Our results indicate that these cells express some markers associated with cardiovascular stemness and retain part of the multipotent properties of embryonic epicardial derivatives, spontaneously differentiating into easy muscle, and fibroblast/myofibroblast-like cells. B,D,E?=?50 m.(EPS) pone.0053694.s001.eps (7.9M) GUID:?43F5E5A0-B957-40F4-B9BE-CC37192A7D88 Figure S2: Quantification of and SMA expression in TGF-induced EPIC cultures. Quantitative PCR confirms the increased expression of – and -SMA in TGF1-treated EPICs (left). TGF2-treated cultures show an increased expression of -SMA but not -SMA (p value<0.05).(EPS) pone.0053694.s002.eps (6.8M) GUID:?7C250A45-E25F-40F6-9F9D-AFE599574ADE Physique S3: Ephrin and Eph EPIC profiling. Expression of Ephrin ligand and ephrin receptor (Eph) in EPICs.(EPS) pone.0053694.s003.eps (8.0M) GUID:?1C00D914-C0A5-4274-82B5-8F5FCFA95161 Physique Naftopidil (Flivas) S4: cEP behaviour on TG-fibrin matrices: proteolytic activity and sprouting. A. cEP spheroids show different proteolytic/sprouting responses when cultured in TG-BPM2 and TG-VEGF fibrin matrices as compared to control experiments (regular fibrin). HUVEC cells are shown as internal control for VEGF activity. B. cEP7 spheroids were embedded into a 3D fibrin matrix with TG-bound-BMP2 and -VEGF121 or soluble bFGF, Wnt3a, Wnt5a, and examined after 48 h. cEP sprouting quantification after the different treatments has been graphically offered. Scale bars: 100 m.(EPS) pone.0053694.s004.eps (10M) GUID:?60C42B90-C22B-43DD-98EF-C75C59943D37 Figure S5: cEP4 zymography and protease inhibitor assays. A. 10% SDS-PAGE gels with 1.5 mg/ml gelatin were used to run cell culture supernatants. Gelatin degradation (48 hours of zymographic reaction) is shown for media from cEP4, EPICs, and proper controls, including simple culture medium, plasmin and supernatant Naftopidil (Flivas) from HT1080 cells (HT1080 is a fibrosarcoma line known to express MMPs after TPA phorbol ester treatment). B. After 24 h cEP4 cells INSR cultured on fibrin gels degrade the substrate and aggregate at the bottom of the culture dish (left, asterisk). Treatment with aprotinin reduces proteolysis and cells remain in the surface of the fibrin gel (arrowheads).(EPS) pone.0053694.s005.eps (5.5M) GUID:?486D67E8-51F8-4F5A-8641-F1E3AE5F89C9 Abstract The non-muscular cells that populate the space found between cardiomyocyte fibers are known as cardiac interstitial cells (CICs). CICs are heterogeneous in nature and include different cardiac progenitor/stem cells, cardiac fibroblasts and other cell types. Upon heart damage CICs soon respond by initiating a reparative response that transforms with time into considerable fibrosis and heart failure. Despite the biomedical relevance of CICs, controversy remains around the ontogenetic relationship existing between the different cell kinds homing at the cardiac interstitium, as well as around the molecular signals that regulate their differentiation, maturation, mutual conversation and role in adult cardiac homeostasis and disease. Our work focuses on the analysis of epicardial-derived cells, the first cell type that colonizes the cardiac interstitium. We present here a characterization and an experimental analysis of the differentiation potential and mobilization properties of a new cell line derived from mouse embryonic epicardium (EPIC). Our results indicate that these cells express some markers associated with cardiovascular stemness and retain part of the multipotent properties of embryonic epicardial derivatives, spontaneously differentiating into easy muscle mass, and fibroblast/myofibroblast-like cells. Epicardium-derived cells are also shown to initiate a characteristic response to different growth factors, to display a characteristic proteolytic expression profile and to degrade biological matrices in 3D assays. Taken together, these data show that EPICs are relevant to the analysis of epicardial-derived CICs, and are a god model for the research on cardiac fibroblasts and the role these cells play in ventricular remodeling in both ischemic or non/ischemic myocardial disease. Introduction Cardiac muscle mass cells (cardiomyocytes) are frequently thought to be the most abundant cell type in the adult heart. However, multiple studies have shown that cardiac chamber walls comprise high numbers of non-myocyte cells. These cells and their milieu (the extracellular space between cardiomyocyte fibers) constitute the cardiac interstitium [1]C[2]. Due to Naftopidil (Flivas) the small relative size of cardiac interstitial cells (CICs) and the enormous contribution of cardiomyocytes to cardiac mass, the proportion of CICs versus cardiac muscle mass cells in the heart is frequently underestimated. In this regard, recent reports suggest that CICs could represent up to a 65% of non-cardiomyocyte cells in the organ [1]C[3]. The biomedical importance of CICs is usually illustrated by their massive involvement in the remodeling of cardiac ventricular walls after myocardial infarction, a phenomenon that is characterized by a progressive fibrosis [4]. This ventricular remodeling entails the initiation of an inflammatory response and the mobilization of CICs. Both phenomena have been described as a normal response of the adult heart to damage [5]. Other acquired cardiac diseases like dilated cardiomyopathy are also characterized by fibrotic disorders [6]. As already indicated, adult CICs are a.