Fig 1: FNDC5 deficiency enhanced palmitate-induced inflammation and NOX4 expression in primary cardiomyocytes (CMs). a Effects of palmitate (PA, 400 µM) treatment on Tnf-a, Il1b and Il6 mRNA levels in CMs of WT and FNDC5-/- mice. b Effects of PA treatment on NOX2 and NOX4 protein level in CMs of WT and FNDC5-/- mice. The measurement was 24 h after PA treatment for measuring the mRNA levels or protein levels. PA: Palmitate; Veh: Vehicle; F-/-: FNDC5-/-. Values are mean ± SEM. *P < 0.05 vs. Veh. †P < 0.05 vs. CMs-WT. n = 3
Fig 2: CA suppressed oxidative stress in mice induced by pressure-overload. (A) SOD activity, (B) MDA production and (C) NADPH activity were assessed in each group (n=15). mRNA levels of (D) NOX2 and (E) NOX4 were also detected in each group. Protein levels of (F) NOX2 and NOX4 and (G) quantitative analysis are presented for each group. (H) Immunohistochemistry staining for 4-HNE and (I) subsequent quantification was performed. *P<0.05 vs. the vehicle + sham group. #P<0.05 vs. the vehicle + AB group. 4-HNE, 4-hydroxynonenal; SOD, superoxide dismutase; MDA, malondialdehyde; NOX2, NADPH oxidase 2; NOX4, NADPH oxidase 4; CA, carnosic acid; AB, aortic banding.
Fig 3: CA protected against cardiac remodelling by regulating the AKT/GSK3ß/NOX4 signalling pathway. (A) Expression of p-AKT, p-GSK3ß and NOX4 were determined in each group (n=15). Quantification analysis of (B) p-AKT/t-AKT, (C) p-GSK3ß/t- GSK3ß and (D) NOX4 protein. *P<0.05 vs. the vehicle + sham group. #P<0.05 vs. the vehicle + AB group. CA, carnosic acid; AB, aortic banding; GSK3ß, glycogen synthase kinase 3 ß; p-, phosphorylated; t-, total; NOX4, NADPH oxidase 4.
Fig 4: Nucleotide-binding oligomerization domain-like receptor 3 (NLRP3) regulated the production of ROS in neonatal rat ventricular myocytes (NRVMs). (A) ROS production of NRVMs transfected with si NLRP3 or si NC and stimulated by ISO or phosphate-buffered saline (PBS) (n = 6). (B–E) Protein expression of NOX2, NOX4, and SOD2 in NRVMs in the indicated group. Western blot images (B) and quantitative results (C–E) (n = 6). *P < 0.05 vs. PBS + siNC, #P < 0.05 vs. ISO + siNC. (F) ROS production of NRVMs transfected with Ad-NLRP3 or Ad-NC and stimulated by ISO or PBS (n = 6). (G–J) Protein expression of NOX2, NOX4, and SOD2 in NRVMs in the indicated group. Western blot images (G) and quantitative results (H–J) (n = 6). *P < 0.05 vs. PBS + Ad-NC, #P < 0.05 vs. ISO + Ad-NC.
Fig 5: Oxidative stress level in corpus cavernosum. (A) Representative western blot results for NOX1, NOX2, and NOX4. (B–D) Relative expression of NOX1, NOX2, and NOX4 compared with ß-actin; data are shown as the fold changes over the control group; n = 4 for each group. (E) MDA concentration in corpus cavernosum was measured with an MDA assay kit; n = 4 for each group. *: p < 0.05 compared with control group. #: p < 0.05 compared with JAK2-/- group. §: p < 0.05 compared with DM group. DM: diabetes mellitus. NOX: NADPH oxidase. MDA: malondialdehyde
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