Dioscin Regulates Endoplasmic Reticulum Stress-induced Apoptosis Of Colon Cancer Cells And Its Mechanism

Objective:Dioscin(Dio)is a natural steroidal saponin compound extracted from Dioscorea plants.It has many pharmacological effects such as improving cardiovascular function,protecting liver function,regulating body immunity,anti-platelet aggregation,and lowering blood lipids.In recent years,dioscin has been found to have a strong anti-tumor effect,but its mechanism has not yet been clarified.This study used Dio to treat different human colon cancer cells,to detect its survival rate under drug stimulation,and to explore its possible mechanism of mediating apoptosis of colon cancer cells.Methods:1.Select the existing colon cancer cells HT29,RKO,HCT116,SW480 and SW620,set control group and drug treatment group,dilute the Dio in the culture medium according to a preset concentration gradient and add it in each group,MTT method was used to detect the relative survival rate of the cell lines when the cells of each group were treated with the drug at various concentrations,and the half inhibitory concentration(IC50)of the cells in each group was calculated by SPSS.2.Two colon cancer cell lines were selected that were most sensitive to Dio,and control group and preset drug concentration treatment group were used to treat the cells for 48 hours,and then the cell cycle was detected by flow cytometry.3.The cells were treated with control group and preset drug concentration treatment group for 48h and treated with IC50 drug concentration for 0,12,24,and 48 h,then the cell apoptosis was detected by flow cytometry.4.The cells were treated with a preset Dio concentration for 48h,and the cell migration was detected by a scratch test.5.The cells were treated with control group and preset drug concentration treatment group for 48h and then western bolt was used to detect the expression of endoplasmic reticulum stress-related molecules PERK,IRE 1?,ATF6,casapsel2,mitochondrial pathway-related molecule Cytochrome C,caspase9 and apoptotic molecules caspase3?Cleaved-caspase3.Results:1.Colon cancer cells HT29,RKO,HCT116,SW480,and SW620 all showed varying degrees of cytostatic effect with increasing drug concentration and prolonged drug treatment time.Their IC50 are HT29(2.27±0.16ug/ml),RKO(9.46±0.23ug/ml),HCT116(1.19±0.24ug/ml),SW480(2.08±0.18ug/ml)and SW620(1.26±0.25ug/ml),of which HCT116 and SW620 are the most sensitive to drugs,their IC50 value is the smallest.2.Compared with control group,the results of flow cytometry showed that after 48 hours of treatment with 0.5?g/mL and 1.0?g/mL dioscin,the proportion of S-phase cells in HCT116 cells was(29.58±0.05)%,(33.96±0.05)%and(38.78±0.02)%,(P<0.05);the apoptosis rates were(0.05±0.00)%,(21.32±0.10)%and(45.97±0.07)%,(P<0.05).The proportion of SW620 cells in S phase were(38.47±0.05)%,(43.77±0.05)%and(56.92±0.05)%,(P<0.05);the apoptosis rates were(0.02±0.01)%and(1.17±0.03)%and(5.2±0.04)%,(P<0.05).3.After treatment with 1.0?g/mL dioscin for 0h,12h,24h and 48h,the apoptosis rates of HCT116 cells were(0.04±0.01)%,(1.59±0.10)%,(3.25±0.03)%,(14.92±0.20)%,(P<0.05);the apoptosis ratios of SW620 cells were(0.10±0.01)%,(3.89±0.16)%,(5.14±0.03)%,(11.31±0.11)%,(F<0.05).4.The scratch test results showed that in HCT116 cells the control group migration rate was(0.37±0.00)%,and the experimental group migration rate was(0.07±0.00)%,(P<0.05);in SW620 cells the control group migration rate was(0.32±0.01)%,the experimental group migration rate was(0.08±0.00)%,(P<0.05).5.Western Bolt results showed that compared with the control group,the endoplasmic reticulum stress-related molecules(PERK,IRE 1?,ATF6,caspase12)in the drug treatment group were highly expressed(P<0.05).6.Compared with the control group,the mitochondrial pathway-related molecule(Cytochrome C,caspase9)in the drug treatment group were highly expressed(P<0.05).7.Compared with the control group,the apoptotic molecules(caspase3?Cleaved-caspase3)in the drug treatment group were highly expressed(P<0.05).Conclusion:l.Dioscin can significantly inhibit the proliferation and migration of colon cancer cells and promote their apoptosis.2.Dioscin can induce the activation of endoplasmic reticulum stress response and mediate apoptosis of colon cancer cells through mitochondrial apoptosis pathway.