QIC2 Inhibits CBS Expression And Induces Apoptosis In Hepato Cell Ular Carcinoma Cells

Objective: To explore the role of cystathionine-beta-synthase?CBS? in hepatoma cells and the mechanism of QIC2 induced apoptosis in hepatoma cells.Methods: RT-PCR and Western blot were used to detect the mRNA and protein levels of CBS in hepatoma cells and normal liver cells; MTT and EdU assay were peformed to determine the effects of downregulation of CBS/H₂ S on proliferation of hepatoma cells; Flow cytometry analysis was used to determine the effect of CBS downregulation on apoptosis of hepatoma cells;Western blot was performed to test the effect of CBS downregulation on the expression of apoptosis related proteins and HO-1 protein; The screening the CBS inhibitor with antitumor activity from compound QICS?QIC0, QIC1, QIC2 and QIC3? was performed by Western blot and MTT assay; MTT method was used to detect the anticancer activity of QIC2 in human hepatoma SMMC-7721 cells, adriamycin and sunitinib as positive controls. EdU assay was used to test the effect of QIC2 on the proliferation of SMMC-7721 cells; flow cytometry was used to analyze cell cycle and apoptosis of hepatoma SMMC-7721 cells; Western blot was performed to detect the effect of QIC2 on CBS, HO-1 and apoptosis related protein expression in SMMC-7721 cells. DCF method was used to detect the intracellular ROS level in SMMC-7721 cells treated with QIC2; DTNB method was used to detect the level of GSH in SMMC-7721 cells treated with QIC2.Results:1. RT-PCR and Western blot results showed that mRNA and protein of CBS in hepatocellular carcinoma SMMC-7721 and HepG2 cells were overexpressed compared withhuman normal liver cells.2. Downregulation of CBS expression or inhibition of CBS activity could inhibit the growth and proliferation of hepatocellular carcinoma SMMC-7721 cells.3. Downregulation of CBS expression or inhibition of CBS activity could induce apoptosis of hepatoma SMMC-7721 cells, increase the ratio of apoptosis related protein Bax/Bcl-2 and activate caspase-3 cascades.4. Downregulation of CBS expression inhibited the expression of HO-1 in SMMC-7721 cells and increased the level of ROS in SMMC-7721 cells.5. QIC2 significantly inhibited the expression of CBS and the growth of SMMC-7721 cells.6. Anticancer activity of QIC2 in SMMC-7721 cells was stronger than the positive control sunitinb, but weaker than the positive control adriamycin.7. QIC2 could inhibit the proliferation of SMMC-7721 cells.8. QIC2 could induce apoptosis of SMMC-7721 cells and activate the caspase-3 cascades.9. QIC2 could inhibit the expression of HO-1 in SMMC-7721 cells, increase the level of intracellular ROS, reduce the level of intracellular GSH, and consequenlty increased the oxidative stress.Conclusion:1. CBS could regulate the growth, proliferation and apoptosis of hepatocellular carcinoma cells.2. The ability of CBS inducing apoptosis in hepatocellular carcinoma cells may be related to oxidative stress.3. QIC2 could inhibit the growth and proliferation of hepatoma SMMC-7721 cells bydownregulating the expression of CBS.4, QIC2 might induce apoptosis of hepatoma SMMC-7721 cells by increasing the oxidative stress.