Evaluation of cytotoxicity of hydroxyapatite nanoparticles on human hepatocellular carcinoma cell line BEL-7402

Objective To evaluate the cytotoxicity of hydroxyapatite (HAP) nanoparticles on human hepatocellular carcinoma cell line BEL 7402 by means of growth inhibition and apoptosis induction in vitro. Methods The hepatocellular carcinoma cell line BEL 7402 was treated with HAP nanoparticles at various concentrations for different duration. Growth suppression was evaluated by MTT method; apoptosis related alterations in morphology were ascertained under cytochemical staining (Hoechst 33258) and transmission electron microscopy (TEM). The apoptotic index was counted under fluorescence microscopy. Flow cytometry (FCM) was used to investigate the apoptotic rate. Results HAP nanoparticles inhibited the growth of hepatoma cells in a dose and time dependent manner, with IC 50 values of 29.30μg/ml after 48h treatment. Under fluorescent staining and transmission electron microscopy, the cells treated with HAP nanoparticles exhibited characteristics of apoptosis including cell membrane shrinkage, cell membrane blebbing, condensation and fragmentation of nuclear chromatin and formation of apoptotic bodies. Flow cytometry analysis showed hypodiploid peaks on histogram, the apoptotic rates at concentrations of 50μg/ml, 75μg/ml, 100μg/ml, 150μg/ml and 200μg/ml were (20.35±2.23)%,(25.35±1.92)%,(29.34±4.61)%,(44.92± 3.78 )% and (53.64±3.49)%, respectively all of which were significantly higher than that of control group (2.23± 0.14)%, but same as (2.73±0.17)%,(3.52±0.35)%,(6.37±0.34)% and (21.64±2.46)% at 12h, 24h, 36h and 48h. Conclusion HAP nanoparticles not only inhibit the proliferation but also induce apoptosis of human hepatocellular carcinoma cell line BEL 7402 with demonstrated strong cytotoxicity.