Carbon black nanoparticles promote endothelial activation and lipid accumulation in macrophages independently of intracellular ROS production.

Yi Cao,Peter Møller,Steffen Loft,Pernille Høgh Danielsen,Martin Roursgaard,Clarissa Menezes Maya-Monteiro

doi:10.1371/journal.pone.0106711

Abstract

Exposure to nanoparticles (NPs) may cause vascular effects including endothelial dysfunction and foam cell formation, with oxidative stress and inflammation as supposed central mechanisms. We investigated oxidative stress, endothelial dysfunction and lipid accumulation caused by nano-sized carbon black (CB) exposure in cultured human umbilical vein endothelial cells (HUVECs), THP-1 (monocytes) and THP-1 derived macrophages (THP-1a). The proliferation of HUVECs or co-cultures of HUVECs and THP-1 cells were unaffected by CB exposure, whereas there was increased cytotoxicity, assessed by the LDH and WST-1 assays, especially in THP-1 and THP-1a cells. The CB exposure decreased the glutathione (GSH) content in THP-1 and THP-1a cells, whereas GSH was increased in HUVECs. The reactive oxygen species (ROS) production was increased in all cell types after CB exposure. A reduction of the intracellular GSH concentration by buthionine sulfoximine (BSO) pre-treatment further increased the CB-induced ROS production in THP-1 cells and HUVECs. The expression of adhesion molecules ICAM-1 and VCAM-1, but not adhesion of THP-1 to HUVECs or culture dishes, was elevated by CB exposure, whereas these effects were unaffected by BSO pre-treatment. qRT-PCR showed increased VCAM1 expression, but no change in GCLM and HMOX1 expression in CB-exposed HUVECs. Pre-exposure to CB induced lipid accumulation in THP-1a cells, which was not affected by the presence of the antioxidant N-acetylcysteine. In addition, the concentrations of CB to induce lipid accumulation were lower than the concentrations to promote intracellular ROS production in THP-1a cells. In conclusion, exposure to nano-sized CB induced endothelial dysfunction and foam cell formation, which was not dependent on intracellular ROS production.

Highlights

Exposure to nanoparticles (NPs) has been suggested to cause vascular health effects with oxidative stress and inflammation as central mechanisms [1]
We hypothesized that oxidatively stressed endothelial cells would be more readily activated and interact more strongly with monocytes or macrophages, and that oxidative stress could further promote the lipid accumulation in macrophages by exposure to NPs. To this end we investigated the effect of exposure to nanosized carbon black (CB) on the activation of endothelial cells by intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1) expression on human umbilical vein endothelial cells (HUVECs) and adhesion of THP-1 monocytes onto HUVECs as well as lipid accumulation in THP-1 macrophages
The viability of both the control and 100 mg/ml CB exposed THP-1 cells were over 90%, and there was no effect of CB exposure

Summary

Introduction

Exposure to nanoparticles (NPs) has been suggested to cause vascular health effects with oxidative stress and inflammation as central mechanisms [1]. It has been shown that exposure of endothelial cells to NPs promotes the expression of ICAM-1 and VCAM-1 as well as adhesion of monocytes onto the endothelial cells [3,4]. The process of endothelial activation might not require oxidative stress, as suggested by increased adhesion molecule expression by NP exposure in a manner not associated with generation of ROS [8,9]. It has been shown that addition of the antioxidant ascorbic acid to the cell culture medium did not alleviate particle-induced ICAM-1 and VCAM-1 expression on human umbilical vein endothelial cells (HUVECs) [10]. NP induced lipid accumulation in rat cells was inhibited by pre-treatment with the antioxidant N-acetylcysteine (NAC) [11]

Methods

Results

Conclusion