Effects Of Different Diets Type And Long Chain Fatty Acid On Milk Fat Synthesis In The Mammary Gland Of Dairy Cows

The objective of this study was to determine the effects of different diets on milk fat synthesis in the mammary gland of dairy cows, and the effects of long chain fatty acids (LCFA) on milk fat synthesis in bovine mammary epithelial cells(BMEC). The results of changed in milk fat synthesis and LCFA in vivo were studied the mechanism of milk fat synthesis in vitro by LCFA added tests. The study in effects of the best combination of LCFA on milk fat synthesis would supply the basis of milk fat metabolism model and would supply the basis in improve milk quality. The study mainly involved in four experiments:1. Thirty multiparous Holstein cows were assigned to a single factor block design with3periods. The diets were:1) Mixed forage treatment (IMF), concentrate-roughage ratio46:54.2) High-concentrate with corn stover (CS1), concentrate-roughage ratio65:35.3) Low-concentrate with corn stover (CS2), concentrate-roughage ratio46:54. Each period lasted21d. At the last7days of each period served as sample collect period. Mammary gland tissue biopsies were randomly performed for half of cows in each group after milking on the last day of experiment. The milk yield was lowest in cows fed with CS2. The milk fat, milk protein and TS were highest in cows fed with IMF. The milk composition of SFA and short FA were highest in cows fed with IMF, and the milk composition of MUFA and LCFA were highest in cows fed with CS2, and the milk compositon of PUFA was highest in cows fed with CS1. The expression of FASN, SCD, LPL and CD36were higher in IMF compared with others.2. We determined the high sequencing technology RNA-Seq to sequence the genes of the mammary tissue in dairy cows and their functions were analyzed in order to study the differences of the transcription level of the mammary tissues in cows. We got good data quality and meeting the requirements for further analysis. We have obtained18828unigene in25classifies in COG involved in877unigene about lipid transport and metabolism, and595unigene about AA transport and metabolism. We have obtained27924unigene in GO classify involved in96295unigene be classified biological process, and77729unigene be classified cellular component, and37950unigene be classified molecular function. Through the differentially expressed genes were enriched lipid metabolism pathway analysis were conducted on fatty acid biosynthesis were higher in IMF and on fatty acid elongate and biosynthesis of unsaturated fatty acids were lower in IMF compared with CS. The results were consistent with the results of previour experiment.3. We determined the cell proliferation on BMEC with different concentration of palmitic acid, stearic acid, oleic acid, linoleic acid and linolenic acid to find the concentration of LCFA did not inhibited in cell proliferation. An orthogonal L16(43) test and L16(45) test were applied to select the optimal combination of UFA and LCFA for increasing TAG accumulation. The combination of UFA was involved in100μM oleic acid,100μM linoleic acid and1.4μM linolenic acid. The combination of LCFA was involved in100μM palmitic acid,5μM stearic acid,100μM oleic acid,25μM linoleic acid and1.2μM linolenic acid. The order of the effect on TAG accumulation ability was C18:3> C18:2>C18:1in UFA combination. The order of the effect on TAG accumulation ability was C18:0> C18:2>C18:3>C18:1> C16:0in LCFA combination. The accumulation of TAG was highest in LCFA combination. The expression of FASN was the lowest, and the expression of PPARG and CD36were the highest in LCFA combination. The expression of SCD and ACSL1were the lowest in control treatment. The expression of AGPAT6and SREBF1were the lowest in UFA combination. The short-, and medium-chain FA was the lowest and UFA in BMEC was the highest in LCFA combination treatment. The SFA in BMEC was the highest in control treatment. The C18:0in culture medium was higher in LCFA combination compared with UFA combination.4. We determined the TAG accumulation in retracement of LCFA combination. The result was consistent with previous experiment which stearic acid is the greatest synthesis in TAG content. The expression of ACACA and FASN were no different among the treatments. The expression of FABP3, ACSL1and SREBF1were highest in retracement of linolenic acid, and the expression of LPL was highest in retracement of palmitic acid. The short-, medium-chain FAs were highest and the LCFA was lowest in control treatment. The SFA in BMEC was highest and the UFA was lowest in retracement of linolenic acid. The LCFA in culture medium was highest in retracement of palmitic acid. The SFA in culture medium was highest and the UFA in culture medium was lowest in retracement of stearic acidIn conclusion, the diet with nutrition balance improved milk yield, milk fat and milk protein and genes expression of milk fat synthesis. The results of milk fat content and composition were verified by transcriptome. In vitro, LCFA can improve TAG content, decreased genes expression of de novo synthesis, increased genes expression of transcription and network regulation in BMEC, and decreased SMFA content, increased UFA content in cells. The stearic acid was the most important fatty acid on TAG synthesis. The LCFA had no significant selectivity in milk fat de novo synthesis. Linolenic acid decreased in genes expression of network regulation and transcription, palmitic acid decreased in LPL gene expression.