Protective Effect Of Ferulic Acid On Liver Injury In Piglets And Its Mechanism

The study aimed to investigate whether ferulic acid could alleviate the side effects of oxidative stress on growth performance and liver function of piglets.In this experiment,a model of oxidative stress in weaned piglets was firstly established by intraperitoneal injection of Diquat to investigate the effect and possible mechanism of ferulic acid in alleviating oxidative stress in piglets.Besides,low birth weight(LBW)piglets were used to further verify the protective effect of ferulic acid on oxidative damage to the liver function.Experiment 1: Effect of ferulic acid on growth performance and liver function of in piglets challenged by diquatIn this experiment,32 crossbred(Duroc ×Landrace × Yorkshire)piglets with an average body weight of 13.24±0.19 kg were selected and randomly divided into 2dietary treatments including control diet(CON)and control diet supplemented with4g/kg ferulic acid(FA).On d 15,piglets in each group were randomly divided into 2sub-groups(sterile saline and Diquat).The four treatment groups were CON,Diquat,Con+FA and Diquat+FA groups.Equal doses of sterile saline and Diquat solution(10mg/kg BW)were injected intraperitoneally.On d 20,after weighing and blood collection,all pigs were slaughtered for sampling.The results were as follows.(1)On 1-14 d,supplementing ferulic acid to the diet significantly increased ADG and decreased F/G in piglets(P<0.05).Diquat-induced oxidative stress significantly reduced ADG,ADFI and final body weight at d 20(P<0.05).Dietary supplementation of ferulic acid significantly improved ADFI and final body weight at d 20 in Diquat challenged piglets(P<0.05).(2)Diquat stress caused significant ballooning of piglet liver cells and severe cytoplasmic vacuolation,significantly increasing serum AST and ALT activity(P<0.05).Dietary supplementation with ferulic acid improves Diquat-induced liver injury and inhibits AST and ALT activity(P<0.05).(3)Under normal conditions,ferulic acid supplementation could significantly decrease serum TC and raise HDL-C and hepatic HL enzymes(P<0.05).Diquat significantly increased serum concentrations of TG and TC,decreased activities of LPL and HL,reduced HDL-C concentration,and decreased hepatic PPARα and CPT1 b m RNA expression(P<0.05).Ferulic acid supplementation inhibited Diquat-induced elevation of serum TG and TC,promoted the restoration of HDL-C concentration and LPL enzyme activity,and increased NEFA concentration and relative PPARα m RNA expression in the liver(P<0.05).(4)Ferulic acid significantly increased CAT and T-AOC in serum and GSP-Px and T-AOC activities in liver of normal piglets.Ferulic acid up-regulated the expression levels of antioxidant genes(HO-1)and proteins(HO-1 and Nrf2)in liver,and decreased the protein expression level of Keap1(P<0.05).Diquat treatment significantly reduced T-AOC capacity in serum and liver,decreased GSH-Px and T-SOD enzyme activities in serum,increased MDA levels in serum and liver,significantly upregulated the relative gene and protein expression of Keap1,and decreased the protein expression level of Nrf2(P<0.05).Under Diquat challenge,ferulic acid supplementation significantly increased serum and liver T-AOC capacity and serum T-SOD activity,decreased serum and liver MDA levels,down-regulated gene and protein expression levels of Keap1,and up-regulated protein expression levels of Nrf2 and HO-1 in the liver(P<0.05).(5)Diquat challenge significantly increased ROS levels in piglet liver,decreased SDH and MDH enzyme activity,and down-regulated mt DNA copy number and TFAM m RNA relative expression in liver(P<0.05).Ferulic acid significantly inhibited Diquatinduced increase in ROS level and increased the relative m RNA expression of hepatic TFAM and ATP5G(P<0.01).(6)Diquat stress significantly increased the rate of late apoptosis in liver cells,increased serum levels of IL-1β,IL-18 and LDH,and upregulated pyroptosis-related genes(Caspase1,GSDMD,IL-1β and IL-18)in the liver of piglets(P<0.05).Ferulic acid reduces late apoptosis in the liver of stressed piglets,suppresses Diquat-induced increases in serum levels of IL-1β,IL-18 and LDH,and downregulates m RNA expression of hepatocyte pyroptosis-related genes(NLRP3,Caspase1,GSDMD,IL-1βand IL-18)(P<0.05).The results showed that Diquat-induced oxidative stress caused impaired liver function and reduced growth performance in weaned piglets.Dietary ferulic acid could induce liver antioxidant capacity,improve lipid metabolism,and alleviate oxidative damage to pig liver.Experiment 2: Protective effect of ferulic acid on liver of piglets with low birth weight(LBW)suffering from oxidative damageIn this experiment,24 piglets of 7 days-old were selected from the piglets with similar physical condition and identical litter date.After 3 d of pre-feeding,LBW piglets were randomly divided into two groups: LBW(2.47 ± 0.28 kg)and LBW+FA(2.53 ± 0.27 kg)(n=8/goup).The NBW(3.53 ± 0.53 kg)and LBW groups were fed basal diet,and the LBW+FA group was the basal diet supplemented with 100 mg/kg FA.On d 22,after weighing and blood collection,pigs were slaughtered for further sampling.The results were as follows.(1)LBW piglets showed disturbed arrangement of liver lobules and some cytoplasmic vacuolation.Serum activities of AST and ALT were significantly higher in LBW piglets than those in NBW piglets(P<0.01).Liver structure was improved in LBW piglets fed ferulic acid,but there was no significant effect on the transaminase activity(P>0.05).(2)Compared with the NBW group,LBW piglets showed a significant increase in serum concentrations of HDL-C and LDL-C,a decrease in liver TG concentration and HL enzyme activity,an increase in NEFA concentration,and a decrease in the relative expression of FASN,LPL and CD36 m RNA in the liver(P<0.05).Supplementation of ferulic acid significantly increased serum HDL-C and LDL-C concentrations,liver TG and TC concentrations and HL enzyme activity,decreased NEFA concentrations,and upregulated the relative expression of PPARα,FASN,LPL and CD36 m RNA(P<0.05).(3)Compared with the NBW group,LBW piglets showed a significant increase in serum MDA,a decrease in serum and liver GSH-Px activity and liver T-SOD enzyme activity,and a decrease in the relative m RNA expression levels of HO-1,SOD1,CAT and GST in the liver(P<0.05).Ferulic acid significantly reduced the serum MDA level,increased CAT enzyme activity in liver and GSH-Px and T-SOD activities in serum and liver,significantly reduced the gene expression level of Keap1 and increased the gene expression level of SOD1 and CAT in LBW piglets(P<0.05).(4)Compared with the NBW group,LBW piglets showed significantly increased serum ROS level,decreased hepatic SDH enzyme activity,decreased hepatic mt DNA copy number and ATP5 G m RNA relative expression,and increased Cyt C m RNA expression(P<0.05).Dietary supplementation with ferulic acid significantly reduced serum and liver ROS level,increased SDH and MDH activity in the liver of LBW piglets,and increased mt DNA copy number and ATP5 G m RNA relative expression(P<0.05).(5)Compared with NBW group,serum LDH levels were elevated in LBW piglets(P=0.08),and the relative expression of ASC,GSDMD and Caspase1 m RNAs in the liver was significantly higher(P<0.05).The addition of ferulic acid to the diet significantly reduced the m RNA expression levels of ASC and GSDMD in the liver of LBW piglets(P<0.05).The results showed that the redox homeostasis system in LBW piglets was impaired,lipid metabolism was abnormal,the liver suffered from oxidative stress,and mitochondrial function was disrupted.Ferulic acid improved the antioxidant capacity of LBW piglets,enhanced mitochondrial function,alleviated cell pyroptosis and regulated lipid metabolism by increasing the antioxidant capacity of LBW piglets.In summary,oxidative stress leads to disorders of lipid metabolism and decreased antioxidant capacity,and impaired mitochondrial function of the liver in piglets,thereby reducing growth performance of piglets.Adding ferulic acid to the diet can improve the antioxidant capacity of piglets,regulate lipid metabolism and mitochondrial function,alleviate liver damage of piglets caused by oxidative stress,and promote production.