Journal: Frontiers in Bioengineering and Biotechnology
Article Title: FOXO4-DRI regulates endothelial cell senescence via the P53 signaling pathway
doi: 10.3389/fbioe.2025.1729166
Figure Lengend Snippet: FOXO4-DRI can alleviate endothelial cell senescence induced by OGD. (A) Western blot analysis was conducted to quantify the changes in protein levels of Ki-67, Lamin B, P21, P16, and γ-H2AX in senescent HUVECs following treatment with FOXO4-DRI or PBS, with n ≥ 4 per group. (B) Representative immunofluorescence analysis (green for Ki-67, blue for DAPI) was performed to assess the number of Ki-67 positive HUVECs in different groups, with n = 5 per group and scale bar = 40 μm. (C) Fluorescent staining was carried out to analyze the content and localization changes of P21 and γ-H2AX in different groups, with n = 5 per group and scale bar = 40 μm. (D) Representative SA-β-GAL staining and quantification were performed to evaluate the number of positive cells in different groups, with n = 5 per group and scale bar = 100 μm. (E) Tube formation assays were conducted to analyze relative tube lengths in different groups, with n = 5 per group and scale bar = 400 μm. (F) Representative DHE staining and quantification were carried out to evaluate the number of DHE-positive cells in different groups, with n = 5 per group and scale bar = 100 μm. (G) Scratch assays were performed to assess endothelial cell migration coverage at 0, 12, 24, and 36 h in different groups, with n = 5 per group and scale bar = 400 μm. Data are represented as mean ± SEM. Statistical analysis assessed by 2-way RM ANOVA. (H) RT-qPCR analysis was performed to detect mRNA levels of IL-6 , IL-8 , IL-1β , and TNF-α in different groups, with n = 3 per group. Data are presented as mean ± SEM and analyzed using a two-tailed Student’s t-test. * indicates P < 0.05, ** indicates P < 0.01, *** indicates P < 0.001.
Article Snippet: In the D-Gal + FOXO4-DRI group, mice also received D-galactose injections and, after 4 weeks, were given intraperitoneal injections of FOXO4-DRI (5 mg/kg, MCE, HY-P4157) every 2 days for 4 weeks, followed by tissue collection.
Techniques: Western Blot, Immunofluorescence, Staining, Migration, Quantitative RT-PCR, Two Tailed Test