Syk-Mediated Translocation of PI3Kδ to the Leading Edge Controls Lamellipodium Formation and Migration of Leukocytes

The non-receptor tyrosine kinase Syk is mainly expressed in the hematopoietic system and plays an essential role in β2 integrin-mediated leukocyte activation. To elucidate the signaling pathway downstream of Syk during β2 integrin (CD11/CD18)-mediated migration and extravasation of polymorphonuclear neutrophils (PMN), we generated neutrophil-like differentiated HL-60 (dHL-60) cells expressing a fluorescently tagged Syk mutant lacking the tyrosine residue at the position 323 (Syk-Tyr323) that is known to be required for the binding of the regulatory subunit p85 of the phosphatidylinositol 3-kinase (PI3K) class IA. Syk-Tyr323 was found to be critical for the enrichment of the catalytic subunit p110δ of PI3K class IA as well as for the generation of PI3K products at the leading edge of the majority of polarized cells. In accordance, the translocation of PI3K p110δ to the leading edge was diminished in Syk deficient murine PMN. Moreover, the expression of EGFP-Syk Y323F interfered with proper cell polarization and it impaired efficient migration of dHL-60 cells. In agreement with a major role of β2 integrins in the recruitment of phagocytic cells to sites of lesion, mice with a Syk-deficient hematopoietic system demonstrated impaired PMN infiltration into the wounded tissue that was associated with prolonged cutaneous wound healing. These data imply a novel role of Syk via PI3K p110δ signaling for β2 integrin-mediated migration which is a prerequisite for efficient PMN recruitment in vivo

Visualization of Src activity at different compartments of the plasma membrane by FRET imaging

SummaryMembrane compartments function as segregated signaling platforms for different cellular functions. It is not clear how Src is regulated at different membrane compartments. To visualize local Src activity in live cells, a FRET-based Src biosensor was targeted in or outside of lipid rafts at the plasma membrane, via acylation or prenylation modifications on targeting tags either directly fused to the biosensor or coupled to the biosensor through an inducible heterodimerization system. In response to growth factors and pervanadate, the induction of Src activity in rafts was slower and weaker, dependent on actin and possibly its mediated transportation of Src from perinuclear regions to the plasma membrane. In contrast, the induction of Src activity in nonrafts was faster and stronger, dependent on microtubules. Hence, Src activity is differentially regulated via cytoskeleton at different membrane compartments

Design, synthesis and biological evaluation of new anticoagulant heterocyclic compounds for rodenticide use

A ce jour, les anticoagulants commerciaux souffrent de deux inconvénients majeurs : leur rémanence avec, pour certains d’entre eux, une demi-vie hépatique proche des 300 jours causant des intoxications secondaires sur les prédateurs des rongeurs, ainsi que le développement de nombreuses mutations génétiques causé par l’utilisation intensive de ces composés, rendant inopérant l’utilisation de certains AVKs commerciaux. Face à ce constat, l’Union Européenne envisage d’interdire l’utilisation de tels composés. La mission prioritaire est donc de trouver un anticoagulant capable de gérer les populations de rongeurs sans affecter leurs prédateurs. Les recherches mises en avant par Adrien Montagut (Thèse 2011-2014) ont permis d’aboutir à une structure type d’anticoagulants, dérivés de la 4-hydroxycoumarine. Actuellement, AMR361 a été testé in vitro sur l’ensemble des mutations de VKORC1 et in vivo sur rats sauvages, et constitue le premier AVK développé qui répond à l’ensemble des caractéristiques du cahier des charges initial. La première partie de mon projet de thèse consistait à compléter l’étude biologique sur le noyau 4-hydroxycoumarine en amenant de la diversité fonctionnelle sur la position para du noyau aromatique. D’un point de vue biologique, l’allongement du bras espaceur sur la chaîne latérale par l’utilisation de fonctions telles que les amides ou amides inversés ou l’introduction d’un groupement diméthyle sur le pont méthylène ont été étudiés afin d’analyser les paramètres d’efficacité et de rémanence. Cependant, la plupart des composés synthétisés appartenant à la famille des 4-hydroxycoumarines font déjà l’objet d’un brevet déposé par l’entreprise Liphatech en 1999. L’étude de nouveaux noyaux, dont certains sont analogues à la 4-hydroxycoumarine, de même que la fonctionnalisation du noyau 4-hydroxycoumarine sur la partie aromatique, a permis l’accès à des structures plus diverses. Ces perspectives originales pour l’innovation ont été introduites pour contourner les brevets déjà existants.To date, commercial anticoagulants suffer from two major inconveniences: their persistence causing secondary poisoning of rodent predators and the development of many genetic mutations caused by the intensive use of these compounds. As a result, the European Union plans to prohibit the use of such compounds. Consequently, the priority task is to find an anticoagulant that can control the rodent populations without affecting their predators. The research of Dr. Adrien Montagut (PhD, 2011-2014) have led to the structure type of an anticoagulant derived from 4-hydroxycoumarin. Currently, AMR361 was tested in vitro on all VKORC1 mutations and in vivo on wild rats. It is the first AVK developed that responds to all the characteristics of the initial specification. The first part of my PhD was to complete the biological study on 4-hydroxycoumarin core by bringing functional diversity on the para position of the aromatic ring. From a biological point of view, the lengthening of the spacer arm on the side chain by use of various functions or the introduction of a dimethyl group on the methylene bridge were studied in order to analyze the effectiveness and persistence parameters. However, most of the synthesized compounds belonging to the family of 4-hydroxycoumarins are already described in a patent filed by Liphatech company in 1999. The study of new cores which are similar to the 4-hydroxycoumarin or the functionalization of the aromatic part of the 4-hydroxycoumarin has provided access to more diverse structures. These original possibilities for innovation have been introduced to circumvent existing patents

Réseaux de transduction stimulés par les récepteurs à activité tyrosine kinase et les récepteurs couplés aux protéines G dans les cellules myométriales (rôle dans l'activation des protéines ERK et impact sur la prolifération cellulaire)

Cette étude concerne les réseaux de signalisation impliqués dans la régulation de la prolifération des cellules musculaires lisses utérines (cellules myomètriales), celle-ci jouant un rôle essentiel dans le contrôle des activités de l'utérus. Nous avons démontré, dans des cellules de myomètre de rate en culture primaire, l'implication des MAP kinases de type ERK dans l'effet mitogène de différents agents: le PDGF, un facteur de croissance qui interagit avec un récepteur à activité tyrosine kinase, l'endothéline-1 (ET-1), un peptide mitogénique qui interagit avec un récepteur couplé aux protéines Gi et Gq dans le myomètre, et le pervanadate (PV), un inhibiteur de protéines tyrosine phosphatases. Nos résultats ont démontré que le PDGF et le PV stimulent les voies PLCg1/InsP3 et ERK qui conduisent à la libération d'acide arachidonique et à la biosynthèse de prostaglandines impliquées dans la production d'AMPc. L'effet inhibiteur de l'AMPc sur l'activation de ERK et la synthèse d'ADN induites par les PDGF et le PV souligne l'existence d'une boucle de rétroinhibition au niveau des réponses médiées par ces deux agents. Nous avons également montré la présence et l'activation par le PV des protéines tyrosine kinases de la famille Src dans les cellules de myomètre. Ces protéines sont impliquées dans l'activation de la PLCg1 et la production d'InsP3 dues au PV, et dans l'activation de ERK par ET-1. En effet, l'activation de ERK par ET-1 met en jeu la stimulation séquentielle de PKC, Src et Ras. Par ailleurs, deux voies de transduction contribuent à l'activation PKC-dépendante de ERK par ET-l: une voie Gq/PLCb/InsP3/PKC conventionnelles et nouvelles, et une voie Gi/PI3-kinase/PKC atypiques. L'ensemble de cette étude contribue à une meilleure compréhension des mécanismes de régulation de la prolifération des cellules de myomètre, qui intervient dans des conditions physiologiques (gestation) mais aussi pathologiques (fibromes) et physiopathologiques (préterme).In this study, we aimed to analyse the signalling pathways involved in the regulation of myometrial cells proliferation which plays an essential role in uterine functions. We demonstrated, in rat myometrial cells in primary culture, the involvement of MAP kinases of the ERK type in the mitogenic effect of various agents: PDGF, a growth factor acting through a receptor tyrosine kinase, endothelin-1 (ET -1), a mitogenic peptide which interacts in the myometrium with receptors coupled to Gi and Gq proteins, and pervanadate (PV), a potent protein tyrosine phosphatase inhibitor. Our results showed that PDGF and PV induced PLC-g1/Ins3 stimulation and ERK activation that both contribute to cAMP production by increasing the release of arachidonic acid and the biosynthesis of prostaglandin. The inhibition of ERK activation and DNA synthesis by cAMP constitutes a potentially important negative feedback loop for PDGF and PV- mediated responses. The presence and the activation by PV of tyrosine kinases of the Src family was also demonstrated in rat myometrial cells. These kinases contributed to the activation of PLCg1 and the production of InsP3 triggered by PV, and to the activation of ERK induced by ET-1. Indeed, we demonstrated that ET-1-mediated ERK activation involves the sequential activation of PKC, Src and Ras. We also showed that two signalling pathways contribute to the PKC-dependant ERK activation induced by ET-1: a Gq-PLCb-InsP3-conventional/novel PKC and a Gi-PI3kinase-atypical PKC pathway. Altogether, the results demonstrate the presence of signalling networks required for the regulation of myometrial cells proliferation which play an essential role in physiological conditions (gestation) as well as pathological (fibroma) and physiopathological (preterm) conditions.ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

3-methylene-2.4-chromandione in situ trapping : introducing molecular diversity on 4-hydroxycoumarin

International audienc

Efficient C-3 reductive alkylation of 4-hydroxycoumarin by dehydrogenative oxidation of benzylic alcohols through ruthenium

International audienceA practical route to prepare 4-hydroxycoumarin derivatives functionalized at the C-3 position is described through a catalytic coupling reaction between 4-hydroxycoumarin and a series of substituted benzylic alcohols. The reaction is conducted in the presence of tris(triphenylphosphine)ruthenium(II) dichloride (5 mol%), KOH (0.2 eq.) in tertamyl alcohol under microwave irradiation at 140 °C for 2 hours

Homogeneous acylation of Cellulose diacetate: Towards bioplastics with tuneable thermal and water transport properties

In this study, we report a simple, non-degrading and efficient homogeneous acylation of cellulose diacetate (CDA) by using a large panel of commercially available acylating aliphatic moieties, differing in their structure (fatty, ramified, bulky, cycloaliphatic, aromatic, more or less spaced from the cellulose backbone), in view of generating a library of well-defined cellulose mixed esters with enhanced thermoplasticity. As reflected by a lowering of the glass temperature (Tg), the covalent grafting confers an improved mobility to the cellulose chains, by disrupting the initial H-bonds. In particular, it appears that the gain in free volume is tailored by the substituent structure and that acylating reagents consisting in a terminal bulky moieties spaced from CDA chains by a linear chain efficiently separate macromolecular chains without generating detrimental stiffening interactions (low Tg around 125 °C). Moreover, free-standing films easily prepared by solvent casting exhibit relevant water transport properties, which are closely dictated and tuned by the water solubility of the cellulose mixed ester

The use of information sources and online databases for writing theses by students of selected studies

The paper presents the results of research on the use of electronic publications for writing theses by first and second degree students of two engineering and technical studies: agriculture and forestry technology, and management and production engineering. 53 first-degree and 175 second-degree students were included in the study. It was found that both first and second degree students use electronic publications on a par with printed ones when writing their theses. First-degree students preferred Google Scholar, while second-degree students preferred Polish scientific publishers. The choice of both the Google Scholar database and the pages of Polish scientific publications resulted from the possibility to search for papers in Polish, which was the main language of publication chosen by both groups of students

Suspended solids and bacteria removal mechanisms in ceramic filter and pervious concrete filter: a review

The need for drinking water with affordable access is increasing nowadays. The poor water quality problems can be solved by several water treatment methods, i.e. ceramic filter (CF) and pervious concrete filter (PCF). Those two mentioned technologies work based on the pores that exist within the filter. This study aimed to review (1) the influence of the material composition of the CF and PCF on hydraulic characteristics (porosity, permeability, pore size, pore structure, pore connectivity/ distribution) and its removal effectiveness, and (2) the removal mechanisms of suspended solids and bacteria based on its hydraulic characteristics. The study reviewed 45 literature, including books, reports, and published articles. The type, mineral, and sources of clay and the type, shape, proportion, and size of combustible materials on CF will affect its hydraulic characteristic. The shape, size, and type of aggregate, the ratio of water to cement, and the ratio of aggregate to cement on PCF will affect its hydraulic characteristics. The removal mechanisms of suspended solids are straining on the surface and trapping on the deadlocked pores. On the other hand, the bacteria removal mechanisms strain the bacteria in the dirt layer and trap the bacteria in the pores