The Mechanism Of Perilipin5 In The Hepatic Lipidosis Of Dairy Cows With Fatty Liver

Fatty liver is a common nutritional metabolic disease in perinatal high-yield dairy cows.Estimates indicate that during the first 4 weeks after parturition 50% or more cows suffer different degrees of triacylglycerol(TAG)accumulation in the liver.The important pathological features of fatty liver cows are hepatic lipid accumulation and high blood concentrations of non-esterified fatty acid(NEFA).The transition dairy cows often showed a negative energy balance caused by the lower intake of dry matter and the increased demand for glucose to support milk production.Negative energy balance initiates fat mobilization and a subsequent increase in blood NEFA concentration.Large amounts of NEFA are esterified into TAG and stored in the form of lipid droplets(LD)in liver,which leads to the development of fatty liver.The surface of the LD is inlaid with a variety of lipoproteins,which play important roles in the stability and metabolic process of LD.Perilipin 5(PLIN5)is a lipoprotein that plays important roles in the development of hepatic steatosis in mice and humans.At present,whether PLIN5 plays a role in development of fatty liver in dairy cows and its underlying molecular mechanisms remains unknown.An in vivo study consisting of healthy and cows with fatty liver was performed to detect the expression of key molecules of lipid synthesis,fatty acid oxidation,very low-density lipoprotein(VLDL)assembly and PLIN5.The results showed that the concentration of NEFA and β-hydroxybutyrate(BHB)in fatty liver cows was higher,while the milk yield and blood glucose concentration were lower,indicating that cows with fatty liver displayed with severe negative energy balance,which initiated fat mobilization.The expression of key molecules of lipid synthesis,sterol regulatory element binding protein-1(SREBP-1),fatty acid synthase(FAS),acetyl-coA carboxylase 1(ACC1),diacylglycerol acyltransferase 1(DGAT1)and diacylglycerol acyltransferase 2(DGAT2)were significantly increased in liver of cows with fatty liver,while key molecules of fatty acid oxidation carnitine palmitoyltransferase-I(CPT-I),carnitine palmitoyltransferase-II(CPT-II)and β-hydroxyacyl-CoA-DH(HADH),and VLDL assembly-related lipoproteins microsomal triglyceride transfer protein(MTP),apolipoprotein B100(ApoB100)and apolipoprotein E(ApoE)expression were significantly decreased.The above results confirmed that lipid synthesis was increased,but fatty acid oxidation and VLDL assembly were decreased in liver of cows with fatty liver.Importantly,PLIN5 was highly expressed in liver of cows with fatty liver,indicating that PLIN5 may play an important role in the development of fatty liver in dairy cows.In vitro,hepatocytes isolated from calves were transfected with PLIN5 overexpression adenovirus.The results showed that PLIN5 overexpression significantly increased the expression of lipid synthesis related molecules SREBP-1,FAS,ACC1,DGAT1 and DGAT2,and inhibited fatty acid oxidation related molecules CPT-I,CPT-II and HADH,and VLDL assembly related molecules MTP,ApoB100 and ApoE expression,which induced TAG accumulation in hepatocytes.These results indicate that PLIN5 can regulate lipid synthesis,fatty acid oxidation and VLDL assembly in bovine hepatocytes.Hepatocytes isolated from calves were treated with different concentration of NEFA.The results showed that NEFA treatment induced the expression of PLIN5 in hepatocytes.Furthermore,NEFA increased the expression of lipid synthesis related molecules SREBP-1,FAS,ACC1,DGAT1 and DGAT2.High concentration NEFA inhibited the fatty acid oxidation related molecules CPT-I,CPT-II and HADH,and VLDL assembly related molecules MTP,ApoB100 and ApoE,which induced TAG accumulation in hepatocytes.Bovine hepatocytes were transfected with PLIN5 knockdown adenovirus,then,the cells were treated with NEFA.The results showed that PLIN5 knockdown attenuated the effect of NEFA on the expression of lipid synthesis molecules,fatty acid oxidation molecules and VLDL assembly molecules,and alleviated the NEFA-induced TAG accumulation.These results indicate that NEFA activates the expression of PLIN5,then promotes the synthesis of fatty acids and TAG,reduces fatty acid oxidation and VLDL assembly,subsequently induces TAG accumulation in hepatocytes.Bovine hepatocytes were co-transfected with PLIN5 overexpression adenovirus and SREBP-1 knockdown adenovirus.The results showed that knockdown of SREBP-1 decreased the expression of lipid synthesis related molecules SREBP-1,FAS,ACC1,DGAT1 and DGAT2,but increased the expression of fatty acid oxidation related molecules CPT-I,CPT-II and HADH,and VLDL assembly related molecules MTP,ApoB100 and ApoE,thereby alleviating PLIN5 overexpression-induced TAG accumulation in hepatocytes.These results indicate that PLIN5 regulates lipid synthesis,fatty acid oxidation and VLDL assembly in bovine hepatocytes through SREBP-1.In summary,the high concentration of fatty acids induced by negative energy balance activates of PLIN5 in the liver,which promotes the expression of SREBP-1,in turn promotes lipid synthesis and inhibits fatty acid oxidation and VLDL assembly,thereby inducing fat accumulation in the liver of cows,leading to development of fatty liver in dairy cows.