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. 2022 Jan 13;5:51. doi: 10.1038/s42003-022-03005-8

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© The Author(s) 2022

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. 2 — a The conditional immortalized ACS-5003 neurons were transiently transfected with either RORA full length (pRORA-2000) or deletion reporter plasmids. After 24 h, the cells were treated with either 5 mM low glucose (LG) or 25 mM high glucose (HG) for 3 days and the relative RORA reporter activities were calculated n = 5. *, P < 0.05, vs. pRORA-2000 group. b The schematic picture for the potential transcriptional binding motif in the range of −300 to 200 (from transcription start site) on the RORA promoter with two potential Oct binding sites marked in red as well as related mutation sites marked in green. c The cells were transiently transfected by either a wild-type RORA reporter construct (pRORA-2000) or single point mutation at the site shown in (b), and then treated with either LG or HG for 3 days, and the RORA reporter activities were calculated n = 5. *, P < 0.05, vs. pRORA-2000 group. d The cells were transiently transfected by RORA full length (pRORA-2000), single mutant, or double mutations as indicated, or infected by SOD2 lentivirus (↑SOD2), and then treated with either LG or HG for 3 days; the RORA reporter activities were then calculated, n = 5. *, P < 0.0001, vs. pRORA-2000/LG group; ¶, P < 0.001, vs. pRORA-2000/HG group. e, f Cells were treated by either 4-day LG plus 4-day LG (LG(4d)+LG(4d)), or 4-day HG plus 4-day LG (HG(4d)+LG(4d)), or infected on day 4 by SOD2 lentivirus (HG(4d)+LG(4d)/↑SOD2); the cells were then used for ChIP analysis: e ChIP analysis by potential transcription factors on the RORA promoter, n = 4; f ChIP analysis by potential histone methylation, n = 4. *, P < 0.0001, vs. LG(4d)+LG(4d)/CTL group. g–i Cells were treated by either LG(4d)+LG(4d)/CTL or HG(4d)+LG(4d)/CTL, or the cells were infected on day 4 by either Oct3/4 expression lentivirus (HG(4d)+LG(4d)/↑Oct3/4) or Oct3/4 knockdown lentivirus (LG(4d)+LG(4d)/shOct3/4); the cells were then harvested for biomedical analysis: g mRNA analysis, n = 4. h Protein quantitation, n = 5. i Representative western blotting pictures for (h). *, P < 0.0001, vs. LG(4d)+LG(4d)/CTL group. Data were expressed as mean ± SEM.