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Fig. 2 Endothelial cell flow maintenance (Flow-M) quiescence stimulus leads to shallow quiescence depth. A, Representative images of human umbilical vein endothelial cell (HUVEC) under static (nonflow) or Flow-M conditions with 5-ethynyl-2′-deoxyuridine (EdU) incorporation and fixation at indicated times post–Flow-M release. Cultures stained for DAPI (4′,6-diamidino-2-phenylindole; white, nuclear mask) and EdU (red, S phase) or phospho-retinoblastoma protein (pRB; blue, interphase). Scale bar, 50 µm. White arrow, flow direction. B, Quantification of percent EdU+ cells with indicated conditions. n=3 replicates, 5 images per condition per replicate. C, Quantification of percent pRB+ cells with indicated conditions. n=3 replicates, 5 images per condition per replicate. D, Representative images of HUVEC under indicated density conditions with EdU incorporation and fixation at indicated times post–high-density release. Cultures stained for DAPI (white, nuclear mask) and EdU (red, S phase) or pRB (blue, interphase). Scale bar, 50 µm. E, Quantification of percent EdU+ cells with indicated conditions. n=3 replicates, 6 images per condition per replicate. F, Quantification of percent pRB+ cells with indicated conditions. n=3 replicates, 6 images per condition per replicate. Statistics, 1-way ANOVA with Tukey multiple comparisons test.