Ischemic stroke in sepsis shock is associated with increased mortality and long-term cognitive impairment. Due to its critical position, the brain vessel recanalization is an emergency. However, in sepsis context, the therapeutic strategies available are limited. Thus, innovative approaches targeting the fibrinolysis impairment are absolutely warranted. Granulocyte-derived Extracellular Vesicles (Gran-EVs) are circulating fibrinolytic carriers whose binding molecules allow them to reach the fibrin clot. The fibrinolytic Gran-EVs were associated with less coagulopathy and a better prognosis in sepsis shock patients. Consistently, in a murine model of sepsis, it was recently observed that the repeated injection of fibrinolytic Gran-EVs significantly 1/increased the mouse fibrinolytic circulating activity 2/reduced the number of thrombi in vital organs and 3/improved mouse survival. In addition, it was shown that Gran-EVs from septic patients lyse a thrombus according to their fibrinolytic potential. Therefore, our hypothesis is that injection of granulocyte-derived EVs with a highly fibrinolytic activity may improve the septic shock prognosis by participating to the organ thrombi resolution in vivo. Two research groups, respectively renowned in EV biology and their impact in the cardiovascular system (C2VN) and neurovascular diseases (PhIND) will join their force to carry out this project whose aims are 1/to produce and characterize gran-EVs with a highly fibrinolytic activity; 2/to evaluate their effect on thrombus resolution in vivo in mouse models of sepsis, and 3/to evaluate their therapeutic potential on sepsis-associated thromboembolic stroke compared to currently available thrombolytic therapies. The ultimate goal of this project is to pave the ground of an innovative therapeutic EV-based strategy in septic shock-associated cerebral stroke relevant for future clinical translation.

Viruses have multiple and globally significant biogeochemical impacts via the metabolic reprogramming and mortality (lysis) of their microbial hosts. Viral lysis modulates food-web dynamics by diverting the living biomass away from higher trophic levels and redirecting it into the microbial loop. This ‘viral shunt’ is one the largest carbon fluxes in the ocean. Viruses also impact nutrient cycling during infection, with the rewiring of host metabolism to support virus production. This reprogramming profoundly alters resource acquisition, carbon and energy metabolisms. Virus research has almost exclusively focused on the study of those that contain DNA genomes. Recently, it has emerged that RNA viruses could account for half of the marine viral communities, yet, we understand very little about their role in ecosystem functioning. BONUS is dedicated to the study of this under-explored component of the biosphere and its biogeochemical impacts on one of the most globally distributed and ecologically successful groups of organisms in the ocean, the diatoms. Diatoms contribute 40% of marine primary production and their silica shells also ballast substantial vertical flux of carbon from the surface to depth. Our 4-year project proposes a thorough study of viral infection of dominant diatom species with contrasting traits (large vs. small-sized) and patterns of occurrence (blooming vs. persistent) to address the hypothesis that infected diatoms are metabolically distinct from uninfected cells and have distinct ecological and biogeochemical fates. To this end, a multidisciplinary team will address four research questions: What are the metabolic functions that respond to viral infection? What is the impact on resource uptake and allocation? What is the fate of infected diatoms? What is the significance of viral infection and metabolic reprogramming in natural diatom populations? This integrative framework should provide novel fundamental mechanistic understanding and direct estimates for assessing the impact of diatom infection on ocean biogeochemistry dynamics. Given the global-scale prominence of RNA viruses and targeted diatom populations, we anticipate that our research will lead to the discovery of important processes that drive the functioning of the ocean.

During the last decade, the development of new techniques for the detection of superoxide radical anion (O2•–) in biological systems has continued to receive increasing attention. Superoxide is positioned upstream in most radical cascades leading to Reactive Oxygen and Nitrogen Species (RONS) and irreversible oxidative damages to biomolecules. Therefore, the ability of rigorous detection of superoxide is of major importance. However, currently, no method fulfills the requirements for the rigorous characterization of the role of O2•– in biological systems and the characterization of RONS in vitro and in vivo remains a challenge. Many probes have been criticized and discarded due to unreliable results, detection artefacts and/or poor reproducibility. For the last decade hydroethidine (HE) has been shown to be a promising probe for the fluorescent detection of O2•–. Unfortunately, the use of HE in biological systems is limited by four main problems: (i) limited understanding of the mechanism of HE reaction with O2•–, (ii) lack of site-specific analogs of HE, (iii) degradation of HE by heme proteins leading to misleading signals, and (iv) requirement of HPLC-based products separation coupled with fluorescence, electrochemical or MS detection. Our teams have been actively involved in developing techniques for the detection and the characterization of RONS (e. g. spin trapping, SOD mimics, fluorescence-based RONS probes) for the last 20 years and their significant contributions have been recognized in the field of free radical/redox biology. The major objectives of this proposal can be summarized as follows: (i) determination of the reaction mechanism of HE with O2•– and other biologically-relevant oxidants, including experimental (kinetics, products characterization) and theoretical studies (DFT calculations), using HE analogs as model systems, (ii) targeting HE type probes to intracellular and extracellular compartments for site-specific detection of O2•–, (iii) supramolecular and other approaches to protect HE from non-specific oxidation by heme proteins, (iv) the development of innovative probes for the direct and specific detection of O2•– by fluorescence, (v) application of optimized probes for detection of O2•– in biological systems. The molecular structure of the probe plays a key role in its performance, including selectivity and specificity. As the development of HE probe was not based on rational design, there is an opportunity to develop new more efficient HE-analogs for the detection and quantification of O2•–. This project involves one French partner (ICR) as well as two foreign partners, led by Dr. Kalyanaraman (Medical College of Wisconsin, Milwaukee, USA) and by Dr. Sikora (Lodz University of Technology, Poland). These partners have already funds to perform the work described in the proposal. Thus, no financial support is requested from ANR for these two groups. All collaborating groups have a strong experience and expertise and are among leaders in their field. Also, the project will capitalize and benefit from well-established and long-term collaborations between the three groups. This project illustrates the need for joint and interdisciplinary effort to push forward the frontiers in the free radical and biomedical fields.

Bronchial mucus is a hydrogel secreted at the surface of the bronchi to protect them from inhaled contaminants. It is continually transported by the collective beat of cilia that cover the bronchial epithelium, and by cough, a violent air flow capable of dragging mucus plugs away. In COPD, expected to be the fourth cause of mortality by 2030, the mucus transport is altered, causing obstructive crises which impose a hospitalisation in emergency room and worsen the disease. Devices based on the rheology of mucus, developed by the industrials participants of this project, allow to monitor the patients and assist their bronchial drainage. The mechanisms of transport at play are nonetheless poorly understood, limiting by the way the potential of these yet empirical approaches. Our objective is to unveil these mechanisms in order to identify the optimal rheological parameters and assistance actions to follow and manage the mucus transport in patients. We will determine the rheological and biochemical properties of pathological mucus and their role in the different mechanisms of natural and assisted transport, at scales ranging from the epithelium to the whole bronchial tree. We will analyse the mechanisms driving the mobilisation of mucus through its interaction with cilia, and their failing in pathological conditions, as well as the couplings between the different levels of complexity (branching, deformability of the bronchi, mucus hydration) and a global mechanical solicitation. We will integrate this fundamental understanding in a new generation of devices, to favour the early detection (by mucus rheology) and the anticipated treatment (by drainage optimisation) of the crises. Beyond its scientific impact, this interdisciplinary project aims in the long term at providing a medico-economic benefit (reduction of hospitalisations, improvement of the patient prognosis) while stimulating the development of the involved industrial partners.

The NanoThermoPockets project aims to develop a novel class of nanotweezers utilizing optically-induced thermal forces. These nanotweezers consist of glass substrates with metallic nanorings atop nanopocket chambers. When illuminated with an infrared laser, a tunable 3D temperature gradient is generated, creating thermal forces that can trap nano-objects as small as 5 nm. This innovative approach will overcome the limitations of conventional optical tweezers, which are constrained by the diffraction limit and cannot trap objects smaller than 100 nm. By exploiting thermal gradient forces, NanoThermoPockets offers several advantages, including a trapping range exceeding 500 nm, simple and reproducible nanofabrication, and seamless integration with optical microscopes and photonic waveguides. The project comprises four work packages, involving multiphysics simulations for trap optimization, rapid prototyping nanofabrication, and experimental techniques to assess temperature profiles and nanoobject responses. Fluorescence correlation spectroscopy will be used to quantify trap stiffness and explore the impact of various parameters on the trapping potential. Integration with optical microscopes and photonic waveguides will be investigated to enhance the versatility of these nanotweezers and do what current optical tweezers cannot do. NanoThermoPockets' significance lies in its ability to manipulate tiny nano-objects with broad applications, such as biotechnology, quantum nanophotonics, and material science. The approach offers a fresh perspective on nanomanipulation, simplifies nanofabrication, enables parallelized spectroscopy, and requires no complex feedback systems. Its integration with single-molecule fluorescence microscopes and photonic waveguides promises exciting possibilities for scientific research and technology development.