Inhibitory Effects And Mechanism Of Nodosin On Growth Of HepG₂Cells

ObjectiveIsodon serra (Maxim.) Hara is a kind of folk herb used for inflammation, antibiosis, anti-tumor and anti-virus etc. Nodosin is one of the extracts from Linearstripe isodon herb. In our experiments, the inhibitory effects and mechanisms of Nodosin on growth of HepG2cells were studied.MethodsHepG2cells were cultured with DMEM to exponential phase of growth. Then the cells were seeded to the wells of six-well plates. After adherence, HepG2cells were treated with different concentrations of Nodosin:1.25μmol/L,2.5μmol/L,5μmol/L,10μmol/L and20μmol/L. The cells without being treated with Nodosin were also set as control. The HepG2cells were also treated with5μmol/L Nodosin for6h,12h,18h,24h, and48h with the same culture methods. The morphology variation of HepG2cells were observed with inverted microscope, Hoechest33258stain, electron microscope. MTT method was used to detect the inhibitory rates of HepG2cells. Flow cytometry was used to detect the apoptotic rates and the expression of Bcl2and Bax proteins. Western blot method was used to evaluate the expression of caspase3protein.Results1.The number of dead HepG2cells increased with Nodosin concentration increasing.2. The number of dead HepG2cells increased with time extension.3. In inverted microscope, drifting cells increased, HepG2cells changed from fusiform shape to round shape and the diastema augmented, especially in5μmol/L and10μmol/L Nodosin groups. In fluorescence microscope and electron microscope, cell shrinkage and nucleus drift were obvious in morphology when Nodosin concentration increased. There were many apoptotic bodies in5μmol/L and10μmol/L Nodosin groups.4. The apoptotic rates increased with Nodosin dose increasing detected with flow cytometer. The number of positive cells of Bax increased with the concentration of Nodosin step-up, while the number of positive cells of Bcl2decreased with the concentration of Nodosin step-up.5. As shown by western blot method, the expression of caspase3protein increased with the dose increasing especially in10μmol/L group. They were in dose-effect relationship.Conclusions1.Nodosin can inhibit the growth of HepG2cells. The effects were in dose-effect and time-effect relationship.2.The inhibition of the growth of HepG2cells was by inducing cell apoptosis due to the decreased Bcl2and increased Bax and caspase3.