Effects Of Croton Alkaloid On The Proliferation And Apoptosis Of Human Gastric Cancer Cells SGC-7901

Objective:To screen digestive system tumor cells that are sensitive to Croton Alkaloid (CA), and to explore their effects on the proliferation and apoptosis of these sensitive tumor cells.Methods:Apply croton alkaloid with different concentrations to human gastric cancer cells SGC-7901, human pancreatic cancer cells PANC-1, human colon cancer cells SW480. Use CCK-8to test the effects of croton alkaloid on the proliferation of different cells after24hours,48hours and72hours respectively. Apply hoechst staining and examine the changes in cell shapes under fluorescence microscope. Use flow cytometer to check the cell apoptosis rate. Examine the status of proliferation and apoptosis related proteins-PCNA, Ki-67, Bax and Fas by immunohistochemistry.Results:1. The inhibition of croton alkaloid on the proliferation of PANC-1cells and SW480cells were irregular and unstable. Its inhibition on the proliferation of SGC-7901cells were significant. The inhibition rate was associated with CA concentration and duration. With the increase in CA concentration, the cell proliferation decreased from97.6%,94.6%,74.9%to86.3%,40.4%,18.1%(p<0.05) after24hours,48hours and72hours respectively, compared with the control group. After applying croton alkaloid to SGC-7901cells for72hours, the IC50was about94.86ug/ml. Take the IC50of100ug/ml, apply to SGC-7901cells, their cell viability decreased from82.08%to10.34%, compared with the control group after24hours,36hours,48hours,60hours and72hours. After24hours and36hours, the p<0.05. After48hours,60hours and72hours, the p<0.01.2. After hoechst staining,the cells showed apoptoticmorphological changes under fluorescence microscope. Some cells became smaller, and cell membrane shrank, showingirregular shape. The nucleus morphology changed significantly. Dense blue fluorescent particles can be observed in the nucleus. Changes like pyknosis and karyorrhexis were discovered, accompanied by the formation of apoptotic bodies. As time passed, the cell apoptosis rate of CA group increased from5.56%to45%, the rate of fluorouracil group increased from5.88%to50%(P<0.05), compared with the control group.3. The early and late stage cell apoptosis were assessed by flow cytometry using AV/PI double-stained method. With the increase in croton alkaloid concentration, the cell apoptosis rate rose from13.46%to40.83%, compared with the control group. The early stage apoptosis rate rose from9.82%to34.92%, and the late stage apoptosis rate rose from3.46%to4.55%(P<0.05).4. The immunohistochemistry showed that with the increase in croton alkaloid concentration, the positive rate of the proliferation related nuclear antigen PCNA and Ki-67decreased from12.3%to1.7%and from23.8%to4.9%respectively (p<0.05). With the increase in croton alkaloid concentration, the positive rate of the apoptosis related proteins Bax and Fasincreased from3.9%to56.7%and from13.3%to65.3%respectively (p<0..05).Conclusion:1. Croton alkaloidhas no significant inhibition on human pancreatic cancer cells PANC-1and human colon cancer cells SW480.2. Croton alkaloid could significantly inhibit the proliferation of human gastric cancer cells (SGC-7901). The effects are associated with concentration and duration. It can be explained by the decrease in PCNA and Ki-67protein expression.3. Croton alkaloid could significantly induce the apoptosis of human gastric cancer cells (SGC-7901). The effects are associated with concentration and duration. It can be explained with the increase in Bax and Fas protein expression.