The pro-apoptotic effect of -tocotrienol was also assessed by measuring the expression of cytochrome c and cleaved PARP-1, both of which were significantly up-regulated in a dose-dependent manner, reaching 36% and 34% increase in protein expression, respectively, with the highest dose of 40 M -tocotrienol (Figure 8). Open in a separate window Open in a separate window Figure 8 Western blot analysis showing the effect of -tocotrienol on the expression levels of apoptosis related proteins in U937 cell line. h reduced the proliferation of U937 and KG-1 cells in a dose-dependent manner with a half inhibitory concentration (IC50) of 29.43 and 25.23 M, respectively. -tocotrienol also induced a dose and time-dependent decrease in the proliferation of both cell lines after 48 h of treatment with IC50s of 22.47 and 24.01 M for U937 and KG-1 cells respectively (Figure 1). Open in a separate window Figure 1 Effect of -tocotrienol on the cell viability of U937 (A) and KG-1 (B) cell lines. U937 and KG-1 were treated with various concentrations of -tocotrienol (0C50 M) for 24 Neu-2000 and 48 h. Cell viability was examined using MTS assay. *, ** and *** indicate 0.05, ? ? 0.001 and ? 0.0001 respectively. 3.2. Effect of -Tocotrienol on the Proliferation of Mesenchymal Stem Cells To test the selectivity of the elicited growth inhibitory effects of -tocotrienol against cancer cells, mesenchymal stem cells (MSCs) were treated with the various concentrations of -tocotrienol for 24 and 48 h. Neu-2000 Cell viability was then examined by MTS reagent. As shown in Figure 2, the cell viability of MSCs was not significantly altered upon -tocotrienol treatment, as compared to control untreated MSCs, except with the highest concentration, 50 M, after Neu-2000 48 h. Neu-2000 This indicates that -tocotrienol can cause cell death in leukemic cell lines with minor effects on normal human cells (Figure 2). All remaining experiments were therefor performed with 24 h exposure, which revealed no cytotoxic effects on normal MSCs. Open in a separate window Figure 2 Effect of -tocotrienol on the cell viability of normal mesenchymal stem cells. MCS cells incubated with various concentrations of -tocotrienol (10, 30 and 50 M) for 24 and 48 h and the cell viabilities were examined using an MTS assay kit. *** indicates ? 0.0001. 3.3. Effect of -Tocotrienol on the Cell Cycle Progression of AML Cell Lines The flow cytometric cell cycle analysis of control untreated U937 cells showed accumulation of the cells in the G0/G1 phase. Treated cells, however, showed a dose-dependent increase in the percentage of deceased cells in the sub-G0/G1 phase of the cell cycle, reaching 63.5% with 50 M Neu-2000 dose of -tocotrienol (Number 3). Similarly, the circulation cytometric cell cycle analyses of KG-1 cells treated with -tocotrienol showed a dose-dependent increase in the percentage deceased cells in the sub-G0/G1 phase, to be 64.5% with 50 M -tocotrienol (Number 4). Open in a separate window Number 3 Effect of -tocotrienol within the cell cycle progression of Rabbit polyclonal to ADI1 U937. (A) Propidium iodide staining and circulation cytometric analysis of cell cycle distribution of U937 cells treated with -tocotrienol for 24 h. The percentage of each cycle was identified using C Flow software. M5: sub-G1, M6: G0-G1 phase, M7: S phase, M8: G2/M phase. (B) Histogram analysis showing the percentage of cell cycle distribution of U937 cells treated with -Tocotrienol. Open in a separate window Number 4 Effect of -tocotrienol within the cell cycle progression of KG-1 cell collection. (A) Propidium iodide staining and circulation cytometric analysis of cell cycle distribution of KG-1 cells treated with -tocotrienol for 24 h. The percentage of each cycle was identified using C Flow software M5: sub-G1, M6: G0-G1 phase, M7: S phase, M8: G2/M phase. (B) Histogram analysis showing the percentage of.