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

. 2022 Oct 2;79(10):537. doi: 10.1007/s00018-022-04529-2

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

© The Author(s) 2022

Open AccessThis 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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/.

PMC Copyright notice

Fig. 4 — Cell density-dependent HIF-1α expression is modulated by Notch1 signaling regulation. A β-GA, GSI, RGDFV-peptide, A205804, or RGD-peptide were administered to high-density cultured U251-MG cells, and HIF-1α expression was evaluated by immunoblot assays. GAPDH was used as the loading control. The data are representative of three individual experiments. B NICD or HIF-1α expression was measured at different confluencies of U251-MG and U373-MG cells by immunoblot assays. Glioblastoma stem cells (GSC) were cultured for 1, 4, or 7 days to establish different sizes of spheroids, after which, NICD or HIF-1α were measured by immunoblot assays. GAPDH was used as the loading control. Relative pixel intensities were measured using ImageJ software. The data are representative of three individual experiments. C, D GSI (20 µM) or recombinant Dll4 (rDll4; 100 mg/L) were added to U251-MG cells in a time-dependent manner, and then NICD or HIF-1α levels were analyzed by immunoblot assays. GAPDH was used as the loading control. Relative pixel intensities were measured using ImageJ software. The data are representative of three individual experiments. E Low- or high-density cultured U251-MG cells were transfected with control or Notch1-targeting siRNA using Lipofectamine 2000™ reagent. After 48 h of siRNA transfection, HIF-1α activity was measured by the luciferase reporter assay (n = 3; Tukey’s post hoc test was applied to detect significant group effects in ANOVA, p < 0.0001; asterisks indicate a significant difference compared to 0% inhibition, ***p < 0.001, n.s.: non-significant). NICD levels were significantly decreased in low and high-density cultured U251-MG cells after si-Notch1 transfections compared to si-Controls (Inset). F Low- or high-density cultured U251-MG cells were maintained for 24 h, after which, the expression levels of NICD, HIF-1α, p-FAK(Y397), FAK, p-AKT, AKT, or GAPDH were evaluated by immunoblot assays. Relative pixel intensities were measured using ImageJ software. The data are representative of three individual experiments