Computational protocol: Effects of 5-Fluorouracil on Morphology, Cell Cycle, Proliferation, Apoptosis, Autophagy and ROS Production in Endothelial Cells and Cardiomyocytes

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Protocol publication

[…] The proportion of apoptotic cells was detected by the Annexin V / 7AAD staining. HUVE or HCM cells were plated at a density of 5x104 cells/well on 6-well plates and grown overnight. The subsequent day the cells were treated with increasing 5-FU concentrations. Controls received DMSO (negative) or Vincristine (positive). Each treatment was in triplicate and three different experiments were performed. After 96 hours, cells were harvested, counted, transferred into flow tubes, pelletted, resuspended in 100 μl of fresh 1X Annexin binding buffer (0.01 M HEPES pH 7.4; 0.14 M NaCl; 2.5 mM CaCl2) plus Annexin V fluorescein isothiocyanate (FITC) and 7AAD peridinin-chlorophyll protein (PerCP). After staining, samples were analyzed by flow cytometry within 1 hour using a FACSCanto (BD). Annexin V+/7AAD± were considered as apoptotic cells. The proportion of apoptotic cells was expressed as a percentage of the total cell number acquired, excluding debris, and analyzed using the BD FACSDiva and FlowJo software. […]

Pipeline specifications

Software tools BD FACSDiva, FlowJo
Application Flow cytometry
Organisms Homo sapiens
Diseases Breast Neoplasms, Cardiovascular Diseases, Colonic Neoplasms, Neoplasms, Stomatitis, Hand-Foot Syndrome, Drug-Related Side Effects and Adverse Reactions
Chemicals Fluorouracil, Oxygen