Metallic Nanoparticles as an Anti-proliferative Activity Against Human Hepatocellular Carcinoma (In-Vitro study)

Abstract

We aim to evaluate cytotoxic effect of gold and silver metallic nanoparticles (AuNPs & AgNPs) 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 SRB and neutral red 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 fragmentation assay 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 HepG-2 with different concentrations of 35 nm diameter of AuNPs did not show alteration of cell morphology after 24 h of cell exposure. Such metallic nanoparticles did not reveal vigorous toxic effect at concentration up to 100 μM after 48 h of cell exposure. Cellular evaluation of AuNPs revealed progressive accumulation at G0/G1 and at G2/M phases of cell cycle. The expression of mRNA of P53, Bak, Bax, BCl2 without expression of mRNA of caspase 3 gene was observed in treated cells with AuNPs , suggesting involvement of intrinsic apoptotic caspase independent pathway. 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, cytotoxicity results revealed that; viability was 58% after cell treatment with 10 μM and decreased to 40.78% after treatment of cells with 1000 μM for 48 h. 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, same as the results were obtained by treating cells with AuNPs, suggesting intrinsic apoptotic casepase independent mechanism but may be induced by different molecules than that exerted by AuNPs. Our engineered gold nanoparticles (35 nm) and silver nanoparticles at size of 22nm showed genotoxic effect on human liver carcinoma cell line HepG-2 through intrinsic apoptotic caspase independent mechanisms. Further quantitative analysis and investigation for the impact of time on genotoxic effect are required before reaching a final conclusion and starting in vivo assays. Key words: Metallic nanoparticles, anti-proliferative activity, HepG2, apoptotic genes expression