Original Articles: 2015 Vol: 7 Issue: 6
Evaluation of cytotoxic effect of metallic nanoparticles in an in vitro liver cancer model
We aim to evaluate cytotoxic effect of silver and gold metallic nanoparticles (AgNPs & AuNPs) on human cucasian hepatocellular carcinoma cell line model (HepG2) and their possible anti-proliferative activity. This new class of engineered nanoparticles with desired physicochemical properties can be applied as new therapeutic approaches against human liver cancer disease. HepG2 was used as a model of human liver cancer cells. Metallic nanoparticles were characterized using UV-visible spectra and transmission electron microscopy (TEM). Cytotoxic effects of metallic nanoparticles on HepG2 cells were followed by colorimetric neutral red and SRB cell viability assays. Further investigation of cytotoxic effect of our nanomaterials were further investigated on a cellular and molecular level using cell cycle analysis , DNA and some apoptotic genes expression on a level of mRNA for P53, Bak, Bax, BCl2 and β actin was served as housekeeping gene. Treatment of HepG2 with different concentrations of 22 nm diameter of AgNPs did not show alteration of cell morphology after 24 h of cell exposure. Also, when cells were treated with high concentration of AgNPs (viability was 78% after cell treatment with 10 μM and decreased to 46% after treatment of cells with 1000 μM). Cellular evaluation of AgNPs revealed progressive accumulation in the S phase of the cell cycle correlating with decreased number of cells in the G2/M phase followed by cellular DNA fragmentation. Extensive evaluation of cytotoxic effect of AgNPs showed mRNA apoptotic genes expression (P53, Bak, Bax, BCl2 ) without expression of mRNA of caspase 3 gene which was expressed in untreated cells, suggesting involvement of intrinsic apoptotic caspase independent pathway. Treatment of HepG-2 with different concentrations of 34 nm diameter of AuNPs did not show alteration of cell morphology after 24 h of cell exposure. Such metallic nanoparticles did not reveal toxic effect at concentration up to 50 μM after 48 h of cell exposure. Cellular evaluation of AuNPs revealed progressive accumulation at G0/G1and at G2/M phases of cell cycle. Also the same results were obtained by treating cells with AgNPs, where the expression of mRNA of P53, Bak, Bax, BCl2 without expression of mRNA of caspase 3 gene was observed in treated cells, suggesting intrinsic apoptotic caspase independent mechanism but may be induced by different molecules than that exerted by AgNPs. Our engineered silver nanoparticles at size of 22nm revealed genotoxic effect on human liver carcinoma cell line HepG-2 through intrinsic apoptotic caspase independent mechanisms. Further quantitative analysis and investigation of impact of time on genotoxic effect are required before reaching a final conclusion and starting in vivo assays.