賴氨酸蛋氨酸配比模式對奶牛乳腺上皮細(xì)胞酪蛋白合成的影響

趙圣國 李喜艷 王加啟

摘 要：以體外培養(yǎng)的奶牛乳腺上皮細(xì)胞為模型，研究賴氨酸蛋氨酸配比模式對體外培養(yǎng)的奶牛乳腺上皮細(xì)胞酪蛋白合成的影響。試驗(yàn)采用因子試驗(yàn)設(shè)計(jì)，以蛋氨酸（0.4、0.5、0.6 mmol/L）和賴氨酸（1.0、1.2、1.4、1.5、1.6 mmol/L）的混合物作為試驗(yàn)處理。試驗(yàn)期為48 h，每個(gè)處理3個(gè)重復(fù)，試驗(yàn)重復(fù)3次。通過ELISA方法檢測添加蛋氨酸和賴氨酸混合物后細(xì)胞中酪蛋白的合成量。得出最佳的賴氨酸蛋氨酸配比模式，并采用Real-Time PCR技術(shù)檢測此水平下乳蛋白基因、與乳蛋白合成轉(zhuǎn)錄和翻譯相關(guān)的基因以及與賴氨酸、蛋氨酸相關(guān)的轉(zhuǎn)運(yùn)載體CAT-1和ASCT-2的表達(dá)。結(jié)果表明：蛋氨酸（0.4 mmol/L）和賴氨酸（1.2 mmol/L）混合添加時(shí)奶牛乳腺上皮細(xì)胞總酪蛋白合成量最高為2.95 ppm，并差異極顯著。當(dāng)賴氨酸和蛋氨酸以 1.2 mmol/L和0.4 mmol/L即3∶1混合添加時(shí)在不影響賴氨酸、蛋氨酸相關(guān)的轉(zhuǎn)運(yùn)載體CAT-1和ASCT-2基因表達(dá)的條件下均能極顯著地促進(jìn)乳蛋白基因的表達(dá)如：CSN1S1、CSN1S2、CSN2、CSN3和LALBA基因表達(dá)均上調(diào)，且CSN1S1和LABA表達(dá)量最高。與乳蛋白轉(zhuǎn)錄相關(guān)的STAT5-JAK2信號通路基因的表達(dá)均顯著上調(diào)；與乳蛋白翻譯相關(guān)的mTOR信號通路的相關(guān)基因除EIF4E-BP1顯著下調(diào)，mTOR和S6K基因的表達(dá)均上調(diào)且mTOR差異極顯著。

關(guān)鍵詞：賴氨酸 蛋氨酸 酪蛋白 合成

Effect of Ratio between Lysine and Methionine on Casein Synthesis in Bovine Mammary Epithelial Cells

Zhao Shengguo Li Xiyan Wang Jiaqi

（Institute of Animal Science of CAAS）

Abstract：The experiment was conducted with a 5×3 factorial design： with lysine （1.0， 1.2， 1.4， 1.5， 1.6 mmol/L in DMEM/F12 medium） and methionine （0.4， 0.5， 0.6 mmol/L in DMEM/F12 medium）. The expression of important genes related to milk protein synthesis process included the genes of the JAK2-STAT5 pathway and the mTOR signaling pathway. All experiments repeated 3 times. The expression of genes and caseins content were detected by RT-qPCR method and Casein ELISA Kit， respectively. Data were analyzed with the PROC GLM and PROC ANOVA. The results showed that when the concentration of lysine was 1.2 mmol/L and the concentration of methionine was 0.4 mmol/L， the content of casein in medium increased significantly and peaked at 2.95 ppm. We measured the genes expression at this ratio （3∶1）.CSN1S1 and LALBA were the most highly expressed genes encoding caseins and increased more than 2-fold，followed by CSN2， CSN3 and CSN2S2， and these genes expression were also up-regulated. The expression of genes related to JAK2-STAT5 pathway were up-regulated， and JAK2 and ELF5 expression increased significantly. The expression of genes related to mTOR signaling pathway was increased， but S6K was not significantly. EIF4E-BP1 expression was significantly down-regulated. The expression of amino acid transporter CAT-1 and ASCT-2 had no change.

Key Words：Lysine；Methionine；Casein；Synthesis

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