三氧化二砷對THP-1細胞及SVEC4-10細胞之毒性機轉探討

Inorganic arsenic is a common environmental contaminant; chronic exposure to arsenic can alter the physiology of various key immune cells, particularly macrophages. The aim of first part research is to elucidate the key parameters associated with arsenic-induced toxicity and investigate the potential and mechanism of -lipoic acid (LA), a potent thioreducant, for reducing the toxicity in human promonocytic THP-1 cells. The result shows that a non-lethal concentration of arsenic trioxide (1 M) significantly induced the expression of heme oxygenase-1(HO-1), a response biomarker to arsenic, without stimulating measurable superoxide production. Co-treatment of cells with the HO-1 competitive inhibitor zinc protoporphyrin (Znpp) potentiated arsenic-induced cytotoxicity, indicating that HO-1 confers a cytoprotective effect against arsenic toxicity. In addition, low concentrations of arsenic trioxide (1 and 2.5 M) markedly inhibited monocyte-to-macrophage differentiation and expression of macrophage markers. Treatment of cells with LA attenuated arsenic trioxide-induced cytotoxicity and HO-1 over-expression and restored the redox state. In addition, LA neutralized arsenic trioxide-inhibition of monocyte maturation into macrophages and reversed the expression and activity of scavenger receptors. In conclusion, the cytotoxicity of arsenic trioxide is associated with an imbalance of the cellular redox state, and LA can protect cells from arsenic-induced malfunctions either through its reducing activity, direct interacting with arsenic or stimulating other unidentified signaling pathways. Chronic arsenic exposure has been linked to an increase risk in vascular diseases. In second part, we proposed the molecular mechanisms through which arsenic causes injuries to blood vessels, we analyzed the effects of arsenic trioxide on the cytotoxicity, intracellular reactive oxygen species (ROS), the expression of related genes and signaling pathways involved in the SVEC4-10 mouse endothelial cells. Arsenic dose-dependently caused SVEC4-10 cell death, which is completely inhibited by α-lipolic acid (LA), a thioreducant, but partially ameliorated by Tiron, a potent superoxide scavenger. The mRNA levels of heme oxygenase-1 (HO-1), interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1) and vascular endothelial growth factor (VEGF) were significantly increased by arsenic. The up-regulation of these can be blocked by LA instead of Tiron, suggesting ROS is not important in their increase. HO-1 competitive inhibitor zinc protoporphyrin improved the cytotoxicty of arsenic in an inverted U dose-response curve, indicating the biphasic hormetic effect of HO-1. HO-1 siRNA decreased VEGF expression in response to arsenic. Arsenic exposure also enhanced NF-E2-related factor 2 (Nrf2) expression, and increased activation of nuclear factor-κB (NF-κB). NF-κB inhibitor Bay 11-7082 reduced arsenic-mediated expression of HO-1 and IL-6. Selective blocking of the MAPK pathways with p38 inhibitor SB203580 significantly decreased arsenic-induced HO-1 and VEGF expression, while JNKs inhibitor SP600125 increased IL-6 expression. These results suggest that in arsenic-treated SVEC4-10 cells, HO-1 expression is mediated through Nrf2-, NF-κB- and p38 MAPK-dependent signaling pathways, and serves as an upstream regulator of VEGF. IL-6 expression is regulated by NF-κB and JNKs. In conclusion, oxidative stress may be associated with arsenic-induced cytotoxicity and endothelial gene up-regulation, but signaling transduction dominates the direct effects of ROS.