Study On Roles Of Mitochondrial And Peroxisomal ?-oxidation In Lipid Nutrient Metabolism Of Nile Tilapia (Oreochromis Niloticus)

Metabolic diseases have widely existed as lipid metabolism disorders in farmed fish,causing detrimental effect on lipid oxidation and nutritional quality.Beta oxidation is the major pathway for the degradation of fatty acids and occurs in two distinct metabolic compartments,mitochondria and peroxisomes.Beta oxidation plays important roles in energy-consuming processes of lipid metabolism and stress resistance.However,little is known about the roles of mitochondria and peroxisomes in nutrient metabolism of fish,and relationship between nutritional quality and?oxidation.Therefore,we firstly explored whether there are some differences between mitochondrial and peroxisomal?oxidation in tilapia fed with different oils enriched with different fatty acids.Then effects of dietary oils on the fillet lipid of tilapia were deep analyzed based on lipidomics.Finally,we studied metabolic characteristics of different fatty acids in mitochondria and peroxisomes in vitro and in vivo,respectively.There are main results and conclusions in the present study as followings:1.Effect of dietary oils with specific fatty acids on lipid metabolism and nutrition valueThe dominant fatty acids?FAs?in oils are often used to explain different nutritional effects of dietary oils in fish.However,the amounts of dominant FAs among oils are different,and the nutritional roles of these important FAs in fish have not been precisely compared at similar levels in feeding trials.In the present study,different amounts of palmitic acid were added to palm oil?PO?,olive oil?OO?,safflower oil?SO?,perilla seed oil?PEO?and fish oil?FO?to obtain comparable amounts?55%of total FAs?of 16:0,18:1n-9,18:2n-6,18:3n-3 and 20:5n-3+22:6n-3,and subsequently fed to Nile tilapia?11.1±0.01g?for 8 weeks.The results showed that PO group obtained the lower growth compared to SO and PEO group.However,OO group obtained higher fat deposition,serum ALT and AST activities,compared to FO.Lipogenesis-related gene expressions were higher in OO group than FO group in liver,muscle and adipose tissue,but there were only few differences in these genes among other groups.Lipid catabolism genes in FO group were higher than other groups.Overall,dietary 18:2n-6,18:3n-3,20:5n-3 and 22:6n-3 were beneficial to normal growth and lipid metabolism.However,high amount of 16:0 depressed growth and,high amount of 18:1n-9 induced lipid deposition and liver damage in Nile tilapia.Taken togehter,these effects are attributed to some differences between mitochondrial and peroxisomal?-oxidation.The nutritional value of fish fillet can be largely affected by dietary oils.However,little is known about how dietary oils modify lipid molecules in fish fillets.Through biochemical and lipidomics assays,this study demonstrated the molecular characteristics of fillet lipids in Nile tilapia fed with different oils for six weeks.High18:2n-6 and low 18:3n-3 deposition in phosphoglycerides resulted high18:2n-6/18:3n-3 ratio in tilapia.Dietary n-3 VLCUFAs intake increased its deposition at sn-1/3 of triglycerides and at sn-2 of phosphatidylcholines.Irrespective of dietary oil,16:0 was distributed preferentially at the outer positions of glycerol backbone.High 18:2n-6 accumulated at sn-2 position for fish fed with n-3 PUFA-enriched oils.High 18:3n-3 deposited at sn-1/3 in TG,sn-1 in phosphatidylethanolamines,while at sn-2 in phosphatidylcholines.Together,dietary oils change the composition and positional distribution of fatty acids on the glycerol backbone,and change nutritional value of fish for human health.2.Analysis of metabolic characteristics of different fatty acids in mitochondriaOur previous studies have already implied that differences between mitochondrial and peroxisomal oxidation might result in different effects of dietary oils on lipid metabolism and fillet nutritional quality of tilapia.Therefore,we studied metabolic characteristics of different fatty acids in mitochondria and peroxisomes,respectively.Firstly,we analyzed metabolic characteristics of different fatty acids in mitochondria in organelles,cell and living fish,respectively.The results showed that PA,OA and LNA increased mitochondrial respiratory,but LA,ARA,EPA and DHA depressed mitochondrial respiratory.The data of fatty acid composition of?oxidation product in mitochondria indicated that mitochondria firstly oxidized OA and PA in Primary hepatocytes with impaired mitochondrial?oxidation.Then,the results was confirmed in vivo.Therefore,the present study used Nile tilapia fed with MD-containing soybean oil?SO?and fish oil?FO?as a fish model,and mimicked actual phenomenon of impaired mitochondrial FA?-oxidation induced by different saturations and chain length fatty acids in farmed fish.The results showed that inhibited mitochondrial oxidation affected the contents of PA,ARA and EPA,instead of their positional distribution and nutritional value.The contents positional distribution and of OA were related to dietary oils,but those of LA and DHA were influenced by impaired mitochondrial oxidation and dietary oils.In conclusion,effects of depressed mitochondrial oxidation on lipid class content,fatty acid composition,the number of differential metabolites in fillet PLs in different ways depending on dietary oil types.To the best of our knowledge,this is the first study to examine the contents,structures and changes of PL molecular species in fish with impaired fat oxidation fed with different oils.3.Analysis of metabolic characteristics of different fatty acids in peroxisomesThe lack of specific inhibitor greatly limits expanding our understanding of peroxisomal oxidation.It was reported that acetylenic acids?TRO and TDYA?were proposed to have the potential to inhibit ACO.Therefore,in the present study,we firstly explored the concentration and treatment time of TRO in primary hepatocytes and then analyzed metabolic characteristics of PA in primary hepatocytes with TRO.The results showed that the concentration and treatment time of TRO were 0.02umol/L and 6 h,respectively.Impaired peroxisomal oxidation increased the VLCFA contents in a short term but affected the utilization of LCFA in a long term.Then,the supplementation of TDYA was determined in vivo in tilapia.The findings were that treatment of TDYA?75mg/kg and 150mg/kg?induced elevated TG,MDA and ALT in serum and liver thereby increasing lipid peroxidation and liver damage.Depressed peroxisomal oxidation did not affect the lipid contents in liver and muscle but changed fatty acid composition.Additionally,TDYA decreased the expressions of genes related to lipid catabolism and lipogenesis in liver,so increased TG and NEFA contents.The results indicated that TDYA inhibited tilapia liver ACO in the concentration of 75mg/kg and 150mg/kg and finally we chose 75mg/kg.Studies indicated that peroxisomal oxidation was induced under conditions of high fat diet which resulted in an increase of hydrogen peroxide and led to decrease mitochondrial oxidation.Inhibited peroxisomal oxidation by the supplementstion of inhibitor could improve oxidative injury in high fat diet.Therefore,we firstly study whether TDYA improves hepatic lipid in tilapia fed a high fat diet.We found that high fat diet increased lipid deposition,which decreased oxidation capacity of lipid and led to liver damage in liver.Treatment of TDYA in high fat diet significantly decreased TG,NEFA and H₂O₂ contents and elevated CAT and SOD activities in serum and liver.Additionally,TDYA did not change the gene expression of lipid metabolism in liver.In conclusion,TDYA improves hepatic lipid by increasing enzyme activities and oxidation capacity,thereby inducing decreased lipid contents in tilapia fed a high fat diet.In conclusion,our previous studies have indicated that differences between mitochondrial and peroxisomal oxidation might result in different effects of dietary oils on lipid metabolism,thereby affecting fillet nutritional quality of tilapia.Impaired mitochondrial oxidation changed PL composition and nutritional value of the fillet in tilapia.Inhibited peroxisomal oxidation increased lipid composition in cell and in living tilapia.This is the first study to investigate comprehensively the roles of mitochondrial and peroxisomal?-oxidation in nutrient metabolism and nutritional quality of fish.Our findings will provide new insight into improment of nutritional quality and selection of dietary oils in fish.