The Effects Of Intestinal Physical Barrier On Conjugated Linoleic Acid Reduced Liver Lipid Accumulation In Grass Carp

In the process of fish culture,high lipid diet could affect the lipid metabolism of liver,then inducing the accumulation of lipid in liver and producing the phenomenon of "nutritional fatty liver".Medical studies found that intestinal factors are involved in the occurrence and development of nonalcoholic fatty liver disease.Then the concept of "gut-liver axis" was put forward.However,there are few reports about the "gut-liver axis" mediated high lipid diet induced fatty accumulation in fish liver.Our previous study showed that adding 2.5%conjugated linoleic acid(CLA)to the feed could reduc e lipid accumulation in the liver of grass carp(Ctenopharyngodon idella).In order to explore whether the concept of "gut-liver axis" is also applicable to fish,and whether the CLA played a lipid-lowering role in grass carp through the "gut-liver axis".Two 12 weeks growth tests were designed:1)There are two experimental diets,the control group(soybean oil 2.8%+fish oil 1.8%),and the high lipid group(soybean oil 6.3%+fish oil 1.8%),were conducted to investigate the effects of high lipid diet on growth performance,related indices of liver and intestinal on grass carp;2)There are two experimental of feed,the high lipid group(same as above),and the CLA group(soybean oil 3.8%+fish oil 1.8%+CLA 2.5%),were conducted to investigate the effects of CLA diet on growth performance,related indices of liver and intestinal on grass carp.The test results are as follows:(1)Growth performance and morphological indicators showed that weight gain rate(WGR)and specific growth rate(SGR)of grass carp in the high lipid group were significantly lower than those in the control group(P<0.05),while liver lipid content,hepatosomatic index(HSI)and viscerosomatic index(VSI)of grass carp in the high lipid group were significantly higher than those in the control group(P<0.05).The results of biochemical indexes showed that the levels of glutamic oxalacetic transaminase(AST),glutamic-pyruvic transaminase(ALT)and triglyceride(TG)in the serum the lipopolysaccharides(LPS)in plasma,and the malondialdehyde(MDA)content in the liver of the high lipid group were significantly higher than those in the control group(P<0.05).The activity of catalase(CAT)in liver was significantly lower than that in the control group(P<0.05).Histopathological observation showed that the high lipid diet increased the number and size of lipid droplets in the liver of grass carp,and liver cells showed lesions and disordered arrangement,nuclear migration cell fibrosis.The lipid metabolism related gene analysis showed that the mRNA expression level of SREBP-1c in the liver of grass carp in the high lipid group was significantly higher than that in the control group(P<0.05),and the mRNA expression level of fatty acid synthase(FAS)in the liver of grass carp in the high lipid group was significantly lower than that m the control group(P<0.05).The immune related gene expression showed that the expression of tumor necrosis factor-α(TNF-α),interleukin-8(IL-8),interleukin-1β(IL-1β),nuclear transcription factor κB P65(NF-κB P65),kinase complex α(IKKα)and kinase complex β(IKKβ)in the liver were significantly lower than that of the control group(P<0.05).(2)The mRNA expression levels of Occludin,Claudin c and Claudin 15a in the proximal intestine of grass carp in the high ipid group were significantly lower than those in the control group(P<0.05).The mRNA expression levels of Occludin,Claudin b,Claudin 3 and Claudin 15a in the mid intestine of grass carps were significantly lower than those in the control group(P<0.05).The mRNA expression levels of Occludin and Claudin 3 in the distal intestine of grass carp in the high lipid group were significantly lower than those in the control group(P<0.05),and the mRNA levels of Claudin 15a was significantly higher than those in the high lipid level group(P<0.05).The mRNA levels of ZO-1 in the proximal mid and distal intestine of the high lipid group higher than those in the control group(P<0.05).Intestinal immune-related gene analysis showed that the mRNA expression level of IL-1β in the proximal intestine of grass carps in the high lipid level group was significantly higher than that in the control group(P<0.05),and the mRNA expression level of TNF-a and IFN-γ2 showed an increasing trernd in the high lipid group(P>0.05).The mRNA expression level of TNF-α and IL-8 in the mid intestine of grass carps in the high lipid group were significantly lower than those in the control group(P<0.05).The mRNA expression level of TNF-a,IL-8 and IFN-y2 in the distal intestine of grass carps in the high lipid group were significantly lower than those in the basal group(P<0.05).The mRNA expression level of NF-κB P65 in the proximal and mid intestine of the high lipid group were significantly lower than those of the control group(P<0.05).The expression level of IKKa in the mid and distal intestine of grass carps in the high lipid group was significantly lower than that in the control group(P<0.05),and IKKβ and IKKy showed a downward trend in the high lipid group(P>0.05).(3)The growth performance and morphological indexes showed that the growth rate of grass carp was not significantly affected by the addition of CLA into the feed(P>0.05).and the lipid content in the liver and HSI in the liver of grass carp in the CLA group were significantly lower than those in the high lipid group(P<0.05).The results of biochemical indexes showed that the contents of AST,ALP,TG and TC in the serum of grass carp in the CLA group were significantly lower than those in the high lipid group(P<0.05),and the contents of total protein(TP),albumin(ALB)in the serum and the level of MDA and CAT in the liver were significantly higher than those in the high lipid group(P<0.05).Histopathological observation showed that the addition of CLA in the high lipid diet reduce the lesion and disorder of the liver cells of grass carp,the nuclear migration and fibrosis of the liver cells.And meanwhile,the size and number of lipid droplets in the liver cells decreased.Analysis of lipid metabolis m related genes showed that mRNA expression level of FAS,ACC,LPL,HSL and PPAR-a in the liver of grass carp were significantly higher than those in high lipid group(P<0.05),and the mRNA expression level of SREBP-1c were significantly lower than those in high lipid group(P<0.05).The immune related gene expression showed that the expression of TNF-α、IL-8、IL-10、IL-1β、PPAR-y and IκBα in the liver of grass carp in CLA group were significantly higher than those in high lipid group(P<0.05),and mRNA level of NF-κB P65、IKKα、IKKβ and IKKγ in the liver were significantly higher than those in high lipid group(P<0.05).(4)Gene analysis of intestinal tight junction protein showed that mRNA expression level of ZO-1 and Occludin in the proximal intestine of grass carp in the CLA group were significantly lower than those in high lipid group(P<0.05),and mRNA expression level of Claudin c were significantly higher than those in high lipid group(P<0.05).In CLA group,the mRNA expression level of ZO-1,Occludin,Claudin 3 and Claudin 12 in mid intestine of grass carp were significantly lower than those in high lipid group(P<0.05),The mRNA expression level of ZO-1,Occludin and Claudin 3 in the distal intestine of grass carp were significantly lower than those in high lipid group(P<0.05),and the mRNA expression level of Claudin c were significantly higher than those in high lipid group(P<0.05).Intestinal immune related gene analysis showed that the mRNA expression level of TNF-α in the proximal,mid and distal intestine of the CLA group was significantly lower than that of high lipid group(P<0.05).The mRNA expression level of PPAR-y in the proximal and mid intestine was significantly higher than that of high lipid group(P<0.05),and the mRNA expression level of IL-8 in the proximal intestine was significantly lower than that of high lipid group(P<0.05).The mRNA expression level of NF-κB P65 and IKKa in the proximal,mid and distal intestines of the CLA group were significantly lower than those of high lipid group(P<0.05),and the mRNA expression level of IκBαand IKKβ in the CLA group were significantly lower than those in the mid intestines of high lipid group(P<0.05).Base on the analysis of the results of above experiment,the accumulation of lipid in liver induced by high lipid diet was closely related to intestinal function of grass carp.Therefore,the "gut-liver axis" theory is also applicable to grass carp.CLA could reduce lipid accumulation in the liver of grass carp.However,CLA did not improve the mRNA expression of intestinal tight junction protein in grass carp.