Character And Regulation Of Fatty Acids Oxidation Metabolism In The Liver Of Ketotic Cows

An underlying mechanism on fatty acid oxidation metabolism capability in theliver of ketotic cows will be brought to light through analyzing the expressing level ofkey enzymes such as acyl-CoA synthetase long-chain (ACSL), carnitinepalmitoyltransferase Ⅰ (CPTⅠ), carnitine palmitoyltransferase Ⅱ (CPTⅡ), acyl-CoAdehydrogenase long-chain (ACADL),3-hydroxy-3-methylglutaryl-CoA synthase(HMGCS), and acetyl-CoA (ACC) in the liver of ketotic cows, and the effect ofmetabolite and hormone on those key enzymes mRNA and protein levels in hepatocytesin vitro. Ⅰn this study, cytobiology and molecular biology were applied to research thealterations of fatty acid oxidation capability in the liver of ketotic cows. The results areas follow:Compared with non-ketotic cows, mRNA abundance of ACSL was increased by5.7-fold in ketotic cows. However, mRNA abundance of CPT Ⅰ, CPT Ⅱ, ACADL,HMGCS, and ACC were decreased by18%,70%,44%,52%, and64%, respectively,compared with non-ketotic cows. ACSL protein levels in the liver of ketotic cows weresignificantly higher, whereas levels of the following enzymes were significantly lower:CPT Ⅱ, ACADL, HMGCS, and ACC. Serum glucose concentration did not appear to beassociated with protein levels of ACSL, CPT Ⅰ, CPT Ⅱ, ACADL HMGCS, or ACC inthe liver of experimental cows. However, serum NEFA concentration was negativelycorrelated with protein levels of CPT Ⅱ, HMGCS, ACADL, and ACC and waspositively correlated with ACSL in experimental cows. Serum BHBA concentrationwas negatively correlated with protein levels of CPT Ⅱ, HMGCS, and ACADL inexperimental cows.Bovine hepatocyte was cultured by the modified two collagenase perfusion. Theeffects of NEFA (0,0.2,0.4,0.8,1.6,3.2mM), BHBA (0,0.5,0.75,1.0,1.5,3.0mM), and glucose (0,1.0,2.0,4.0,8.0,16.0mM) on mRNA andprotein expression of ACSL, CPTⅠ, CPTⅡ, ACADL, HMGCS, ACC in hepatocyte were detected by real-time PCR and ELISA. The results showed that: with increasedconcentration of NEFA, ACSL, ACADL, HMGCS transcription and translation levelswere increased, and ACC transcription and translation levels were decreased in culturedbovine hepatocytes, transcription was decreased with NEFA but translation levels wasnot changed, the transcription and translation of CPTⅠwere increased with lowerconcentration of NEFA and then decreased with increasing concentration of NEFA inthe media, and there was no obvious change in the CPTⅡtranscription and translation.With increased concentration of BHBA, ACSL transcription and translation levels wereincreased, and CPTⅠ, HMGCS transcription and translation were decreased in culturedbovine hepatocytes. There was no obvious change in the CPTⅡ, ACADL, ACCtranscription and translation levels with increasing BHBA concentrations in the culturemedia of bovine hepatocytes. With increased concentration of glucose, ACCtranscription and translation levels were increased, and CPTⅡ, HMGCS transcriptionand translation were decreased in cultured bovine hepatocytes, CPTⅡ transcriptionlevels were increased with glucose but translation levels was not changed. Thetranscription and translation of ACSL were increased with lower concentration ofglucose and then decreased with increasing concentration of glucose in the media. Therewas no obvious change in the ACADL transcription and translation levels withincreasing glucose concentrations.Bovine hepatocyte was cultured by the modified two collagenase perfusion. Theeffects of insulin (0,5,10,20,50,100nM), glucagon (0,0.01,0.1,1,10,100nM), and leptin (0,2.5,5,10,50,100ng/mL) on mRNA and proteinexpression of ACSL, CPTⅠ, CPTⅡ, ACADL, HMGCS, ACC in hepatocyte weredetected by real-time PCR and ELISA. The results showed that: with increasedconcentration of insulin, ACC transcription and translation levels were increased, andACSL, CPTⅠ, ACADL transcription and translation levels were decreased in culturedbovine hepatocytes, CPTⅡ transcription was decreased with insulin but translationlevels was not changed, and there was no obvious change in the HMGCS transcriptionand translation. With increased concentration of glucagon, ACSL, CPTⅠ transcriptionand translation levels were increased, and HMGCS, ACC transcription and translationlevels were decreased in cultured bovine hepatocytes, CPTⅡ transcription was increased with glucagon but translation levels was not changed, and there was noobvious change in the ACADL transcription and translation.With increasedconcentration of leptin, ACSL, CPTⅠ, CPTⅡ, ACADL transcription and translationlevels were increased, and ACC transcription and translation levels were decreased incultured bovine hepatocytes, and there was no obvious change in the HMGCStranscription and translation.As above-mentioned, fatty acid β-oxidation capability was altered in the liver ofketotic cows. Increased activation and decreased β-oxidation of fatty acid suggests morefatty acyl-CoA esterification into triacylglyceride, which may lead to liver lipidosis.High serum NEFA and BHBA concentrations play key roles in metabolism of fatty acidin ketotic cows. The intermediate metabolic products such as NEFA, BHBA, glucoseand hormeones such as insulin, glucagon, leptin could regulate the fatty acid oxidationmetablism capability through promoting or inhibiting the expression of fatty acidoxidation-related enzymes.