淀粉消化位点重分配调控奶牛糖代谢途径的分子机制

It is a long held belief that feeding starch in forms that increase digestion in the small intestine and glucose absorption will benefit the cows in terms of energetic efficiency and production response, but energy corrected milk did not support the conclusion. Two experiments are conducted to investigate the molecular mechanism of site of starch digestion on key hepatic gluconeogenesis and glucose transporters enzymes expression and activities, the effects of glucose, insulin, NEFA and BHBA on the glucose and fat metabolism, the optimal ratio of starch degradation in the rumen for maximum milk production. In experiment 1, eight cows in early lactation fed low starch concentrates are continuously infused into the rumen or abomasum at 1600 g/d corn starch for 21 d in a double 4×4 Latin square design. The treatments based on the infusion ratio of rumen to abomasum are as follows: a）RSAW：1600 g/d:0;（b）RSAS25%: 1200 g/d :400 g/d；（c）RSAS50%： 800 g/d:800 g/d（d）RWAS100%：0:1600 g/d. In experiment 2, eight cows in early lactation fed normal starch concentrates are continuously infused into the abomasum with water or 800, 1600, or 2400 g of corn starch for 14 d in a double 4×4 Latin square design. The glucose tolerance test and insulin tolerance test will be conducted to investigate the insulin resistance and glucose metabolism. The liver and subcutaneous fat in tail head will be analysed the key hepatic gluconeogenesis (PC,PEPCK, FBPase and G6-Pase) enzymes expression and activities. Milk production and composition will be recorded to search the effect of site of starch digestion. Fecal and urinal samples are collected to investigate the digestibility of starch.

通过奶牛真胃精准梯度灌注淀粉技术（通过瘤胃瘘管、无真胃瘘管）、真胃无损采样技术、肝脏活体采样技术和牛奶体细胞检测葡萄糖转运载体技术等，运用分子生物学、营养代谢组学和基因功能芯片检测技术研究淀粉消化位点重分配调控奶牛糖代谢途径的分子机制。重点是糖异生关键酶（PC、PEPCK、FBPase和G6-Pase等）和乳腺葡萄糖转运蛋白（GLUT1等）mRNA表达丰度和活性的变化规律，阐明外源刺激（胰岛素和葡萄糖）及代谢中间产物（NEFA和BHBA）对糖异生的调控机理，为从基因水平和蛋白质水平揭示内源和外源葡萄糖供能机制奠定分子生物学基础和理论依据，为生产中确定降解淀粉和过瘤胃淀粉最佳比例，提高奶牛生产性能和健康水平提供理论依据。

从泌乳奶牛瘤胃降解率、泌乳性能、瘤胃发酵、消化代谢、血液指标、肝脂基因、葡萄糖耐受试验等方面探讨了玉米淀粉能量走向及过瘤胃淀粉的最适添加量。选用6头（3头初产牛，3头经产牛）带有永久性瘤胃瘘管的荷斯坦泌乳奶牛，采用重复3×3拉丁方设计来研究不同位点（瘤胃或真胃）灌注淀粉对泌乳奶牛采食量、消化代谢、泌乳性能、肠道发酵性能、血液代谢、肝脏脂肪基因及葡萄糖耐受试验的影响。试验分为三个处理：对照组不灌注淀粉，瘤胃灌注组向瘤胃内灌注淀粉800 g/d，真胃灌注组向真胃内灌注淀粉800 g/d，分别在试验的第7天和第14天采集样品。研究结果表明：背膘厚度的增加表明了灌注淀粉降低了脂肪组织的动员而刺激了脂质合成，PPAR-γ mRNA的降低和GLUT-4 mRNA的增加都表明了泌乳奶牛的脂质累积。真胃灌注的淀粉直接到达小肠改善能量供应，因此对葡萄糖的耐受能力更强。总体说来，800 g/d淀粉的灌注对泌乳奶牛能量分配无影响，真胃额外供应800 g/d的淀粉能够降低脂质动员，淀粉有进入脂肪合成通路的可能性。.选用4头带有永久性瘤胃瘘管的荷斯坦二胎泌乳奶牛，采用4×4拉丁方设计来研究真胃灌注不同浓度的淀粉对泌乳后期奶牛采食量、消化代谢、泌乳性能、肠道发酵性能、血液代谢、肝脏脂肪基因及葡萄糖耐受试验的影响。淀粉的灌注量为0、800、1600、2400 g/d，每个试验期为16天，分别在试验的第7天和第14天采集样品。研究结果表明：真胃灌注1600 g/d的淀粉，奶牛肝脏中PEPCK-C的mRNA表达（1.45）显著高于对照组（0.82）和2400 g/d淀粉真胃灌注组（0.71），与800 g/d淀粉真胃灌注组差异不显著（1.34），四组奶牛肝脏PEPCK-C的表达呈先上升后下降的变化规律。胰岛素受体mRNA（P = 0.167）和GLUT-4 mRNA（P = 0.116）丰度都随着淀粉灌注量的增加呈现先上升后下降的趋势，且都是800 g/d最高；而PPAR-γ则呈先下降后上升的趋势（P = 0.104），1600 g/d最低。真胃灌注淀粉组的葡萄糖峰浓度较高，有先上升后下降的二次变化趋势（P = 0.115）。800 g/d淀粉真胃灌注奶牛的胰岛素CR值最高，显著高于2400 g/d淀粉真胃灌注的奶牛，CR值随着淀粉真胃灌注量的增加呈先上升后下降的二次关系（P = 0.0