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CYTO_21152_sm_SuppFig1.tif3762KSupplementary Figure 1. Phenotype characterization of aortic pericytes in culture. Rat aorta pericytes present a myo-endothelial phenotype in culture with DMEM-low medium and 10% FBS, under the phase contrast microscopy show a great amount of stress fibers (A). Confocal microscopy of pericytes reveals the stress fibers by anti-smooth muscle α-actin (red) and DAPI show the nuclei (blue). Bar 50μm. (B). Flow cytometry histogram of pericyte marker Thy-1/CD90. Cells obtained from adult rat aorta explants express Thy-1/CD90, a pericyte and mesenchymal stem cell marker. Flow cytometry analysis indicated that 75% of the cell population obtained from adult rat aorta explants express Thy-1/CD90 (marked by the arrow) and 25% is superposed with the negative control (gray curve). These results indicate that the cell culture from aorta explant is enriched with pericyte cells with stem cell potential (C). Pericytes expressing markers from cells obtained with adult rat aorta explants. PCR reactions demonstrated that the cell culture obtained from adult rat aorta explants express pericyte markers αSMA, PDGFRα and PDFGRβ. Nestin expression indicates an undifferentiated state. GAPDH is internal control of a house-keeping gene expression (D). Optical density of gel amplicon products of each gene are represented in a histogram compared to GAPDH expression (considered as 100 % expression) (E).
CYTO_21152_sm_SuppMov1.avi2448KSupplementary Movie 1. Videos demonstrating time kinetics of [Ca2+]i responses of pericytes differentiated in the presence of RA alone. Cells induced to differentiation in the presence of RA alone (Video 1) or with RA and the differentiation cocktail (Video 2) were analyzed by single-cell calcium imaging. Cells were stimulated first by KCl [100 mM], then followed by NMDA 100 μM]. Fluo-3 fluorescence was collected at 515–530 nm using sampling rate of one image per second. For calculation of free cytosolic calcium concentration ([Ca2+]i), ionophore (4-Br-A23187) [5 μM] followed by EGTA [10 mM] were used to determine Fmax and Fmin fluorescence values, respectively.
CYTO_21152_sm_SuppMov2.avi1477KSupplementary Movie 1.
CYTO_21152_sm_SuppInfo.doc38KSupplementary Information.
MIFlowCyt-checklist.doc45KMIFlowCyt_checklist

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