Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2021 Apr 1;23(4):413–423. doi: 10.1038/s41556-021-00637-6

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

PMC Copyright notice

Extended Data Fig. 1 — a, Immunoblot analysis of quiescence-associated protein markers in HUVECs transduced with a FOXO1^A3 (AdFOXO1^A3) or control (AdCtrl) adenovirus. A FLAG antibody was used to validate the expression of the FLAG-tagged FOXO1^A3 mutant. Tubulin served as loading control. b, Confocal images showing decreased EdU-incorporation in HUVECs transduced with AdFOXO1^A3. The analysis was performed 24h after transduction. DAPI was used to identify endothelial nuclei. c, Quantification of EdU-incorporation in AdCtrl and FOXO1^A3-expressing HUVECs. Values represent the percentage of EdU-labelled ECs, (n=6, 10 independent samples for AdCtrl and AdFOXO1^A3). d, 2HG levels in AdCtrl- and AdFOXO1^A3-transduced HUVECs measured by LC-MS, (n=4 independent samples). AU, arbitrary units. e, Immunoblot analysis of HUVECs transduced with a doxycycline-inducible control- (iLentiCtrl) or FOXO1^A3-encoding lentivirus (iLentiFOXO1^A3) showing expression of the FLAG-tagged FOXO1^A3 mutant after doxycycline (Dox) treatment for 48h. The asterisk (*) denotes an unspecific protein detected by the FLAG antibody. f, Immunoblot analysis showing that the FOXO1-induced quiescence signature is reversible. HUVECs transduced with iLentiCtrl or iLentiFOXO1^A3 were treated with Dox for 48h, after which Dox was removed from the culture media. HUVECs were then cultured for additional 48h. g, Quantitative RT-PCR (RT-qPCR) showing increased expression of canonical FOXO1 target genes in dense or sparse HUVEC cultures. Values are normalized to β-actin and represent fold-change regulation relative to control, (n=3 independent samples). Western blot data in a, e and f were from the respective experiment, processed in parallel, and are representative of at least three independent experiments. c, d and g, Data represent mean ± s.e.m.; a two-tailed unpaired t-test was used; *P<0.05; **P<0.01; ***P<0.001; ****P<0.0001. The numerical data, unprocessed western blots and P values are provided as source data.

Source data