title: Leucine and histidine independently regulate milk protein synthesis in bovine mammary epithelial cells via mTOR signaling pathway.
abstract: The aim of this study is to investigate the effects of leucine (Leu) and histidine (His) on the expression of both the mammalian target of rapamycin (mTOR) signaling pathway-related proteins and caseins in immortalized bovine mammary epithelial cells (CMEC-H), using a single supplement through Western blotting. The Earle's balanced salt solution (EBSS) was set as the control group and other treatment groups, based on the EBSS, were added with different concentrations of Leu or His, respectively. The results showed that, compared with the control group, the expression of caseins and the phosphorylation of mTOR (Ser(2481)), Raptor (Ser(792)), eIF4E (Ser(209)), and eEF2 (Thr(56)) increased with the Leu concentrations ranging from 0.45 to 10.80 mmol/L (P<0.01). The P-4EBP1 (Thr(37)) at 10.80 mmol/L Leu, and P-RPS6 (Ser(235/236)) at 5.40 to 10.80 mmol/L Leu all decreased. Similarly, the His supplementation from 0.15 to 9.60 mmol/L increased the expression of αs2-casein, β-casein, κ-casein, P-mTOR (Ser(2481)), P-Raptor (Ser(792)), P-S6K1 (Thr(389)), P-4EBP1 (Thr(37)), P-eIF4E (Ser(209)), and P-eEF2 (Thr(56)) (P<0.01) in CMEC-H, whereas the αs1-casein expression was only reduced at 9.60 mmol/L His, G protein β subunit-like protein (GβL) at 0.15 and 9.60 mmol/L His, and P-RPS6 at 4.80 to 9.60 mmol/L His. Our linear regression model assay suggested that the αs1-casein expression was positively correlated with P-mTOR (P<0.01), P-S6K1 (P<0.01), and P-eEF2 (P<0.01) for the addition of Leu, while the expressions of β-casein (P<0.01) and κ-casein (P<0.01) were positively correlated with P-eEF2 for the addition of His. In conclusion, the milk protein synthesis was up-regulated through activation of the mTOR pathway with the addition of Leu and His in CMEC-H.
authors.affiliation: Ministry of Agriculture-Milk Risk Assessment Laboratory, Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193, China; College of Animal Science and Technology, Gansu Agricultural University, Lanzhou 730070, China; Ministry of Agriculture-Milk and Dairy Product Inspection Center, Beijing 100193, China; State Key Laboratory of Animal Nutrition, Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
authors.forename: Hai-na
authors.initials: HN
authors.lastname: Gao
authors.forename: Han
authors.initials: H
authors.lastname: Hu
authors.forename: Nan
authors.initials: N
authors.lastname: Zheng
authors.forename: Jia-qi
authors.initials: JQ
authors.lastname: Wang
mesh_terms: D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology; D006639:Histidine / Q000008:administration & dosage*; D007930:Leucine / Q000008:administration & dosage*; D008321:Mammary Glands, Animal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008894:Milk Proteins / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology
chemical_list: D008894:Milk Proteins; D006639:Histidine; D058570:TOR Serine-Threonine Kinases; D007930:Leucine
keywords: Bovine mammary epithelial cells; Casein; Histidine; Leucine; Western blotting; mTOR