summary of current projects
Investigation of the role of the transcriptional co-regulator RIP-140 in endothelial homeostasis
Dr Damien Calay (BHF-funded post-doctoral fellow)
Receptor-interacting protein 140 (RIP140) is a transcriptional co-regulator highly expressed in metabolic tissues like the adipose tissue, the skeletal muscle and the liver. In these tissues, RIP140 regulates lipid and glucose metabolism, by acting as a co-repressor for most members of nuclear receptor family of transcription factors, like PPARs or ERs, through its binding to transcriptional repressors such as HDAC or CtBP. On the other hand, RIP140 can bind the transcriptional activator CBP and positively regulate the transcription factor NFkB in murine macrophages under LPS treatment, controlling inflammation. Our project aims to determine the regulation of the transcriptional co-regulator RIP140 in human primary endothelial cells under inflammatory conditions (TNFa, IL-1b, LPS) and its potential role in the regulation of inflammatory responses in these cells. Overexpression and silencing by RNA interference approaches have revealed that RIP140 is regulated under inflammatory conditions in endothelial cells and is an important regulator of the endothelial inflammatory response. Understanding the precise role of RIP140 in endothelial cells, particularly its exact regulatory functions in inflammation, might contribute to a better comprehension of endothelial dysfunction, an early and common feature of many vascular diseases. In this context, RIP140 might emerge as a potentially interesting therapeutic target.
Unravelling the molecular mechanisms of autoantibody-mediated endothelial dysfunction in systemic lupus erythematosus and antiphospholipid syndrome
Dr Charis Pericleous (Arthritis Research UK Career Development Fellow)
Endothelial injury leading to dysfunction is central to the development of cardiovascular disease (CVD) in many autoimmune rheumatic diseases. Antiphospholipid syndrome (APS) and systemic lupus erythematosus (SLE) in particular carry a significant burden of associated CVD in young people. APS accounts for one in six strokes in patients under 50 years old and carries an almost 3-fold greater risk of atherosclerosis compared to matched controls. SLE is the archetypal example for autoimmune-mediated accelerated CVD, with patients less than 40 years old having a 5.6-fold increased risk of atherosclerosis and at less than 50 years, a 5.9-fold greater coronary heart disease risk. Circulating IgG autoantibodies found in patients with APS and SLE are key players in autoimmune-mediated vasculopathy, however the underlying molecular mechanisms involved are ill-defined. My work aims to establish how autoantibodies disrupt intracellular homeostatic and cytoprotective signalling while enhancing pathogenic responses, and how such perturbations at the molecular level influence endothelial function. Ultimately, my goal is to identify novel therapeutic targets that will allow the development of targeted treatments to improve vascular outcomes in patients with autoimmune rheumatic disease.
Investigation of the molecular mechanisms underlying PKCε-mediated modulation of NF-κB signalling to selectively promote vascular endothelial homeostasis
Dr Garrick Wilson (BHF-funded post-doctoral fellow)
We have established an important role for PKCε in endothelial homeostasis and specifically cell survival, resistance to injury and inflammation. A key factor in our understanding has been the observation that PKCε selectively modulates NF-κB transcriptional responses to induce cytoprotective genes, while suppressing pro-inflammatory pathways. Using a cell molecular and in vivo approach, we propose to identify the molecular mechanisms underlying selective targeting of NF-κB cytoprotective transcripts by PKCε. A key aim will be to determine the influence of shear stress patterns on the activation of PKCε and NF-κB, and the subsequent induction of protective genes. Analyses will include investigation of post-translational modifications and epigenetic mechanisms, the study of physiological PKCε agonists and novel PKCε targets including CREB-1 and Nrf2, and demonstration of the functional importance of PKCε-NF-κB signalling in the vascular endothelium in vivo. Ultimately, the value of targeting PKCε therapeutically will be determined by understanding its associated signalling pathways. We anticipate that this approach will eventually lead to identification of optimal targets and novel therapeutics for vascular inflammation, endothelial dysfunction and atherosclerosis.
Analysis of extracellular vesicle interactions with vascular endothelium under physiological shear stress to determine their role in endothelial injury, cytoprotection and as a therapeutic target
Dr Allan Kiprianos (BHF-funded post-doctoral fellow)
Extracellular vesicles (EVs), including exosomes and microvesicles (MVs), are important for cell-cell communication. MVs have been implicated in endothelial cell (EC) injury, inflammatory responses and atherogenesis. However, their impact on EC function may also be protective and questions remain concerning the real influence of circulating EVs on vascular endothelial function.Using physiological and pathophysiological in vitro and in vivo models, the aims of the current proposal are: (i) To compare the ability of EVs to bind endothelium under distinct shear stress conditions, to identify pro-atherogenic and protective outcomes, and to determine the effect of therapeutic conditioning. (ii) To identify molecular pathways regulating pro-inflammatory/proapoptotic and protective actions of MVs and exosomes, focusing on intracellular signalling and miRNAs, and (iii) To translate findings using in vivo and ex vivo human and murine models. We propose that the relationship between EVs, endothelial dysfunction and cardiovascular disease suggests therapeutic modulation of EVs may be beneficial, and that a detailed understanding of the functional outcomes of EV interactions with the endothelium is a critical next step.
The role of extracellular vesicles in the regulation of endothelial homeostasis and dysfunction during systemic inflammatory disease
Marie Lang (NHLI-funded PhD student
Our group is interested in the role of the vascular endothelium in the accelerated development and progression of cardiovascular disease in patients with systemic inflammatory rheumatic diseases. This project addresses the question how chronic systemic inflammation increases the susceptibility of the endothelium to atherogenesis. Our focus lies particularly on the effects of extracellular vesicles (EV) on endothelial (dys‑)function and their potential therapeutic modulation. EV comprise a heterogeneous group of small, cell-derived particles, which function as inter-cellular messengers in health and disease states. EV derived from endothelial cells (EC) are more abundant in patients with cardiovascular and systemic inflammatory diseases. We are currently investigating the mechanisms of endothelial-derived EV interaction with EC under inflammatory conditions. To explore vasculo-protective therapeutic strategies, we aim to examine the effects of methotrexate (MTX), a routinely given disease-modifying anti-rheumatic drug (DMARD), on EV phenotype, content and functionality and cyto-protective endothelial signaling pathways involving AMP-activated kinase (AMPK) and cAMP response element binding (CREB).
Relationship between protein kinase Cε and transcriptional co-regulator RIP140 in endothelial to cardiomyocyte cross-talk during vascular inflammation
Jerome Fourré (BHF-funded PhD student)
Hypothesis: RIP140 induction during chronic inflammation predisposes to endothelial and cardiomyocyte dysfunction. This can be reversed by PKCε activation, which favours translocation of RIP140 from the nucleus to the cytoplasm resulting in reduced NF-κB activation, enhanced mitochondrial biogenesis and protection against ER stress and apoptosis.
Aim 1: To determine the relationship between PKCε and RIP140 in EC and cardiomyocytes and to establish the mechanisms involved in their ability to influence NF-κB activation.
Aim 2: To investigate the effects of increased nuclear expression of RIP140 on the function of endothelial cells and cardiomyocytes, and on the cross-talk between them during chronic inflammation.
Aim 3: To determine the sub-cellular localisation of RIP140 in EC and cardiomyocytes of PKCε-/-and RIP140 Tg mice and to assess susceptibility to pro-inflammatory stimuli and ER stress.
The research focuses on the activity of endothelial cells and cardiomyocytes in the inflammatory diseases that can alter the functions of the heart. By suppressing or overexpressing RIP140 in vitro cultures Jerome aims to understand the effects of this protein on electrophysiological responses, cell survival and structural properties. This involves techniques such as RT-qPCR, flow cytometry, high temporal or spatial resolution microscopy, optical mapping and patch-clamping.
An important aspect this research is also to study cardiac inflammation in a multicellular environment, with a particular interest on myocyte-endothelial cell cross-talk. Co-culture models will be used to investigate how the inflamed endothelium can impair the functions of the cardiomyocytes and vice versa.