Idiopathic dilated cardiomyopathy (iDCM) is the most common cause of heart failure in the young/middle age population. A genetic basis is found in 20-50% of iDCM cases. Unfortunately, there is no effective therapy to reverse iDCM. The heart is a metabolically demanding organ and mitochondria are paramount to the survival and activity of cardiomyocytes. In a former collaboration, partners of this consortium identified imbalanced mitochondrial dynamics (fusion and fission) and mitochondrial fragmentation as an underlying cause of iDCM in mice. A cardiac metabolic switch was involved in the iDCM pathogenesis. More importantly, a simple dietary approach based on high fat diet was able to inhibit this metabolic switch and prevent iDCM. The proposed work program seeks to uncover the nature of these cardioprotective dietary elements in small and large animal models with the ultimate aim of developing a novel therapy to treat iDCM in humans. Mice with overt iDCM will be fed diets composed of different substrates (fatty acids, carbohydrates, and proteins) and different compositions of fatty acids in order to pinpoint the dietary components responsible for cardioprotection. Porcine iDCM models will expand the translational relevance of these discoveries. Screening of iDCM patients for germline mutations affecting mitochondrial dynamics will be tested in vitro to evaluate the impact on mitochondrial fusion and fission. Overall, this project will provide the foundation for future clinical trial testing a simple, cost-effective therapy aimed at curbing the heart failure epidemic.

DDysregulation of fat metabolism in liver and adipose tissues is a hallmark of insulin resistance. White and brite/beige adipose tissues, and liver produce proteins and lipids with systemic action on insulin sensitivity. The regulation of the pathways involved in the synthesis, release and use of fatty acids shall influence the secretory capacities of these organs to promote systemic effects on insulin sensitivity. Survey of human data shows that fatty acids are not the only metabolic villains in insulin resistance. There are major gaps in the understanding of the impact of modified fat metabolism in adipocytes and hepatocytes on insulin sensitivity. Therefore, HepAdialogue aims at identifying new mediators of insulin sensitivity which production is influenced by key nodes of regulation in adipose and hepatic lipid metabolic pathways. The partners have identified three proteins that play such a role. In white adipocytes, hormone-sensitive lipase (HSL) independently of its role in adipose tissue lipolysis physically interacts with the glucose-responsive transcription factor, ChREBP, to control de novo lipogenesis and fat cell insulin signaling. ChREBP also influences insulin sensitivity through its major role in the control of de novo fatty acid synthesis in the liver. The nuclear receptor PPARalpha controls fat oxidation. In adipose tissue, it also promotes white-to-brite conversion of fat cells. In the liver, PPARalpha acts as a free fatty acid sensor during adipose tissue lipolysis and controls ketogenesis. Unexpectedly, PPARalpha shares a common transcriptional target with ChREBP, the hepatokine FGF21 which controls brown and white adipose tissue metabolism. Identification of lipids and proteins with putative endocrine action has been initiated by the Partners. In HepAdialogue, it will be completed though combined lipidomics, proteomics and transcriptomics analyses of adipocytes and hepatocytes in vitro and, of fat and liver from adipose-specific and hepatocyte-specific knock out mice in vivo. This part of the work is supported by already established models and preliminary unpublished data. Following validation on complementary models of a short list of secreted lipid and protein species, the influence of adipose factors on hepatocyte glucose metabolism and insulin signaling will be investigated. Conversely, we will study the influence of hepatic factors on adipocyte glucose metabolism and insulin signaling. Regulation of the production of the novel endocrine factors will be investigated in preclinical models of type 2 diabetes and tissue samples from human cohorts. Various pharmacological and nutritional approaches known to impact on the metabolic nodes of lipid metabolism will be used to manipulate the production and levels of the identified factors. Upon project completion, we aim at having identified and validated new secreted molecules with confirmed bioactivity in cell culture. Seeking industrial partnership, the next step will be to test whether the compounds themselves, their precursors or some inhibitors may have the capacity to reverse insulin resistance and associated metabolic disorders and/or to induce browning of white adipose tissues in vivo.

The clinical spectrum of cognitive disorders (CD) varies widely from Intellectual Disability (ID) to Autism Spectrum Disorder (ASD) and is estimated to affect 1-3% of the population. Genetic evidence indicates that one major functional group of CD-related proteins corresponds to proteins that are enriched at synaptic compartments, defining the concept of synaptopathies. While many studies have focused on the involvement of neurons in the pathology, the multi-partite synapse questions the contribution of the glia in CD, thus urging the study of the astrocyte-to-neuron bidirectional communication in cellular and animal models. This project is based on the functional characterization of the interactions between astroglia and neurons in the context of synaptopathies. We will focus our studies on Oligophrenin1 gene, which mutations are associated with ID, ASD or schizophrenia. The Ophn1 gene encodes a RhoGAP protein that is expressed not only in neurons, but also in astrocytes during development and in adult. At the synapse, its neuronal functions have been largely reported, but its roles in astroglial cells are still unexplored. We have shown in a mouse model of the pathology that ophn1 loss is responsible for endocytosis defects in astrocytes together with hyperactivation of RhoA/ROCK and PKA pathways. Preliminary data suggest that loss of ophn1 function in glia also contributes to the dendritic spine phenotype observed in neurons. The cellular and molecular defects in astrocytes deserve a full characterization of the consequences of ophn1 inactivation in these cells. In this context, we will address the two following scientific questions: 1. What are the cell autonomous and non-cell autonomous consequences of oligophrenin1 loss of function in astrocytes? (i) To decipher its function in astrocytes, we will establish primary astroglial cultures from constitutive KO brain and monitor their morphology, adhesion capacities and vesicular trafficking. Given the well-known function of ophn1 on synapses, we will investigate the synaptic densities on co-culture systems of WT neurons on KO astrocytes and vice versa. Since we have access to iPS cells developed from cutaneous fibroblasts of OPHN1 mutated ID patients, we will differentiate them into glia or neurons to allow us to extend and confirm our results to humans. (ii) Beside these in vitro studies, conditional floxed alleles of ophn1 will be crossed to neuronal or astroglial Cre driver lines. Double transgenic animals (Flox;Cre) will be assessed in behavioral tests and we will compare their phenotypes to previously reported constitutive models. (iii) In parallel, we will perform stereotaxic injections of Cre expressing lentiviral vectors in floxed animals in order to study in only few cells the consequences of ophn1 inactivation. We will analyze astroglia cells at the molecular level (markers), cellular level (morphology) and functional level (electrophysiological recording and Ca2+ imaging). 2. Does restoring or improving astroglial functions can rescue some of the ID-linked phenotype observed in mice? We will tackle this question using two different approaches either genetic or pharmacologic. Since Ophn1 has been shown to negatively regulate the RhoA/ROCK signaling pathway, we will test whether reducing RhoA levels can restore ophn1 function in astrocytes from the conditional mouse model. Alternatively, we will treat conditional KO mice with drugs previously identified as beneficial in in vitro or in vivo systems. Our consortium aims to decipher the astroglial function of ophn1 a molecule involved in synaptopathy and to highlight the astrocyte contribution to the pathology. Our goal is ambitious because we have a comprehensive and multi-disciplinary strategy to explore Ophn1 mouse ID models. Through this comprehensive study from cell to the animal, we also hope to uncover new pharmacological approaches targeting the glial cells to improve the learning capacities of ID patients.