The Vitamin E Derivative Gamma Tocotrienol Promotes Anti-Tumor Effects in Acute Myeloid Leukemia Cell Lines
"> Figure 1
<p>Effect of γ-tocotrienol on the cell viability of U937 (<b>A</b>) and KG-1 (<b>B</b>) cell lines. U937 and KG-1 were treated with various concentrations of γ-tocotrienol (0–50 µM) for 24 and 48 h. Cell viability was examined using MTS assay. *, ** and *** indicate <span class="html-italic">p</span> < 0.05, <span class="html-italic">p</span> < 0.001 and <span class="html-italic">p</span> < 0.0001 respectively.</p> "> Figure 2
<p>Effect of γ-tocotrienol on the cell viability of normal mesenchymal stem cells. MCS cells incubated with various concentrations of γ-tocotrienol (10, 30 and 50 µM) for 24 and 48 h and the cell viabilities were examined using an MTS assay kit. *** indicates <span class="html-italic">p</span> < 0.0001.</p> "> Figure 3
<p>Effect of γ-tocotrienol on the cell cycle progression of U937. (<b>A</b>) Propidium iodide staining and flow cytometric analysis of cell cycle distribution of U937 cells treated with γ-tocotrienol for 24 h. The percentage of each cycle was determined using C Flow software. M5: sub-G1, M6: G0-G1 phase, M7: S phase, M8: G2/M phase. (<b>B</b>) Histogram analysis showing the percentage of cell cycle distribution of U937 cells treated with γ-Tocotrienol.</p> "> Figure 4
<p>Effect of γ-tocotrienol on the cell cycle progression of KG-1 cell line. (<b>A</b>) Propidium iodide staining and flow cytometric analysis of cell cycle distribution of KG-1 cells treated with γ-tocotrienol for 24 h. The percentage of each cycle was determined using C Flow software M5: sub-G1, M6: G0-G1 phase, M7: S phase, M8: G2/M phase. (<b>B</b>) Histogram analysis showing the percentage of cell cycle distribution of KG-1 cells treated with γ-tocotrienol.</p> "> Figure 5
<p>Effect of γ-tocotrienol on cell death in U937 cells. (<b>A</b>) Apoptosis of U937 cells was measured by flow cytometry following dual staining with annexin V-FITC (FL1-H) and propidium iodide (FL2-H). Cells were treated with γ-tocotrienol 24 h. Cells were then analyzed using the C Flow software: the lower left quadrant shows cells which are negative for both propidium iodide (PI) and annexin (normal cells). The upper left quadrant shows only PI positive cells (necrotic). The lower right quadrant shows annexin positive cells (early apoptotic). The upper right quadrant shows annexin and PI positive cells (late apoptotic cells). (<b>B</b>) Histogram analysis showing the percentage of cells in each quadrant before and after treatment with γ-tocotrienol.</p> "> Figure 6
<p>Effect of γ-tocotrienol on cell death of KG-1 cells. (<b>A</b>) Apoptosis of KG-1 cells was measured by flow cytometry following dual staining with annexin V-FITC (FL1-H) and propidium iodide (FL2-H). Cells were treated with γ-tocotrienol for 24 h. Cells were then analyzed using the C Flow software. (<b>B</b>) Histogram analysis showing the percentage of cells in each quadrant before and after treatment with γ-tocotrienol.</p> "> Figure 7
<p>Effect of γ-tocotrienol on DNA fragmentation in U937 (<b>A</b>) and KG-1 <b>(B</b>) cell lines. Results show a dose-dependent increase in apoptotic enrichment factor upon treatment with γ-tocotrienol. Data represent mean ± standard error of the mean (SEM) from three independent experiments. *** indicates a significantly different mean values compared with control cells with <span class="html-italic">p</span> < 0.0001.</p> "> Figure 8
<p>Western blot analysis showing the effect of γ-tocotrienol on the expression levels of apoptosis related proteins in U937 cell line. (<b>A</b>) Representative western blotting images for the expression of Bax, Bcl-2, caspase-3, cytochrome c, and cleaved PARP-1 in U937 cells following treatment with γ-tocotrienol for 24 h. (<b>B</b>) Quantification analysis of the western blots. (<b>C</b>) Quantification of Bax to Bcl2 ration. Values represent the percent expression relative to control, normalized to β-actin, for a total of three western blots. *, ** and *** indicate <span class="html-italic">p</span> < 0.05, <span class="html-italic">p</span> < 0.001 and <span class="html-italic">p</span> < 0.0001 respectively.</p> "> Figure 8 Cont.
<p>Western blot analysis showing the effect of γ-tocotrienol on the expression levels of apoptosis related proteins in U937 cell line. (<b>A</b>) Representative western blotting images for the expression of Bax, Bcl-2, caspase-3, cytochrome c, and cleaved PARP-1 in U937 cells following treatment with γ-tocotrienol for 24 h. (<b>B</b>) Quantification analysis of the western blots. (<b>C</b>) Quantification of Bax to Bcl2 ration. Values represent the percent expression relative to control, normalized to β-actin, for a total of three western blots. *, ** and *** indicate <span class="html-italic">p</span> < 0.05, <span class="html-italic">p</span> < 0.001 and <span class="html-italic">p</span> < 0.0001 respectively.</p> "> Figure 9
<p>Effect of γ-tocotrienol on ROS level in KG-1 cells. TBHP was used in combination with γ-tocotrienol as a positive control in order to magnify the scale of change. γ-tocotrienol decreased ROS production in KG-1 cells in a dose-dependent manner. Data represent mean ± SEM from three independent experiments. *** indicates <span class="html-italic">p</span> < 0.0001.</p> ">
Abstract
:1. Introduction
2. Materials and Methods
2.1. Cell Culture
2.2. Isolation and Culture of Mesenschnymal Stem Cells (MSCs) from Rat Bone Marrow
2.3. Drug Preparation
2.4. Cell Viability Assay
2.5. Cell Cycle Analysis
2.6. Apoptosis Detection
2.7. DNA Fragemnetation using Cell Death ELISA
2.8. Western Blotting
2.9. Detecteion of Reacrtive Oxygen Species
2.10. Statistical Analysis
3. Results
3.1. Effect of γ-Tocotrienol on the Proliferation of AML Cell Lines
3.2. Effect of γ-Tocotrienol on the Proliferation of Mesenchymal Stem Cells
3.3. Effect of γ-Tocotrienol on the Cell Cycle Progression of AML Cell Lines
3.4. Effect of γ-Tocotrienol on Apoptosis in AML Cell Lines
3.5. Effect of γ-Tocotrienol on DNA Fragmentation in AML Cell Lines
3.6. Effect of γ-Tocotrienol on the Expression of Proteins involved in Pro-Apoptotic and Anti-Proliferative Pathways
3.7. Effect of γ-Tocotrienol on ROS Production in KG-1 Cell Line
4. Discussion
5. Conclusions
Author Contributions
Funding
Conflicts of Interest
References
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Ghanem, P.; Zouein, A.; Mohamad, M.; Hodroj, M.H.; Haykal, T.; Abou Najem, S.; Naim, H.Y.; Rizk, S. The Vitamin E Derivative Gamma Tocotrienol Promotes Anti-Tumor Effects in Acute Myeloid Leukemia Cell Lines. Nutrients 2019, 11, 2808. https://doi.org/10.3390/nu11112808
Ghanem P, Zouein A, Mohamad M, Hodroj MH, Haykal T, Abou Najem S, Naim HY, Rizk S. The Vitamin E Derivative Gamma Tocotrienol Promotes Anti-Tumor Effects in Acute Myeloid Leukemia Cell Lines. Nutrients. 2019; 11(11):2808. https://doi.org/10.3390/nu11112808
Chicago/Turabian StyleGhanem, Paola, Annalise Zouein, Maya Mohamad, Mohammad H. Hodroj, Tony Haykal, Sonia Abou Najem, Hassan Y. Naim, and Sandra Rizk. 2019. "The Vitamin E Derivative Gamma Tocotrienol Promotes Anti-Tumor Effects in Acute Myeloid Leukemia Cell Lines" Nutrients 11, no. 11: 2808. https://doi.org/10.3390/nu11112808