Evaluation of the cytotoxicity, cell-cycle arrest, and apoptotic induction by Euphorbia hirta in MCF-7 breast cancer cells

Context: Euphorbia hirta L. (Euphorbiaceae) has been used as a folk remedy in Southeast Asia for the treatment of various ailments. Objective: The current study evaluates the cytotoxicity, cell-cycle arrest, and apoptotic induction by E. hirta in MCF-7 breast cancer cells. Materials and methods: Cytotoxic activity of methanol extract of whole part of E. hirta was determined by the MTT assay at various concentrations ranging from 1.96 to 250.00 mu g/mL in MCF-7 cells. Cell morphology was assessed by light and fluorescence microscopy. Apoptosis and cell-cycle distribution were determined by annexin V staining and flow cytometry. DNA fragmentation, caspase activity, and reactive oxygen species (ROS) assays were performed using the commercially available kits. To identify the cytotoxic fraction, E. hirta extract was subjected to bioassay-guided fractionation. Results: Euphorbia hirta exhibited significant inhibition of the survival of MCF-7 cells and the half inhibitory concentration (IC50) values was 25.26 mu g/mL at 24h. Microscopic studies showed that E. hirta-treated cells exhibited marked morphological features characteristic of apoptosis. Euphorbia hirta extract also had an ignorable influence on the LDH leakage and generating intracellular ROS. The flow cytometry study confirmed that E. hirta extract induced apoptosis in MCF-7 cells. Euphorbia hirta also resulted in DNA fragmentation in MCF-7 cells. Moreover, E. hirta treatment resulted in the accumulation of cells at the S and G(2)/M phases as well as apoptosis. The caspase activity study revealed that E. hirta extract induced apoptosis through the caspase-3-independent pathway by the activation of caspase-2, 6, 8, and 9. Euphorbia hirta hexane fraction, namely HFsub4 fraction, demonstrated highest activity among all the fractions tested with an IC50 value of 10.01 mu g/mL at 24h. Discussion and conclusion: This study revealed that E. hirta induced apoptotic cell death and suggests that E. hirta could be used as an apoptosis-inducing anticancer agent for breast cancer treatment with further detailed studies.