Cadmium induces migration of colon cancer cells : the roles of reactive oxygen species, p38 and Cyclooxygenase-2

Abstract

Background: Increasing evidence shows that exposure to environmental pollutants such as heavy metals is positively correlated with colorectal malignancy. Cadmium (Cd) is a heavy metal contaminant whose toxicity has been strongly associated with different types of cancer including colorectal cancer (CRC). Despite this, the underlying molecular mechanisms of Cd-induced CRC remain obscure. Methods-Aims: HT-29 human adenocarcinoma cells were employed to the study the effects of acute low-level Cd exposure (100 nM) on their migratory (scratch assay) and proliferative (MTT) capacity. To this end, Luciferase reporter assay was used to assess COX-2 transcriptional activity, and Western blotting was used to detect p38 Mitogen Activated Protein Kinase (MAPK) and Akt phosphorylation as well as Cyclooxygenase-2 (COX-2) expression in the presence or absence of specific inhibitors. Prostaglandin E2 (PGE2) levels were measured using Enzyme Linked Immunosorbent Assay (ELISA). Finally, reactive oxygen species (ROS) formation was assessed using Dihydroethidium (DHE) stain. Results: Our results show that the migratory capacity of cells treated with Cd were significantly higher than vehicle-treated cells. Cd caused a time-dependent increase in COX-2 transcription and protein expression. Treatment with celecoxib (10 µM), a COX-2 selective inhibitor, significantly reduced Cd-induced migration. Because ROS and p38 are implicated in COX-2 expression and migration, we determined if Cd modulates their levels. Indeed, Cd increased levels of ROS and phosphorylation of p38. Importantly, Cd-induced COX-2 expression and migration were significantly abolished when cells were pre-treated with N-Acetyl-Cysteine NAC (10 mM), a ROS scavenger, or SB202190 (10 µM), a specific p38 inhibitor. Furthermore, p38 phosphorylation was inhibited by NAC, suggesting that ROS acts through p38 to upregulate COX-2. In line with this, Cd (100 nM) caused a significant time dependent increase in PGE2 levels. This Cd-increased PGE2 was abrog