| The objective of this study was to investigate the effects of stearic acid, oleic acid, linoleic acid andlinolenic acid on triacylglycerol (TAG) accumulation and the effects of concentration and ratio changeof the four18-carbon fatty acids (18-C FAs) on TAG accumulation, and to elucidate which FA is themost important determinant of TAG accumulation when the four18-C FAs were existed in mediumsimultaneously.1. The experiment was to study the effects of18-C FAs on cell proliferation in bovine mammaryepithelial cells. The cells were cultured with different concentration of stearic acid, oleic acid, linoleicacid or linolenic acid for24h at38℃, and then the MTT experiment was performed. Theconcentrations of18-C FAs were all0,0.78,1.56,3.125,6.25,12.5,25,50,100,200,400μmol/L,respectively. The results showed that cell proliferation was inhibited significantly when the cell culturedwith200or400μmol/L stearic acid, oleic acid, linoleic acid or linolenic acid (P<0.05or P<0.01).2. The bovine mammary epithelial cells were cultured with0,20,40,60,80,100μmol/L stearicacid, oleic acid, linoleic acid or linolenic acid for24h, respectively. Then TAG contents in mediumwere detected by Triglyceride Quantitation Kit. The mRNA expression of lipogenic genes was detectedby RT-qPCR after18-C FAs (100μmol/L) performed on cells for24h, respectively. The results showedas follows:1) the contents of TAG in medium were increased in a concentration-dependent mannerfrom0to100μmol/L.2) Compared with the control, the four18-C FAs were all upregulated the mRNAexpression of CD36, FABP4, ACSL1(P<0.05), which involved in fatty acids uptake, trafficking andactivation, while FABP3, ACSS2were downregulated (P<0.05);18-C FAs were all downregulated themRNA expression of ACACA, FASN, SCD (P<0.05), which involved in de no FA synthesis and FAdesaturation; among the genes involved in mono-, di-and tri-acylglycerol synthesis, only mRNAexpressions of DGAT1after stearic acid performed, DGAT2after oleic acid, linoleic acid or linolenicacid performed, GPAM after stearic acid, oleic acid or linoleic acid performed were increased (P<0.05);mRNA expression of regulation gene of trascription PPARG, PPARGC1A were upregulated after stearicacid performed, but mRNA expressions of SREBF1ã€SCAP were downregulated after oleic acid, linoleicacid or linolenic acid performed.3. An orthogonal L16(45) test was applied to select the optimal combination of18-C FAs forincreasing TAG accumulation, and study their contribution to TAG accumulation. The experimentalfactors were stearic acid, oleic acid, linoleic acid and linolenic acid. The concentrations of18-C FAswere all25,50,75and100μmol/L. TAG contents in cells and medium were tested by TriglycerideQuantitation Kit after different combination performed for24h. Then the mRNA expression oflipogenic genes was detected by RT-qPCR and the fatty acids composition was tested by gaschromatography (GC) method after the optimal combination of18-C FAs performed for24h. Theresults showed that the optimal combination was100μmol/L stearic acid,100μmol/L oleic acid,50μmol/L linoleic acid,75μmol/L linolenic acid, and stearic acid was the most important determinant of TAG accumulation. Compared with control, the optimal combination of18-C FAs upregulated themRNA expression of CD36, ACSL1, FABP4, GPAM, DGAT1, DGAT2and PPARG (P<0.05), butdownregulated the mRNA expression of ACSS2, FABP3, FASN, ACACA, SCD, SREBF1, and SCAP(P<0.05). Besides that, the addition of18-C FAs decreased short-and medium-chain fatty acids inmedium relative to the control (P<0.05).In conclusion, TAG accumulation can be increased as the concentration increase of18-C FAs,separately, and stearic acid was the most important determinant of TAG accumulation when the four18-C FAs were all available. The mixture of18-C FAs have important effects on lipid metabolismthrough the regulation of SREBF1and PPARG, included inducing the pathway of long chain fatty acidsutilization (uptake, trafficking and activation), TAG synthesis, and inhibiting de novo fatty acidsynthesis pathway. |
