Low‐intensity pulsed ultrasound prevents prolonged hypoxia‐induced cardiac fibrosis through HIF‐1α/DNMT3a pathway via a TRAAK‐dependent manner

Abstract

Hypoxia-induced cardiac fibrosis is an important pathological process in cardiovascular disorders. This study aimed to determine whether low-intensity pulsed ultrasound (LIPUS), a novel and safe apparatus, could alleviate hypoxia-induced cardiac fibrosis, and to elucidate the underlying mechanisms. Hypoxia (1% O₂ ) and transverse aortic constriction (TAC) were performed on neonatal rat cardiac fibroblasts and mice to induce cardiac fibrosis, respectively. LIPUS irradiation was applied for 20 minutes every 6 hours for a total of 2 times in vitro, and every 2 days from 1 week before surgery to 4 weeks after surgery in vivo. We found that LIPUS dose-dependently attenuated hypoxia-induced cardiac fibroblast phenotypic conversion in vitro, and ameliorated TAC-induced cardiac fibrosis in vivo. Hypoxia significantly upregulated the nuclear protein expression of hypoxia-inducible factor-1α (HIF-1α) and DNA methyltransferase 3a (DNMT3a). LIPUS pre-treatment reversed the elevated expression of HIF-1α, and DNMT3a. Further experiments revealed that HIF-1α stabilizer dimethyloxalylglycine (DMOG) hindered the anti-fibrotic effect of LIPUS, and hampered LIPUS-mediated downregulation of DNMT3a. DNMT3a small interfering RNA (siRNA) prevented hypoxia-induced cardiac fibrosis. Results also showed that the mechanosensitive protein-TWIK-related arachidonic acid-activated K⁺ channel (TRAAK) messenger RNA (mRNA) expression was downregulated in hypoxia-induced cardiac fibroblasts, and TAC-induced hearts. TRAAK siRNA impeded LIPUS-mediated anti-fibrotic effect and downregulation of HIF-1α and DNMT3a. Above results indicated that LIPUS could prevent prolonged hypoxia-induced cardiac fibrosis through TRAAK-mediated HIF-1α/DNMT3a signalling pathway.