Abstract
BACKGROUND: Cardiomyocyte oxidative stress significantly contributes to the progression of hypertension-induced heart failure, highlighting the need for targeted therapies. We developed a novel peptide, NPA7, that coactivates the GC-A (guanylyl cyclase A)/cGMP and MasR (Mas receptor)/cAMP pathway. This study aimed to test NPA7's ability to inhibit oxidative stress by modulating the p62 (Sequestosome 1)-KEAP1 (Kelch-like ECH-associated protein 1)-NRF2 (nuclear factor erythroid 2-related factor 2) pathway in human cardiomyocytes (HCMs) and a rat model of hypertension. METHODS: Oxidative stress was induced in HCMs using H(2)O(2) with phosphate-buffered saline or NPA7 treatment. Intracellular reactive oxygen species levels were assessed via dihydroethidium staining. Western blotting analysis measured p62, KEAP1, and NRF2 protein levels, while GSH/GSSG (glutathione/glutathione disulfide) ratios and antioxidant gene expression were analyzed. HCMs were transfected with small interfering RNA targeting GC-A, MasR, or p62 before NPA7 and H(2)O(2) treatment. In vivo, spontaneously hypertensive rats received saline or NPA7, with normotensive Wistar Kyoto rats as control and cardiac oxidative stress, KEAP1 protein levels, NOX2 (NADPH oxidase 2), and p67 (NADPH oxidase subunit p67-phox) mRNA levels were measured. RESULTS: NPA7 reduced H(2)O(2)-induced reactive oxygen species levels and increased GSH/GSSG ratio in HCMs. Silencing GC-A (guanylyl cyclase A receptor) and MasR (Mas receptor) reversed NPA7's effects. NPA7 activated the KEAP1-NRF2 pathway, enhancing NRF2's antioxidant target gene expression. In p62 knockdown HCMs, NPA7-induced KEAP1 degradation and NRF2 activation were diminished. Reactive oxygen species levels were elevated in spontaneously hypertensive rat versusWistar Kyoto rats' hearts, however, NPA7 treatment reduced myocardial reactive oxygen species, suppressed KEAP1 protein, and decreased NOX2 and p67 mRNA levels. CONCLUSIONS: NPA7 exhibits antioxidant properties in HCMs and spontaneously hypertensive rat hearts by targeting GC-A and MasR through the p62-KEAP1-NRF2 pathway, supporting a novel therapeutic approach against cardiovascular disease-related oxidative stress.