Amorphous silica nanoparticles trigger vascular endothelial cell injury through apoptosis and autophagy via reactive oxygen species-mediated MAPK/Bcl-2 and PI3K/Akt/mTOR signaling

Abstract

Environmental exposure to silica nanoparticles (SiNPs) is inevitable due to their widespread application in industrial, commercial, and biomedical fields. In recent years, most investigators focus on the evaluation of cardiovascular effects of SiNPs in vivo and in vitro. Endothelial injury and dysfunction is now hypothesized to be a dominant mechanism in the development of cardiovascular diseases. This study aimed to explore interaction of SiNPs with endothelial cells, and extensively investigate the exact effects of reactive oxygen species (ROS) on the signaling molecules and cytotoxicity involved in SiNPs-induced endothelial injury. Significant induction of cytotoxicity as well as oxidative stress, apoptosis, and autophagy was observed in human umbilical vein endothelial cells following the SiNPs exposure (P<0.05). The oxidative stress was induced by ROS generation, leading to redox imbalance and lipid peroxidation. SiNPs induced mitochondrial dysfunction, characterized by membrane potential collapse, and elevated Bax and declined bcl-2 expression, ultimately leading to apoptosis, and also increased number of autophagosomes and autophagy marker proteins, such as LC3 and p62. Phosphorylated ERK, PI3K, Akt, and mTOR were significantly decreased, but phosphorylated JNK and p38 MAPK were increased in SiNPs-exposed endothelial cells. In contrast, all of these stimulation phenomena were effectively inhibited by N-acetylcysteine. The N-acetylcysteine supplement attenuated SiNPs-induced endothelial toxicity through inhibition of apoptosis and autophagy via MAPK/Bcl-2 and PI3K/Akt/mTOR signaling, as well as suppression of intracellular ROS property via activating antioxidant enzyme and Nrf2 signaling. In summary, the results demonstrated that SiNPs triggered autophagy and apoptosis via ROS-mediated MAPK/Bcl-2 and PI3K/Akt/mTOR signaling in endothelial cells, and subsequently disturbed the endothelial homeostasis and impaired endothelium. Our findings may provide experimental evidence and explanation for cardiovascular diseases triggered by SiNPs. Furthermore, results hint that the application of antioxidant may provide a novel way for safer use of nanomaterials.

Keywords:

• nanoparticle
• cytotoxicity
• mechanism
• oxidative stress

Supplementary materials

Figure S1 Lysosome impairment of HUVECs exposed to the SiNPs (50 μg/mL) for 24 hours.

Notes: LysoTracker Red (Beyotime) was used for the staining of lysosome in HUVECs. Compared with the control group, the significant decrease of fluorescence intensity in the SiNPs group indicated that SiNPs induced lysosome impairment in HUVECs.

Abbreviations: HUVECs, human umbilical vein endothelial cells; SiNPs, silica nanoparticles.

Figure S2 TEM images of HUVECs exposed to the SiNPs (50 μg/mL) for 24 hours.

Notes: (A) Control group. (B) The magnification of selected area of control showed evidently intact mitochondria. (C) The induction of mitochondrial swelling and cristae rupturing and disappearance after SiNPs exposure (black arrows), and also SiNPs deposition in mitochondria. (D and E) Severe mitochondrial swelling in SiNPs-treated endothelial cells (black arrows), and (E and F) autophagosome including cytoplasmic material, especially impaired mitochondria (white arrow).

Abbreviations: TEM, transmission electron microscopy; HUVECs, human umbilical vein endothelial cells; SiNPs, silica nanoparticles.

Acknowledgments

This work was supported by the National Natural Science Foundation of China (No 81102095, No 81202242, No 81573176, No 81230065), General Program of Beijing Natural Science Foundation (7162021, 7162022), Science and Technology Development Program of the Beijing Municipal Commission of Education (KM201510025005), and Opening Project of Beijing Key Laboratory of Metabolic Disorders Related Cardiovascular Diseases (2015DXWL01).

Disclosure

The authors report no conflicts of interest in this work.