Fig. 4: Flt-1 downstream signalling affects LAT1 and NRF2 signalling and cellular oxidative stress.
From: LAT1-NRF2 axis controls sFlt-1/PlGF imbalance and oxidative stress in preeclampsia
A Immunoblots of phosphorylated and total Flt-1, phosphorylated and total Akt, LAT1, and alpha-tubulin (A-Tub) in control, LAT1 deficient and SC79 (Akt activator) treated cells. B NFE2L2 (NRF2) mRNA levels and GSH/GSSG ratio in cells treated with Wortmannin C NFE2L2 (NRF2) mRNA levels, and GSH/GSSG ratio in cells deficient of LAT1 or treated with SC79. D NFE2L2 (NRF2) and SLC7A5 (LAT1) mRNA levels, and GSH/GSSG ratio in cells overexpressing PlGF or Flt-1 that have been previously depleted of Flt-1 or PlGF, respectively. E–G Viability, NFE2L2 (NRF2) and SLC7A5 (LAT1) mRNA levels, and GSH/GSSG ratio of PLECs treated for 72 h with conditioned media (CM) from control (siPool), LAT1- and NRF2 deficient HTR-8/SVneo cells. H Proposed mechanism of the LAT1 and NRF2 interaction. LAT1 depletion causes amino acid deficiency and further activates ATF4. Activation of ATF4 upregulates Flt-1 (and also sFlt-1) and downregulates PlGF levels. This decreases Flt-1 signalling, causing less Akt-mediated NRF2 activation. Reduced NRF2 activity leads to lower levels of NRF2 itself, but also of its target genes including LAT1. This ultimately leads to more oxidative stress in the cell. The scheme describes the experimental steps underlying the working model, the circular nature of the mechanism, and that increased oxidative stress is a secondary effect of reduced NRF2 function and impaired amino acid uptake in PE placentas. In addition, as shown in Figs. 1I, 2, and Fig. 4A–D, LAT1 and NRF2 strongly correlate and interact regardless of whether PE is present or not. Created in BioRender. Gundacker, C. (2025) https://BioRender.com/boezau2 Data represent mean value ± SD. A The experiment was repeated at least three times with similar results. B–D N = 3 (biological replicates); E–G N = 5 (biological replicates). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Two-sided unpaired t test was used in (B). ANOVA followed by Dunnett´s or Tukey´s multiple comparisons test was used in (C–G).
