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Biomedical subjects

P Gallay

Publications and source records attributed to P Gallay.

At least 19 recordsLinked to original sources
PubMed2001

A randomized double-blind trial of intravenous trimetazidine as adjunctive therapy to primary angioplasty for acute myocardial infarction.

BACKGROUND: Despite high patency rates, primary angioplasty for myocardial infarction does not necessarily result in optimal myocardial reperfusion and limitation of infarct size. Experimentally, trimetazidine limits infarct size, decreases platelet aggregation, and reduces leukocyte influx into the infarct zone. To assess trimetazidine as adjunctive therapy to primary angioplasty for acute myocardial infarction a prospective, double-blind, placebo-controlled pilot trial was performed. METHODS: 94 patients with acute myocardial infarction were randomized to receive trimetazidine (40 mg bolus followed by 60 mg/day intravenously for 48 h) (n=44) or placebo (n=50), starting before recanalization of the infarct vessel by primary angioplasty. Patients underwent continuous ST-segment monitoring to assess return of ST-segment deviation to baseline and presence of ST-segment exacerbation at the time of vessel recanalization. Infarct size was measured enzymatically from serial myoglobin measurements. Left ventricular angiography was performed before treatment and repeated at day 14. RESULTS: Blinded ST segment analysis showed that despite higher initial ST deviation from baseline in the trimetazidine group (355 (32) vs. 278 (29) microV, P=0.07), there was an earlier and more marked return towards baseline within the first 6 h than in the placebo group (P=0.014) (change: 245 (30) vs. 156 (31) microV respectively, P=0.044). There was a trend towards less frequent exacerbation of ST deviation at the time of recanalization in the trimetazidine group (23.3 vs. 42.2%, P=0.11). There was no difference in left ventricular wall motion at day 14, or in enzymatic infarct size. There was no side effect from treatment. Clinical outcomes were similar between groups. CONCLUSION: Trimetazidine was safe and led to earlier resolution of ST-segment elevation in patients treated by primary angioplasty for acute myocardial infarction.

Adolescent↗
PubMed1997

HIV-1 infection of nondividing cells through the recognition of integrase by the importin/karyopherin pathway.

The karyophilic properties of the HIV-1 nucleoprotein complex facilitate infection of nondividing cells such as macrophages and quiescent T lymphocytes, and allow the in vivo delivery of transgenes by HIV-derived retroviral vectors into terminally differentiated cells such as neurons. Although the viral matrix (MA) and Vpr proteins have previously been shown to play important roles in this process, we demonstrate here that integrase, the enzyme responsible for mediating the integration of the viral genome in the host cell chromosome, can suffice to connect the HIV-1 preintegration complex with the cell nuclear import machinery. This novel function of integrase reflects the recognition of an atypical bipartite nuclear localization signal by the importin/karyopherin pathway.

Cell Differentiation↗
PubMed1997

Long-term pacemaker dependency after radiofrequency ablation of the atrioventricular junction.

This prospective study was conducted to determine the percentage of patients with long-term pacemaker dependency after successful radiofrequency ablation of the atrioventricular junction. Abrupt inhibition of the pacemaker was performed 13.5 +/- 8.1 months after ablation in 59 patients. A > or =5-second asystole was considered to indicate pacemaker dependency. Pacemaker dependency was present in 18 patients. Absence of escape rhythm immediately after ablation was strongly associated with a higher incidence of long-term pacemaker dependency. The following variables were not associated with pacemaker dependency: age, presence of cardiac disease, presence of preablation bundle branch block, number of radiofrequency applications, a bilateral approach for ablation, and continuation of antiarrhythmic therapy after ablation. We concluded that (1) long-term pacemaker dependency is present in 30.5% of the patients after successful atrioventricular junction radiofrequency ablation and (2) absence of escape rhythm immediately after ablation predicts long-term pacemaker dependency.

Aged↗
PubMed1997

The Nef protein of human immunodeficiency virus type 1 enhances serine phosphorylation of the viral matrix.

The human immunodeficiency virus type 1 matrix (MA) protein is phosphorylated during virion maturation on its C-terminal tyrosine and on several serine residues. Whereas MA tyrosine phosphorylation facilitates viral nuclear import, the significance of MA serine phosphorylation remains unclear. Here, we report that MA serine but not tyrosine phosphorylation is strongly enhanced by Nef. Mutations that abrogated the membrane association of Nef and its ability to bind a cellular serine/threonine kinase greatly diminished the extent of virion MA serine phosphorylation. Correspondingly, a protein kinase coimmunoprecipitated with Nef could phosphorylate MA on serine in vitro, producing a phosphopeptide pattern reminiscent of that of virion MA. Recombinant p21-activated kinase hPAK65, a recently proposed relative of the Nef-associated kinase, achieved a comparable result. Taken together, these data suggest that MA is a target of the Nef-associated serine kinase.

Gene Products, gag↗
PubMed1997

Human immunodeficiency virus matrix tyrosine phosphorylation: characterization of the kinase and its substrate requirements.

During virus assembly, a subset of human immunodeficiency virus (HIV) matrix (MA) molecules is phosphorylated on C-terminal tyrosine. This modification facilitates infection of nondividing cells by allowing for the recruitment of the karyophilic MA into the viral core and preintegration complex. MA tyrosine phosphorylation is accomplished by a cellular protein kinase which is incorporated into virions. In this study, we have investigated the nature of this enzyme as well as the determinants of MA necessary for its phosphorylation. Employing an in vitro kinase assay, we found that the MA tyrosine kinase activity is present in various cultured cell lines including CEM and SupT1 T-lymphoid cells, Namalwa B cells, 293 and CV-1 kidney fibroblasts, and P4 HeLa cells. In addition, it could be detected in platelets, macrophages, and activated peripheral blood lymphocytes (PBLs) but not in erythrocytes and resting PBLs isolated from human blood. Subcellular localization of the kinase activity by cell fractionation demonstrated that it is enriched in cellular membranes. In HIV type 2 (HIV-2) particles, the MA tyrosine kinase is associated with the inner leaflet of the viral membrane, while the tyrosine-phosphorylated MA is localized to the core. Individual mutations of each of the last eight residues immediately upstream of the C-terminal tyrosine (Y132) of HIV-1 MA did not prevent Y132 phosphorylation, suggesting that the kinase does not require a highly specific sequence adjacent to the C-terminal tyrosine. Confirming this, we found that the MA of murine leukemia virus, the sequence of which is only moderately homologous to that of HIV-1 and HIV-2 MA, is also C-terminally tyrosine phosphorylated.

3T3 Cells↗
PubMed1996

In vivo gene delivery and stable transduction of nondividing cells by a lentiviral vector.

A retroviral vector system based on the human immunodeficiency virus (HIV) was developed that, in contrast to a murine leukemia virus-based counterpart, transduced heterologous sequences into HeLa cells and rat fibroblasts blocked in the cell cycle, as well as into human primary macrophages. Additionally, the HIV vector could mediate stable in vivo gene transfer into terminally differentiated neurons. The ability of HIV-based viral vectors to deliver genes in vivo into nondividing cells could increase the applicability of retroviral vectors in human gene therapy.

Amino Acid Sequence↗
PubMed1996

Role of the karyopherin pathway in human immunodeficiency virus type 1 nuclear import.

The interaction of the human immunodeficiency virus type 1 (HIV-1) nucleoprotein complex with the cell nuclear import machinery is necessary for viral replication in macrophages and for the establishment of infection in quiescent T lymphocytes. The karyophilic properties of two viral proteins, matrix (MA) and Vpr, are keys to this process. Here, we show that an early step of HIV-1 nuclear import is the recognition of the MA nuclear localization signal (NLS) by Rch1, a member of the karyopherin-alpha family. Furthermore, we demonstrate that an N-terminally truncated form of Rch1 which binds MA but fails to localize to the nucleus efficiently blocks MA- but not Vpr-mediated HIV-1 nuclear import. Correspondingly, NLS peptide inhibits the nuclear migration of MA but not that of Vpr and prevents the infection of terminally differentiated macrophages by vpr-defective virus but not wild-type virus. These results are consistent with a model in which Rch1 or another member of the karyopherin-alpha family, through the recognition of the MA NLS, participates in docking the HIV-1 nucleoprotein complex at the nuclear pore. In addition, our data suggest that Vpr governs HIV-1 nuclear import through a distinct pathway.

Amino Acid Sequence↗
PubMed1996

[Mid-term outcome of escape rhythm after radiofrequency ablation of the atrioventricular junction].

UNLABELLED: Mid-term outcome of the underlying escape rhythm developed after radiofrequency ablation of the atrio-ventricular junction was studied in 50 consecutive patients (28 women and 22 men with a mean age of 66.2 +/- 9.6 years). The escape rhythm was assessed immediately after ablation and after 13.7 +/- 8 months. At the end of ablation: an escape rhythm was present in 38 patients (76%), with a mean rate of 40.7 +/- 9.7 beats/min and a QRS morphology identical to the preablation QRS morphology in 22 patients (58%). At follow-up: an escape rhythm was present in 37 patients (74%), with a slower mean rate of 36.4 +/- 6.8 beats/min (p < 0.05) and an unchanged QRS morphology in 87.5% of the patients. Patients presenting with an escape rhythm at follow-up were more frequently found to have a postablation escape rhythm (p < 0.01). Escape rhythm presence at follow-up was not influenced by age, presence of a cardiac disease, continuation of an antiarrhythmic treatment after ablation, use of a bilateral approach for ablation or number of radiofrequency applications. CONCLUSION: after abrupt inhibition of the stimulation, an escape rhythm was present only in 74% of the patients 13.7 +/- 8 months after atrio-ventricular junction radiofrequency ablation. QRS morphology was identical to the preablation morphology in 57% of the patients.

Aged↗
PubMed1995

HIV nuclear import is governed by the phosphotyrosine-mediated binding of matrix to the core domain of integrase.

The karyophilic properties of the viral matrix (MA) protein govern HIV nuclear import in nondividing cells such as macrophages. A critical regulator of this process is the C-terminal tyrosine phosphorylation of MA during virus maturation. Here, we reveal the mechanism of this phenomenon, by demonstrating that tyrosine phosphorylation induces the binding of MA to integrase (IN). This leads to the incorporation of MA molecules into virus cores, and subsequently into uncoated viral nucleoprotein complexes. A direct interaction between tyrosine-phosphorylated MA and the central domain of IN can be demonstrated in vitro. It is blocked by phosphotyrosine, indicating that IN recognizes the phosphorylated C-terminal residue of MA. These results explain how the karyophilic potential of MA is conferred to the HIV nucleoprotein complex.

Base Sequence↗
PubMed1995

HIV-1 infection of nondividing cells: C-terminal tyrosine phosphorylation of the viral matrix protein is a key regulator.

The HIV-1 matrix (MA) protein contains two subcellular localization signals with opposing effects. A myristoylated N-terminus governs particle assembly at the plasma membrane, and a nucleophilic motif facilitates import of the viral preintegration complex into the nucleus of nondividing cells. Here, we show that myristoylation acts as the MA dominant targeting signal in HIV-1 producer cells. During virus assembly, a subset of MA is phosphorylated on the C-terminal tyrosine by a virion-associated cellular protein kinase. Tyrosine-phosphorylated MA is then preferentially transported to the nucleus of target cells. An MA tyrosine mutant virus grows normally in dividing cells, but is blocked for nuclear import in terminally differentiated macrophages. MA tyrosine phosphorylation thus reveals the karyophilic properties of this protein within the HIV-1 preintegration complex, thereby playing a critical role for infection of nondividing cells.

Amino Acid Sequence↗
PubMed1995

Characterisation of bovine lipopolysaccharide binding protein and the in vivo acute phase response to Pasteurella haemolytica Type A.

Lipopolysaccharide binding protein (LBP) is a liver-derived acute phase protein which is implicated in modulating the host responses to lipopolysaccharide (LPS) from Gram-negative bacteria. LBP interacts with circulatory LPS to form complexes which bind to the CD14 receptor or cells of the monocytic lineage and neutrophils resulting in their activation. This causes the release of mediators and cytokines which are responsible for initiating the acute phase response. LBP-like activity has now been identified in bovine serum and in this study LBP has been purified from acute phase bovine serum using ion exchange chromatography. On sodium dodecyl sulphate polyacrylamide electrophoresis, bovine LBP demonstrated a single band with a molecular mass of 58 kDa. Bovine LBP enhanced the binding of LPS to human monocytes while enzymatic removal of the CD14 receptor abrogated this interaction. Furthermore, bovine LBP increased the sensitivity of monocytes to LPS by at least 100-fold. Depletion of LBP by means of antibodies to bovine LBP inhibited the serum mediated LPS binding to monocytes. Antibodies to rabbit LBP or recombinant human LBP did not cross-react with bovine LBP. Studies on the kinetics of LBP activity in calves during the acute phase response demonstrated a four-fold increase in the serum concentration 36 h after a single intratracheal inoculation of Pasteurella haemolytica A1. The findings of this study indicate that cattle possess a LPS detection mechanism comparable to that described in man and experimental animals in which LBP forms complexes in serum with circulatory LPS enhancing the signal to the immune system to mount a host response. The isolation of LBP will allow further investigations into LBP-mediated responses to LPS in cattle.

Acute-Phase Proteins↗
PubMed1995

Lipopolysaccharide binding protein as a marker of inflammation in synovial fluid of patients with arthritis: correlation with interleukin 6 and C-reactive protein.

OBJECTIVE: To determine levels of lipopolysaccharide binding protein (LBP) in serum and in synovial fluid (SF) of patients presenting with various articular disorders [degenerative arthritis, rheumatoid arthritis (RA), reactive arthritis (ReA)] and to correlate these levels with C-reactive protein (CRP) and interleukin 6 (IL-6), 2 markers of the acute phase response. METHODS: LBP was measured by a radioimmunoassay made up of lipopolysaccharide (LPS) to capture LBP and radiolabelled anti-LBP antibodies to detect LBP. LBP was also measured for its ability to present fluorescein isothiocyanate LPS (FITC-LPS) to human monocytes. CRP was measured by nephelometry and IL-6 bioassay. RESULTS: Levels of LBP in serum and in SF were significantly higher in patients with RA and ReA than in the control group of degenerative arthropathies. In the latter group, LBP values were similar to those found in controls. Serum LBP values correlated positively with SF LBP values. LBP values also correlated with CRP and IL-6 levels measured in SF. Functionally, LBP was found to be active and able to present LPS to monocytes, resulting in tumor necrosis factor-alpha (TNF-alpha) release upon LPS challenge. CONCLUSION: These in vitro data support the observation that LBP could play a major role in local joint disorders. Our results also strengthen the view that LBP may be a new marker of synovial inflammation.

Acute-Phase Proteins↗
PubMed1994

Mode of action of anti-lipopolysaccharide-binding protein antibodies for prevention of endotoxemic shock in mice.

Lipopolysaccharide (LPS)-binding protein (LBP) has been shown to regulate the response of monocytes to LPS in vitro. In a previous study, polyclonal anti-LBP IgGs were found to protect D-galactosamine-sensitized mice against a lethal endotoxemic shock induced by a low challenge of LPS or lipid A when administered simultaneously with endotoxin. In the present study, we investigated the mode of action of these anti-LBP IgGs. In vitro, we demonstrated that they interfere with LPS binding to monocytes or polymorphonuclear cells in different ways: by the mere prevention of binding of LPS to LBP thus preventing the binding of LPS to CD14, or by reacting with LPS-LBP complexes thus mediating their binding to complement or Fc receptors on monocytes and on polymorphonuclear cells. In vivo, we demonstrated that anti-LBP IgGs afforded protection against lethal endotoxemic shock by one of two mechanisms. First, LBP blockade by pretreatment with anti-LBP IgG allowed protection against a low dose of LPS (100 ng). This protection occurred despite LPS levels in blood similar to those in control mice but in the absence of detectable tumor necrosis factor (TNF). This demonstrated that anti-LBP IgG could block the LBP-mediated TNF release upon LPS challenge. In contrast, anti-LBP IgG did not afford protection in mice not sensitized with D-galactosamine and challenged with high-dose LPS (1 mg), confirming that LPS at high concentrations could stimulate cells independently of the LBP pathway. Second, anti-LBP treatment administered simultaneously with LPS challenge protected mice against both low and high doses of LPS. Unlike after pretreatment with anti-LBP IgG, this protection was accompanied by a decrease of circulating LPS, suggesting that anti-LBP IgG in these conditions facilitated clearance of LPS probably by clearing LPS-LBP complexes. These data and the fact that LBP binds to all LPS through lipid A suggest that antibody directed to LBP could be a candidate for therapeutic strategies in endotoxemic shock.

Acute-Phase Proteins↗
PubMed1994

Radioimmunoassay versus flow cytometric assay to quantify LPS-binding protein (LBP) concentrations in human plasma.

LPS-binding protein (LBP) is an acute phase protein present in plasma, that has been shown to play a major role in sensitizing monocytes to LPS. We describe here two assays to quantify LBP in plasma. The first assay made use of monophosphoryl lipid A to capture LBP in human plasma, and LBP was detected by radiolabeled anti-LBP IgG. The second assay measured LBP by flow cytometry, using LBPs ability to present LPS to the CD14 receptor present on monocytes. Both assays had a similar level of sensitivity, that allowed the quantification of LBP in human plasma.

Acute-Phase Proteins↗
PubMed1994

Interferon gamma receptor deficient mice are resistant to endotoxic shock.

Antibody neutralization studies have established interferon gamma (IFN-gamma) as a critical mediator of endotoxic shock. The advent of IFN-gamma receptor negative (IFN gamma R-/-) mutant mice has enabled a more direct assessment of the role of IFN-gamma in endotoxin (lipopolysaccharide [LPS]-induced shock. We report that IFN gamma R-/- mice have an increased resistance to LPS-induced toxicity, this resistance manifesting well before the synthesis and release of LPS-induced IFN-gamma. LPS-induced lymphopenia, thrombocytopenia, and weight loss seen in wild-type mice were attenuated in IFN gamma R-/- mice. IFN gamma R-/- mice tolerated 100-1,000 times more LPS than the minimum lethal dose for wild-type mice in a D-galactosamine (D-GalN)/LPS model. Serum tumor necrosis factor (TNF) levels were 10-fold reduced in mutant mice given LPS or LPS/D-GalN. Bone marrow and splenic macrophages from IFN gamma R-/- mice had a four- to sixfold decreased LPS-binding capacity which correlated with similar reduction in CD14. Serum from mutant mice reduced macrophage LPS binding by a further 50%, although LPS binding protein was only 10% reduced. The expression of TNF receptor I (p55) and II (p75) was identical between wild-type and mutant mice. Thus, depressed TNF synthesis, diminished expression of CD14, and low plasma LPS-binding capacity, in addition to blocked IFN-gamma signaling in the mutant mice, likely to combine to manifest in the resistant phenotype of IFN gamma R-/- mice to endotoxin.

Animals↗
PubMed1994

Cytokine regulation of TNF-alpha mRNA and protein production by unprimed macrophages from C57Bl/6 and NZW mice.

It is well known that bacterial lipopolysaccharide (LPS) induces the synthesis of tumor necrosis factor alpha (TNF-alpha) and other inflammatory cytokines by primary monocytes and macrophages and that the Th1 lymphokines, interleukin-2 (IL-2) and interferon-gamma (IFN-gamma), augment this response. We investigated the ability of IL-2 and IFN-gamma to induce the production of TNF-alpha mRNA and protein independently of LPS and the modulation of this response by macrophage colony-stimulating factor (M-CSF) and IL-10. We found that IL-2 and IFN-gamma were both able to induce the accumulation of TNF-alpha mRNA, albeit with slower kinetics than LPS, and that they acted synergistically. However, very little TNF bioactivity was secreted by lymphokine-stimulated macrophages unless LPS was also added. This finding underscores the importance of translational effects in the control of TNF production. M-CSF and IL-10 strongly inhibited TNF production at the level of both mRNA and bioactivity but had no effect on the production of IL-6. Bone marrow-derived or thioglycollate-elicited macrophages from the NZW mouse strain, which have been reported to be deficient in their ability to produce TNF, were at least as responsive to LPS or lymphokines as those taken from the C57Bl/6 strain and were similarly affected by M-CSF and IL-10. Therefore, the genetic defect of NZW mice is not a primary deficiency in TNF production.

Animals↗