PS-BPB01-6: PROTECTIVE NOX4 EFFECT IN ENDOTHELIAL CELLS INVOLVES ACTIVATION OF TRANSIENT RECEPTOR POTENTIAL MELASTATIN 2 (TRPM2) CATION CHANNEL
Bibliographic record
Abstract
Objective: In endothelial cells, Nox4 is the most prevalent isoform and, in contrast to other Noxes, which generate superoxide O2, Nox4 produces mainly H2O2. Low H2O2 levels have been described to lower blood pressure, which suggests that a certain level of reactive oxygen species (ROS) can have protective effects on the cardiovascular system. We demonstrated that TRPM2 is a target of H2O2 in vascular cells. TRPM2 is a redox-sensitive Ca2+ channel and once activated leads to Ca2 influx. Endothelial dysfunction involves a reduction in eNOS activation by Ca2+ and is potentiated in the absence of Nox4. As TRPM2 channel is a H2O2 sensor and regulates Ca2+ entry, we questioned whether protective Nox4 effect in endothelial cells involves the H2O2-TRPM2-Ca2+ axis, with consequent regulation of NO signalling. Design and Methods: WT and TTRhRen hypertensive mice were crossed with Nox4 KO mice. Vascular function was measured by wire myograph in resistance arteries. ROS and H2O2 generation were assessed by lucigenin and amplex red, and Ca2+ influx by fluorescence microscopy in rat aortic endothelial cells (RAEC). eNOS activation was assessed by immunoblotting. Results: Blood pressure was significantly increased in TTRhRen (130.3 ± 7.0 mmHg) and TTRhRen/Nox4 KO mice (141.3 ± 18 mmHg) versus control mice (98.1 ± 8.0 mmHg). TTRhRen mice had reduced endothelium-dependent relaxation (Emax: WT 83.5 ± 4.03 vs TTRhRen 59.1 ± 3.5); an effect worsened by Nox4 KO (37.9 ± 5.4), p < 0.05. The TRPM2 activator, ADPR, improved vascular relaxation in TTRhRen/Nox4 KO mice (75.9 ± 7.7); an effect recapitulated by H2O2 (74.2 ± 15.4), p < 0.05. Ang II-induced ROS and H2O2 generation in RAEC, effect reversed by Nox4 siRNA and followed by increased Ca2+ influx (AUC - Ca2+: 20440.3333 ± 1870.9), important in eNOS activation. Ang II-induced Ca2+ influx in endothelial cells was reduced by TRPM2 siRNA, TRPM2 inhibitors (AUC - Ca2+: 8-br-cADPR 15232.2 ± 1052.1; Olaparib 14952.6 ± 843.2 and 2-APB 13270.8333 ± 277.4, p < 0.05) and by Nox4 siRNA and Nox4 inhibitor GKT137831 (AUC - Ca2+: GKT 15067.5 ± 255.6). Ang II-induced eNOS activation (Ser1177) (%: 72.3 ± 24.7) and NO release were blocked by Nox4 and TRPM2 sirNA, GKT137831, PEG-catalase and the TRPM2 inhibitor 8-br-cADPR (p < 0.05). Conclusions: We defined a novel molecular pathway in endothelial cells, where Nox4-induced H2O2 production activates PARP/TRPM2 signalling followed by Ca2+ influx, eNOS activation and NO release. The absence of Nox4 impairs Ca2+ homeostasis leading to endothelial dysfunction, an effect exacerbated in hypertension. The protective role of Nox4 corroborates the idea that the use of antioxidants based on the sole evidence that decreased oxidative stress is cardioprotective, it is not effective.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.001 | 0.002 |
| Insufficient payload (model declined to judge) | 0.005 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".