Abstract:

Objective

To investigate the biological toxic effect and mechanisms of gold nanorods （AuNRs） on A549 cells.

Methods

Normal cultured A549 cells were treated with different concentrations of AuNRs for 6， 12 or 24 hours for subsequent detection. CCK-8 test and LDH test were used to observe cell viability and cell membrane damage. The distribution and morphology of AuNRs in cells were observed by transmission electron microscope. Western blot was used to detect the expression of autophagy related proteins ATG16， ATG4， Beclin1， LC3-Ⅱ and P62. ROS was detected by laser scanning confocal microscope. The levels of GSH/GSSG and T-AOC were detected by kits. One way ANOVA was performed after the homogeneity test of data variance and Student- Newman-Keuls test was used for pairwise comparison between different groups.

Results

CCK-8 results showed that compared with the 0 μg/mL， the cell viability of the 4 μg/ mL AuNRs [（80.05 ± 3.45）% vs （100 ± 2.47）%］ was decreased at 6 h after treated with AuNRs （P ＜0.01）. The cell viability of 2 μg/mL and 4 μg/mL AuNRs [（71.36 ± 3.87）% and （39.47 ± 4.16）%vs （100 ± 3.12）%］ were decreased （P ＜ 0.01） at 12 h after treated with AuNRs； The cell viability of 1 μg/mL， 2 μg/mL and 4 μg/mL AuNRs [（91.83 ± 1.98）%， （56.53 ± 3.57）% and （28.65 ± 3.09）%vs （100 ± 2.34）%］ were decreased （P ＜ 0.01） at 24 h after treated with AuNRs. LDH test showed that the leakage of LDH was increased significantly （P ＜ 0.01）. Transmission electron microscopy showed that AuNRs could enter A549 cells and exist in the cytoplasm and some membrane vesicles in the form of single particles or aggregates 6 h after treatment with 4 μg/mL AuNRs. Western blot showed that after treated with different concentrations of AuNRs （0， 0.5， 1， 2， 4 μg/mL）for 6 h， the expression levels of ATG16， ATG4， Beclin1 and LC3- Ⅱ proteins were significantly increased， while the expression level of autophagy substrate protein P62 was significantly decreased in A549 cells（P ＜ 0.01）. After treated with 100 μmol/L MnTBAP or 10 mmol/L NAC， the increased expression of ROS and LC3 induced by AuNRs could be reversed， and the content of T-AOC [（92.37 ± 8.49）%vs （68.46 ± 6.38）%， （110.49 ± 7.39）% vs （51.26 ± 7.39）%］ and GSH/GSSG [（92.56 ± 5.52）%vs （62.48 ± 5.52）% ， （104.49 ± 4.84）% vs （57.39 ± 4.84）%］ were increased （all P ＜ 0.05）.

Conclusion

AuNRs can enter A549 cells and induce cytotoxicity effect， which in a dose-dependent and time-dependent manner. The mechanism may be related to the reduction of antioxidative stress ability of A549 cells caused by AuNRs.

Key words: Gold nanorods, Cytotoxicity, Autophagy, Oxidative stress, ROS