Coordination between NADPH oxidase and vascular peroxidase 1 promotes dysfunctions of endothelial progenitor cells in hypoxia-induced pulmonary hypertensive rats.

Previous studies have demonstrated that NADPH oxidase (NOX)/vascular peroxidase (VPO1) pathway - mediated oxidative stress plays an important role in the pathogenesis of multiple cardiovascular diseases. This study aims to evaluate the correlation between NOX/VPO1 pathway and endothelial progenitor cells (EPCs) dysfunctions in hypoxia-induced pulmonary hypertension (PH). The rats were exposed to 10% hypoxia for 3 weeks to establish a PH model, which showed increases in right ventricle systolic pressure, right ventricular and pulmonary vascular remodeling, acceleration in apoptosis and impairment in functions of the peripheral blood derived - EPCs (the reduced abilities in adhesion, migration and tube formation), accompanied by up-regulation of NOX (NOX2 and NOX4) and VPO1. Next, normal EPCs were cultured under hypoxia to induce apoptosis in vitro. Consistent with the in vivo findings, hypoxia enhanced the apoptosis and dysfunctions of EPCs concomitant with an increase in NOX and VPO1 expression, hydrogen peroxide (H2O2) and hypochlorous acid (HOCl) production; these phenomena were attenuated by NOX2 or NOX4 siRNA. Knockdown of VPO1 showed similar results to that of NOX siRNA except no effect on NOX expression and H2O2 production. Based on these observations, we conclude that NOX/VPO1 pathway-derived reactive oxygen species promote the oxidative injury and dysfunctions of EPCs in PH, which may contribute to endothelial dysfunctions in PH.

Wang EL ，Jia MM ，Luo FM ，Li T ，Peng JJ ，Luo XJ ，Song FL ，Yang JF ，Peng J ，Liu B ... -  《-》

Vascular peroxidase 1 mediates hypoxia-induced pulmonary artery smooth muscle cell proliferation, apoptosis resistance and migration.

Reactive oxygen species (ROS) play essential roles in the pulmonary vascular remodelling associated with hypoxia-induced pulmonary hypertension (PH). Vascular peroxidase 1 (VPO1) is a newly identified haeme-containing peroxidase that accelerates oxidative stress development in the vasculature. This study aimed to determine the potential role of VPO1 in hypoxia-induced PH-related vascular remodelling. The vascular morphology and VPO1 expression were assessed in the pulmonary arteries of Sprague-Dawley (SD) rats. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 4 (NOX4) and VPO1 expression and HOCl production were significantly increased in hypoxic rats, which also exhibited obvious vascular remodelling. Furthermore, a hypoxia-induced PH model was generated by exposing primary rat pulmonary artery smooth muscle cells (PASMCs) to hypoxic conditions (3% O2, 48 h), which significantly increased the expression of NOX4 and VPO1 and the production of HOCl. These hypoxic changes were accompanied by enhanced proliferation, apoptosis resistance, and migration. In PASMCs, hypoxia-induced changes, including effects on the expression of cell cycle regulators (cyclin B1 and cyclin D1), apoptosis-related proteins (bax, bcl-2, and cleaved caspase-3), migration promoters (matrix metalloproteinases 2 and 9), and NF-κB expression, as well as the production of HOCl, were all inhibited by silencing VPO1 with small interfering RNAs. Moreover, treatment with HOCl under hypoxic conditions upregulated NF-κB expression and enhanced proliferation, apoptosis resistance, and migration in PASMCs, whereas BAY 11-7082 (an inhibitor of NF-κB) significantly inhibited these effects. Collectively, these results demonstrate that VPO1 promotes hypoxia-induced proliferation, apoptosis resistance, and migration in PASMCs via the NOX4/VPO1/HOCl/NF-κB signalling pathway.

You B ，Liu Y ，Chen J ，Huang X ，Peng H ，Liu Z ，Tang Y ，Zhang K ，Xu Q ，Li X ，Cheng G ，Shi R ，Zhang G ... -  《-》

Magnesium Lithospermate B Derived from Salvia miltiorrhiza Ameliorates Right Ventricle Remodeling in Pulmonary Hypertensive Rats via Inhibition of NOX/VPO1 Pathway.

Right ventricle (RV) remodeling is a major pathological feature in pulmonary arterial hypertension (PAH). Magnesium lithospermate B (MLB) is a compound isolated from the roots of Salvia miltiorrhiza and it possesses multiple pharmacological activities such as anti-inflammation and antioxidation. This study aims to investigate whether MLB is able to prevent RV remodeling in PAH and the underlying mechanisms. In vivo, SD rats were exposed to 10% O2 for 21 d to induce RV remodeling, which showed hypertrophic features (increases in the ratio of RV weight to tibia length, cellular size, and hypertrophic marker expression), accompanied by upregulation in expression of NADPH oxidases (NOX2 and NOX4) and vascular peroxidase 1 (VPO1), increases in hydrogen peroxide (H2O2) and hypochlorous acid (HOCl) production and elevation in phosphorylation levels of ERK; these changes were attenuated by treating rats with MLB. In vitro, the cultured H9c2 cells were exposed to 3% O2 for 24 h to induce hypertrophy, which showed hypertrophic features (increases in cellular size and hypertrophic marker expression). Administration of MLB or VAS2870 (a positive control for NOX inhibitor) could prevent cardiomyocyte hypertrophy concomitant with decreases in NOX (NOX2 and NOX4) and VPO1 expression, H2O2 and HOCl production, and ERK phosphorylation. Based on these observations, we conclude that MLB is able to prevent RV remodeling in hypoxic PAH rats through a mechanism involving a suppression of NOX/VPO1 pathway as well as ERK signaling pathway. MLB may possess the potential clinical value for PAH therapy.

Li T ，Peng JJ ，Wang EL ，Li NS ，Song FL ，Yang JF ，Luo XJ ，Liu B ，Peng J ... -  《-》

Suppression of NADPH oxidase attenuates hypoxia-induced dysfunctions of endothelial progenitor cells.

NADPH oxidases (NOX) - derived reactive oxygen species (ROS) contribute to oxidative injury in hypoxia-induced pulmonary arterial hypertension. This study aims to evaluate the status of NOX in endothelial progenitor cells (EPCs) under hypoxic condition and to determine whether NOX inhibitors could attenuate hypoxia-induced dysfunctions of EPCs. EPCs were isolated from peripheral blood of SD rats and subjected to hypoxia (O2/N2/CO2, 1/94/5) for 24 h. The cells were collected for β-galactosidase or Hoechst staining, or for functional analysis (migration, adhesion and tube formation). The NOX expression, activity and H2O2 content in EPCs were measured. The results showed that hypoxia treatment promoted EPC senescence and apoptosis, accompanied by the deteriorated functions of EPCs (the reduced abilities in adhesion, migration and tube formation), as well as an increase in NOX2 and NOX4 expression, NOX activity and H2O2 production, these phenomena were attenuated by NOX inhibitors. Furthermore, administration of catalase could also improve the functions of hypoxia-treated EPCs. Based on these observations, we conclude that NOX-derived ROS contributes to the dysfunctions of EPCs under hypoxic condition. Thus, suppression of NOX may provide a novel strategy to improve endothelial functions in hypoxia-relevant diseases.

Liu B ，Ren KD ，Peng JJ ，Li T ，Luo XJ ，Fan C ，Yang JF ，Peng J ... -  《-》

Involvement of vascular peroxidase 1 in angiotensin II-induced hypertrophy of H9c2 cells.

Oxidative stress has been implicated in cardiac hypertrophy and heart failure. Vascular peroxidase 1 (VPO1), a peroxidase in the cardiovascular system, uses the hydrogen peroxide (H2O2) derived from co-expressed NADPH oxidases (NOX) to produce hypochlorous acid (HOCl) and catalyze peroxidative reactions. Our previous studies showed that VPO1 contributes to the vascular smooth muscle cell proliferation and endothelial dysfunction in spontaneous hypertensive rats (SHRs); however, the role of VPO1 in cardiomyocytes hypertrophy is still uninvestigated. The present study was therefore undertaken to examine the role of VPO1 in the angiotensin II-induced cardiac hypertrophy, and the underlying mechanism by which VPO1 regulates the redox signaling. As compared to WKY rats, the SHRs exhibited increased myocyte cross sectional area, enhanced Nox2 and VPO1 expression level in cardiac tissue, and an increased Ang II level in plasma. In cultured H9c2 cell line, Ang II increased the hypertrophy-related gene (BNP/ANF) expression and the cellular surface area, which was attenuated by knocking down of VPO1 via VPO1 siRNA or pharmacological inhibition of NOX/VPO1 pathway. Moreover, the enhanced hypochlorous acid (HOCl) production and phosphorylation of ERK1/2 was suppressed by VPO1 knockdown. Furthermore, the protective role of VPO1 siRNA transfection on H9c2 cardiomyocytes hypertrophy was abrogated on the HOCl stimulation, and the phosphorylated ERK1/2 expression level was found also upregulated after HOCl stimulation. In conclusion, these results suggest that the Nox2/VPO1/HOCl/ERK1/2 redox signaling pathway was implicated in the pathogenesis of Ang II-induced cardiac hypertrophy.

Yang W ，Liu Z ，Xu Q ，Peng H ，Chen L ，Huang X ，Yang T ，Yu Z ，Cheng G ，Zhang G ，Shi R ... -  《-》