Size-dependent cytotoxicity of Fe₃O₄ nanoparticles induced by biphasic regulation of oxidative stress in different human hepatoma cells

Figures & data

Figure 1 Characterization of Fe₃O₄ NPs.

Notes: TEM images of Fe₃O₄ NPs with the diameter of 6 nm (A), 9 nm (B), and 14 nm (C). (D) The size distribution images of Fe₃O₄ NPs; the inserted image is the image of Fe₃O₄ NPs aqueous solution. (E) The zeta potential image of Fe₃O₄ NPs.

Abbreviations: NPs, nanoparticles; TEM, transmission electron microscope.

Figure 2 Cellular uptake of Fe₃O₄ NPs with different diameter sizes in human hepatoma SK-Hep-1 cells (A and B) and Hep3B cells (C and D) after exposure for 24 hours. Fe₃O₄ NPs-treated cells were determined by flow cytometry.

Notes: Cells incubated in medium only were used as control. Results correspond to mean ± SE of three independent experiments with triplicates in each case. ***P<0.001 compared to the control without Fe₃O₄ NPs treatment; ^##P<0.01, ^###P<0.001 compared to the corresponding Fe₃O₄ NPs treatment at 200 µg/mL; ^&&P<0.01, ^&&&P<0.001 between the indicated groups.

Abbreviations: fcs, flow cytometers; NPs, nanoparticles; SE, standard error; SSC, side scatter.

Figure 3 Internalization and localization of Fe₃O₄ NPs with different diameters in human hepatoma SK-Hep-1 cells after exposure for 24 hours.

Notes: Cells treated with Fe₃O₄ NPs were determined by TEM. Fe₃O₄ NPs are enclosed in vesicles.

Abbreviations: NPs, nanoparticles; TEM, transmission electron microscope.

Figure 4 Effect of Fe₃O₄ NPs on mitochondrial function of human hepatoma SK-Hep-1 cells (A) and Hep3B cells (B).

Notes: Mitochondrial function of cells treated with various concentrations of Fe₃O₄ NPs with different diameters for 24 hours was measured by the CCK-8 assay. Data are presented as mean ± SE of three independent experiments with six replicates in each case. *P<0.05, **P<0.01, ***P<0.001 compared to the control without Fe₃O₄ NPs treatment.

Abbreviations: CCK-8, Cell Counting Kit-8; NPs, nanoparticles; SE, standard error.

Figure 5 Influence of Fe₃O₄ NPs on the induction of ROS in human hepatoma SK-Hep-1 cells (A and B) and Hep3B cells (C and D).

Notes: Fe₃O₄ NPs-treated cells were determined by flow cytometry. Cells incubated in medium only were used as control. Results correspond to mean ± SE of three independent experiments. *P<0.05, **P<0.01, ***P<0.001 compared to the control without Fe₃O₄ NPs treatment; ^##P<0.01, ^###P<0.001 compared to the corresponding Fe₃O₄ NPs treatment at 200 µg/mL; ^&&P<0.01, ^&&&P<0.001 between the indicated groups.

Abbreviations: fcs, flow cytometers; NPs, nanoparticles; ROS, reactive oxygen species; SE, standard error.

Figure 6 Induction of apoptosis and necrosis in Fe₃O₄ NPs-treated human hepatoma SK-Hep-1 cells (A) and Hep3B cells (B).

Notes: Cells exposed to Fe₃O₄ NPs with different diameters for 24 hours were determined by flow cytometry. Cells incubated in medium only were used as control. Results correspond to mean ± SE of three independent experiments.

Abbreviations: NPs, nanoparticles; SE, standard error; FITC, fluorescein isothiocyanate; PE, phycoerythrin; PI, propidium iodide.

Figure 7 Fe₃O₄ NPs regulate the activities of Akt and MAPK in human hepatoma cells.

Notes: The expressions of Akt, Erk, and p38 were measured using Western blot in Hep3B cells exposed to Fe₃O₄ NPs with different diameters for 24 hours. Cells incubated in medium only were used as control.

Abbreviation: NPs, nanoparticles.

Figure 8 Influence of ROS scavenger on Fe₃O₄-induced necrosis and plasma membrane damage.

Notes: (A and B) ROS scavenger SOD and CAT suppressed Fe₃O₄ NPs-induced necrosis in human hepatoma SK-Hep-1 cells (A) and Hep3B cells (B). Cells exposed to Fe₃O₄ NPs with different diameters for 24 hours were determined by flow cytometry. Cells incubated in medium only were used as control. (C–F) Effect of Fe₃O₄ NPs on cellular membrane damage of human hepatoma SK-Hep-1 cells (C and E) and Hep3B cells (D and F). Cellular membrane damage of cells treated with different concentrations of Fe₃O₄ NPs with different diameters for 24 hours was measured by the LDH leakage assay. In some experiments, cells were incubated with SOD and CAT. Data are presented as mean ± SE of three independent experiments with triplicates in each case. **P<0.01, ***P<0.001 compared to the control without Fe₃O₄ NPs treatment; ^##P<0.01, ^###P<0.001 compared to the corresponding Fe₃O₄ NPs treatment at 200 µg/mL; ^&P<0.05, ^&&&P<0.001 between the indicated groups.

Abbreviations: CAT, catalase; LDH, lactate dehydrogenase; NPs, nanoparticles; ROS, reactive oxygen species; SE, standard error; SOD, superoxide dismutase.