| Background Hepatocellular carcinoma(HCC)is one of the most common malignant tumors in China,ranking on the fifth incidence rate and on the second death rate among tumor-related deaths.Although there are many ways to treat liver cancer,including chemotherapy,radiotherapy,surgery and molecular targeted therapy,the overall survival benefit is limited.Currently,chemotherapy is still the main choice of patients with advanced liver cancer.However,the cytotoxicity and drug resistance are the main problems that limit chemotherapy in the treatment of liver cancer.Therefore,exploring new and effective therapeutic drugs and finding new clinical treatment strategies are imperative for the treatment of liver cancer.In recent years,due to the mild side effects and obvious treatment effect of Huaier aqueous extract,Huaier aqueous extract has received extensive attention in recent years,especially in the aspect of anti-tumor.Currently Huaier aqueous extract is widely used in a variety of clinical cancer treatment,including liver cancer,breast cancer,colon cancer,stomach cancer.Huaier aqueous extract has achieved certain effects whether it is applied alone or in combination with chemotherapy or radiotherapy.However,the specific anti-tumor mechanism of the Huaier aqueous extract is not clear.Under certain conditions,such as inflammation,oxidative stress,endoplasmic reticulum stress is conducive to malignant transformation of cells,promote tumor cell proliferation and reduce apoptosis.Endoplasmic reticulum stress can also promote tumor cell survival,proliferation and promote tumor recurrence by regulating apoptosis,autophagy and induction of drug resistance in tumor cells.However,in the anti-tumor effect of Huaier aqueous extract,the role of endoplasmic reticulum stress has not been reported until now.This study intends to explore the antitumor mechanism of Huaier aqueous extract and to explain the role of endoplasmic reticulum stress in this process.Objective: 1.To observe the effect of Huaier aqueous extract on human hepatocellular carcinoma cell proliferation and apoptosis 2.To observe the effect of endoplasmic reticulum stress on the proliferation and apoptosis of human hepatocellular cells and to explore its mechanism.Methods: 1.Establish different concentration Huaier aqueous extract.HCC cells were co-cultivated with Huaier aqueous extract for 24 hours.The cell inhibition rate was detected by MTT method.2.Hepatoma cells were co-cultured with different concentrations of Huaier aqueous extract for 24 hours,and then flow cytometry was used to detect the apoptosis rate of hepatoma cells.Western blot was used to detect the protein expression of Bax,Bcl-2 GRP78,IRE1 and PERK proteins in hepatoma cells treated with different concentration of Huaier aqueous extract.4.Pretreatment of hepatocellular carcinoma cells with endoplasmic reticulum stress inhibitor 4-phenylbutyric acid,and then hepatocellular carcinoma cells were stimulated with Huaier aqueous extract.The cell viability of each group was detected by MTT assay.5.Flow cytometry and Western blot were used to detect the incidence of apoptosis and the protein expression of Bax and Bcl-2 in hepatocellular carcinoma cells in the control group,the Huaier aqueous extract group,the 4-phenylbutyric acid group,the 4-phenylbutyrate+Huaier aqueous extract group.Result: 1.In vitro experiments showed that Huaier aqueous extract can effectively inhibit the proliferation of hepatocellular carcinoma cells,and the inhibitory efficiency is also increased with the increase of the concentration of Huaier aqueous extract.The IC50 of on Hep3 B cells was 4.2mg/m L,and the IC50 of Hep G-2 was 5.5mg/m L.2.Huaier aqueous extract can effectively induce the apoptosis of hepatocellular carcinoma cells,and with the increase of the concentration of Huaier aqueous extract,the incidence of apoptosis increased.Western blot showed that with the increase of the concentration of Huaier aqueous extract,the levels of Bcl-2 protein in Hep3 B and Hep G-2 cells decreased significantly while the level of Bax protein increased.3.Huaier aqueous extract can induce endoplasmic reticulum stress in hepatocellular carcinoma cells,and with the increase of the concentration of Huaier aqueous extract,the expression of GRP78,IRE1 and PERK protein in hepatoma cells Hep3 B and Hep G-2 increased.4.The results of cell activity test showed that compared with the control group,no significant changes were observed in the hepatoma cells of 4-phenylbutyrate(500μM)group.In the group of 4-phenylbutyrate(500μM)+ Huaier aqueous extract(6mg/m L),the activity of hepatoma cells was significantly decreased compared with the group of the Huaier aqueous extract(6mg/m L).5.Compared with the control group,cell apoptosis results did not change significantly in 4-phenylbutyrate(500μM)group.Compared with the Huaier aqueous extract(6mg/m L)group,the apoptosis rate of hepatocellular cells in 4-phenylbutyric acid(500μM)+ Huaier aqueous extract(6mg/m L)group was significantly increased.Western blot showed that compared with Huaier aqueous extract(6mg/m L),the expression of Bax protein was significantly increased and Bcl-2 protein expression levels decreased in Huaier aqueous extract(6mg/m L)+ 4-phenylbutyrate(500μM)groupConclusions: 1 Huaier aqueous extract can inhibit the proliferation of hepatoma cells and induce apoptosis of hepatocellular carcinoma.2 Huaier aqueous extract may induce endoplasmic reticulum stress in hepatocellular carcinoma cells.3 Inhibition of ER stress enhanced apoptosis induced by Huaier aqueous extract in HCC cells. |
