Incorporation of ifosfamide into various essential oils -based nanoemulsions ameliorates its apoptotic effect in the cancers cells

Abstract

The chemotherapeutic drugs, loaded in nanocarriers, have recently attracted the pharmaceutical industries due to their limited adverse side effects. The objective of the current study was to incorporate the ifosfamide (IFO) into two different essential oils-based nanoemulsions, lemon (LEM-IFO) and salvia (SAL-IFO). The antiproliferation activities of the resulted formulas were evaluated in the MCF-7 breast cancer cells and HeLa cervical cancers cells. The cytotoxic effect of the NE formulas was detected by the MTT assay, DAPI stain and light microscopy. The z-average diameters range of LEM-IFO and SAL-IFO, determined by the zetasizer, were 49.15-61.81 nm and 56.64-64.62 nm, respectively. The half maximal inhibitory concentration (IC₅₀) of LEM-IFO and SAL-IFO, applied into the HeLa cells, were 0.165 ± 0.025 and 0.141 ± 0.035 mM, respectively, whereas the IC₅₀ of LEM-IFO and SAL-IFO subjected into the MCF-7 cells were 0.200 ± 0.005 mM and 0.270 ± 0.025 mM, respectively. The IC₅₀ of the free IFO was markedly larger than LEM-IFO and SAL-IFO when applied into MCF-7 cells (9.20 ± 2.01 mM) and HeLa cells (7.69 ± 1.88 mM). Among the tested formulas, LEM-IFO and SAL-IFO have the greatest apoptotic effect on the MCF-7 and HeLa cells, respectively. Solubilizing the IFO in the essential oils-based NE has ameliorated the antitumor efficacy of IFO.

Figure 1

Pseudo ternary phase diagrams constructed at different weight fractions of lemon oil () or salvia oil (), water, and a surfactant mixture blended at a fixed ratio of 2:1 of Tween 80 to Span 20, respectively.

Figure 2

The percentages of cell viability of HeLa and MCF-7 cells treated for 24 h at different concentration of the tested formulas. Error bars represent the standard deviation for n = 3. The levels of the differences between the formulas at each concentration are ranked as very highly (***P < 0.001), highly (**0.001 ≤ P < 0.01) and (*0.01 ≤ P < 0.05) significant.

Figure 3

Light Microscopy images of HeLa cells subjected into different formulas for 24 h at their IC₅₀s (n = 3). Images were magnified at 400x. The arrows colored green, orange and red displayed the membrane blebbing with collapse nucleus, the enlarged cells and the digestive vesicles, respectively.

Figure 4

Light Microscopy images of MCF-7 cells subjected into different formulas for 24 h at their IC₅₀s (n = 3). Images were magnified at 400x. The arrows colored orange, red and green represented condensed chromatin, extracellular apoptotic bodies and ghost cells, respectively.

Figure 5

The nuclear morphology changes of HeLa cells stained with DAPI and treated with different concentrations of the drug formulas (n = 3) for 24 h.

Figure 6

The nuclear morphology changes of MCF-7 cells stained with DAPI and treated with different concentrations of the drug formulas (n = 3) for 24 h.

Figure 7

The cell counts at different concentrations of the tested formulas measured from the fluorescent microscopic images stained with DAPI (Figs 5 and 6). Error bars represent the standard deviation for n = 3. The levels of the differences between the formulas at each concentration are ranked as very highly (***P < 0.001), highly (**0.001 ≤ P < 0.01)) and (*0.01 ≤ P < 0.05) significant.