Activation of endothelial cells after exposure to ambient ultrafine particles: The role of NADPH oxidase |
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Authors: | Yiqun Mo Rong Wan Sufan Chien Qunwei Zhang |
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Institution: | a Department of Environmental Health and Occupational Health Sciences, School of Public Health and Information Sciences, University of Louisville, 485 E. Gray Street, Louisville, KY 40209, USA b Department of Surgery, University of Louisville, Louisville, KY, USA |
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Abstract: | Several studies have shown that ultrafine particles (UFPs) may pass from the lungs to the circulation because of their very small diameter, and induce lung oxidative stress with a resultant increase in lung epithelial permeability. The direct effects of UFPs on vascular endothelium remain unknown. We hypothesized that exposure to UFPs leads to endothelial cell O2 − generation via NADPH oxidase and results in activation of endothelial cells. Our results showed that UFPs, at a non-toxic dose, induced reactive oxygen species (ROS) generation in mouse pulmonary microvascular endothelial cells (MPMVEC) that was inhibited by pre-treatment with the ROS scavengers or inhibitors, but not with the mitochondrial inhibitor, rotenone. UFP-induced ROS generation in MPMVEC was abolished by p67phox siRNA transfection and UFPs did not cause ROS generation in MPMVEC isolated from gp91phox knock-out mice. UFP-induced ROS generation in endothelial cells was also determined in vivo by using a perfused lung model with imaging. Moreover, Western blot and immunofluorescence staining results showed that MPMVEC treated with UFPs resulted in the translocation of cytosolic proteins of NADPH oxidase, p47phox, p67phox and rac 1, to the plasma membrane. These results demonstrate that NADPH oxidase in the pulmonary endothelium is involved in ROS generation following exposure to UFPs. To investigate the activation of endothelial cells by UFP-induced oxidative stress, we determined the activation of the mitogen-activated protein kinases (MAPKs) in MPMVEC. Our results showed that exposure of MPMVEC to UFPs caused increased phosphorylation of p38 and ERK1/2 MAPKs that was blocked by pre-treatment with DPI or p67phox siRNA. Exposure of MPMVEC obtained from gp91phox knock-out mice to UFPs did not cause increased phosphorylation of p38 and ERK1/2 MAPKs. These findings confirm that UFPs can cause endothelial cells to generate ROS directly via activation of NADPH oxidase. UFP-induced ROS lead to activation of MAPKs through induced phosphorylation of p38 and ERK1/2 MAPKs that may further result in endothelial dysfunction through production of cytokines such as IL-6. Our results suggest that endothelial oxidative stress may be an important mechanism for PM-induced cardiovascular effects. |
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Keywords: | UFPs ultrafine particles MPMVEC mouse pulmonary microvascular endothelial cells ROS reactive oxygen species DPI diphenyleneiodonium chloride CAT catalase NAC l(+)-cysteine" target="_blank">N-acetyl-l(+)-cysteine H2-DCFDA 2&prime 7&prime -dichlorodihydrofluorescein diacetate DCF dichlorofluorescein HE hydroethidine MAPK mitogen-activated protein kinase |
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