Transcriptomic and proteomic profiles of vascular cells involved in human abdominal aortic aneurysm

Diseases & disorder

Expression and implication of clusterin in left ventricular remodeling after myocardial infarction

International audienceBACKGROUND: Left ventricular remodeling (LVR) after myocardial infarction is associated with an increased risk of heart failure and death. In spite of a modern therapeutic approach, LVR remains relatively frequent and difficult to predict in clinical practice. Our aim was to identify new biomarkers of LVR and understand their involvement in its development.METHODS AND RESULTS:Proteomic analysis of plasma from the REVE-2 study (Remodelage Ventriculaire)-a study dedicated to the analysis of LVR which included 246 patients after a first anterior myocardial infarction-identified increased plasma levels of CLU (clusterin) in patients with high LVR. We used a rat model of myocardial infarction to analyze CLU expression in the LV and found a significant increase that was correlated with LVR parameters. We found increased CLU expression and secretion in primary cultures of rat neonate cardiomyocytes hypertrophied by isoproterenol. Silencing of CLU in hypertrophied neonate cardiomyocytes induced a significant decrease in cell size, ANP (atrial natriuretic peptide), and BNP (B-type natriuretic peptide) expression, associated with a decreased ERK (extracellular signal-regulated kinase) 1/2 activity, suggesting a prohypertrophic role of CLU. We then confirmed a significant increase of both intracellular p-CLU (precursor form of CLU) and m-CLU (mature form of CLU) in failing human hearts. Finally, the circulating levels of CLU (secreted form) were increased in patients with chronic heart failure who died from cardiovascular cause during a 3-year follow-up (n=99) compared with survivors (n=99).CONCLUSIONS: Our results show for the first time that plasma CLU levels are associated with LVR post-myocardial infarction, have in part a cardiac origin, and are a predictor of early death in heart failure patients

Proteomic Profiling of Macrophages by 2D Electrophoresis

International audienc

Modifications in Rat Plasma Proteome after Remote Ischemic Preconditioning (RIPC) Stimulus: Identification by a SELDI-TOF-MS Approach

<div><p>Remote ischemic preconditioning’s (RIPC) ability to render the myocardium resistant to subsequent prolonged ischemia is now clearly established in different species, including humans. Strong evidence suggests that circulating humoral mediators play a key role in signal transduction, but their identities still need to be established. Our study sought to identify potential circulating RIPC mediators using a proteomic approach. Rats were exposed to 10-min limb ischemia followed by 5- (RIPC 5′) or 10-min (RIPC 10′) reperfusion prior to blood sampling. The control group only underwent blood sampling. Plasma samples were isolated for proteomic analysis using surface-enhanced laser desorption and ionization - time of flight - mass spectrometry (SELDI-TOF-MS). A total of seven proteins, including haptoglobin and transthyretin, were detected as up- or down-regulated in response to RIPC. These proteins had previously been identified as associated with organ protection, anti-inflammation, and various cellular and molecular responses to ischemia. In conclusion, this study indicates that RIPC results in significant modulations of plasma proteome.</p></div

Serum MMP-8: a novel indicator of left ventricular remodeling and cardiac outcome in patients after acute myocardial infarction.

OBJECTIVE: Left ventricular (LV) remodeling following myocardial infarction (MI) is characterized by progressive alterations of structure and function, named LV remodeling. Although several risk factors such as infarct size have been identified, LV remodeling remains difficult to predict in clinical practice. Changes within the extracellular matrix, involving matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs), are an integral part of left ventricular (LV) remodeling after myocardial infarction (MI). We investigated the temporal profile of circulating MMPs and TIMPs and their relations with LV remodeling at 1 year and clinical outcome at 3 years in post-MI patients. METHODS: This prospective multicentre study included 246 patients with a first anterior MI. Serial echocardiographic studies were performed at hospital discharge, 3 months, and 1 year after MI, and analysed at a core laboratory. LV remodeling was defined as the percent change in LV end-diastolic volume (EDV) from baseline to 1 year. Serum samples were obtained at hospital discharge, 1, 3, and 12 months. Multiplex technology was used for analysis of MMP-1, -2, -3, -8, -9, -13, and TIMP-1, -2, -3, -4 serum levels. RESULTS: Baseline levels of MMP-8 and MMP-9 were positively associated with changes in LVEDV (P = 0.01 and 0.02, respectively). When adjusted for major baseline characteristics, MMP-8 levels remained an independent predictor LV remodeling (P = 0.025). By univariate analysis, there were positive relations between cardiovascular death or hospitalization for heart failure during the 3-year follow-up and the baseline levels of MMP-2 (P = 0.03), MMP-8 (P = 0.002), and MMP-9 (P = 0.03). By multivariate analysis, MMP-8 was the only MMP remaining significantly associated with clinical outcome (P = 0.02). CONCLUSION: Baseline serum MMP-8 is a significant predictor of LV remodeling and cardiovascular outcome after MI and may help to improve risk stratification

Increased clusterin levels after myocardial infarction is due to a defect in protein degradation systems activity.

Clusterin (CLU) is induced in many organs after tissue injury or remodeling. Recently, we show that CLU levels are increased in plasma and left ventricle (LV) after MI, however, the mechanisms involved are not yet elucidated. On the other hand, it has been shown that the activity of the protein degradation systems (PDS) is affected after MI with a decrease in ubiquitin proteasome system (UPS) and an increase in macroautophagy. The aim of this study was to decipher if the increased CLU levels after MI are in part due to the alteration of PDS activity. Rat neonate cardiomyocytes (NCM) were treated with different modulators of UPS and macroautophagy in order to decipher their role in CLU expression, secretion, and degradation. We observed that inhibition of UPS activity in NCM increased CLU mRNA levels, its intracellular protein levels (p-CLU and m-CLU) and its secreted form (s-CLU). Macroautophagy was also induced after MG132 treatment but is not active. The inhibition of macroautophagy induction in MG132-treated NCM increased CLU mRNA and m-CLU levels, but not s-CLU compared to NCM only treated by MG132. We also demonstrate that CLU can be degraded in NCM through proteasome and lysosome by a macroautophagy independent pathway. In another hand, CLU silencing in NCM has no effect either on macroautophagy or apoptosis induced by MG132. However, the overexpression of CLU secreted isoform in H9c2 cells, but not in NCM decreased apoptosis after MG132 treatment. Finally, we observed that increased CLU levels in hypertrophied NCM and in failing human hearts are associated with proteasome inhibition and macroautophagy alteration. All these data suggest that increased CLU expression and secretion after MI is, in part, due to a defect of UPS and macroautophagy activities in the heart and may have a protective effect by decreasing apoptosis induced by proteasome inhibition

Interplay between phosphorylation and O-GlcNAcylation of sarcomeric proteins in ischemic heart failure Running title: Interplay between phosphorylation and O-GlcNacylation of sarcomeric proteins

International audiencePost-translational modifications (PTMs) of sarcomeric proteins could participate to left ventricular (LV) remodeling and contractile dysfunction leading in advanced heart failure (HF) with altered ejection fraction. Using an experimental rat model of HF (ligation of left coronary artery) and phosphoproteomic analysis, we identified an increase of desmin phosphorylation and a decrease of desmin ON -acetylglucosaminylation (O-GlcNAcylation). We aim to characterize interplay between phosphorylation and O-GlcNAcylation for desmin in primary cultures of cardiomyocyte by specific O-GlcNAcase (OGA) inhibition with thiamet G and silencing O-GlcNAc transferase (OGT) and, in perfused heart perfused with thiamet G in sham-and HF-rats. In each model, we found an efficiency of O-GlcNAcylation modulation characterized by the levels of O-GlcNAcylated proteins and OGT expression (for silencing experiments in cells). In perfused heart, we found an improvement of cardiac function under OGA inhibition. But none of the treatments either in in vitro or ex vivo cardiac models, induced a modulation of desmin, phosphorylated and O-GlcNAcylated desmin expression, despite the presence of O-GlcNAc moities in cardiac desmin. Our data suggests no interplay between phosphorylation and O-GlcNAcylation of desmin in HF post myocardial infarction. The future requires finding the targets in heart involved in cardiac improvement under thiamet G treatment

Lim Domain Binding 3 (Ldb3) Identified as a Potential Marker of Cardiac Extracellular Vesicles

International audienceExtracellular vesicles (EVs) are considered as transporters of biomarkers for the diagnosis of cardiac diseases, playing an important role in cell-to-cell communication during physiological and pathological processes. However, specific markers for the isolation and analysis of cardiac EVs are missing, imposing limitation on understanding their function in heart tissue. For this, we performed multiple proteomic approaches to compare EVs isolated from neonate rat cardiomyocytes and cardiac fibroblasts by ultracentrifugation, as well as EVs isolated from minced cardiac tissue and plasma by EVtrap. We identified Ldb3, a cytoskeletal protein which is essential in maintaining Z-disc structural integrity, as enriched in cardiac EVs. This result was validated using different EV isolation techniques showing Ldb3 in both large and small EVs. In parallel, we showed that Ldb3 is almost exclusively detected in the neonate rat heart when compared to other tissues, and specifically in cardiomyocytes compared to cardiac fibroblasts. Furthermore, Ldb3 levels, specifically higher molecular weight isoforms, were decreased in the left ventricle of ischemic heart failure patients compared to control groups, but not in the corresponding EVs. Our results suggest that Ldb3 could be a potential cardiomyocytes derived-EV marker and could be useful to identify cardiac EVs in physiological and pathological conditions

Chapter Transcriptomic and proteomic profiles of vascular cells involved in human abdominal aortic aneurysm

Diseases & disorder