Effect Of Fatty Acids And Hormone Supplementation On Milk Protein And Milk Fat Synthesis In Bovine Mammary Epithelial Cells

This study investigated the effects of milk protein and milk fat synthesis in bovine mammary epithelial cells(BMECs)through addition different concentrations of short-chain fatty acids(acetic acid,?-hydroxybutyrate),long-chain fatty acids(oleic acid,linoleic acid,linolenic acid)and hormones(prolactin,growth hormone,insulin)in two-dimensional and three-dimensional cultures,which provide theoretical basis for regulating the synthesis of milk components.This thesis consists of two trials.The first trial studied addition short-chain(SCFA)and long-chain fatty acids(LCFA)in two-dimensional cultures mode to detect the relative proliferation rate(RGR)of BMECs,the amount of synthesis of triglycerides(TAG)and the expression of genes associated with milk protein and milk fat synthesis.The test was divided into 5 groups.No fatty acids were added to the control group.The concentrations of acetic acid and ?-hydroxybutyric acid added in groups I and II were both 2.0:1.0,the concentration ratios of oleic acid,linoleic acid and linolenic acid were 2.0:13.3:1.0 and 9.6:7.4:1.0,respectively;The ratio of acetic acid and ?-hydroxybutyric acid added in group III and IV was 1.0:1.0,the concentration ratios of oleic acid,linoleic acid and linolenic acid were 2.0:13.3:1.0 and 9.6:7.4:1.0,respectively.The total concentration of short-chain fatty acids(SCFA)and long-chain fatty acids(LCFA)added in each group was 14.541 mmol/L.Selecting the appropriate concentration of fatty acid to perform the experiment in three-dimensional cultures.The trial included three groups,a control group and the group I and II.The second trial studied the effects of addition SCFA,LCFA,and hormones on BMECs in the medium of the two-dimensional cultures.The trial was divided into three groups.The control group did not add fatty acids and hormones.The concentration of acetic acid and ?-hydroxybutyric acid added in the group I was 2.0.: 1.0,the concentration ratio of oleic acid,linoleic acid,and linolenic acid was 2.0:13.3:1.0,the concentration ratio of PRL,INS and GH was 10.0:1.0:3.0;The concentration of acetic acid and ?-hydroxybutyric acid added in the group II was 1.0:1.0,and the concentration ratio of oleic acid,linoleic acid,and linolenic acid was 9.6:7.4:1.0,the concentration ratio of PRL,INS,and GH was 10.0:1.0:3.0.Selecting the appropriate concentration of fatty acids and hormone to perform in three-dimensional cultures,and the effects of different culture modes on the expression of TAG content and milk protein and milk fat synthesis-related genes in BMECs werecompared.The results of first trial study showed that in the two-dimensional culture mode,the synthesis of RGR and TAG of the BMECs in the experimental group was significantly higher than that in the control group,the RGR in the group I was the highest,and the TAG synthesis was the highest;Compared with the control group,the group I significantly increased gene expression of 4EBP1,m TOR,S6K1 and CSN3,the group IV significantly increased gene expression of 4EBP1,S6K1,AKT and CSN3;the gene expression of STAT5 was significantly decreased;Compared with the control group,the gene expression of DGAT2 in the experimental group was significantly increased,and the gene expression of FASN was significantly decreased.In the three-dimensional culture mode,the amount of TAG synthesis in the control group and the group I was significantly higher than that in the two-dimensional culture;the gene expression levels of m TOR,AKT,CSN1S1 and CSN3 in the groups I and II were significantly higher in the three-dimensional culture than that in the control group;The expression of milk fat synthesis-related genes in BMECs was significantly higher than that in two-dimensional cultures;the experimental results showed that concentrations of acetic acid,?-hydroxybutyrate,oleic acid,linoleic acid,linolenic acid was 9.6mmol/L,4.8mmol/L,17.3?mol/L,115.03?mol/L,8.65?mol/L and 7.2 mmol/L,7.2 mmol/L,75.2?mol/L,58?mol/L,7.8?mol/L,respectively.which can have a good promotive effect on the expression levels of milk protein and milk fat synthesis-related genes in BMECs.The results of the second trial showed that in the two-dimensional culture mode,the synthesis of RGR and TAG of the experimental group was significantly higher than that in the control group;the gene expression of the S6K1,m TOR,C-Case2 and CSN3 in the group I was higher than in the control group,the gene expression of 4EBP1,m TOR,AKT in group II was significantly higher than that in the control group;The groups I and II significantly increased the gene expression of DGAT2 and the group I significantly promoted the gene expression of PPARG;Compared with the control group,the gene expression of PRLR was significantly increased in the group I.In the three-dimensional culture mode,the synthesis of RGR and TAG in the BMECs of the experimental group was significantly higher than that in the two-dimensional culture,the gene expression levels of AKT and CSN3 were significantly higher than in the control group;In the three-dimensional culture mode,the expression of milk protein,milk fat and hormone-related receptor genes of the experimental group was significantly higher than that of two-dimensional culture.The result show that concentrations of acetic acid,?-hydroxybutyrate,oleic acid,linoleic acid,linolenic acid,PRL,INS,GH was 9.6mmol/L,4.8mmol/L,17.3?mol/L,115.03?mol/L,8.65?mol/L,50nmol/L,5nmol/L,15nmol/L,respectively.which can have a good promotive effect on the expression levels of milk protein and milk fat synthesis-related genes in BMECs.