| BACKGROUNDVitamin E (VE) is a general term used indiscriminately to refer to a group of naturally existing compounds called tocopherols and tocotrienols as well as synthetic Vitamin E, and acetate and succinate derivatives of both natural and synthetic a-tocopherol. As a fat-soluble membrane antioxidant, VE plays various physiological functions. Vitamin E succinate (VES) is the most effective derivative form of VE in cancer prevention and therapy. VES has been shown to be a widespread potent growth inhibitor of human cancer cells, but there are few papers about the effects and mechanisms of VES in lymphoid leukemia cell.The oncogenesis and development of tumor are associated with the unbalance of cyclins - cyclin dependent kinases (CDKs) - cyclin dependent kinase inhibitors (CKIs) system that regulates cell cycle. Cyclin-dependent kinase-2 (CDK2) is one of the most important proteins of cell cycle. And the changes and effects of CDK2 in VES-induced growth inhibition in cancer cells are known little.The oncogenesis and development of tumor are not only associated with cancer cell proliferation but also apoptosis. Bcl-2 and Bax is a couple of important regulatory proteins, which plays negative or positive function in inducing cell apoptosis. Also there are only limited results to understand the role of VES on the ratio of the expressions of Bcl-2 and Bax.C-Jun NH2-termmal kinase (JNK) is one of mitogen-activated protein kinases (MAPKs) in mammal. It maybe play an important role in cancer cell apoptosis induced by VES, although so far there is no date to confirm.PURPOSE(1)To determine the effects of VES in lymphoid leukemia cell in vitro.(2)To explore the roles of CDK2, Bcl-2, Bax and INK played in the signal transduction in VES-induced growth inhibition in lymphoid leukemia cell. MATERIALS AND METHODS(1) Fluorescence microscope was used to observe the morphological changes of apoptosis in Raji cells induced by VES. The cell growth curve, apoptosis ratio and the procession of cell cycle were assayed by cell counting and PI staining and FCM.(2) The changes of expressions of CDK2, Bcl-2 and Bax were assayed by immunocytochemical staining.(3) The effects of VES on the expression and activation of JNK were assayed by Western Blotting analysis.RESULTS(1) Both VES and VE could inhibit Raji cells growth in vitro. VES was more effective than VE, which inhibited cells growth in dose-dependent and time-dependent manners. After being treated with 5μg/ml, 10μg/ml and 20 μg/ml VES for 48h, the cellular growth inhibitory rate were respectively 28.33 %,45.83% and 85.83%. After being treated with 20μg/ml VES for 24h, 48h and 72h, the cellular growth inhibitory rate were respectively 30.16%, 85.83 % and 100%.(2)With FCM, the G1 sub-peak obviously showed on the DNA histogram, which suggested VES could induce apoptosis in Raji cells in dose-dependent and time-dependent manners. After being treated with 5μg/ml, 10μg/ml and 20μg/ml VES for 24h, the percents of apoptosis were respectively 23.9 %, 32.0% and 46.2%. After being treated with 20μg/ml VES for 24h, 48h and 72h, the percents of apoptosis were respectively 46.2%, 68.5% and 89.1 %. VES was more effective to induce apoptosis in Raji cells.(3) The percent of cells in the G0/G1-phase was increased and the cells in the S-phase was decreased after being treated with VES, which suggestedcells were arrested in G1-phase.(4) During growth inhibition induced by VES in Raji cells, the expressions of CDK2 and Bcl-2 were down-regulated, as the expression of Bax was increased.(5) In Raji cells treated with VES, the level of phosphorylated form of JNK increased at 6h and reached peak level at 12h, which continuing through 24h after treatment initiation. The level of phosphorylation state-independent JNK was not changed.CONCLUSIONS(1) VES could inhibit cell growth and induce apoptosis in Raji cells in vitro.(2) The changes of the expression of CDK2 and cell cycle may be related to the mechanisms of an...
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