Induction of Caspase-dependent Apoptosis in Rat Bone Marrow Mesenchymal Stem Cells Due to Di-2-Ethylhexyl Phthalate Toxicity was Found to Arrest the Cell Cycle at the G1 Stage

Background: Di-(2-ethylhexyl) phthalate (DEHP) is used as a plasticizer in polyvinyl chloride products which is widely utilized. Previously we found, DEHP reduced the viability and proliferation ability of bone marrow mesenchymal stem cells (BMSCs). Objective: In the present study, the mechanism of DEHP toxicity was investigated. Methods: Rat BMSCs were cultured up to 3^rd passage and their viability was determined after treatment with 100 and 500 μM of DEHP for 24 and 48 hours. The levels of sodium, potassium, and calcium as well as induction of apoptosis were investigated. Using flow cytometry, cell cycle analysis was performed and the expression of genes involved in the cell cycle was evaluated using reverse transcriptase-PCR. Data were analyzed and p < 0.05 was taken as the level of significance. Results: Although the viability and electrolyte level of BMSCs were not affected with 100 μM of DEHP, this environmental pollution induced caspase-dependent apoptosis in a concentration-dependent manner. In both of the concentrations, DEHP arrests the cell cycle at the G0/G1 phase, and the expression of Cdk2 and Cdk4 was significantly reduced whereas an over-expression of P53 was observed. However, the expression of the raf1 gene remained unchanged. Conclusion: DEHP induces caspase-dependent apoptosis in BMSCs and arrests the cell cycle due to the reduction of Cdk2 and Cdk4 expression via over-expression of P53.