## 3 Results
### Anticancer Effects of CsE and NmE  
#### 3.3.1 CsE and NmE  
Both extracts inhibited the growth of all investigated cancer cells in different dose-dependent manners with remarkable disease in cell viability (Figures 4 and 5) associated with cytotoxicity increase. The IC\({}_{50}\) values between _Nm_E and _Cs_E are greatly different indicating the higher efficacy of _Nm_E in the inhibition of HepG2, MCF-7, and Caco-2 cancer cells proliferation. The IC\({}_{50}\) values for _Nm_E and _Cs_E on HepG2 cells were 10.33 and 271.48 \(\upmu\)g/mL, respectively, while on MCF-7, they were 12.48 and 467.19 \(\upmu\)g/mL, respectively, whereas on Caco-2, cells were 10.52 and 1570.82 \(\upmu\)g/mL, respectively (Table 2). Due to the _Nm_E potentiality as an antiproliferative active agent, it was chosen for further investigations. Figure 4: Effects of _Nm_E on HepG2, MCF-7, and Caco-2 cell viability after 48 h. Cells were incubated with the indicated concentration of _Nm_E while the percentage of cell viabilities were determined by MTT. Values used for plotting are means of triplicates, with each concentration tested in 7–8 wells. Statistical differences from control cultures are shown as bar graphs with error bars representing the means \(\pm\) SD; **_p_ < 0.01, ***_p_ < 0.001 and ***_p_ <0.0001 vs. control (DMSO-treated) cells. Figure 5: Effects of _Cs_E on HepG2, MCF-7, and Caco-2 cell viability after 48 h. Cells were incubated with the indicated concentration of _Cs_E while the percentage of cell viabilities were determined by MTT. Values used for plotting are means of triplicates, with each concentration tested in 7–8 wells. Statistical differences from control cultures are shown as bar graphs with error bars representing the means \(\pm\) SD; **_p_ < 0.01, ***_p_ < 0.001 and ***_p_ <0.0001 vs. control (DMSO-treated) cells. 3.2 NmE Induced Apoptosis and Increased Bax, Cleavage PARP, and Caspase-3 Expressions in Cancer Cells  
In HepG2, _NmE_ in a dose-dependent manner induced significant early and late apoptosis causing complete cell death (Figures 6 and 7), indicated by the elevated percentage of PI stain that binds to the genetic material (DNA and RNA) inside the damaged dead cells. In addition, the _NmE_ apoptotic effect is confirmed through the expression of some regulatory proteins that are involved in either the extrinsic or intrinsic apoptotic pathways. _NmE_ increased the level of the pro-apoptotic protein Bax, cleavage PARP, and caspase-3, while it decreased the level of the anti-apoptotic protein BCI2 in HepG2, MCF  
3.3 NmE Induced G0/G1-Phase Cell Cycle Arrest and Decreased Cyclins D1 and E1 and CDK6 Expressions in Cancer Cells  
The cell cycle profile analysis revealed significant dose-dependent increases in the number of cells in the G0/G1 phase of the cell cycle in _Nm_E-exposed HepG2 cells compared to no-treatment HepG2 cells control (Figures 9 and 10). The G0/G1 phase distribution of HepG2 cells after treatment with _Nm_E (0, 6.25, 12.5, and 25 \(\upmu\)g/mL) was 45.8%, 49.83%, 59.67%, and 67.56%, respectively (Figure 10). The increases in the G0/G1 phase cell population were accompanied by decreases in the G2/M and S phase cell populations of  
Figure 8: Effects of _Nm_E on protein expression of BCI2, Bax, cleaved PARP, and cleaved Caspase-3 in HepG2, MCF-7, and Caco-2 cells, which were treated with each _Nm_E concentration and harvested 48 h after treatments. The immunoblots shown are representative of three independent experiments, which all gave similar results, cropped and unedited images are provided here. Bars represent the means \(\pm\) SD. * \(p\) < 0.05, ** \(p\) < 0.01, *** \(p\) < 0.001 and *** \(p\) <0.0001 vs. control (DMSO-treated) cells. Figure 7: Cell death percentage induced by _Nm_E in HepG2 cells: (**a**) Ctrl. HepG2, (**b**) _Nm_E (6.25 \(\upmu\)g/mL)/HepG2, (**c**) _Nm_E (12.5 \(\upmu\)g/mL)/HepG2, and (**d**) _Nm_E (25 \(\upmu\)g/mL)/HepG2.