Zinc oxide nanoparticles induce necroptosis and inhibit autophagy in MCF-7 human breast cancer cells

Several studies have shown the toxic effect of zinc oxide nanoparticles (NZnO) on various cancer cells. This study aimed to assess the cellular death mechanism of the NZnO in MCF-7 human breast cancer cells. The cells were pretreated with zVAD (apoptosis inhibitor), 3-MA (an autophagy inhibitor), and Nec (a necroptosis inhibitor) for two hours, followed by their exposure to NZnO for 48 h. In the presence of Nec and 3-MA, NZnO significantly reduced proliferation and viability of MCF-7 cells while the apoptotic index and Bax/Bcl-2 ratio was significantly increased, compared to the only NZnO-treated cells. In the only NZnO-treated cells expression of Caspases-3 and Caspase-8 were up-regulated. NZnO with Nec and 3-MA could significantly increase expression of Caspase-3 and Caspase-9, but not Caspase-8 in the MCF-7 cells. NZnO in presence of zVAD and 3-MA significantly increased expression of RIPK1, RIPK3, and MLKL genes, compared to the only NZnO-treated cells. Furthermore, viability and proliferation of MCF-7 cells in NZnO with the apoptosis and autophagy inhbitors were lower than the other experimental groups. NZnO with apoptosis and necroptosis inhibitors could significantly increase cell viability, cell proliferation, and relative expression of autophagy-related genes such as LC3-II, Beclin-1, and ATG5, compared to the only NZnO-treated. According to the results of this study, the high toxicity of NZnO toward breast cancer cells mainly depends on activation of necroptosis and suppression of autophagy processes.