Benzene metabolism occurs primarily in the liver
Benzene metabolism occurs primarily in the liver where benzene is converted into phenol, catechol, hydroquinone (HQ). HQ is further converted to 1,4-benzoquinone (1,4-BQ) in the bone marrow which is the primary organ of benzene toxicity (Bolton et al., 2000). 1,4-BQ is an important benzene metabolite which has a significant role in benzene-induced myelotoxicity/myeloid neoplasms (Son et al., 2016). Benzene metabolites can induce intracellular formation of ROS. Studies shown that 1,4-BQ can lead to elevated levels of ROS (Shen et al., 1996).
ROS attack biological macromolecules, such as lipid, DNA, and proteins to cause oxidative damage (Shen et al., 1996), which plays a role in controlling various pathological processes (Diebold and Chandel, 2016; Kalashnikova and Mazar, 2017). Studies indicates that excessive ROS cause oxidative stress and thus destroy cellular homeostasis or cause defective repair of ROS-induced damage, contributing to bone marrow failure and acute myeloid leukemia (AML) (Richardson et al., 2015). Accumulating evidence shows that oxidative stress, especially ROS, has a great influence on stem cell characteristics and function (Ludin et al., 2014).
HIF-1a is reduced in the bone marrow cells in benzene exposed mice (Meng et al., 2016) and involved in modulating cell response to ROS and metabolism. Thus, it is hypothesized that overexpression of HIF-1a may play a protective role in benzene-induced toxicity via its target genes. To verify this hypothesis, we established a stable HIF-1a overexpression K562 cell line and investigated the effects of overexpression of HIF-1a on the cytotoxicity of the potent predominant benzene metabolite 1,4-BQ and the related mechanisms.
Materials and methods
Discussion Benzene exposure suppress WIN 18446 of hematopoietic progenitor cells (Yoon et al., 2001). 1,4-BQ can induce cell apoptosis and cell cycle arrest at G0/G1 phase. Cell cycle arrest may participate in DNA repair and cell survival (Hustedt and Durocher, 2016). Zhang et al. demonstrated that G6PD inhibition could cause G2 arrest thus enhancing 1,4-BQ-induced oxidative damage in K562 cells (Zhang et al., 2016). In addition, G2 arrest of the cell cycle can also activate mitochondria-dependent apoptotic events (Kim et al., 2016). HIF-1a is essential for cell cycle quiescence regulation in HSCs (Takubo et al., 2010). This present study found that overexpression of HIF-1α caused significant higher percentage of cells in G2 phase than that in HIF-1α control cells, suggesting that HIF-1α may regulate DNA damage repair and apoptosis through its role in cell cycle. The ROS and HIF-1α signaling have long been known to be involved in various disease processes, including inflammatory diseases, ischemia injury and tumor (Movafagh et al., 2015). ROS can destroy cell membranes and has cellular toxicity, and induce cell death. At 20 μM 1,4-BQ treatment group, significant increases in ROS level and the expression of HIF-1α were observed in NC cells. The difference in the relationship of ROS and HIF-1α between our previous in vivo finding and present in vitro result may due to various oxygen environments. The role of ROS on HIF-1Α regulation under hypoxia and normoxia is different. In hypoxia, with the ROS increasing the accumulation of HIF-1α should be decreased, but under normoxia HIF-1α increased (Movafagh et al., 2015). It was reported that ROS regulated HIF-1α expression in the cells specifically via H2O2 levels (Xia et al., 2007). In 1,4-BQ exposure, the main ROS production are hydrogen peroxide (H2O2) and hydroxylradicals (OH.) (Shen et al., 1996). Thus, 1,4-BQ induced ROS, mainly H2O2, is responsible for the expression of HIF-1Α. After 1,4-BQ exposure, we also found that the increase of ROS is accompanied by decreased Nox4 protein level. Nox family of NADPH oxidases is a significant source of ROS in cells, and Nox4 releases hydrogen peroxide (H2O2) (Nisimoto et al., 2014). The low level of Nox4 is helpful to reduce intracellular Nox4-derived ROS generation. This is a good way to reduce the cytotoxicity of ROS. In addition, our results showed that overexpression of HIF-1a reduced ROS levels in 1,4-BQ exposed K562 cells. Loss of HIF-1a was reported to induce cellular ROS production and lead to tumor cell death (Saito et al., 2015). Here our results indicated that overexpression of HIF-1a could lower ROS levels in K562 cells under 1,4-BQ exposure.