Effects Of Tripchlorolide On Proliferation And Apoptosis Of Mammalian Cells

To explore the mechanisms controlling cell cycle and apoptosis, an antitumor drug, tripchlorolide (TC), was applied to Chinese hamster ovary (CHO) cells. Application of TC induced cell cycle arrest and subsequent cell death of CHOC400 cells. Early-G1 treatment with TC severely delayed the progression of G1 phase, which might be due to downregulation of CDK2 kinase activity and of E2F-downstream-gene transcription. Studies on the state of Rb phosphorylation revealed there were two types of Rb phosphorylation in cell cycle: ERK-dependent and ERK-independent phosphorylation. In normal cells, the ERK-dependent Rb phosphorylation was predominant, while in early-G1 treated CHO cells the ERK-independent Rb phosphorylation prevailed. The aberrant Rb phosphorylation might contribute to the decreased transcription of E2F downstream genes.TC treatment at early-S phase induced the activation of intra-S-phase checkpoint and apparent delay of S-phase progression, while TC treatment at middle-S-phase did not, suggesting the sensitivity of early- or middle-S-phase cells to DNA damage was quite different. Analysis on DNA replication patterns demonstrated that the activation of intra-S-phase checkpoint inhibited the DNA replication initiated from late origins. Long-duration treatment of TC caused time- and dose-dependent apoptosis of CHOC400 cells. TC treatment induced translocation of Bax to mitochondria, degradation of Bcl-2 and release of cytochrom c from mitochondria. Overexpression of Bcl-2-EGFP in CHO cells inhibited the changes above and blocked the apoptosis induced by TC. These results demonstrated that TC induced apoptosis of CHO cells by mitochondrial pathway.