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Role of P2 receptors for nucleotides in septic shock

Research unit

UMR_S 949 - Biologie et pharmacologie des plaquettes sanguines : hémostase, thrombose, transfusion (BPPS)
10, rue Spielmann - BP 36 67065 Strasbourg Cedex

Group

Name: Biologie et pharmacologie des plaquettes sanguines : hémostase, thrombose, transfusion.

Group leader: GACHET Christian - christian.gachet@efs-alsace.fr

Group leader's phone: 03 88 21 25 25

Website: Visit website

Group organization:
- Chercheurs: 4
- ITA: 8
- Doctorants: 3
- Post-Docs: 2
- Autres: 2

Publications of the team linked to the topic (3 last years):
1) Hechler B, Maître B, Magnenat S, Heim V, El Mdawar M-B, Gachet C and de la Salle H (2016) Platelets are dispensable for immunological transfusion-related acute lung injury in the mouse. J Thromb Haemost. (sous presse)
2) Hechler B and Gachet C (2015) Purinergic Receptors in Thrombosis and Inflammation. Arterioscler Thromb Vasc Biol 35(11): 2307-2315.
3) Maitre B, Magnenat S, Heim V, Ravanat C, Evans RJ, de la Salle H, Gachet C and Hechler B (2015) The P2X1 receptor is required for neutrophil extravasation during lipopolysaccharide-induced lethal endotoxemia in mice. J Immunol 194(2): 739-749.

About PhD

PhD Director: GACHET Christian - christian.gachet@efs.sante.fr

Phone: 03 88 21 25 25

Junior advisor: HECHLER Béatrice

Co-tutely: non

Co-Director: non

About PhD topic :

Title: Role of P2 receptors for nucleotides in septic shock

Project: Septic shock is an acute systemic inflammatory syndrome following host response to pathogens, leading to intense cell activation and multiple organ dysfunction syndrome (MODS). So far, the mainstay of treatment remains a general supportive care, including fluid resuscitation, mechanical ventilation, without pharmacologic agents with demonstrated improvement of the outcome. The proinflammatory response to sepsis leads to activation of the coagulation system resulting in bacterial containment, but also in thrombotic microangiopathy, through uncontrolled thrombin and fibrin generation, platelet activation and consumption, which may evolve toward disseminated intravascular coagulation (DIC). Thrombocytopenia negatively impacts patient prognosis at day 28 and may therefore stand as an early predictor of death. Among the numerous activators of inflammatory cells and platelets, extracellular nucleotides (ADP and ATP) play a central role. ADP and ATP, released from dying cells and damaged tissues or in response to inflammatory stimuli, act as “danger signals” or DAMP “damage-associated molecular pattern” which display potent immune-enhancing activity, while at sites of vascular injury, adenine nucleotides, massively released by activated platelets, contribute to both hemostasis and thrombus formation by greatly amplifying most of the platelet responses. Three distinct P2 receptors are present on blood platelets: the P2Y1 and P2Y12 receptors are G protein-coupled ADP receptor subtypes, while the P2X1 ligand-gated cation channel is activated by ATP. The P2Y12 receptor is targeted by the widely used antithrombotic drug clopidogrel (Plavix®) and prasugrel (Efient®). Beyond platelets, these P2 receptors are present on various blood cell types (neutrophils, monocytes/macrophages) and on cells of the vessel wall (endothelial and smooth muscle cells). The role of these P2 receptors during the septic shock remains to be evaluated. The aim of the project is to evaluate the role of the P2Y1, P2Y12 and P2X1 receptors during septic shock.
We will use a model of cecal ligation and puncture (CLP)-induced experimental sepsis in mice, available in our laboratory. The role of each P2 receptor will be evaluated using mice deficient for each of these P2 receptors and available in our laboratory. In addition, pharmacological inhibitors of each receptor subtype will be evaluated in this model. In order to determine the respective contribution of these P2 receptor subtypes present on circulating blood cells and in the vascular wall, mice with tissue-specific invalidation of each P2 receptor will be evaluated in the model of septic shock.
In each case, blood, various organs will be harvested for further ex vivo analysis. Briefly, plasmatic markers of inflammation (cytokine and chemokine levels), coagulation and fibrinolysis, as well as platelet activation markers will be assessed. Leukocyte and platelet activation states will be evaluated by measurement of surface activation markers and platelet-leukocyte aggregates by flow cytometry. Multiple organ injuries and inflammation will be characterized by histopathological examination and evaluation of neutrophil and platelet recruitment and NET production by immunohistochemical analysis of tissue sections.
Overall, this experimental approach should clarify the specific role of the P2Y1, P2Y12 and P2X1 receptors in septic shock. The most relevant steps will represent potential preferred targets for new and efficient therapeutic strategies.

Wished skills: General knowledge in cellular biology, highly motivated, elevated scientific rigor.

Expertises which will be acquired during the training: Expertise in the field of platelet physiology, infection, inflammation. Technical expertise in surgery, injection and collection of blood and organs, histology, immunohistochemistry, flow cytometry.