ABSTRACT
Objective: The callus induction and subculture condition of Taxus chinensis var. mairei were optimized in the experiment. High Taxol-yielding callus was obtained, and the correlation between expression patterns of Taxol biosynthesis related genes and Taxol content was defined. Methods: Optimal hormone composition and concentration for induction and subculture of callus derived from T. chinensis var. mairei were briefly studied. The growth characteristics and Taxol content of calli derived from T. chinensis var. mairei (including vitro embryos, juvenile stems, and juvenile buds), T. media (vitro embryos) and T. brevifolia (vitro embryos) were compared. Expression of Taxol biosynthesis related genes in the above different calli was analyzed. Results MS Medium supplemented with 3.0 mg/L 2,4-D was suitable for callus induction of T. chinensis var. mairei, the induced rate is up to 92%. Subculture medium supplemented with 2.0 mg/L 2,4-D and 1.0 mg/L 6-BA was suitable for Taxol biosynthesis of T. chinensis var. mairei. Under the same culture conditions, Taxol content of calli derived from vitro embryos of T. chinensis var. mairei was the highest and had reached 0.027% of callus dry weight. The genes of Taxol biosynthesis related enzymes - geranylgeranyl diphosphate synthase (GGPPS), taxadiene synthase (TASY), taxane-10β-hydroxylase (T10βH), 10-deacetylbaccatin III-β-10-O-acetyltransferase (DBAT), phenylalanine aminomutase (PAM), and 3′-N-debenzoyltaxol N-benzoyltransferase (DBTNBT), were expressed at the high level in the high Taxol-producing callus. Conclusion: Vitro embryos can be used as the first choice expiant source to obtain high Taxol-yielding callus. To improve gene expression level of GG-PPS, TASY, T10βH, DBAT, PAM, and DBTNBT would promote Taxol biosynthesis.
ABSTRACT
OBJECTIVE: A major limiting factor in human cancer chemotherapy is toxicity in normal cells and tissues. Our goal was to determine whether normal proliferating cells could be protected from chemotherapeutic agents by taking advantage of the differential drug sensitivity of cell cycle G1 checkpoint in normal and cancer cells. METHODS: Normal peripheral blood mononuclear cells (PBMC) and ovarian cancer cell lines (OVCAR- 3 and SKOV-3) were initially treated with 10 nM of staurosporine for 48 hours. After removal of staurosporine contained media, both PBMC and ovarian cancer cells were treated with 20 nM of paclitaxel for 24 hours. Cells were then allowed to recover in drug-free medium for 4 days. The DNA contents and cell cycle changes were detected by FACScan flow cytometer in the cells harvested whenever the medium was changed. RESULTS: After pretreatment of ovarian cancer cell lines (OVCAR-3 and SKOV-3) with 10 nM of staurosporine followed by treatment with 20 nM of paclitaxel, both OVCAR-3 and SKOV-3 cells were selectively arrested in G2M phase of cell cycle by paclitaxel and they resumed their proliferative cycle to some extents after the drugs were removed and cultured with fresh media. However. pretreatment with 10 nM of staurosporine protected normal circulating PBMC that had been induced to proliferate in vitro with phytohemagglutinin from paclitaxel. Staurosporine-induced arrest of PBMC in G0/G1 phase was reversible, and arrested cells tolerated 10 nM of paclitaxel in culture. CONCLUSION: OVCAR-3 and SKOV-3 cancer cells can be targeted specifically with paclitaxel, following staurosporine-mediated, selective and reversible G0/G1 arrest in PBMC.
Subject(s)
Humans , Cell Cycle , Cell Line , Cytoprotection , DNA , Drug Therapy , Ovarian Neoplasms , Paclitaxel , StaurosporineABSTRACT
Objective To investigate the effects of taxol on QBC939 and its mechanism. Methods The cell proliferation was assessed by MTT assays; cell cycle kinetics and apoptosis were analyzed by flow cytometry and microscopic examination. Results Taxol inhibited the cell growth in concentration and time-dependent manners. The cell showed S and G2/M arrest and apoptosis. Conclusion Taxol suppresses the growth of QBC939 cells in vitro by causing cell-cycle arrest, and apoptosis of the cells.The mechanisms of taxol will provide theoretical guidance for clinical treatment.