Biomedical research institute
     

    Research project 1

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    Molecular mechanisms controling neutrophil apoptosis in the resolution of inflammation


    Figure 1: Role of neutrophils in innate immunity and inflammation


    Historically, neutrophils have been viewed only as professional phagocytic cells that are able to phagocytose and destroy infectious agents. Indeed, they are key anti-infectious actors in host defense but can mediate tissue damages (Witko-Sarsat et al Lab Invest 2000). However, it is now clear that the role of neutrophils go far beyond phagocytosis and pathogen killing. Following successive steps of adhesion, chemotaxis and migration, can phagocytose invading pathogens and mobilize their microbicidal effector molecules, including reactive oxygen species, antibiotic proteins and proteinases. This neutrophil activation triggers an apoptosis program to ultimately signal their phagocytosis to macrophages (Witko-Sarsat et al Trends Immunol 2011). The recognition and the subsequent engulfment of apoptotic neutrophils by macrophages is a key event of the resolution of inflammation, which can be associated with autoimmunity or inflammatory.  In addition, this safe disposal of apoptotic neutrophils generate anti-inflammatory signals to complete the inflammation resolution (Kantari et al. Contrib Microbiol, 2008; Martin et al Swiss Med Wkly. 2015 Feb 21;145:w14056 )

     

    Figure 2 : Neutrophil apoptosis is a pivotal step in the resolution of inflammation


    There are many different mechanisms responsible for the balance between apoptosis and survival of neutrophils and disruptions of these mechanisms has the potential to lead to uncontrolled inflammation. Our group has identified an unexpected anti-apoptotic role for PCNA. In proliferating cells, PCNA is a critical factor in DNA synthesis and repair. In neutrophils which are devoid of proliferative capacities, PCNA is exclusively cytosolic.
    (Witko-Sarsat et al 2011 Trends Immunol; De Chiara et al Frontiers Immunol 2012).

     

    Figure 3 : Control of neutrophil survival by cytoplasmic PCNA


    PCNA is a trimeric protein (in blue on the cartoon) which can build a scaffold by its association with several proteins (Witko-Sarsat et al. J Exp Med 2010). In the neutrophil, PCNA is exclusively cytosolic and its scaffold is adapted to the different states of neutrophil physiology:

    1) Under the steady state survival (represented in the center), PCNA is associated with “steady-state partners” such as pro-apoptotic proteins (procaspases) 2) Under inflammation or G-CSF treatment or hypoxia, de novo synthesis of “inducible partners” (for exemple p21/waf1) will modify the scaffold to promote survival. 3) Under non-inflammatory conditions, physiologic apoptosis is accompanied by a proteasomal degradation of PCNA with caspase activation. 4) Phagocytosis also modulates the PCNA scaffold but the role of PCNA in neutrophil fate seems to be highly dependent on the type of bacteria phagocytosed. (Witko-Sarsat & Ohayon Immunol Reviews in press)

     

    Figure 4: PCNA : a potential target to promote neutrophil apoptosis in cystic fibrosis


    Cystic fibrosis is characterized by a chronic infection by Pseudomonas aeruginosa and a destructive neutrophil-dominated pulmonary inflammation. Neutrophils functions are altered in CF and might be considered as potential targets to dampen inflammation Editorial J Innate Immunity 2013

    For instance, we have observed that neutrophils from CF patients have a delayed apoptosis associated with an increased expression of PCNA.

    Roscovitine, a synthetic inhibitor of cyclin-dependent kinase modulating innate immunity (Meijer et al J Innate Immunity 2016) and triggering neutrophil apoptosis is tested in clinical trial in CF ((Meijer et al J Innate Immunity 2016 : article which is the choice of the editor) (Editorial J Innate Immunity 2016)

    These studies are supported by the patient associations Vaincre la Mucoviscidose and ABCF2 Mucoviscidose

     

    Figure 5 : PCNA possess a nuclear export sequence (NES) and can be exported from the nucleus during differentiation.

    We have also provided evidence that the monomeric PCNA can have anti-apoptotic activity (De Chiara et al J Leukocyte Biol 2013).


    Figure 6: Cytosolic PCNA is associated with chemotherapy resistance in HL-60 cells: a potential target to promote apoptosis in cancer cells ?

    This nuclear-to-cytosolic relocalization occurred which occured at the end of granulocyte differentiation under normal conditions, is observed in the promyelocytic HL-60 cells resistant to daunorubicin to favor their survival (Ohayon et al Manuscript submitted for publication).










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