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. 2023 Mar 10;14:1323. doi: 10.1038/s41467-023-37094-8

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© The Author(s) 2023

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/.

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Fig. 1 — a Diagram illustrating the de novo serine biosynthesis pathway derived from glycolysis. 3-PG, 3-phosphoglycerate; P-Pyr, 3-phosphohydroxypyruvate; P-Ser, phosphoserine; Ser, serine. b GSEA profiles showing senescence-regulated genes were significantly enriched in glycine, serine and threonine metabolism. Analysis of PHGDH, PSAT1 and PSPH expression in different passages of HUVECs by RT-qPCR (c) and immunoblots (d). e Analysis of the relative intracellular serine levels in different passages of HUVEC cells (P10, P20 and P30) when cultured in serine-free medium. f Analysis of ATF4 expression in HUVECs during senescence by RT-qPCR and immunoblots. g RT-qPCR analysis of ATF4 and PHGDH transcription in siCtrl and siATF4 young (P10) and senescent (P20) HUVECs. h Analysis of the relative intracellular serine levels in siCtrl and siATF4 HUVECs when cultured in serine-free medium. i Effect of serine (Ser) and glycine (Gly) on HUVECs senescence as determined by SAHF formation (DAPI staining) and SA-β-gal staining. HUVECs (young, P10; senescent, P20) were cultured in serine-free medium and then treated with 300 μM serine or glycine for 48 h. Right panel: Quantification of the percentage of SAHF-positive and SA-β-gal positive cells. j RT-qPCR analysis of PHGDH, P21 and HMGB1 transcription in siCtrl and siPHGDH HUVECs when cultured in serine-free medium. k Effect of PHGDH knockdown on the senescence of HUVECs (young, P10; senescent, P20) when cultured in serine-free medium as determined by SAHF formation (DAPI staining) and SA-β-gal staining. Right panel: Quantification of the number of SAHF-positive and SA-β-gal positive HUVECs. l Effect of serine (Ser) on HUVECs senescence in control (siCtrl), PHGDH knockdown (siPHGDH) and ATF4-knockdown (siATF4) HUVECs. Cells were cultured in serine-free medium and then treated with 300 μM serine for 24 h. For h, j, l, HUVECs (P10) were used in the experiments. For c, e–j, data represent means ± SE; n = 3 independent experiments. For i, j, n = 4 independent experiments. Two-sided t-tests were used for statistical analysis. For d, a typical example of three biological replicates is shown.