Study On Toxicity Of Hepatic Cells Induced By Gold Nanoparticles

Gold nanoparticles (GNPs) have been broadly used in multiple biological and smedical fields, such as tumor therapy, disease diagnosis and detection, biological sensor; and so on. The direct impact of this material to health and environment is of great concern to the population with the increasing of human exposure.In this study, the monodisperse GNPs coated with lysine (d=18 nm, c=5000.0μg/mL) were made in the traditional Frens's synthesis. Human hepatic cell line HL-7702 was used as model and exposed to different concentration (312.5, 625.0, 1250.0, 2500.0 and 5000.0μg/mL) of GNPs for 24 h. The cytotoxicity of GNPs nanoparticles was observed by MTT assay; the lipid peroxidation effects were examined by biochemical; the changes of mitochondria membrane potential (MMP) and intracellular reactive oxygen species (ROS) were observed by confocal laser microscope; the change s of cell cycle and cell apoptosis was detected by flow cytometry (FCM). Results showed as follow:1. The cytotoxicity of HL-7702 induced by GNPsMTT assay was performed to detect the cytotoxicity after exposure to GNPs (312.5, 625.0, 1250.0, 2500.0 and 5000.0μg/mL) for 24 h, there was significant differences between exposure and negative control group (P<0.001). The results showed that the survival rate decreased with the increase dosage of GNPs and suggested that GNPs had the inhibitory effect on HL-7702 cell proliferation.2. ROS changes of HL-7702 induced by GNPsThe effects of GNPs on the generation of intracellular ROS were determined by laser confocal microscope after exposure to GNPs (312.5, 625.0, 1250.0, 2500.0 and 5000.0μg/mL) for 24 h. The results showed that ROS decreased with the increase dosage of GNPs, and there was significant differences between exposure and negative control group (P<0.001).3. Changes of mitochondrial membrane potential of HL-7702 induced by GNPsRhodamine-123 staining and FCM were used to detect the changes of mitochondrial membrane potential. After exposure to GNPs (312.5, 625.0, 1250.0, 2500.0 and 5000.0μg/mL) for 24 h, mitochondrial membrane potential decreased with the increase of GNPs, and there were significant differences between exposure groups and negative control groups (P<0.001).4. Lipid peroxidation of HL-7702 induced by GNPsAfter treatment with GNPs (312.5, 625.0, 1250.0, 2500.0 and 5000.0μg/mL) for 24 h, SOD,GSH-Px activity decreased with the increase dosage of GNPs, MDA content increased with the increase dosage of GNPs, there were significant differences between exposure groups and negative control group respectively (P<0.001). The results suggested that GNPs could induce lipid peroxidation on HL-7702 cells.5. Apoptosis of HL-7702 induced by GNPsHL-7702 cells were stained with Annexin V/PI and subsequently analyzed by flow cytometry to confirm the apoptosis. After exposure to GNPs (312.5, 625.0, 1250.0, 2500.0 and 5000.0μg/mL) for 24 h, a dose-dependent increasing in apoptotic rate of exposure groups was found when compared to negative control groups (P<0.001)6.Cell cycle changes of HL-7702 induced by GNPsHL-7702 cells were also stained with PI to analyze cell cycle by flow cytometry. After 24 h exposure, the percentage of cells in G0/G1 phage increased while S phage decreased with the increasing of GNPs (P<0.001). All suggested that GNPs could induce G0/G1 phase arrest, decreased cells population in S phase, accordingly interfering the process of DNA synthesis.In conclusion, GNPs could induce the cell toxic effects on HL-7702 cells, including decreasing the activity of anti-oxidative enzyme (GSH-Px, SOD), increasing the concent of MDA, inducing lipid peroxidation; increasing ROS; decreasing mitochondrial membrane potential, impacting the progress of cell cycle; inducing cell apoptosis. The research had important theoretical meaning in studying the toxic effects of GNPs on liver and related mechanism, which would provide experimental basis for safety evaluation of GNPs.