BioMed Research International / 2014 / Article / Fig 1

Research Article

Effects of Crude Extracts from Medicinal Herbs Rhazya stricta and Zingiber officinale on Growth and Proliferation of Human Brain Cancer Cell Line In Vitro

Figure 1

Combination of CAERS and CFEZO acted synergistically to inhibit cell proliferation and colony formation in U251 cells. The U251 cells were seeded, at a density of 104/well in 96-well plates and treated with the indicated concentrations of CAERS and/or CFEZO for displayed time intervals. The inhibition of cell proliferation was assessed by the tetrazolium salt WST-1 kit as detailed in Section 2. The experiments were repeated five times in triplicate, and cell viabilities at each dose of extract(s) were expressed in terms of percent of control and reported as the mean ± SD. (b) Left histogram: multiple comparisons for effect of different doses’ categories on U251 cells using post hoc test (Dunnett T3). All reported P values are 2-tailed; **correlation is significant at the 0.01 level (2-tailed); ***correlation is significant at the 0.001 level (2-tailed); a = significant differences compared to control; b = significant differences compared to the lowest dose, 10 μg/mL. Compared to control, doses in 10, 20, 40, and 50 μg/mL showed a significantly lower percentage of viable U251 cells ( **, ***, ***, and ***, resp.). We observed another significant variation between dose 10 μg/mL and doses 40 and 50 μg/mL, with a stronger suppression of GBM cells with the two higher doses (40 and 50 μg/mL) ( **, and ***, resp.). Right histogram: multiple comparisons for effect of different durations on U251 viable cells using post hoc test (LSD). All reported P values are 2-tailed. *Correlation is significant at the 0.05 level (2-tailed). Compared to one-day duration, three-day duration of treatment showed a significantly lower percentage of viable U251 cells ( *). The table (on the right) shows regression of dose and duration with percent U251 viable cells. A multiple linear regression analysis was used (Enter Selection Procedure).***Correlation is significant at 0.001 level (2-tailed). Dose had a stronger negative impact than that of duration, in predicting the % U251 viable cells, (Beta = −0.84*** and Beta = −0.34*** resp.). The R 2 indicates that 81.3% of the variation in % U251 viable cells could be explained by these two variables (dose and duration). (c) The MCF-7, HeLa, and HF-5 cell lines were seeded and treated with the indicated concentrations of CAERS and/or CFEZO for displayed time intervals. The experiments were repeated five times in triplicate, and cell viabilities at each dose of extract(s) were expressed in terms of percent of control and reported as the mean ± SD. (d) U251 cells were seeded onto a 6-well plate at 1000 cells/well and treated with the indicated concentrations of CAERS and/or CFEZO as detailed in Section 2. The colonies were counted under a dissection microscope and the experiment was repeated three times. (e) CAERS and/or CFEZO acted synergistically to inhibit anchorage-independent growth in U251 cells in growth in soft agarose assays. U251 cells were plated, in triplicate, in 0.35% soft agarose and treated with CAERS (25 μg/mL) or CFEZO (25 μg/mL) and a combination of CAERS (5 μg/mL) and CFEZO (5 μg/mL). After 2 weeks, the colonies were stained with 0.0005% crystal violet and photographed using a digital camera coupled to a Carl Zeiss inverted microscope. Representative images of colonies in soft agar are shown.
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