantibodies: charge, lipid reactivity, Fab'2 fragments activity and complement fixation.

Polyspecific natural autoantibodies (NAAb) are antibodies present in normal unimmunized animals and are able to react with very dissimilar antigens (Ag). To better delineate the characteristics of polyspecificity, we subjected monoclonal NAAb to four different immunochemical studies: (1) Two-dimensional gel electrophoresis performed on eight NAAb did not reveal any obvious relationship between charge and antigen specificities; (2) NAAb widely polyspecific on proteins and nucleic acid were reactive with lipids bearing either phosphate, sulfate or carboxyl polar groups; (3) pepsin digestion of polyspecific IgM NAAb yielded Fab'2 fragments which maintained their multireactivities, but exhibited a decrease in reactivity as compared to that seen with monospecific mAb (induced); (4) two different assays were used to analyse the complement fixation ability of IgM NAAb. While very weak or no complement fixation was observed with a classical complement fixation test (fluid phase), when a complement enzyme immunoassay was used where Ag is immobilized on a solid phase, polyspecific NAAb fixed reproducible and easily detectable amounts of complement.

Animals↗
PubMed1988

Comparative analysis of natural antibody specificities among hybridomas originating from spleen and peritoneal cavity of adult NZB and BALB/c mice.

A preliminary experiment showed that the supernatants of in vitro cultured peritoneal cells (rich in Ly-1 B cell subset shown to secrete most IgM autoantibodies against bromelain-treated mouse red blood cells (BrMRBC) and DNA) from different mouse strains did not contain any significant antibody activity against DNA and cytoskeleton proteins, although the presence of anti-BrMRBC antibodies was clearly evident. Therefore, we investigated comparative natural antibody (NAb) specificities against an antigen panel (DNA, cytoskeleton proteins, IgG, bovine serum albumin (BSA), BrMRBC, trinitrophenyl (TNP), and trimethylammonium (TMA) haptens) among Ig-secreting hybridoma collections from the splenic (158) and peritoneal (230) immune compartments of autoimmune New Zealand black (NZB) and lipopolysaccharide (LPS)-stimulated BALB/c mice. The data showed: (i) isotypic restriction (mu and gamma 3 only), predominance of TMA ion-reactive (including BrMRBC) but negligible anti-DNA-reactive antibody specificities, and lack of simultaneous polyspecific widespread reactivity (i.e. at least four or more antigens) against DNA and cytoskeleton proteins in the peritoneal cavity; (ii) predominance of simultaneous widespread polyspecific reactivity against DNA and cytoskeleton proteins but negligible or no TMA hapten-reactive antibody specificities in the spleen. These observations reflect certain differences in the B cell repertoire of peritoneal cavity (rich in Ly-1 B cells) compared with spleen. The NAb against BrMRBC and those reactive with DNA and cytoskeleton proteins, which have been suggested to be secreted by the Ly-1 B cell subset, are distinguishable on the basis of the presence of separate recurrent idiotypes and preferential localization of B lymphocytes directed against these autoantigens.

Animals↗
PubMed1987

[Microdetermination of pyrazinamide and its metabolites (pyrazinoic acid, 5-hydroxypyrazinoic acid, 5-hydroxypyrazinamide and pyrazinuric acid) in plasma and urine with liquid chromatography].

The method reported here for determining pyrazinamide and its metabolites (2-pyrazinoic acid, 5-hydroxypyrazinamide, 5-hydroxypyrazinoic acid and pyrazinuric acid) consists of diluting urine or acid deproteinisation of serum followed by chromatography on a cation-exchange column. The column length and the detection system (ultraviolet or fluorimetry) allow for a very good separation of the different compounds; the sensitivity of the method makes it suitable for pharmacokinetic studies.

Chromatography, High Pressure Liquid↗
PubMed1987

Novel V genes encode virtually identical variable regions of six murine monoclonal anti-bromelain-treated red blood cell autoantibodies.

The variable (V) region sequences of six immunoglobulin M (IgM, kappa) monoclonal autoantibodies that recognize bromelinized isologous red blood cells, obtained by fusions of peritoneal cells from NZB or CBA/J nonimmunized mice with BALB/c myeloma cells, were determined by direct mRNA sequencing. The V regions of the light chains (VL) are almost identical with one another, as are the V regions of the heavy chains (VH), which, however, differ by six linked-base substitutions, depending on the strain of mice producing the autoantibodies. Such variations may reflect allelic differences. The VH segments determined have no obvious correspondence to any VH genes identified so far. They may belong to the small VH group 4, where 73% homology, at the most, can be calculated at the protein level for codons 1 to 94. Alternatively, the VH regions may be members of a new group of VH sequences not previously found. The V kappa regions appear closely homologous to members of the V kappa-9 subgroup of myeloma proteins of unknown antigen-binding specificity. The joining segments, J kappa and JH, used by the autoantibodies investigated, originate from the J kappa 2 and JH1 germ-line gene segments, respectively. The nine base-long diversity segments, D, derive from one member of the germ-line D gene SP2 family.

Animals↗
PubMed1987

Age-related natural antibody specificities among hybridoma clones originating from NZB spleen.

In the studies presented here, age-related natural antibody specificities have been investigated in the autoimmune NZB mouse strain by cell fusion. The monoclonal immunoglobulins (MIg) secreted by productive hybridoma clones were examined for their antibody activities against a panel of antigens, including single- and double-stranded DNA, actin, tubulin, myosin, bromelain-treated mouse red blood cells (BrMRBC) and TNP-BSA, employing both direct and competitive enzyme immunoassays. The antibody specificities against this panel of antigens were strikingly frequent among hybridoma clones from neonatal NZB (49%) mice, compared to normal BALB/c neonates (8.8%) shown earlier. Among neonatal hybridomas with known antigen reactivities, 73% of the clones exhibited polyspecific binding. In contrast, the majority of hybridomas from 5- and 7-month-old NZB spleen reacted monospecifically (76%) with the antigens tested. Such a characteristic reactivity pattern reflects an age-related evolution of B-cell repertoire expression. Unlike normal BALB/c mice, a high frequency of monospecific TNP-hapten-reactive clones (75%) was noticed among hybridomas of known antigen reactivities from 5- and 7-month-old NZB-strain mice, an age when autoimmune haemolytic anaemia sets in. In conclusion, an elevated frequency of autoreactive clones among neonates (49%) and an aberrant expression of TNP-reactive clones in adults seem to be an outward signal of certain discrepancies at the level of B-cell repertoire expression in autoimmune NZB-strain mice.

Aging↗