The Research On Apoptosis Induction And Mechanism Of Unsaturated Fatty Acids From Rice Bran Oil To Human Liver Hepatocellular Carcinoma HepG2

Rice bran oil with high nutritional value,which is consumpted regularly can reduce blood fat,cholesterol,regulate blood lipid metabolism,improve cardiovascular and cerebrovascular diseases,antioxidant,anti-cancer,etc.,has a broad consumer prospects,which is due to rice bran oil rich in biological activity Ingredients,especially oleic acid,linoleic acid and other unsaturated fatty acids.So in this study,the apoptosis-inducing effect of unsaturated fatty acids from rice bran oil to liver hepatocellular carcinoma(HepG2)was measured and preliminary analysis of its mechanism.In this study,MTT method was used to determine changes in HepG2 and normal liver cells(HL-7702)growth curve;Inverted microscope and fluorescence microscope were used for observation of the change on HepG2 and HL-7702 cells morphology;Scratch test and Transwell chamber migration assay were used to measure the ablility of HepG2 cells migration and attack.The results showed that the inhibition ability of unsaturated fatty acids which were under certain concentrations from rice bran oil to HepG2 was obviously(P<0.05).When the concertration of RBO,OA and LA reached more than 0.25 mmol/L,0.2 mmol/L and 0.1 mmol/L,but the effect of inhibition was not evident for HL-7702 cells;Result of light microscope showed that cells became smaller cell body,intercellular space increased.Nucleus became blurred,cell membrane cell fell off shrinkage phenomenon in treated groups,but the emergence of morphology was not evident for HL-7702 cells;The migration experiment results show that the treated groups can inhibite it's athletic ability and invasion ability in different degrees,the athletic migration inhibitory rate was about 15%,the invasion migration inhibitory rate was about 50%.This part shows that rice bran oil and its unsaturated fatty acids on normal liver cells no significant proliferation inhibition,and liver cancer cells have significant inhibition of proliferation.In contrast to normal liver cells,the effect of rice bran oil and its unsaturated fatty acids on the growth of hepatocarcinoma cells was established,which laid the foundation for the following research.On the basis of the above research,in order to investigate the reason of the inhibition of growth and proliferation of hepatocarcinoma cells by rice bran oil and its unsaturated fatty acids,the apoptosis of HepG2 cells before and after treatment with rice bran oil and its unsaturated fatty acids was studied by flow cytometry;The change of HepG2 cells cycle was detected by flow cytometry;The expression of Caspase-3 in hepatocellular carcinoma cell line HepG2 was detected by Western blot;The changes of mitochondrial membrane potential in HepG2 cells was detected by flow cytometry before and after treatment with rice bran oil and its unsaturated fatty acids used rhodamine 123.The result of apoptotic rateswere that:The HepG2 cells treated with rice bran oil,oleic acid and linoleic acid was 31.5%,48.9%and 37.5%,respectively,the apoptotic rate was significantly higher than that of the control group;Cell cycle distribution test showed that rice bran oil unsaturated fatty acids can cause significant apoptosis of liver cancer cells,but not cause cell cycle arrest to achieve the effect of growth inhibition;The Western blot results were shows that the expression of Caspase-3 protein in the whole protein of hepatocellular carcinoma cells was up-regulated by rice bran oil and its unsaturated fatty acid treatment;The results of mitochondrial membrane potential showed that HepG2 cellsmitochondrial membrane potential loss withrice bran oil and its unsaturated fatty acid treatment;This part shows that rice bran oil and its unsaturated fatty acids can induce apoptosis of liver cancer cells to inhibit its growth and proliferation.On the basis of the above research,in order to further explore the mechanism of rice bran oil and its unsaturated fatty acids on the induction of apoptosis in hepatocellular carcinoma cells,the expression of GRP78,CHOP and Caspase-12 in rice bran oiland its unsaturated fatty acids were studied by Western blot,in order to researching and exploring anticancer mechanism under the different time pionts;The expression of apoptotic markers Bax and Bcl-2 in HepG2 cells were detected by Western blot.From the endoplasmic reticulum stress pathway,it was shown that rice bran oil and its unsaturated fatty acid was induced by endoplasmic reticulum stress initiation protein GRP78 at 6 h after treatment,sustained stress and induced endoplasmic reticulum stress apoptotic protein CHOP was up-regulated after 12 h,and the expression of Caspase-12 was up-regulated at 24 h,which induced apoptosis.The effect of rice bran oil and its unsaturated fatty acid on the apoptosis of hepatocellular carcinoma cells and the mechanism of rice bran oil and its unsaturated fatty acids on the apoptosis of hepatocarcinoma cells were studied.In the review,rice bran oil and its unsaturated fatty acids in vitro on liver cancer cells HepG2 significant apoptosis induction,and can effectively reduce the migration of liver cancer cells,while the normal liver cells HL-7702 hadn't significant effect.It is suggested that rice bran oil and its unsaturated fatty acids can activate the endoplasmic reticulum stress-related apoptosis pathway,and induce apoptosis of HepG2.This will further develop the use of rice bran oil that provides research-based data,and provide a reference for finding and developing a new type of natural anti-cancer substances.It also provides the theoretical basis for the application of unsaturated fatty acids from rice bran oil in the food and pharmaceutical area.