PubMed Health⌕ Search

Biomedical subjects

S Ouellet

Publications and source records attributed to S Ouellet.

16 recordsLinked to original sources
PubMed2001

Identification of three new DRB3* (DRB3*0106, DRB3*0107 and DRB3*02022) alleles.

Three novel DRB3* alleles were identified using CANTYPE reverse hybridization assay. The initial unusual hybridization patterns of DRB3-specific polymerase chain reaction (PCR)-amplified DNA from each subject were confirmed by cloning and sequencing analysis. DRB3*0106 allele is identical to DRB3*0101 except for a single nucleotide substitution (CTG-->GTG) changing codon 38 from Leu to Val. This polymorphism is commonly found in DRB3*03 alleles. Compared with DRB3*0202, DRB3*02022 contains a single silent nucleotide substitution (AAT-->AAC, both encoding for Asn) at codon 77. This polymorphism is also present in DRB3*0204 allele. The new DRB3*0107 allele has a sequence unique to DRB3 alleles. From codon 5 to codon 36 the sequence is identical to that of DRB3*0101 allele. From codon 37 to codon 87 the sequence of DRB1*0107 allele is identical to that of DRB3*0202. This sequence would thus explain the CANTYPE(R) DRB3-specific unusual pattern of reactions. The new DRB3*0107 could have arisen from a gene conversion between DRB3*0101 and DRB3*0202 alleles, but the DRB3*0106 and the DRB3*02022 may have been generated by a point mutation event. The DRB3*0107 allele was identified in a Caucasoid individual. The ethnic origin of the subjects carrying the other two alleles are unknown. The three alleles presented here were only identified once, in a total population of 49,000.

Alleles↗
PubMed2001

Peroxynitrite mediates cytokine-induced IL-8 gene expression and production by human leukocytes.

Recent studies indicate that nitric oxide (NO) or related compounds may regulate the production of interleukin (IL)-8, a potent proinflammatory chemokine. Here we report that peroxynitrite (ONOO(-)) formed by a reaction of NO with superoxide mediates IL-8 gene expression and IL-8 production in IL-1beta- and TNF-alpha-stimulated human leukocytes in whole blood. The NO synthase inhibitors aminoguanidine and N(G)-nitro-L-arginine methyl ester blocked nuclear accumulation of activator protein-1 (AP-1) and nuclear factor (NF)-kappaB in both polymorphonuclear (PMN) and mononuclear leukocytes and inhibited IL-8 mRNA expression and IL-8 release by approximately 90% in response to IL-1beta and TNF-alpha. Enhanced ONOO(-) formation was detected in granulocytes, monocytes, and lymphocytes after challenge with IL-1beta or TNF-alpha. The addition of ONOO(-) (0.2-80 microM) to whole blood increased nuclear accumulation of AP-1 and NF-kappaB in PMN and mononuclear leukocytes and augmented IL-8 mRNA expression and IL-8 production in a concentration-dependent fashion. Pyrrolidine dithiocarbamate, an inhibitor of NF-kappaB activation, attenuated approximately 70% of IL-8 release evoked by IL-1beta, TNF-alpha, or ONOO(-). These results indicate that ONOO(-) formation may underlie the action of cytokines towards IL-8 gene expression in human leukocytes.

Antioxidants↗
PubMed2000

The anti-inflammatory peptides, antiflammins, regulate the expression of adhesion molecules on human leukocytes and prevent neutrophil adhesion to endothelial cells.

Antiflammin-1 and antiflammin-2 are nonapeptides corresponding to the region of highest similarity between glucocorticoid-inducible proteins lipocortin-1 and uteroglobin. We have studied whether antiflammins could affect expression of adhesion molecules on human leukocytes and coronary artery endothelial cells (HCAEC) and binding of neutrophils (PMNs) to HCAEC. Although neither antiflammin-1 nor antiflammin-2 affected expression of adhesion molecules on resting PMNs, monocytes, and lymphocytes in whole blood, they attenuated changes in L-selectin and CD11/CD18 expression evoked by platelet-activating factor or interleukin-8 with IC(50) values of 4-20 micromol/l. The maximum inhibition was similar to those seen with human recombinant lipocortin-1 (100 microgram/ml). Unlike dexamethasone (100 nmol/l), the antiflammins had little effect on LPS-stimulated expression of E-selectin and ICAM-1 on HCAEC. Consistently, culture of HCAEC with dexamethasone, but not with antiflammins, decreased PMN binding to endothelial cells. Preincubation of PMNs with antiflammins markedly decreased their adhesion to LPS-activated HCAEC. Inhibition of adhesion was additive with function blocking anti-E-selectin and anti-L-selectin antibodies, but was not additive with anti-CD18 antibody. These results show that antiflammins inhibit PMN adhesion to HCAEC by attenuating activation-induced up-regulation of CD11/CD18 expression on leukocytes, and suggest that antiflammins may represent a novel therapeutic approach in blocking leukocyte trafficking in host defense and inflammation.

Adult↗
PubMed1998

Identification of new DRB1*07 (DRB1*0703), DRB1*08 (DRB1*0817) and two DRB3* (DRB3*0302 and DRB3*01014) alleles.

We report here the identification of four novel DRB alleles using a reverse hybridization (CANTYPE) assay. Molecular cloning and sequencing confirmed the initial unusual hybridization patterns. All four new alleles were detected during routine HLA typing for the Canadian Unrelated Bone Marrow Donor Registry. DRB1*0703 is identical to DRB1*0701 except for a single nucleotide substitution (AGA-->AGT), changing codon 29 from Arg to Ser, a so far undetected DRB polymorphism. DRB1*0817 differs from DRB1*0801 by a single nucleotide substitution (TAC-->TTC), changing codon 47 from Tyr to Phe. This polymorphism has not, until now, been identified in DRB1*08 alleles. Compared with DRB3*0301, DRB3*0302 contains a single nucleotide substitution (TAC-->CAC) at codon 30, changing the encoded Tyr to His. This polymorphism is typical for DRB3*02 alleles. DRB3*01014 is identical to DRB3*0101 except for a single silent nucleotide substitution (GGG-->GGA) at codon 84. This polymorphism has previously only been described for the DRB1*15012 allele. DRB1*0817, DRB3*0302 and DRB3*01014 may have arisen from gene conversion, but DRB1*0703 most likely was generated by a point mutation event. The DRB3*0302 allele was detected in two unrelated subjects, while the other three have each only been detected once.

Alleles↗
PubMed1998

Identification of novel DRB1*13 (DRB1*1333), DRB1*04 (DRB1*0426) and DRB5* (DRB5*0109) alleles.

Three novel HLA class II alleles (DRB1*1333, DRB1*0426, DRB5*0109) are described here. The 3 novel alleles were initially detected as previously unidentified SSO hybridization patterns using the CANTYPE reverse hybridization assay. Sequences were determined by cloning/sequencing. DRB1*1333 is identical to DRB1*1303 except for a single nucleotide substitution (ACC-->AAC), changing codon 77 from Thr to Asn. This polymorphism is typical for DRB1*03 alleles. DRB1*0426 is identical to DRB1*0401 except for a single nucleotide substitution (GCC-->ACC) at codon 58, changing the encoded Ala to Thr. DRB5*0109 is identical to DRB5*0101, except for a single nucleotide substitution (GAC-->AAC), changing codon 70 from Asp to Asn. Both latter polymorphisms were so far undetected in DRB alleles. DRB1*1333 could have arisen from a gene conversion event, but DRB1*0426 and DRB5*0109 most likely were generated by point mutation events. For all 3 alleles, the sequence was confirmed by the original hybridization pattern (DRB1*1333) or by hybridization to a newly designed probe (DRB1*0426 and DRB5*0109). Ethnic backgrounds were Lebanese for DRB1*1333 and Caucasian for DRB1*0426 and DRB5*0109.

Alleles↗
PubMed1998

A novel DRB3 allele (DRB3*0208), a new allelic variant of DRB1*1502 (DRB1*15023) and two new DQB1 (DQB1*03012 and DQB1*0614) alleles.

Four novel HLA Class II alleles were identified using CANTYPE reverse hybridization assay. The initial unusual SSO hybridization patterns were confirmed by cloning and sequencing analysis. DRB3*0208 allele is identical to DRB3*0202 except for three nucleotide substitutions (GAT-->AGC) changing codon 57 from Asp to Ser. This polymorphism has so far been undetected in DRB3 alleles. DRB1*15023 differs from DRB1*15021 by a single silent nucleotide substitution (AAC-->AAT, both encoding for Asn) at codon 33. This polymorphism has not, until now, been identified in DRB alleles. Compared with DQB1*03011, the novel DQB1*03012 contains a single silent nucleotide substitution (GCA-->GCG, both encoding for Ala) at codon 38. Finally, DQB1*0614 allele is identical to DQB1*0603 except for a single nucleotide substitution (TAC-->TTC), changing codon 9 from Tyr to Phe. Polymorphisms observed here in the DQB1*03012 and DQB1*0614 alleles are present in several of the known DQB1 alleles. DRB3*0208, DQB1*03012 and DQB1*0614 may have arisen from gene conversion, but the DRB1*15023 most likely was generated by a point mutation event. DQB1*0614 was detected in three related subjects, while each of the other three new alleles has only been detected once.

Alleles↗
PubMed1996

Elimination of neuroblastoma and small-cell lung cancer cells with an anti-neural cell adhesion molecule immunotoxin.

BACKGROUND: The development of immunotoxins has been hampered by difficulties, particularly in solid tumors, of finding appropriate target antigens and of linking sufficiently potent toxins. PURPOSE: We evaluated the tissue specificity of an immunotoxin, N901-blocked ricin (N901-bR), and assessed its potential for eliminating neural cell adhesion molecule (NCAM)-positive tumor cells in conditions appropriate for in vitro purging, prior to autologous stem cell transplantation, and its potential for myelosuppression. N901-bR consists of a monoclonal antibody (MAb), N901, directed against CD56, an antigen of the family of NCAMs, covalently linked to blocked ricin as the cytotoxic effector moiety. METHODS: The tissue specificity of the N901 MAb and the N901-bR immunotoxin was tested against a wide array of human tumor tissues and normal human tissues by immunohistochemical staining. The cytotoxic activity of N901-bR was tested against both small-cell lung cancer (SCLC) cells and neuroblastoma cells, either alone or among normal bone marrow mononuclear cells, and the efficacy of this treatment to specifically eliminate these cells was evaluated in a limiting dilution assay. In addition, normal bone marrow mononuclear cells were incubated with N901-bR, and the toxic effects of the immunotoxin on normal hematopoietic progenitors was evaluated. RESULTS: N901 and N901-bR exhibited specificity for several neoplasms of neuroectodermal origin, including SCLC and neuroblastoma. Staining of normal tissues was essentially limited to various neuroendocrine cells, cardiac muscle cells, and cells in peripheral nerve tissue. We observed a time- and dose-dependent elimination of tumor cells in vitro, with three logs (i.e., > 99.9%) of malignant cells being killed following only 5 hours of exposure to 10 nM N901-bR. Unconjugated N901 MAb specifically blocked the elimination of NCAM-positive cells by N901-bR, whereas neither an isotype-matched control MAb nor galactose (the ligand of native ricin) had any effect on the activity of the immunotoxin, confirming the specificity of its cytotoxic activity. Importantly, N901-bR used under optimal conditions for in vitro tumor cell depletion was not toxic to hematopoietic precursors. CONCLUSIONS: N901-bR has the properties required to target CD56, an antigen present not only on cells from a large number of cancers of neuroendocrine origin, but also on some important normal tissues. In addition, treatment with this immunotoxin results in the highly effective and specific elimination of neuroblastoma and SCLC cells and does not affect normal hematopoietic progenitors. IMPLICATIONS: N901-bR may have clinical utility for purging of neuroblastoma cells and SCLC cells before autologous stem cell transplantation. Further toxicology studies are warranted to assess the potential of N901-bR for in vivo administration.

Antibodies, Monoclonal↗
PubMed1996

Distinctive profile of alveolar macrophage-derived cytokine release induced by fibrogenic and nonfibrogenic mineral dusts.

Groups of 7 Wistar rats each received a single intratracheal instillation of either saline (control), UICC chrysotile B asbestos (5 mg), or very short 4T30 chrysotile asbestos fibers (5 mg). Five animals in each group were killed at 1, 3, and 6 wk posttreatment and analyzed by bronchoalveolar lavage (BAL) for BAL cell populations and cytokine production in conjunction with histopathological assessment of lung tissue. Chrysotile B and short 4T30 chrysotile fibers induced chronic inflammatory reactions characterized by alveolar macrophage (AM) accumulation that resulted, respectively, in lung fibrosis and resolving granuloma. Alveolar macrophages (AM) obtained from rats treated with UICC chrysotile B and short 4T30 chrysotile produced enhanced levels of interleukin-1 (IL-1) and interleukin-6 (IL-6), both spontaneously and in response to lipopolysaccharide (LPS). A different pattern of response was observed for tumor necrosis factor-alpha (TNF-alpha). Fibrogenic chrysotile B caused biphasic changes characterized by significant inhibition of LPS-induced TNF-alpha release by AM 1 and 3 wk after treatment, followed by stimulation of spontaneous and LPS-induced TNF-alpha at 6 wk. In contrast, no significant change in spontaneous and LPS-induced TNF-alpha release was seen with AM from animals with resolving granuloma (4T30 group). Thus, modulation of AM-derived TNF-alpha was correlated under these conditions with the fibrogenic potential of asbestos dusts. These data support a role for TNF-alpha in fibrosis and suggest that TNF-alpha may represent a useful marker of lung damage induced by fibrogenic dusts.

Animals↗
PubMed1994

IFN-gamma up-regulates platelet-activating factor receptor gene expression in human monocytes.

Human monocytes constitutively express the platelet-activating factor receptor (hPAF-R). In this report, we have investigated the modulation of hPAF-R by IFN-gamma. Treatment of monocytes with IFN-gamma caused a time- and concentration-dependent accumulation of hPAF-R mRNA. In contrast, IFN-alpha was inactive. The effect of IFN-gamma was rapid, evident by 1 h of stimulation and reaching a maximum after 2 h. The high level of hPAF-R mRNA was maintained for at least 24 h. Flow cytometry analysis revealed that monocytes treated with IFN-gamma had a two- to sixfold increase in PAF receptor expression at the cell surface, when compared with untreated cells. The increase in hPAF-R expression was associated with an augmented response of IFN-gamma-treated cells to PAF in terms of cytosolic calcium ([Ca2+]i) variations. The IFN-gamma-dependent accumulation of hPAF-R mRNA was not due to the stabilization of hPAF-R mRNA, as shown by unchanged hPAF-R mRNA t1/2. Pretreatment of monocytes with actinomycin D, however, completely abrogated the effect of IFN-gamma, suggesting a transcriptional regulation. Moreover, the up-regulation of hPAF-R mRNA by IFN-gamma was independent of de novo protein synthesis since cycloheximide, an inhibitor of protein synthesis, did not affect this up-regulation. These studies are the first report showing that IFN-gamma regulates hPAF-R gene expression in human monocytes, by a mechanism suggesting transcriptional regulation. This may represent a prototypic example of regulation by lymphocyte-derived cytokines of lipid mediator receptors in myeloid cells, thus adding a novel element in the interrelationship between immune and inflammatory responses.

Base Sequence↗
PubMed1993

Bidirectional modulation of TNF-alpha production by alveolar macrophages in asbestos-induced pulmonary fibrosis.

Bronchoalveolar lavage (BAL) cells were isolated from rats 1, 3, and 6 weeks after a single intratracheal instillation of saline, UICC chrysotile asbestos (5 mg), or silica (5 mg). In asbestos-exposed rats, the pulmonary response was characterized by a significant increase in the number of alveolar macrophages (AMs) and the appearance of fibrotic lesions within 1 week. By contrast, mixed macrophage and neutrophil accumulations were observed in the silica group without evidence of fibrosis. Tumor necrosis factor-alpha (TNF-alpha) production by lipopolysaccharide (LPS)-stimulated BAL cells from asbestos-treated rats was significantly lower than controls 1 and 3 weeks after exposure. However, by 6 weeks higher levels of TNF-alpha production were noticeable in this group. Decreases in LPS-induced TNF-alpha production were also observed with BAL cells from silica-treated animals at all time points studied. Lower levels of TNF-alpha were not related to decreased BAL cell viability or the presence of a significant proportion of neutrophils in the silica group. Furthermore, biphasic changes in TNF-alpha production seen in the asbestos group were correlated with concomitant decreases (3 weeks) and increases (6 weeks) in levels of TNF-alpha mRNA in AMs. These data indicate that lower levels of TNF-alpha resulted from inhibition at the gene expression level and provide evidence for bidirectional modulation of TNF-alpha production by AMs during inflammatory reactions.

Animals↗
PubMed1992

Kinetic properties of type-II ATP diphosphohydrolase from the tunica media of the bovine aorta.

The kinetic properties of type-II ATP diphosphohydrolase are described in this work. The enzyme preparation from the inner layer of the bovine aorta, mostly composed of smooth muscle cells, shows an optimum at pH 7.5. It catalyzes the hydrolysis of tri- and diphosphonucleosides and it requires either Ca2+ or Mg2+ for activity. It is insensitive to ouabain (3 mM), an inhibitor of Na+/K(+)-ATPase, to tetramisole (5 mM), an inhibitor of alkaline phosphatase, and to Ap5A (100 microM), an inhibitor of adenylate kinase. In contrast, sodium azide (10 mM), a known inhibitor for ATPDases and mitochondrial ATPase, is an effective inhibitor. Mercuric chloride (10 microM) and 5'-p-fluorosulfonylbenzoyl adenosine are also powerful inhibitors, both with ATP and ADP as substrates. The inhibition patterns are similar for ATP and DP, thereby, supporting the concept of a common catalytic site for these substrates. Apparent Km and Vmax, obtained with ATP as the substrate, were evaluated at 23 +/- 3 microM and 1.09 mumol Pi/min per mg protein, respectively. The kinetic properties of this enzyme and its localization as an ectoenzyme on bovine aorta smooth muscle cells suggest that it may play a major role in regulating the relative concentrations of extracellular nucleotides in blood vessels.

Animals↗