Studies Of Isomycin Derivant On Inducing Apoptsis Of Human Hepatoma HepG-2 Cell Line

Objective:To investigate the efffects of isomycin derivant on inhibiting proliferation and inducing apoptosis of human hepatoma HepG-2 cell line, and to provide the evidence of theory and experiment for the traditional Chinese medicine.Methods:MTT assay was used to detected the proliferation inhibiting rate and cytotoxic effect of HepG-2 cell line; Flow cytometry was used to measure cell cycle and apoptosis rate. Morphological survey by inverted microscope, hematoxylin-eosin staining, AO/EB double staining and fluorescence microscope show the change of human hepatoma HepG-2 cell line with isomycin derivant. Immunocytochemistry was used to demonstrate the express of Caspase-3,Caspase-8,Caspase-9,Survivin,Livin,NF-κB,Bcl-2,Bax,Fas.Results:(1) MTT assay showed that isomycin dervant by different concentrations could inhibite the growth of human hepatoma HepG-2 cell line in a doseage and time dependent manner.(2) Flow cytometry showed that isomycin dervant could induce the apoptosis of human hepatoma HepG-2 cell line and sub-dipioid peak was showed after treated with isomycin derivant. isomycin dervant could induce cell cycle disturbance with the increase of G0/G1 phase in dose-and time-dependent manners. The apoptosis rate was increased compared with the control group. (3) Inverted microscope view showed that compared with the control group, HepG-2 cell growed slowly, adherency loose; Hematoxylin-eosin staining showed nucleus of HepG-2 cell with isomycin dervant exhibited pyknosis, some of the cell showed apoptosis and necrosis; AO/EB double staining and fluorescence microscope showed the chromatin of HepG-2 cell in control group display green colour, but the early apoptosis cell display yellow green and the late apoptosis cell display orange colour. Immunocytochemistry showed that the expression of Caspase-8,Caspase-9 Caspase-3,Fas and Bax proteins increased, the expression of Survivin,Livin,NF-κB,Bcl-2 protein decreased after treated with isomycin derivant (P<0.05)Conclusions:(1) Isomycin dervant could inhibite the growth of human hepatoma HepG-2 cell line in a doseage and time dependent manner, and arrested in G0/G1 phase.(2) Isomycin dervant could induce the apoptosis of human hepatoma HepG-2 cell line.(3) Isomycin dervant induce the apoptosis of human hepatoma HepG-2 cell line may through the mitochondrial pathway and the death receptor-mediated pathway.