Aging refers to the process by which cells, tissues and organs deteriorate as they grow older. Since this time-dependent decay often interferes with normal function, aging represents a major challenge for an organism survival. Demographic data unambiguously indicates that world’s population is getting older leading to an increased incidence of age-related diseases. Consequently, aging has become a top priority in biomedical research. In humans, consequences of aging are particularly devastating in the nervous system. Thus, aging is commonly associated with functional deficits ranging from mild impairments to global loss of brain functions such as in dementia. However, it is well established that brain aging is not the same for all individuals. Indeed, a subset of older individuals maintains optimal brain abilities (physiological aging) whereas others experience a measurable functional decline (pathological aging) supporting the notion that the brain possesses some adaptive mechanisms to counterbalance the detrimental effects of aging. Nonetheless, the nature of these mechanisms remains largely unknown. MicroRNAs (miRNAs) are small (~18-24 nucleotide) non-coding RNAs, which regulate gene expression post-transcriptionally by inhibiting translation or inducing degradation of target messenger RNAs (mRNAs). Many miRNAs appear to be disrupted during cell senescence, aging and/or disease. However, so far, only few miRNAs have been associated to age-related alterations in cellular and organ functions. Recent results support the notion that miRNAs contribute to age-dependent brain deficits. Thus, it has shown that the abundance of key neuronal miRNAs, such as miR-124, decreases with age. Additional findings obtained by the applicant indicate that restoring miR-124 levels in a mouse model of frontotemporal dementia, leads to a partial rescue of deficits in social behavior. Here, we propose to thoroughly investigate the role of miRNAs in the process of brain aging. Using a multidisciplinary approach, the objectives of this proposal are the following: 1. Exploring the molecular mechanisms leading to age-dependent decay of miRNAs contents in the brain. Using in vitro assays and cell culture models, we seek to determine the molecular factors responsible for the age-dependent reduction in miRNAs levels, using miR-124 as a paradigmatic example. 2. Obtaining a detailed pattern of molecular changes linked to physiological and pathological aging. Young and aged mice will be first tested at multiple behavioral tasks and classified into different functional categories. Brain regions related to the behavioral tasks as well as control areas will be subjected to: i) RNA extraction and transcriptome analysis (miRNAs and mRNAs); and ii) protein isolation followed by quantitative proteomics. Data from miRNA sequencing will be analyzed using a novel bioinformatics approach taking into account the behavioral performance at different tasks, the anatomical region and the concomitant changes in downstream targets at both mRNA and protein level. Overall, this analysis will provide a comprehensive description of the molecular alterations associated to aging as well as their potential correlations to functional preservation/loss. 3. Functional validation of novel miRNAs involved in the aging process. Using the data from our screening, the involvement of candidate miRNAs in physiological/pathological aging will be systematically tested in vivo using intracerebral injections of customized adeno-associated vectors (AAVs). These vectors, incorporating the latest advances in genome editing technology (e.g. CRISPR/Cas9), would enable to deliver targeted genetic modifications to specific neuronal networks. AAVs represent, therefore, a simple, safe and straightforward procedure to manipulate the levels of candidate miRNAs in vivo. The effect of altering miRNA levels in vivo will be then analyzed at the morphological, electrophysiological and behavioral level.

According to the World Health Organization (WHO), the priority diagnostic needs for human African trypanosomiasis (HAT) are a test for gambiense HAT (gHAT) to identify individuals to receive widened treatment and a test for rhodesiense HAT (rHAT) usable in peripheral health facilities. The ERASE project will develop rapid diagnostic tests (RDT) compliant with the respective WHO target product profiles. For gHAT, current RDTs have inadequate specificity. ERASE will develop a 2nd generation antibody detection gHAT RDT with improved diagnostic performance, incorporating the best selected recombinant antigens. The gHAT RDT performance will be evaluated in 3 clinical trials in epidemiologically different gHAT foci in West- and Central Africa. Manufacturing of the gHAT RDT will be transferred to an African SME (TRL8/9). Diagnosis of rHAT relies on microscopy, resulting in underdetection. For rHAT, ERASE will identify target antigens and develop an antigen capture RDT using the innovative Affimer technology. After evaluation of its clinical performance on biobanked specimens and in a clinical trial, the rHAT RDT will be ready for industrialisation (TRL6). Mapping of health centres, forecasting of RDT consumption and implementation costs, and social research will pave the way for future rHAT RDT introduction. The consortium consists of commercial, non-for profit, academic and governmental partners, including an SME with experience in HAT RDT commercialisation, and will ensure successful RDT development and sustainable production and implementation. Both RDTs will support HAT elimination as targeted by the WHO neglected tropical diseases roadmap. The gHAT RDT will be applicable in a “test & treat” strategy to rapidly stop gHAT transmission. The rHAT RDT will revolutionize rHAT management, allowing faster diagnosis and safer treatment. It will strengthen rHAT surveillance, lead to faster detection of outbreaks and will facilitate elimination of rHAT as a public health problem.

Many studies have evidenced that peaks of air pollution are a cause of mortality in humans by promoting cardiovascular and pulmonary diseases. While the many consequences of acute intoxication with high levels of pesticides, carbon particles or metal derivatives are well documented, the more subtle, long term effects caused by ambient air pollution remain difficult to evaluate. Among the major pollutants in air, soils and water, recent epidemiological studies have suggested a possible link between exposure to pesticides and type 2 diabetes. Thus, ingestion of food contaminated with pesticides has been associated to impaired glucose regulation, endocrine disruption and obesity. However, it has not been clearly established whether lower levels of pesticides found in aerial particulate matter (PM) may be a new risk factor for insulin resistance and progression of diabetes mellitus because reliable experimental models are still not available. Ultrafine particles having diameter ranges < 2.5 µm (PM2.5) or even smaller are very active components of today's atmospheres and are practically non-removable once inhaled. PM25 have been shown to be responsible for a wide array of pathologies, including inflammation, endothelial and mitochondrial dysfunction, and cardiac diseases due to their deep penetration within the bronchial cells. Regarding the mechanisms of action at the molecular level, PM25 promote oxidative processes, leading to protein oxidation and/or causing genotoxicity. For instance, lipophilic toxicants in volatile form can accumulate within mitochondria to trigger redox cycles highly detrimental to the cardiorespiratory system. For decades, members of our Laboratory (ICR-SMBSO) have been involved in the study of the various facets of the pathophysiological effects of oxidative stress, from the design of sensitive, highly specific and targetable probes and improved analytical procedures for protein oxidation and inflammation assessment, and their use in animal studies and clinical trials. In the present project encompassing chemistry, biology, environment and health fields Dr. Sophie Thétiot-Laurent will led a subgroup of ICR-SMBSO, with the cooperation of all other members of the Team. Together they will apply their know-how to investigate the chronic effects of low levels of pesticides aerosols on mitochondrial and cardiac dysfunction, oxidative stress and insulin and metabolic disorders in experimental animals. Central to the project are: i)a unique atmosphere-mimicking exposure device recently developed in the Laboratory and adapted for animal housing to produce aerosols of nanosized (= 1 µm) and concentration-controlled particles. This will allow chronic exposure to environmentally-relevant pollutants (1-20 ng/m3). ii)EPR measurements at low frequency (L-band). Combining this technology, already available at ICR-SMBSO, with a series of novel vectorized probes will define a new research axis aimed at monitoring the redox state and pH changes in vivo. To summarize, new mitochondria targeted probes will be synthesized and applied to investigations of metabolic, oxidative stress and histological endpoints using EPR and improved analytical methods. Below, the chronology of the 2-years project: 1-design (subcellular targeting, pKa adjustment, search for low toxicity), synthesize and apply (test tube to in vivo) new probes with specific spectroscopic fingerprints for investigating pollution-induced redox dysfunction in vivo 2-expose animals to controlled atmospheres (size, composition, concentration) of selected pesticides under realistic conditions (e.g. 2 hours/day) 3-run a proof of principle in vivo experiment aiming to assess the effect of pesticide exposure on the course of symptoms and pathological features of metabolic, inflammatory, and cardiac disorders in animals. This step will include the set up of analytical assays, and designing molecular sensors to monitor oxidative stress and systemic inflammation at real-time.

Reducing agents with neutral organic structures and exceptionally negative redox potentials have received a renewed interest especially since the astonishing advances of their reactivity in organic chemistry. Strong organic electron donors (OEDs) are capable of spontaneous single- or double-electron transfer to organic substrates under mild and homogeneous conditions. The OEDs represent serious rivals to highly aggressive metal-based reducers and emerge as an attractive novel source of reducing electrons. Thus far, they have been scarcely studied and approaches involving OED-promoted electron transfer steps are not sufficiently exploited despite their synthetic potential and their tunability. Consequently, their application scope remains quite narrow. In this context, it is of utmost importance to fully master the chemistry of organic reducers and to establish their fields of application. To address the important shortcomings of their applications, this research proposal aims first at developing novel libraries of organic electron donors able to overcome current boundaries. More importantly, the preparation of air-stable precursors, easily in-situ activated, will extend their practicality. The development of OED-promoted redox catalytic cycles will further fulfill the need for atom-economy and latent approaches, often necessary in the manufacturing industries.Their structures will be diversified and evaluated in order to better understand the factors governing single- or double-electron transfer as well as their reducing power. The diversity of the synthesized donors will thus provide a privileged database to investigate the relationships between molecular structure and reactivity. The second aspect of the project will be dedicated to the exploitation of the OED-generated active species in the reduction of challenging substrates and to the exploration of unprecedented applications. A comprehensive study of their scope and limitations, as well as of the involved mechanism will supply chemists with an exhaustive guide of their capabilities and spread their application panel in radical chemistry. This also falls into a context of searching new reduction methodologies, combining efficiency, modularity and high selectivity, and following the requirement of the sustainable chemistry. Reduction reactions are much less mastered than the oxidative processes extensively used for the fabrication of many everyday objects. With the decline of fossil sources, it is urgent to develop alternative routes to these fundamental compounds. In addition, a special focus will be given to the domain of material chemistry through the use of organic electron donors as initiators of polymerization. We have demonstrated that OEDs represent an unique tool for efficient, simple and room temperature polymerization process, responding to energy-friendly, cost-efficient and secure technical specifications. Their high group tolerance makes them fully compatible with the synthesis of a large range of polymers of wide industrial importance. The additional reducible functional groups accessible with our novel series of organic electron donors will extend the process to the metal-free polymerization of a whole new range of monomers under mild conditions and without the need for co-initiators. To go further with this highly innovative process, a detailed comprehensive mechanistic study will help to i) understand the initiation and chain propagation pathways and ii) prepare optimized OED structures as polymerization initiators. This will illustrate their undeniable potential as efficient and versatile reagents to the scientific and industrial community. In term of social and economic benefits, the financial support of this project will allow us to collect the decisive results necessary to reach the high technology readiness levels required by the European research program Horizon 2020 and access to the pre-industrialization step of our concept.