Figure 2.

Apoptosis and cell cycle in vitro study (mouse mammary carcinoma BJMC3879luc2 cells). (A) Cell viability was significantly lower in mouse mammary carcinoma BJMC3879luc2 cells treated with more than 12 μM α-mangostin for 24 or 48 hours (**P < 0.01). Five samples from each dosage of α-mangostin were examined. The IC50 concentration was determined to be 12 μM; therefore, 12 μM α-mangostin and 24 hour-incubation was used for all in vitro studies. (B) Caspase activities were evaluated using the luminescence assay. Activities of caspase-3, caspase-8 and caspase-9, but not caspase-12, were significantly elevated in BJMC3879luc2 cells treated with 12 μM α-mangostin for 24 hours (*P < 0.05 or **P < 0.01). Six samples from the control and eight samples from α-mangostin-treated cells were used for measurement of caspase-3 activity, and three samples from each group were used for activity measurements of the other caspases. (C) Cytochrome c in the cytosolic fraction, as determined by ELISA, was significantly increased in BJMC3879luc2 cells treated with α-mangostin for 24 hours compared to control levels (*P < 0.05). Three samples each from the control and α-mangostin-treated cell cultures were examined. (D) Western blots of Bid (22 kDa) in BJMC3879luc2 cells treated with or without α-mangostin for 24 hours were similar (upper panel). Cleaved Bid (15 kDa) was not observed after α-mangostin treatment. β-Actin served as the internal control (lower panel). (E) In BJMC3879luc2 cells treated with α-mangostin for 24 hours, cell viability was significantly increased by 10 or 100 μM of the broad-spectrum caspase inhibitor z-VAD-fmk, the caspase-3 specific inhibitor Ac-DNLD-CHO, the caspase-8 specific inhibitor z-IETD-fmk, and the caspase-9 specific inhibitor z-LETD-fmk (**P < 0.01). (F) Cell-cycle analysis showed that α-mangostin induced arrest in the G1-phase and inhibition of cells entering the S-phase in metastatic mouse mammary carcinoma BJMC3879luc2 cells (**P < 0.01). Data are presented as mean ± SD. Ac: acetyl; CHO: aldehyde; ELISA: enzyme-linked immunosorbent assay; fmk: fluoromethyl ketone; z: N-benzyloxycarbonyl.

Shibata et al. BMC Medicine 2011 9:69   doi:10.1186/1741-7015-9-69
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