Study On Biofunctional Mechanism Of The Occurrence And Development Of Colon Cancer With Polyunsaturated Fatty Acids Supplementation

Colorectal cancer is one of the most common digestive tract tumors.With the improvement of people’s living standards,the morbility of colorectal cancer showed an increasing trend in China.The high incidence of colon cancer has been attributed to the replacement of the traditional,high-fiber,low-fat,and low-calorie diet by a high fat,low-fiber,and calorie dense diet.This suggests that dietary intervention may form an innovative way to decrease the occurrence of colorectal cancer.Studies have shown that changes in diet fatty acids composition,raising the ratio of polyunsaturated fatty acids and decreasing the ratio of saturated fatty acids would help in reducing the incidence of tumor.Therefore,the action mechanism of PUFAs on colon cancer is worthy of in-depth discussion.In the present work,we examined the growth inhibition effect of PUFAs on LoVo and RKO colon cancer cells in vitro,and further mechanisms were studied in the effects of PUFAs on the mitochondrial dysfunction and fatty acid metabolism.On the other hand,the effects of different types of fatty acids on the gut environment and preventing colorectal carcinoma were explored in vivo.Experiments in vitro were conducted on human colon cancer LoVo and RKO cells,and cellular injury model was successfully constructed by six PUFAs.It was observed that linoleic acid(LA),alpha-linolenic acid(ALA),arachidonic acid(AA),eicosapentaenoic acid(EPA),docosahexaenoic acid(DHA),γ-linolenic acid,(GLA)induced apoptosis of the colon cancer cells LoVo and RKO at concentrations above 120μM,but had little effect on HUVEC.The semi-differentiated colon cancer cell line RKO was more sensitive to the cytotoxic action of PUFAs compared to the undifferentiated colon cancer cell line LoVo.Based on the results of MTT,we selected the most effective concentrations of various fatty acids for further mitochondrial dysfunction and fatty acid metabolism studies as depicted below:ALA(150μM),EPA(150μM),DHA(150 gM),LA(150 μM),GLA(300 μM),AA(150μM),5-FU(10 μM)to test on LoVo cells and ALA(140μM),EPA(120μM),DHA(120 μM),LA(120 μM),GLA(200 μM),AA(120 μM),5-FU(0.4 μM)to test on RKO cells.The effects were found as follow:(1)The effect of different fatty acids on mitochondrial dysfunction.We showed that n-3 PUFAs(ALA,EPA,DHA)and n-6 PUFAs(LA,GLA,AA)fatty acids induce apoptosis of LoVo and RKO cells by a combination of mechanisms that include enhanced generation of ROS,accumulation of intracellular Ca2+,and a decrease in ATP level,an increase in Bax/Bcl-2 ratio,followed by attenuation of mitochondrial membrane potential via small pores present in mitochondrial membrane,accompanied by cytochrome c and AIF release to the cytosol,activation of initiator and executioner caspases.PUFAs induced apoptosis of colon cancer cells through a mitochondrial dependent pathway.(2)We investigated the biotransformation of PUFAs in LoVo and RKO cells and effect of PUFAs metabolites on cells.Oil red staining showed that PUFAs were stored as lipid droplets in LoVo and RKO cytoplasm.Supplementation with PUFAs ratio of unsaturated fatty acid and increased ratio of unsaturated fatty acid significantly by GC analysis.In addition,tumoricidal action of PUFAs on colorectal LoVo and RKO cancer cells in vitro was associated with suppressed expression of COX-2,ALOX5,mPGES and decreased synthesis of PGE2,LTB4,increased formation of LXA4.Experiments in vivo were conducted on C57BL/6 mice fed with diets supplemented with fish oil,sunflower oil or flaxseed oil.We investigated the colon microbe momposition,the amount of short chain fatty acids and bile acids of feces in mice and the lipid metabolism regulation effect of three distinct fatty acids on mice to analyze the effect of PUFAs on improving gastrointestinal health and preventing colorectal carcinoma.The results showed that PPARa and OSTa mRNA level increased significantly in three PUFAs supplementation group with the effect of acceleration metabolism of fat.Supplementation of fish oil and sunflower oil had increased the concentrations of the ratio of n-3/n-6 PUFAs in colon tissues,avoiding oxidative stress damage of colon tissues.Moreover,PCR-DGGE results demonstrated that composition of bacteria in C57BL/6J mice colonic content had been changed in fish oil,flaxseed oil,and sunflower oil supplementation groups in comparison with control group.Fish oil and flaxseed oil can promote Eubacterium,Novosphingobium,and Hymenobacte.Bacteria diversity in mice colonic content treated with sunflower oil was less than that of the fish oil and flaxseed oil groups.Fish oil rather than flaxseed oil or sunflower oil dietary consumption increased the butyric acid producing bacteria Eubacterium,and total SCFA levels,particularly,the n-butyric acid level.Furthermore,fish oil,flaxseed oil and sunflower oil consumption presented lower ratios of fecal secondary to primary bile acids,reduced the fecal pH.Therefore,we propose that oils rich in PUFAs may have contribution on improving gastrointestinal health and preventing colorectal carcinoma.