We have addressed the role of Rho-kinase in the different steps of thrombin receptor agonist peptide (TRAP)-induced platelet activation. Interestingly, under physiological conditions, incubation of platelets with increasing concentrations... more
We have addressed the role of Rho-kinase in the different steps of thrombin receptor agonist peptide (TRAP)-induced platelet activation. Interestingly, under physiological conditions, incubation of platelets with increasing concentrations of the specific Rho-kinase inhibitor Y-27632 resulted in a dose-dependent reversion of the aggregation induced by 10 microM TRAP, without affecting serotonin secretion. Addition of Y-27632 after three minutes of TRAP stimulation, when the maximal aggregation was reached, resulted in a rapid disaggregation of platelets. Accordingly, the early peak of myosin light chain (MLC) phosphorylation induced by TRAP was not affected by Y-27632 but its sustained phosphorylation, observed during the irreversible phase of aggregation, was dependent of Rho-kinase activity. The rapid decrease in MLC phosphorylation upon Y-27632 treatment correlated well with the specific disappearance of myosin heavy chain from the cytoskeleton and preceded platelet disaggregation...
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Phosphoinositides play a central role in many physiological processes by assisting the recruitment of proteins to membranes through specific phosphoinositide-binding motifs. How this recruitment is coordinated in space and time is not... more
Phosphoinositides play a central role in many physiological processes by assisting the recruitment of proteins to membranes through specific phosphoinositide-binding motifs. How this recruitment is coordinated in space and time is not well understood. Here we show that BIN1/M-Amphiphysin2, a protein involved in T-tubule biogenesis in muscle cells and frequently mutated in centronuclear myopathies, clusters PtdIns(4,5)P2 to recruit its downstream partner dynamin. By using several mutants associated with centronuclear myopathies, we find that the N-BAR and the SH3 domains of BIN1 control the kinetics and the accumulation of dynamin on membranes, respectively. We show that phosphoinositide clustering is a mechanism shared by other proteins that interact with PtdIns(4,5)P2, but do not contain a BAR domain. Our numerical simulations point out that clustering is a diffusion-driven process in which phosphoinositide molecules are not sequestered. We propose that this mechanism plays a key role in the recruitment of downstream phosphoinositide-binding proteins.
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The oral Bruton's tyrosine kinase inhibitor, ibrutinib, has recently demonstrated high efficiency in patients with relapsed B-cell malignancies. Occurrence of bleeding events has been reported in a subgroup of ibrutinib-treated... more
The oral Bruton's tyrosine kinase inhibitor, ibrutinib, has recently demonstrated high efficiency in patients with relapsed B-cell malignancies. Occurrence of bleeding events has been reported in a subgroup of ibrutinib-treated patients. We demonstrate that ibrutinib selectively inhibits platelet signaling and functions downstream of the collagen receptor glycoprotein VI and strongly affects firm platelet adhesion on von Willebrand factor (VWF) under arterial flow. A longitudinal study of 14 patients indicated a correlation between occurrence of bleeding events and decreased platelet aggregation in response to collagen in platelet-rich plasma and firm adhesion on VWF under arterial flow. The addition of 50% untreated platelets was sufficient to efficiently reverse the effects of ibrutinib, and platelet functions recovered after treatment interruption as physiological platelet renewal occurred. These data have important clinical implications and provide a basis for hemostasis man...
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Pretreatment of intact platelets with cytochalasin D prevented actin polymerization and cytoskeleton reorganization induced by thrombin, but did not affect platelet aggregation. Under these conditions, synthesis of phosphatidylinositol... more
Pretreatment of intact platelets with cytochalasin D prevented actin polymerization and cytoskeleton reorganization induced by thrombin, but did not affect platelet aggregation. Under these conditions, synthesis of phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P2) stimulated by thrombin was strongly inhibited, while production of phosphatidic acid was unaffected. The inhibitory effect of cytochalasin D was not observed when platelet aggregation was prevented by the RGDS
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Thrombin-induced accumulation of phos- phatidylinositol 3,4-bisphosphate (Ptdlns(3,4)P2) but not of PtdIns(3,4,5,)P3 is strongly correlated with the relocation to the cytoskeleton of 29 % of the p85o~ regulatory subunit of... more
Thrombin-induced accumulation of phos- phatidylinositol 3,4-bisphosphate (Ptdlns(3,4)P2) but not of PtdIns(3,4,5,)P3 is strongly correlated with the relocation to the cytoskeleton of 29 % of the p85o~ regulatory subunit of phosphoinositide 3-kinase (Ptdlns 3-kinase) and is accompanied by a significant increase in Ptdlns 3-kinase activity in this subcellular fraction. Actually, Ptdlns(3,4)P2 accumulation and Ptdlns 3-kinase, pp60 ~-s'~, and p125 eA~ translocations
Research Interests: Cytoskeleton, Signal Transduction, Biological Sciences, Thrombin, Humans, and 13 morePhosphorylation, Actin Cytoskeleton, Platelet activation mechanism, Integrins, Enzyme, Physical Interaction, Multienzyme complexes, Actin Filaments, Tyrosine, Focal Adhesion Kinase, Cell Adhesion Molecules, Tyrosine Phosphorylation, and Cumulant
In this study, we characterised the mechanisms of Rac GTPase activation in human platelets stimulated by two physiological agonists, either thrombin, acting through membrane receptors coupled to heterotrimeric G-proteins, or collagen... more
In this study, we characterised the mechanisms of Rac GTPase activation in human platelets stimulated by two physiological agonists, either thrombin, acting through membrane receptors coupled to heterotrimeric G-proteins, or collagen which is known to mobilise a tyrosine kinase-dependent pathway. Both agonists induced a rapid activation of Rac that was not significantly affected by the inhibition of integrin αIIbβ3 engagement.
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Over the last few years, many reports have extended our knowledge of the inositol lipid metabolism and brought out some exciting information about the location, the variety and the role of phosphoinositides (PIs). Besides the so-called... more
Over the last few years, many reports have extended our knowledge of the inositol lipid metabolism and brought out some exciting information about the location, the variety and the role of phosphoinositides (PIs). Besides the so-called “canonical PI pathway” leading to the production of phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2), the precursor of the intracellular second messengers inositol 1,4,5-trisphosphate and diacylglycerol (DAG), many
Research Interests: Signal Transduction, Cellular Signalling, Humans, Animals, Cell Signalling, and 14 moreLipid metabolism, Actin Cytoskeleton, Enzyme, High Pressure Liquid Chromatography, Time Factors, Metabolic pathway, Vesicle trafficking, Quntitative Thin Layer Chromatography, Cell Surface Markers, Second Messengers, Phosphatidic acid, Biochemistry and cell biology, Phospholipase C, and Cell Membrane
Platelets are critical for the maintenance of the integrity of the vascular system and are the first line of defence against haemorrhage. When they encounter a subendothelial matrix exposed by injury to a vessel, platelets adhere, are... more
Platelets are critical for the maintenance of the integrity of the vascular system and are the first line of defence against haemorrhage. When they encounter a subendothelial matrix exposed by injury to a vessel, platelets adhere, are activated, and become adhesive for other platelets so that they aggregate. αIIb/β3, a platelet-specific integrin, is largely prominent amongst the adhesion receptors and
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Daunorubicin induces apoptosis in myeloid leukemia cells by activation of neutral sphingomyelinase and ceramide generation occurring 4–10 min after daunorubicin addition. We show here that daunorubicin is able to increase the... more
Daunorubicin induces apoptosis in myeloid leukemia cells by activation of neutral sphingomyelinase and ceramide generation occurring 4–10 min after daunorubicin addition. We show here that daunorubicin is able to increase the phosphoinositide 3-kinase activity and enhance intracellular phosphoinositide 3-kinase lipid products prior to ceramide generation. Daunorubicin activates Akt, a downstream phosphoinositide 3-kinase effector. Interestingly, the phosphoinositide 3-kinase inhibitors wortmannin and
Research Interests: Kinetics, Enzyme Inhibitors, Apoptosis, Cell line, Acute Myeloid Leukemia, and 12 moreHumans, Drug Resistance, Uv Radiation, High Performance Liquid Chromatography, Akt, Chromones, Tumor Necrosis Factor–α (TNF), Cell Survival, Ceramides, Biochemistry and cell biology, Insulin Receptor Substrate, and Acute Disease
The effects of cell adhesion on leukemia cell proliferation remain poorly documented and somehow controversial. In this work, we investigated the effect of adhesion to fibronectin on the proliferation of acute myeloid leukemia (AML) cell... more
The effects of cell adhesion on leukemia cell proliferation remain poorly documented and somehow controversial. In this work, we investigated the effect of adhesion to fibronectin on the proliferation of acute myeloid leukemia (AML) cell lines (U937 and KG1a) and CD34+ normal or leukemic primary cells. We observed an increased rate of proliferation of AML cells when adhered to fibronectin,
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Research Interests: Genetics, Phospholipids, Membrane Trafficking, Humans, Mutation, and 17 moreMice, Animals, Neurons, Peripheral Neuropathy, Mouse Model, Rats, Degeneration, Peripheral nerves, Peripheral Nerve, Schwann Cells, PLoS Genetics, CHarcot Marie Tooth, Autosomal Recessive, Charcot Marie Tooth Disease, Axons, Schwann Cell, and Myelin Sheath
The Listeria monocytogenes InlB protein is a 630-amino-acid surface protein that mediates entry of the bacterium into a wide variety of cell types, including hepatocytes, fibroblasts and epithelial cells such as Vero, HEp-2 and HeLa... more
The Listeria monocytogenes InlB protein is a 630-amino-acid surface protein that mediates entry of the bacterium into a wide variety of cell types, including hepatocytes, fibroblasts and epithelial cells such as Vero, HEp-2 and HeLa cells. Invasion stimulates host proteins tyrosine phosphorylation, PI 3-kinase activity and rearrangements in the actin cytoskeleton. We previously showed that InlB is sufficient for entry of InlB-coated latex beads into cells and recent results indicate that purified InlB can stimulate PI 3-kinase activity and is thus the first bacterial agonist of this lipid kinase. In this study, we identified the region of InlB responsible for entry and stimulation of signal transduction events. Eight monoclonal antibodies directed against InlB were raised and, of those, five inhibited bacterial entry. These five antibodies recognized epitopes within the leucine-rich repeat (LRR) region and/or the inter-repeat (IR) region. InlB-staphylococcal protein A (SPA) fusion proteins and recombinant InlB derivatives were generated and tested for their capacity to mediate entry into cultured mammalian cells. All the InlB derivatives that carried the amino-terminal 213-amino-acid LRR region conferred invasiveness to the normally non-invasive bacterium L. innocua or to inert latex beads and the corresponding purified polypeptides inhibited bacterial entry. In addition, the 213-amino-acid LRR region was able to stimulate PI 3-kinase activity and changes in the actin cytoskeleton (membrane ruffling). These properties were not detected with purified internalin, another invasion protein of L. monocytogenes that displays LRRs similar to those of InlB. Taken together, these results show that the first 213 amino acids of InlB are critical for its specific properties.
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Research Interests: Physiology, Apoptosis, Diarrhea, Protein Structure and Function, Humans, and 15 moreMutagenesis, Animals, Salmonella Typhimurium, The, Medical Physiology, Actin Cytoskeleton, Anions, Rats, Epithelial cells, HeLa cells, Experimental Medicine, Hydrolysis, General Physiology, Tight Junctions, and Vacuoles
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Receptor or nonreceptor tyrosine kinases (TKs) are known to play an important role in leukemogenesis. Here we studied the level of protein tyrosine phosphorylations in a series of fresh AML samples and evaluated the effect of TK... more
Receptor or nonreceptor tyrosine kinases (TKs) are known to play an important role in leukemogenesis. Here we studied the level of protein tyrosine phosphorylations in a series of fresh AML samples and evaluated the effect of TK inhibitors. Compared with normal hematopoietic progenitors, a high level of tyrosine phosphorylation was detected in most acute myeloid leukemia (AML) samples. The Src family kinases (SFKs) appeared constitutively activated in most cases, including in the CD34(+)CD38(-)CD123(+) compartment as revealed by the level of phosphorylated tyrosine 416. Lyn was the major SFK family member expressed in an active form in AML cells where it was abnormally distributed throughout the plasma membrane and the cytosol as opposed to normal hematopoietic progenitors. The SFK inhibitor, PP2, strongly reduced the global level of tyrosine phosphorylations, inhibited cell proliferation, and induced apoptosis in patient samples without affecting normal granulomonocytic colony forming units. Moreover, silencing Lyn expression by small interfering RNA in primary AML cells strongly inhibited proliferation. Interestingly, a link between Lyn and the mTOR pathway was observed as PP2 and a Lyn knockdown both affected the phosphorylation of mTOR targets without inhibiting Akt phosphorylation. Lyn should be considered as a novel pharmacologic target for AML therapy.
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Glycosphingolipid- and cholesterol-enriched membrane microdomains, called rafts, can be isolated from several mammalian cells, including platelets. These microdomains appear to play a critical role in signal transduction in several... more
Glycosphingolipid- and cholesterol-enriched membrane microdomains, called rafts, can be isolated from several mammalian cells, including platelets. These microdomains appear to play a critical role in signal transduction in several hematopoietic cells, but their function in blood platelets remains unknown. Herein, we first characterized the lipid composition, including the fatty acid composition of phospholipids, of human platelet rafts. Then their role in platelet activation process was investigated. Interestingly, thrombin stimulation led to morphological changes of rafts correlating with the production of lipid second messengers in these microdomains. Indeed, we could demonstrate for the first time that a large part of the stimulation-dependent production of phosphatidic acid and phosphoinositide 3-kinase products was concentrated in rafts. Moreover, cholesterol depletion with methyl-beta-cyclodextrin disrupted platelet rafts, dramatically decreased the agonist-dependent production of these lipid signaling molecules, and impaired platelet secretion and aggregation. Cholesterol repletion restored the physiological platelet responses. Altogether our data indicate that rafts are highly dynamic platelet membrane structures involved in critical signaling mechanisms linked to the production of lipid second messengers. The demonstration of phosphatidylinositol 3,4,5-trisphosphate production in rafts may have general implications for the understanding of the role of this key second messenger found ubiquitously in higher eucaryotic cells.
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The physiological roles of the class II phosphoinositide 3-kinases (PI3Ks) and their contributions to phosphatidylinositol 3-monophosphate (PI3P) and PI(3,4)P2 production remain elusive. Here we report that mice heterozygous for a... more
The physiological roles of the class II phosphoinositide 3-kinases (PI3Ks) and their contributions to phosphatidylinositol 3-monophosphate (PI3P) and PI(3,4)P2 production remain elusive. Here we report that mice heterozygous for a constitutively kinase-dead PI3K-C2α display aberrant platelet morphology with an elevated number of barbell-shaped proplatelets, a recently discovered intermediate stage in the final process of platelet production. Platelets with heterozygous PI3K-C2α inactivation have critical defects in α-granules and membrane structure, which are associated with modifications in megakaryocytes. These platelets are more rigid and unable to form filopodia following stimulation. Heterozygous PI3K-C2α inactivation in platelets led to a significant reduction in the basal pool of PI3P and a mislocalization of several membrane skeleton proteins known to control the interactions between the plasma membrane and cytoskeleton. These alterations had repercussions on the performance...
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Retinoic acid (RA) induces cell growth arrest and differentiation through two families of nuclear receptors, the RARs and the RXRs. The phosphoinositide 3-kinase (PI3K)/Akt pathway also plays key roles in these processes, that is, cell... more
Retinoic acid (RA) induces cell growth arrest and differentiation through two families of nuclear receptors, the RARs and the RXRs. The phosphoinositide 3-kinase (PI3K)/Akt pathway also plays key roles in these processes, that is, cell cycle progression, cell differentiation and cell survival. We report that, in mouse embryocarcinoma cells (F9 cells), RA induces an early activation of PI3K and Akt
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FcgRIIA, the only Fcg receptor present in platelets, is involved in heparin-associ- ated thrombocytopenia (HIT). Recently, adenosine diphosphate (ADP) has been shown to play a major role in platelet activation and aggregation induced by... more
FcgRIIA, the only Fcg receptor present in platelets, is involved in heparin-associ- ated thrombocytopenia (HIT). Recently, adenosine diphosphate (ADP) has been shown to play a major role in platelet activation and aggregation induced by FcgRIIA cross-linking or by sera from HIT patients. Herein, we investigated the mechanism of action of ADP as a cofactor in FcgRIIA-dependent platelet activation, which is
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Phosphoinositides represent a major class of lipids specifically involved in the organization of signaling cascades, maintenance of the identity of organelles and regulation of multiple intracellular trafficking steps. We previously... more
Phosphoinositides represent a major class of lipids specifically involved in the organization of signaling cascades, maintenance of the identity of organelles and regulation of multiple intracellular trafficking steps. We previously reported that phosphatidylinositol 5-monophosphate (PI5P), produced by the Shigella flexneri phosphatase IpgD, is implicated in the endosomal sorting of the epidermal growth factor receptor (EGFR). Here, we show that the adaptor protein TOM1 is a new direct binding partner of PI5P. We identify the domain of TOM1 involved in this interaction and characterize the binding motif. Finally, we demonstrate that the recruitment of TOM1 by PI5P on signaling endosomes is responsible for the delay in EGFR degradation and fluid-phase bulk endocytosis. Taken together, our data strongly suggest that PI5P enrichment in signaling endosomes prevents endosomal maturation through the recruitment of TOM1, and point to a new function of PI5P in regulating discrete maturation...
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Class IA phosphoinositide 3-kinase β (PI3Kβ) is considered a potential drug target in arterial thrombosis, which is a major cause of death worldwide. Here we show that a striking phenotype of mice with selective p110β deletion in the... more
Class IA phosphoinositide 3-kinase β (PI3Kβ) is considered a potential drug target in arterial thrombosis, which is a major cause of death worldwide. Here we show that a striking phenotype of mice with selective p110β deletion in the megakaryocyte lineage is thrombus instability at a high shear rate, which is an effect that is not detected in the absence of p110α in platelets. The high shear rate-dependent thrombus instability in the absence of p110β is observed both ex vivo and in vivo with the formation of platelet emboli. Moreover, PI3Kβ is required for the recruitment of new platelets to a growing thrombus when a pathological high shear is applied. Treatment of human blood with AZD6482, a selective PI3Kβ inhibitor, phenocopies p110β deletion in mouse platelets, which highlights the role of the kinase activity of p110β. Within the growing platelet thrombus, p110β inactivation impairs the activating phosphorylations of Akt and the inhibitory phosphorylation of GSK3. In accord with...
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PtdIns5P is a lipid messenger acting as a stress-response mediator in the nucleus, and known to maintain cell activation through traffic alterations upon bacterial infection. Here, we show that PtdIns5P regulates actin dynamics and... more
PtdIns5P is a lipid messenger acting as a stress-response mediator in the nucleus, and known to maintain cell activation through traffic alterations upon bacterial infection. Here, we show that PtdIns5P regulates actin dynamics and invasion via recruitment and activation of the exchange factor Tiam1 and Rac1. Restricted Rac1 activation results from the binding of Tiam1 DH-PH domains to PtdIns5P. Using an assay that mimics Rac1 membrane anchoring by using Rac1-His and liposomes containing Ni(2+)-NTA modified lipids, we demonstrate that intrinsic Tiam1 DH-PH activity increases when Rac1 is anchored in a PtdIns5P-enriched environment. This pathway appears to be general since it is valid in different pathophysiological models: receptor tyrosine kinase activation, bacterial phosphatase IpgD expression and the invasive NPM-ALK(+) lymphomas. The discovery that PtdIns5P could be a keystone of GTPases and cytoskeleton spatiotemporal regulation opens important research avenues towards unravel...
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Flavonoids provide a large number of interesting natural compounds that are consumed daily and exhibit more or less potent and selective effects on some signaling enzymes as well as on the growth and proliferation of certain malignant... more
Flavonoids provide a large number of interesting natural compounds that are consumed daily and exhibit more or less potent and selective effects on some signaling enzymes as well as on the growth and proliferation of certain malignant cells in vitro. Among the identified signal transducers, phosphoinositide 3-kinase (PI 3-kinase) and protein kinase C (PKC) are now considered key players in many cellular responses including cell multiplication, apoptosis, and transformation. Despite their lack of strict specificity, some flavonoids provide valuable bases for the design of analogues that could be used to specifically block particular isoforms of PI 3-kinase or PKC and their downstream-dependent cellular responses.
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Activation of the Wnt/beta-catenin pathway has recently been shown to be crucial to the establishment of leukemic stem cells in chronic myeloid leukemia. We sought to determine whether beta-catenin was correlated to clonogenic capacity... more
Activation of the Wnt/beta-catenin pathway has recently been shown to be crucial to the establishment of leukemic stem cells in chronic myeloid leukemia. We sought to determine whether beta-catenin was correlated to clonogenic capacity also in the acute myeloid leukemia (AML) setting. Eighty-two patients were retrospectively evaluated for beta-catenin expression by Western blot. beta-Catenin was expressed (although at various protein levels) in 61% of patients, and was undetectable in the remaining cases. In our cohort, beta-catenin expression was correlated with the clonogenic proliferation of AML-colony forming cells (AML-CFC or CFU-L) in methylcellulose in the presence of 5637-conditioned medium, and more strikingly with self-renewing of leukemic cells, as assessed in vitro by a re-plating assay. In survival analyses, beta-catenin appeared as a new independent prognostic factor predicting poor event-free survival and shortened overall survival (both with P<0.05). Furthermore, variations in beta-catenin protein levels were dependent on post-transcriptional mechanisms involving the Wnt/beta-catenin pathway only in leukemic cells. Indeed, beta-catenin negative leukemic cells were found to increase beta-catenin in response to Wnt3a agonist in contrast to normal counterparts. Altogether, our data pave the way to the evaluation of Wnt pathway inhibition as a new rationale for eradicating the clonogenic pool of AML cells.
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Focal adhesion kinase (FAK) is a nonreceptor tyrosine kinase playing an important role in cell motility and survival. However, very little is known about FAK in normal and leukemic myeloid cells. In this study, FAK protein expression and... more
Focal adhesion kinase (FAK) is a nonreceptor tyrosine kinase playing an important role in cell motility and survival. However, very little is known about FAK in normal and leukemic myeloid cells. In this study, FAK protein expression and mRNA were detected in 25 of 60 cases (42%) of acute myeloid leukemia (AML). Whereas FAK was expressed in 46% of CD34
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Relapses following chemotherapy are a major hindrance to patients’ survival in acute myeloid leukemia (AML). To investigate the role of the hematopoietic niche in the chemoresistance of leukemic cells, we examined two pathways: one... more
Relapses following chemotherapy are a major hindrance to patients’ survival in acute myeloid leukemia (AML). To investigate the role of the hematopoietic niche in the chemoresistance of leukemic cells, we examined two pathways: one mediated by adhesion molecules/integrins, and the other by soluble factors of the morphogen Wnt pathway. In our study, both the adhesion of leukemic blasts to fibronectin
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Depending on their structure, flavonoids display more or less potent inhibitory effects on the growth and proliferation of certain malignant cells in vitro, and these effects are thought to be due to inhibition of various enzymes. We... more
Depending on their structure, flavonoids display more or less potent inhibitory effects on the growth and proliferation of certain malignant cells in vitro, and these effects are thought to be due to inhibition of various enzymes. We investigated the inhibitory action of fourteen flavonoids of different chemical classes on phosphatidylinositol 3-kinase a (PI 3-kinase a) activity, an enzyme recently shown
Research Interests: Flavonoids, Comparative Study, Signal Transduction, In Vitro, Kinase inhibitor, and 11 moreBiochemical, Enzyme, Epidermal Growth Factor Receptor, Quercetin, BIOCHEMICAL PHARMACOLOGY, Biological activity, Polyphenol, Flavonols, Structure activity Relationship, Thin Layer Chromatography, and Tyrosine Kinase
The grape-derived polyphenol resveratrol is anti-proliferative for human liver myofibroblasts, which may be beneficial for the treatment of liver fibrosis. However, its mechanism of action is ill understood. Here, we have studied how... more
The grape-derived polyphenol resveratrol is anti-proliferative for human liver myofibroblasts, which may be beneficial for the treatment of liver fibrosis. However, its mechanism of action is ill understood. Here, we have studied how resveratrol interfered with signaling pathways used by epidermal or platelet-derived growth factors to induce the proliferation of these cells. We found that resveratrol inhibited epidermal growth factor or platelet-derived growth factor-induced DNA synthesis. Resveratrol did not, however, decrease epidermal growth factor receptor autophosphorylation or activation of extracellular regulated kinases, but strongly inhibited the phosphorylation of Akt and of its substrate forkhead related transcription factor. This suggested that resveratrol inhibited epidermal growth factor-induced mitogenic signaling through inhibition of the phosphatidylinositol 3-kinase /Akt pathway. The phosphatidylinositol 3-kinase inhibitor LY 294002, also, inhibited epidermal growth factor-dependent DNA synthesis and Akt phosphorylation but did not decrease extracellular regulated kinases phosphorylation. In contrast, resveratrol inhibited platelet-derived growth factor-stimulated receptor autophosphorylation and every subsequent signaling step. Resveratrol did not directly inhibit phosphatidylinositol 3-kinase activity measured on immunoprecipitates from epidermal growth factor-stimulated myofibroblasts, but it strongly reduced the autophosphorylation of the phosphatidylinositol 3-kinase downstream target phospho-inositide-dependent kinase-1 that phosphorylates Akt. We, thus, show that resveratrol has growth factor-specific effects: it inhibits platelet-derived growth factor signaling via reduced receptor activation, whereas it reduces epidermal growth factor-dependent DNA synthesis via inhibition of the phosphatidylinositol 3-kinase/Akt pathway, possibly through inhibition of phospho-inositide-dependent kinase-1 activity.
Research Interests: Enzyme Inhibitors, Flavonoids, Signal Transduction, Humans, Polyphenols, and 17 moreLiver Cirrhosis, Liver, Liver Fibrosis, ERK, Transcription Factor, Epidermal Growth Factor, Epidermal Growth Factor Receptor, Phenols, Stilbenes, Vitis, Mechanism of action, Cell Proliferation, Polyphenol, Growth Factor, Biochemistry and cell biology, Dna Synthesis, and Myoblasts
Phosphoinositides play a central role in the control of several cellular events including actin cytoskeleton organization. Here we show that, upon infection of epithelial cells with the Gram-negative pathogen Shigella flexneri, the... more
Phosphoinositides play a central role in the control of several cellular events including actin cytoskeleton organization. Here we show that, upon infection of epithelial cells with the Gram-negative pathogen Shigella flexneri, the virulence factor IpgD is translocated directly into eukaryotic cells and acts as a potent inositol 4-phosphatase that specifically dephosphorylates phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P(2)] into phosphatidylinositol 5-monophosphate [PtdIns(5)P] that then accumulates. Transfection experiments indicate that the transformation of PtdIns(4,5)P(2) into PtdIns(5)P by IpgD is responsible for dramatic morphological changes of the host cell, leading to a decrease in membrane tether force associated with membrane blebbing and actin filament remodelling. These data provide the molecular basis for a new mechanism employed by a pathogenic bacterium to promote membrane ruffling at the entry site.