Linearol's Cell Cycle Effects

A total of 10⁴ cells were treated with 91 and 182 μΜ for the T98 cell line, and 98 and 196 μΜ for the U87 cell line of linearol. Untreated cells were used as negative control having less than 1% of DMSO. At least three independent experiments were performed, and all samples were run in triplicates. Cells were treated with linearol at its IC50 and 2IC50 values. Flow cytometric analysis was performed 72 h post-treatment with linearol. For the DNA cell cycle, cells were treated with trypsin, centrifuged, washed with PBS twice and then incubated with PI (Propidium Iodide) working solution (50 µg/mL PI, 20 mg/mL RNase A, and 0.1% Triton X-100, Sigma-Aldrich, St. Louis, MO, USA) for 15 min at 37 °C in the dark. With the use of a flow cytometer (Omnicyt Flow Cytometer, Cytognos, Athens, Greece), the PI fluorescence of 10⁴ individual nuclei was determined. Then, the fractions of cells in G0/G1, S, G2/M and sub-G0/G1 phase were analyzed.