NiONP-Induced Oxidative Stress and Mitochondrial Impairment in an In Vitro Pulmonary Vascular Cell Model Mimicking Endothelial Dysfunction.

The development and use of nanomaterials, especially of nickel oxide nanoparticles (NiONPs), is expected to provide many benefits but also has raised concerns about the potential human health risks. Inhaled NPs are known to exert deleterious cardiovascular side effects, including pulmonary hypertension. Consequently, patients with pulmonary hypertension (PH) could be at increased risk for morbidity. The objective of this study was to compare the toxic effects of NiONPs on human pulmonary artery endothelial cells (HPAEC) under physiological and pathological conditions. The study was conducted with an in vitro model mimicking the endothelial dysfunction observed in PH. HPAEC were cultured under physiological (static and normoxic) or pathological (20% cycle stretch and hypoxia) conditions and exposed to NiONPs (0.5-5 µg/cm2) for 4 or 24 h. The following endpoints were studied: (i) ROS production using CM-H2DCF-DA and MitoSOX probes, (ii) nitrite production by the Griess reaction, (iii) IL-6 secretion by ELISA, (iv) calcium signaling with a Fluo-4 AM probe, and (v) mitochondrial dysfunction with TMRM and MitoTracker probes. Our results evidenced that under pathological conditions, ROS and nitrite production, IL-6 secretions, calcium signaling, and mitochondria alterations increased compared to physiological conditions. Human exposure to NiONPs may be associated with adverse effects in vulnerable populations with cardiovascular risks.

NiONPs-induced alteration in calcium signaling and mitochondrial function in pulmonary artery endothelial cells involves oxidative stress and TRPV4 channels disruption.

In New Caledonia, anthropic activities, such as mining, increase the natural erosion of soils in nickel mines, which in turn, releases nickel oxide nanoparticles (NiONPs) into the atmosphere. Pulmonary vascular endothelial cells represent one of the primary targets for inhaled nanoparticles. The objective of this in vitro study was to assess the cytotoxic effects of NiONPs on human pulmonary artery endothelial cells (HPAEC). Special attention will be given to the level of oxidative stress and calcium signaling, which are involved in the physiopathology of cardiovascular diseases. HPAEC were exposed to NiONPs (0.5-150 µg/cm2) for 4 or 24 h. The following different endpoints were studied: (i) ROS production using CM-H2DCF-DA probe, electron spin resonance, and MitoSOX probe; the SOD activity was also measured (ii) calcium signaling with Fluo4-AM, Rhod-2, and Fluo4-FF probes; (iii) inflammation by IL-6 production and secretion and, (iv) mitochondrial dysfunction and apoptosis with TMRM and MitoTracker probes, and AnnexinV/PI. Our results have evidenced that NiONPs induced oxidative stress in HPAEC. This was demonstrated by an increase in ROS production and a decrease in SOD activity, the two mechanisms seem to trigger a pro-inflammatory response with IL-6 secretion. In addition, NiONPs exposure altered calcium homeostasis inducing an increased cytosolic calcium concentration ([Ca2+]i) that was significantly reduced by the extracellular calcium chelator EGTA and the TRPV4 inhibitor HC-067047. Interestingly, exposure to NiONPs also altered TRPV4 activity. Finally, HPAEC exposure to NiONPs increased intracellular levels of both ROS and calcium ([Ca2+]m) in mitochondria, leading to mitochondrial dysfunction and HPAEC apoptosis.

Lung Antioxidant Depletion: A Predictive Indicator of Cellular Stress Induced by Ambient Fine Particles.

Regulations on ambient particulate matter (PM) are becoming more stringent because of adverse health effects arising from PM exposure. PM-induced oxidant production is a key mechanism behind the observed health effects and is heavily dependent on PM composition. Measurement of the intrinsic oxidative potential (OP) of PM could provide an integrated indicator of PM bioreactivity and could serve as a better metric of PM hazard exposure than PM mass concentration. The OP of two chemically contrasted PM2.5 samples was compared through four acellular assays, and OP predictive capability was evaluated in different cellular assays on two in vitro lung cell models. PM2.5 collected in Paris at a site close to the traffic exhibited a systematically higher OP in all assays compared to PM2.5 enriched in particles from domestic wood burning. Similar results were obtained for oxidative stress, expression of antioxidant enzymes, and pro-inflammatory chemokine in human bronchial epithelial and endothelial cells. The strongest correlations between OP assays and cellular responses were observed with the antioxidant (ascorbic acid and glutathione) depletion (OPAO) assay. Multivariate regression analysis from OP daily measurements suggested that OPAO was strongly correlated with polycyclic aromatic hydrocarbons at the traffic site while it was correlated with potassium for the domestic wood burning sample.

Stretch-activated Piezo1 Channel in Endothelial Cells Relaxes Mouse Intrapulmonary Arteries.

In intrapulmonary arteries (IPA), endothelial cells (EC) respond to mechanical stimuli by releasing vasoactive factors to set the vascular tone. Piezo1, a stretch-activated, calcium-permeable channel, is a sensor of mechanical stress in EC. The present study was undertaken to investigate the implication of Piezo1 in the endothelium-dependent regulation of IPA tone and potential involvement of Piezo1 in pulmonary hypertension, the main disease of this circulation. IPA tone was quantified by means of a myograph in control Piezo1+/+ mice and in mice lacking endothelial Piezo1 (EC-Piezo1-/-). Endothelial intracellular calcium concentration ([Ca2+]i) and nitric oxide (NO) production were measured, in mouse or human EC, with Fluo-4 or DAF-FM probe, respectively. Immunofluorescent labeling and patch-clamp experiments revealed the presence of Piezo1 channels in EC. Yoda1, a Piezo1 agonist, induced an endothelium-dependent relaxation that was significantly reduced in pulmonary arteries in EC-Piezo1-/- compared with Piezo1+/+ mice. Yoda1 as well as mechanical stimulation (by osmotic stress) increased [Ca2+]i in mouse or human EC. Consequently, both stimuli increased the production of NO. NO and [Ca2+]i increases were reduced in EC from Piezo1-/- mice or in the presence of Piezo1 inhibitors. Furthermore, deletion of Piezo1 increased α-adrenergic agonist-mediated contraction. Finally, in chronically hypoxic mice, a model of pulmonary hypertension, Piezo1 still mediated arterial relaxation, and deletion of this channel did not impair the development of the disease. The present study thus demonstrates that endothelial Piezo1 contributes to intrapulmonary vascular relaxation by controlling endothelial [Ca2+]i and NO production and that this effect is still present in pulmonary hypertension.