This study describes the fabrication of nanostructured silver-doped zinc ferrite nanoparticles (AgxZn1-xFe2O4, where x = 0.0 and 0.05) using a chemical co-precipitation method. Field-emission scanning electron microscopy (FE-SEM) analysis was performed to investigate the surface characteristics and particle size of the pure and Ag-doped nanoparticles. X-ray diffraction (XRD) patterns confirmed the formation of a single-phase cubic structure for the zinc ferrite nanoparticles. The anti-cancer potential of the nanoparticles was evaluated using the MTT assay (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) against human breast cancer cells (MCF-7). The results showed that (AgxZn1-xFe2O4, where x = 0.05) nanoparticles with x = 0.05 exhibited significant cytotoxicity. At a concentration of 100 µg/mL, these nanoparticles reduced cell viability to 26.27%, with a calculated LC50 value (concentration lethal to 50% of cells) of 41.30 µg/mL after 24 hours of treatment. Notably, the LC50 value for (AgxZn1-xFe2O4, where x = 0.05) nanoparticles in normal L929 fibroblast cells was over four times higher compared to MCF-7 cancer cells, indicating a degree of selectivity for cancer cells. Further investigation revealed that (AgxZn1-xFe2O4, where x = 0.05) treatment significantly increased the generation of reactive oxygen species (ROS) within the cancer cells. This was confirmed by fluorescence intensity measurements, which showed a substantial increase from 1260.61 (control) to 15600 for cells treated with (AgxZn1-xFe2O4, where x = 0.05) nanoparticles. This included flow cytometry analysis of apoptosis (programmed cell death), migration assays to evaluate metastasis potential, measurement of antioxidant enzyme activity (SOD and catalase) to understand the impact on the cellular antioxidant system, indirect ELISA to detect caspase activity (a marker of apoptosis), cell cycle analysis, and finally, real-time PCR analysis to determine the mRNA expression of p53, a tumor suppressor gene. The apoptosis assay confirmed a significant increase in apoptotic cells upon treatment with (AgxZn1-xFe2O4, where x = 0.05) nanoparticles. This activity is likely mediated by the generation of ROS and subsequent oxidative stress within the cancer cells. These findings highlight the potential of these nanoparticles as a promising therapeutic strategy for cancer treatment.
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