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Alain Bernard

Publications and source records attributed to Alain Bernard.

29 records · Page 2Linked to original sources
PubMed2003

Involvement of a CD47-dependent pathway in platelet adhesion on inflamed vascular endothelium under flow.

Resting platelet adhesion to inflammatory vascular endothelium is thought to play a causal role in secondary thrombus formation or microcirculatory disturbance after vessel occlusion. However, though adhesion receptors involved in platelet-matrix interactions have been extensively studied, the molecular mechanisms involved in platelet-endothelium interactions are incompletely characterized and have been mainly studied under static conditions. Using human platelets or platelets from wild-type and CD47-/- mice in whole blood, we demonstrated that at low shear rate, CD47 expressed on human and mouse platelets significantly contributes to platelet adhesion on tumor necrosis factor-alpha (TNF-alpha)-stimulated vascular endothelial cells. Using the CD47 agonist peptide 4N1K and blocking monoclonal antibodies (mAbs), we showed that CD47 binds the cell-binding domain (CBD) of endothelial thrombospondin-1 (TSP-1), inducing activation of the platelet alphaIIbbeta3 integrin that in turn becomes able to link the endothelial receptors intercellular adhesion molecule 1 (ICAM-1) and alphavbeta3. Platelet CD36 and GPIbalpha are also involved because platelet incubation with blocking mAbs directed against each of these 2 receptors significantly decreased platelet arrest. Given that anti-CD47 treatment of platelets did not further decrease the adhesion of anti-CD36-treated platelets and CD36 is a TSP-1 receptor, it appears that CD36/TSP-1 interaction could trigger the CD47-dependent pathway. Overall, CD47 antagonists may be potentially useful to inhibit platelet adhesion on inflamed endothelium.

Animals↗
PubMed2002

The chaperone protein 14-3-3 interacts with 3BP2/SH3BP2 and regulates its adapter function.

Lymphocyte stimulation by immunoreceptors is achieved through the activation of multiple signaling pathways leading to cytokine gene transcription. Adapter proteins are critical signaling components that can integrate multiple pathways by allowing the assembly of multimolecular signaling complexes. We previously showed that the cytoplasmic adapter 3BP2 (also known as SH3BP2) promotes NFAT/AP-1 transcriptional activities in T cells through the activation of Ras- and calcineurin-dependent pathways. However, the molecular mechanisms by which 3BP2/SH3BP2 regulates cell signaling and activation remain poorly documented. In this study, using a combination of yeast two-hybrid analysis and biochemical approaches, we present evidence for a physical interaction between 3BP2 and the chaperone protein 14-3-3. This interaction was direct and constitutively detected in yeast and in mammalian cells. Phorbol ester, pervanadate, and forskolin/isobutylmethylxanthine stimulations enhanced this interaction, as well as co-expression of constitutive active mutants of serine/threonine kinases, including protein kinase C. We found that dephosphorylation of 3BP2 by alkaline phosphatase disrupted its interaction with 14-3-3 and that 3BP2 was a substrate of purified protein kinase C in vitro, suggesting that the phosphorylation of 3BP2 by upstream kinases was required for 14-3-3 binding. Using deletion mutants of 3BP2, two 14-3-3 binding domains were mapped to two proline-rich (residues 201-240 and 270-310) domains of 3BP2. These domains were shown to contain two 14-3-3 consensus binding motifs. We identified residues Ser(225) and Ser(277) of 3BP2 as being essential for interaction with 14-3-3 family proteins, optimal 3BP2 serine phosphorylation, and then for 3BP2-dependent function. Indeed, a 3BP2 mutant protein incapable of binding 14-3-3 showed increased capacity to stimulate NFAT transcriptional activities, suggesting that 14-3-3 binding to 3BP2 negatively regulates 3BP2 adapter function in lymphocytes.

14-3-3 Proteins↗
PubMed2002

CD99 isoforms expression dictates T cell functional outcomes.

CD99, a unique integral membrane protein present on the surface of all human T cells, has previously been shown to regulate cell function and fate. In peripheral T cells, it triggers immediate activation of alpha4b1 integrin and cell arrest on inflamed vascular endothelium, whereas it mediates an apoptotic signal in double-positive thymocytes undergoing the selection process. Two isoforms of CD99 exist, a long form corresponding to the full-length protein and a short form harboring a deletion in the intracytoplasmic segment. Here, we show that while peripheral T cells display exclusive expression of the long form, double-positive thymocytes express both isoforms. Moreover, differential expression of these two CD99 molecules can lead to distinct functional outcomes. Expression of the long form in a CD99-deficient Jurkat T cell line is sufficient to promote CD99-induced cell adhesion, whereas coexpression of the two isoforms is required to trigger T-cell death. When coexpressed, the two proteins form covalent heterodimers, which locate within glycosphingolipidic rafts and induce sphingomyelin degradation. Cholesterol depletion experiments show that this localization is required for the induction of apoptosis. Thus, the surface expression pattern of CD99 isoforms determines T-cell functional outcomes.

12E7 Antigen↗
PubMed2002

Signaling through ZAP-70 is required for CXCL12-mediated T-cell transendothelial migration.

Transendothelial migration of activated lymphocytes from the blood into the tissues is an essential step for immune functions. The housekeeping chemokine CXCL12 (or stroma cell-derived factor-1alpha), a highly efficient chemoattractant for T lymphocytes, drives lymphocytes to sites where they are highly likely to encounter antigens. This suggests that cross-talk between the T-cell receptor (TCR) and CXCR4 (the CXCL12 receptor) might occur within these sites. Here we show that the zeta-associated protein 70 (ZAP-70), a key element in TCR signaling, is required for CXCR4 signal transduction. The pharmacologic inhibition of ZAP-70, or the absence of ZAP-70 in Jurkat T cells and in primary CD4(+) T cells obtained from a patient with ZAP deficiency, resulted in an impairment of transendothelial migration that was rescued by the transfection of ZAP-70. Moreover, the overexpression of mutated forms of ZAP-70, whose kinase domain was inactivated, also abrogated the migratory response of Jurkat T cells to CXCL12. In contrast, no involvement of ZAP-70 in T-cell arrest on inflammatory endothelium under flow conditions or in CXCL12-induced actin polymerization was observed. Furthermore, CXCL12 induced time-dependent phosphorylation of ZAP-70, Vav1, and extracellular signal-regulated kinases (ERKs); the latter were reduced in the absence of functional ZAP-70. However, though a dominant-negative Vav1 mutant (Vav1 L213A) blocked CXCL12-induced T-cell migration, pharmacologic inhibition of the ERK pathway did not affect migration, suggesting that ERK activation is dispensable for T-cell chemotaxis. We conclude that cross-talk between the ZAP-70 signaling pathway and the chemokine receptor CXCR4 is required for T-cell migration.

CD4-Positive T-Lymphocytes↗
PubMed2002

Vav1 couples T cell receptor to serum response factor-dependent transcription via a MEK-dependent pathway.

The Vav family of guanine nucleotide exchange factors for Rho family GTPases plays a critical role in lymphocyte proliferation, gene transcription, and cytoskeleton reorganization following immunoreceptor stimulation. However, its role in immediate early gene activation is unclear. In this study, we have investigated the mechanisms by which Vav1 can regulate c-fos serum response element transcriptional activity. We show that T cell antigen receptor (TCR) stimulation induces the phosphorylation of serum response factor (SRF) on serine 103 and increases the binding of SRF complexes on serum response element in a MEK- and p38-dependent pathway. The physiological relevance of our findings is supported by the inhibition of the interleukin-2 gene transcriptional activity by a dominant negative SRF mutant. Overexpression of Vav1, which partially mimics TCR stimulation, promotes SRF-dependent transcription, and dominant negative Vav1 mutants block SRF activation by TCR. SRF activation by Vav1 occurs through a signaling cascade consisting of Rac1/Cdc42 and the serine/threonine kinases Pak1 and MEK, but independently of the phosphatidylinositol 3-kinase pathway. Interestingly, Vav2 also enhances SRF through Rho GTPases, suggesting that Vav proteins are general regulators of SRF activation in lymphocytes. This report establishes Vav proteins as a direct link between antigen receptors and SRF-dependent early gene expression.

Antibodies, Monoclonal↗
PubMed2002

Early CD4(+) T cell recovery in human immunodeficiency virus-infected patients receiving effective therapy is related to a down-regulation of apoptosis and not to proliferation.

This prospective study investigated the contributions of apoptosis and proliferation of CD4(+) T cells obtained by the introduction of a new antiretroviral treatment for human immunodeficiency virus infection. Virus load; T cell counts; apoptosis of T cell subsets, including naive cells; and proliferation were determined from treatment initiation to the third month in a cohort of patients. An increase in CD4(+) T cell count > or = 100 cells/microL over baseline was considered to be a satisfactory immune reconstitution. Sixty-nine patients completed the protocol, 22 of whom met our definition of a satisfactory immune reconstitution, showing a significantly more pronounced reduction in spontaneous CD4(+) T cell apoptosis at month 1 as well as month 3, compared with the other patients. In contrast, neither Fas-induced apoptosis down-regulation nor Fas-induced increased proliferation capacity was associated with a satisfactory immune reconstitution. Down-regulation of CD4(+) T cell apoptosis by antiretroviral treatment is the main mechanism associated with early CD4(+) T cell increase.

Adult↗
PubMed2002

The two-signal model of T-cell activation after 30 years.

The two-signal model of T-cell activation is still valid after 30 years. The recent understanding of the first signal intricacy and its tight relationship with the second signal have thrown decisive light on T-cell activation processes and the complex molecular events that occur on the surface and within the T cell. Furthermore, the recognition of numerous accessory pathways that, in addition to the CD28 and CD40 pathways, operate to manage antigen-presenting cell T cell cooperation in view of lymphocyte activation, disclose the exquisite and numerous regulatory events that compose the second signal.

Animals↗
PubMed2002

Early and late mortality after pleurodesis for malignant pleural effusion.

BACKGROUND: The purpose of this study is to analyze morbidity and mortality and to determine the relative contribution of each of these potential prognosis variables for predicting morbidity and mortality in patients after pleurodesis by thoracotomy or thoracoscopy. METHODS: Between March 1, 1996, and January 31, 2001, a total of 70 patients underwent pleurodesis for recurrent malignant pleural effusion. Thoracoscopy was performed in 54 patients (77%); pleurodesis was achieved by pleural abrasion (n = 15), pleurectomy (n = 5), and talc insufflation (n = 34). Thoracotomy was performed in 16 patients (23%) who also needed pleurectomy and decortication for a trapped lung. RESULTS: Postoperative complications occurred in 24 patients (34%). Factors adversely affecting morbidity with univariate analysis included: three or four metastatic sites (p = 0.003), and thoracotomy (p = 0.009). Factors adversely affecting morbidity with multivariate analysis included: thoracotomy (p = 0.0005) and number of metastatic sites (p = 0.007). Six patient deaths (8.6%) occurred during hospitalization. Factors adversely affecting in-hospital mortality with univariate analysis included: Eastern Cooperative Oncology Group Performance Status 2 to 3 (p = 0.001), lower preoperative serum hemoglobin (p = 0.001), lower preoperative serum albumin (p = 0.0001), and thoracotomy (p = 0.03). Factors adversely affecting in-hospital mortality with multivariate analysis included: preoperative serum albumin less than 60 g/L (p = 0.007) and ECOG Performance Status 2 to 3 (p = 0.008). Twelve patients (17%) died within 90 days after surgery. Factors adversely affecting 3-month mortality with univariate analysis included: ECOG Performance Status 2 to 3 (p = 0.001), lower preoperative serum hemoglobin (p = 0.03), higher preoperative white cells (p = 0.03), lower preoperative serum albumin (p = 0.03), and preoperative thoracentesis more than once per month (p = 0.03). Factors adversely affecting 3-month mortality with multivariate analysis included: ECOG Performance Status 2 to 3 (p = 0.01), preoperative thoracentesis more than once per month (p = 0.03), three or four metastatic sites (p = 0.02), and preoperative white blood cell count > or = 12,000/mm3 (p = 0.03). CONCLUSIONS: Thoracotomy is not indicated in patients with a malignant effusion because of poor survival, a high frequency of complications, and prolonged hospital stay. Pleurodesis thoracoscopy is indicated in patients with good performance status coupled with good nutrition.

Aged↗
PubMed2002

Completion pneumonectomy: factors affecting operative mortality and cardiopulmonary morbidity.

BACKGROUND: The purpose of this report is to analyze preoperative and perioperative factors affecting operative mortality and cardiopulmonary morbidity after a completion pneumonectomy. METHODS: We retrospectively reviewed all patients who underwent completion pneumonectomy from January 1985 through September 1998 at the Mayo Clinic in Rochester, MN. Factors affecting operative mortality and postoperative morbidity and were analyzed using univariate and multivariate analysis. RESULTS: There were 115 patients (73 men and 42 women), with a median age of 64 years (range, 12 to 83 years). Indication for pneumonectomy was benign disease in 57 patients (49.6%), lung cancer in 51 (44.3%) and metastatic disease in 7 (6.1%). There were 24 deaths (mortality 20.9%, 95% CI 13.9% to 29.4%). Mortality for patients undergoing completion pneumonectomy for benign disease, lung cancer, and metastatic cancer was 26.3%, 17.6%, and 0%, respectively (p = 0.24). Factors adversely affecting mortality with univariate analysis included advanced age (p = 0.004), preoperative corticosteriod use (p = 0.01), decreased preoperative diffusion capacity of lung to carbon monoxide (p = 0.01), intraoperative blood transfusion (p = 0.04), and excessive crystalloid infusion within the first 12 hours (p = 0.01) and 24 hours (0.03) postoperatively, respectively. Factors adversely affecting mortality with multivariate analysis included advanced age (p = 0.001), preoperative corticosteriod use (p = 0.002), and low preoperative hemoglobin (p = 0.02). Cardiopulmonary complications occurred in 72 patients (63.7%). Factors adversely affecting morbidity with univariate analysis included benign disease (p = 0.002), decreased preoperative diffusion capacity of lung to carbon monoxide (p = 0.04), bronchial stump reinforcement (p = 0.0001), and excessive crystalloid infusion within the first 12 hours (p = 0.006) and 24 hours (p = 0.02) postoperatively, respectively. Factors adversely affecting morbidity with multivariate analysis included advanced age (p = 0.005) and bronchial stump reinforcement (p = 0.001). CONCLUSIONS: Multiple factors adversely affect operative mortality and cardiopulmonary morbidity after completion pneumonectomy. Although completion pneumonectomy remains a high-risk procedure, especially for benign disease, it still should be considered a treatment option in selected patients.

Adolescent↗
PubMed2002

Relaxant effects of selective phosphodiesterase inhibitors on U46619 precontracted human intralobar pulmonary arteries and role of potassium channels.

We examined the influence of K+ channel antagonists on the vasorelaxation induced by theophylline (non selective PDEI), siguazodan (PDE3I), rolipram (PDE4I) and zaprinast (PDE5I) in human intralobar pulmonary arteries. All PDEI tested induced a concentration-dependent relaxation with theophylline being significantly (p < 0.05) more efficient and rolipram more potent than PDE5I and PDE3I (Emax values, expressed as a percentage of maximal relaxation by papaverine 10(-4)M, were 92% +/- 2%, 84% +/- 8%, 90% +/- 4% and 99% +/- 1%, and pD2 values were 7.30 +/- 0.35, 6.14 +/- 0.25, 5.86 +/- 0.17, and 4.85 +/- 0.47 for rolipram, siguazodan, zaprinast and theophylline, respectively). 4-Aminopyridine (4-AP, Kv, voltage dependent channel blocker, 1 mM) induced a significant increase (+17% p < 0.05) of U46619-induced vasoconstriction whereas the other K+-channels blockers, glibenclamide (KATP channels, 1 microM) charybdotoxin (predominant BKCa, large conductance Ca2+-sensitive K+ channels, 0.1 microM) and apamine (SKCa, small conductance, 0.3 microM) were without effect. The concentration response curves (CRC) for rolipram were significantly shifted to the right by glibenclamide (1 microM), charybdotoxin (0.1 microM) and 4-AP (1 mM). The CRC for siguazodan was significantly displaced to the right by 4-AP. None of the potassium channel blockers displaced the CRC for zaprinast and theophylline. Apamine was without effect on the CRC for all the PDEI used in this study. (1) PDE3, 4 and 5 are functionally present in human intralobar pulmonary arteries; (2) the vasoconstriction induced by U46619 is downregulated by 4-aminopyridine sensitive-K+ channels; (3) the relaxant effects of rolipram (PDE4I) are partly mediated through KATP, BKCa, and Kv potassium channels and those of siguazodan (PDE3I) by Kv potassium channels.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗