Figure 3.

S1P activates STAT3 signaling to promote the transcription of Gpx4 and Slc7a11. a. Membrane S1PR1/2/3 levels in H9C2 cells and NRVMs were analyzed by Western blot; NaKATPase loading control; n=6. b. Western blot analysis and quantification of p-STAT3, STAT3 and nuclear p-STAT3 levels in H9C2 cells and NRVMs; LaminB loading control; n=6. c. IGV visualization of STAT3 binding events on the Gpx4 and Slc7a11 promoters, derived from ChIP-seq data in the GEO database (GSE117164). d. Schematic of the top five predicted STAT3 binding sites on the Gpx4 promoter, as predicted by JASPAR, with confirmed binding sites and their respective mutations shown in red. e. Schematic of the top five predicted STAT3 binding sites on the Slc7a11 promoter, as predicted by JASPAR, with confirmed binding sites and their respective mutations shown in red. f. Pearson correlation analysis of Stat3 and Gpx4 mRNA expression in the human left ventricle from the GEPIA database. g. Pearson correlation analysis of Stat3 and Slc7a11 mRNA expression in the human left ventricle from the GEPIA database. h. Quantitative analysis of Gpx4 mRNA expression in H9C2 cells and NRVMs measured by quantitative reverse transcription polymerase chain reaction (qRT-PCR); n=6. i. Quantitative analysis of Slc7a11 mRNA expression in H9C2 cells and NRVMs measured by qRT-PCR; n=6. All data are means ± standard deviations. Statistical analysis involved one-way ANOVA followed by Tukey's post-hoc test. Lines indicate comparisons between samples, and asterisks denote statistical significance (*P < 0.05, **P < 0.01, ***P < 0.001).