Fibrocalcific Aortic Valve Stenosis (AVS), characterized by a narrowing of the aortic valve opening, leads to an obstacle to the blood flow through the aortic valve during left ventricular ejection. In industrialized countries, this disease affects 2 to 6% of people aged above 65 years and its prevalence is likely to double within the next 50 years due to an increasingly ageing population. Initially described as a degenerative process related to valve repeated aggression by blood flow, the AVS pathophysiology is now understood to be an active process akin to atherosclerosis involving initiating factors, endothelial dysfunction, inflammatory responses and angiogenesis leading to valve remodeling and calcification. Although important progress has been made in understanding the pathophysiology of this disease, there is currently no pharmacological treatment available to alter the natural course of the disease. While the onset of symptoms indicates a poor prognosis, the only effective therapy for SVA is valve replacement by a mechanical prosthesis or bioprosthesis. In industrialized countries, bioprosthesis are mainly used. These prostheses do not require anticoagulant treatment but present limited lifespan (10 years), mainly due to progressive calcification and degeneration of the tissue. The mechanisms involved in this context seem close to those observed in AVS. A healthy aortic valve is mainly composed of valvular interstitial cells (VIC) which are highly dynamic and plastic. Thus, these cells can differentiate into calcific osteoblast-like cells or into perivascular cells, as shown in our preliminary results. A preliminary analysis of normal and fibrocalcific human aortic valves and VIC isolated from these valves indicates that the retinoic acid pathway via its nuclear receptors is implicated in the process of AV calcification. The objectives of the RETINAVS project are to study more precisely the role of the retinoic acid pathway in AVS development and bioprosthesis degeneration by focusing on the calcification and angiogenesis processes, to highlight new factors modulating these processes and to identify, in the retinoic acid pathway, innovative pharmacological targets that will be validated by in vitro and in vivo preclinical approaches (animal models of aortic valve calcification and bioprosthesis degeneration). The RETINAVS project combines 3 major assets : 1) an ongoing collection of human fibrocalcific and normal aortic valves but also explanted bioprostheses. Studies on normal valves are very scarce in the literature and the access to such samples is a great advance to better understand the processes underlying the progression of the disease, from normal to pathological valves, 2) the gathering of biologists, cardiologists specialists in atherosclerosis, AVS and angiogenesis, cardiac surgeons specialists in valvulopathy and basic researchers involved in the deciphering of the role of the retinoid acid pathway and nuclear receptors in cardiovascular diseases (UMR Inserm 1011) and in progenitor cells characterization, of their angiogenic potential and ability to induce vascular remodeling (UMR-S Inserm 1140). The successful collaboration between partners for almost 10 years ensures the success of this ambitious translational project at the interface between fundamental and clinical research; 3) the combination of studies on human samples and animal models already developed in 2 teams. Our ambition is to identify retinoic acid and its associated pathways as potential new therapeutic targets in AVS and to define a new paradigm in pathophysiology of this disease and valvular bioprosthesis degeneration that represents a real challenge of public health.

The management of the acute ischemic stroke (AIS) remains a major challenge for modern medicine. The main goal of AIS treatment is to recanalize the occluded artery by removing the thrombus causing the ischemia as quickly as possible. There are two treatment options: intravenous thrombolysis (IVT) and mechanical thrombectomy (MT), both of which have significant limitations. IVT aims at dissolving the thrombus by administering tissue plasminogen activator (t-PA), a drug that targets the thrombus fibrin network. However, IVT has a limited recanalization efficacy (75% success). Yet, MT requires access to highly specialized stroke centers, and is only indicated for patients with large artery strokes of the anterior circulation (20-30% of strokes) Therefore, there is still significant potential and a critical need for improvement of AIS recanalization therapy. Within the framework of the ANR axes C9 and C10 (translational research and Biomedical Innovation), we aim to study a new therapeutic strategy that would increase the efficacy of IVT, thus improving the outcome of a larger number patients. Our hypothesis, based on our preliminary data, is that carbamylation, a type of post-translational modification, occurs in AIS thrombi and limits the efficacy of IVT and MT. Our goal is therefore to demonstrate 1) that carbamylation of AIS thrombi negatively impacts their response to IVT and MT, and 2) that its reversal with a recombinant decarbamylase could provide the basis for the development of a new adjuvant treatment to improve recanalization therapy in AIS.

Improving early recanalization and preventing neurovascular damage at the acute phase of ischemic stroke are the main objectives of current stroke research. In contrast, much less attention is being paid to the possibility of reversing and repairing post-stroke neurological damage. Previous studies, including ours, have indicated that pro-angiogenic and immunomodulation-based therapies can stimulate post-stroke cerebral remodeling and healing, thus decreasing infarct volume, improving sensorimotor deficit but also preventing cognitive decline. Remarkably, although the healing properties of platelets have long been known and taken advantage of in regenerative medicine, data on the role of platelets in post-stroke cerebral plasticity are scarce. Our previous studies and preliminary results show that in addition to storing large amounts of growth factors, platelets are the main source of various factors involved in post-stroke recovery, including brain-derived neurotrophic factor, angiopoietin-1, TGF-beta, serotonin and dopamine. In fact, our results obtained from a cohort of thrombocytopenic patients show that low platelet counts are associated with a significant decrease in those factors' plasma levels. In addition, our results obtained in experimental stroke models in mice indicate that platelets play a vital role in the days following an ischemic stroke, and that platelet inhibition by aspirin as used in secondary prevention post- Stroke worsens cognitive decline. In this context, our translational research project aims at determining the mechanistic contribution of platelets to cognition and post-stroke cerebral plasticity. This is of utmost importance since antiplatelet therapy is the main treatment given to stroke patients for secondary prevention.

Endothelial damage and coagulation activation at the lung microvascular level may play an important role in the physiopathology of the COVID-19 ARDS. The project aims to prospectively investigate both bedside pulmonary physiological markers (mainly alveolar dead-space) and biological markers of coagulopathy and endothelial dysfunction in COVID-19 and non-COVID-19 ARDS patients. In December 2019, an outbreak of pneumonia caused by a new coronavirus occurred in Wuhan and spread rapidly throughout China, with an evolution towards a global pandemic. Originally called new coronavirus 2019 (2019-nCoV), the virus was later officially named Coronavirus 2 of Severe Acute Respiratory Syndrome (SARS-CoV-2) by WHO. On 30 January 2020, WHO declared the SARS-CoV-2 outbreak as a public health emergency of international concern. Compared to SARS-CoV, which caused an outbreak of SARS in 2003, SARS-CoV-2 has a higher transmission capacity. The clinical manifestations of severe forms of SARS-CoV-2 are dominated by respiratory symptoms, leading in the more severe cases to Acute Respiratory Distress syndrome (ARDS). Understanding the impairments caused by SARS-CoV-2 to the respiratory and vascular systems and the underlying mechanisms is of an utmost importance. A coagulopathy is found in severe cases of SARS-CoV-2 infection, including significantly higher levels of D-dimers in severe forms and fatalities as compared to survivors. Accordingly, the hypothesis of microthrombosis at the organ level was first proposed for renal injury. High levels of creatinine were associated with higher levels of D-Dimers, which could suggest a micro-thrombotic origin for kidney failure. Thereafter, pulmonary thrombotic complications were reported both at the pulmonary arterial level and at the rich capillary lung level. Endothelial dysfunction (triggered by endothelial cells viral infection) and its microthrombotic consequences may thus play a major role in the respiratory physiopathologic process. Indeed, the SARS-CoV-2 receptor (ACE2) is strongly expressed in lung cells, preferentially alveolar type II cells, but also lung endothelial cells. Infection of endothelial cells could cause a lesion of the endothelium but also an activation that can trigger the activation of coagulation. Circulating endothelial cells (CECs) are markers of severe endothelial lesions and may act as non-invasive markers of pulmonary vascular dysfunction during SARS-CoV-2 infection, beside other biological markers of endothelial dysfunction. CECs are defined by a number of phenotypic and functional morphological criteria, which differentiate them from endothelial progenitor cells. Physiological bedside respiratory markers, such as physiological dead-space and more importantly alveolar dead-space, extracted from the advanced pulmonary monitoring performed in ARDS patients, could be highly correlated to such markers. Accordingly, the translational project proposes to study different aspects of the underlying pathophysiology of SARS-CoV-2 ARDS and to support specific therapeutic approaches. We propose to fully study in SARS-CoV-2 ARDS patients: (i) bedside pulmonary physiological parameters during the ICU clinical course, with alveolar dead-space as a major study parameter (ii) coagulation and fibrinolytic systems and (iii) endothelium activation and senescence. This work will contribute to a better description of the SARS-CoV-2 ARDS pathophysiology and should allow identifying patients' profile in which curative anticoagulant therapy or even thrombolytic treatments could be considered. Additionally, specific mechanical ventilator settings could also be proposed.

Gain-of-function mutations in the mechanotransductor Piezo1 are involved in more than 90% of hereditary xerocytosis (XH), a rare hemolytic pathology which physiopathology is not well understood until now. We published in 2019 the largest series of patients and described precisely their clinical and biological phenotype, associating venous and arterial thrombosis occurring after splenectomy, hemolysis (quasi-constant), iron overload occurring independently of any transfusion and finally perinatal edema. For a long time, XH has been perceived as an essentially "mature red cell" pathology, but a growing number of data have shown an expression and function of PIEZO1 in other cell types than erythrocytes, including lymphatic valves, endothelial cells, macrophages, megakaryocytes and platelets, and erythroid precursors. Our hypothesis is that the phenotype of patients with hereditary xerocytosis reflects an abnormal expression/function of mutated PIEZO1 in these different cell types. We propose a collaborative project involving a diagnostic reference centre and four research laboratories to study the expression and function of PIEZO1 and the consequences of its pathological activation in the cellular systems impacted in HX. The first step of this project will be the phenotypic characterisation of patients and the functional study of the identified mutations, by electrophysiology, carried out in the laboratory of Dr Shang Ma, an international expert in this field. The consequences of these mutations leading to abnormal activation of PIEZO1 will then be studied in three systems: (i) erythropoiesis and the red blood cell (haemolysis, dyserythropoiesis) (ii) megakaryocytes, platelets and endothelial cells (frequent thrombosis after splenectomy) (iii) enterocytes, which may be involved in the iron overload described in these patients. Each of these tasks will be led by a specialist in the theme, in close interaction with the other members of the consortium. The research teams will share three experimental models: (i) haematopoietic, endothelial and enterocytic cells cultivated in vitro and in which the activation and expression of PIEZO1 will be modulated, (ii) primary haematopoietic cells from patients and (iii) a knock-in mouse model reproducing the recurrent gain-of-function R2456H mutation described in the patients, already generated by one of the partners. This collaborative project based on complementary expertise and shared tools will allow, via an original multi-systemic approach, the identification of new therapeutic targets for patients for whom treatment options are still lacking. The consortium attaches a particular importance to share the results with patients that will be recruited in the bi-centric clinical trial and included in the COStho cohort, led by one of the partners. These patients will be actively involved in the progress of the research through an annual virtual meeting organised by the consortium.