谷氨酰胺缓解断奶犊牛断奶应激及其机理研究

发布时间：2018-09-05 06:29

【摘要】：本论文通过在断奶犊牛日粮中添加过瘤胃谷氨酰胺(Gin),来探讨Gin对断奶犊牛生长性能、免疫能力、抗氧化能力、小肠肠道结构、肝细胞和肠道上皮细胞自噬的影响,从细胞水平来研究Gin缓解犊牛断奶应激的作用机理。试验一Gin对断奶前后犊牛生长性能和生理代谢指标的影响选择75头出生日期、体重相近的公犊牛进行饲喂试验,将犊牛随机分为5组：对照组饲喂常规日粮,4个处理组分别添加日采食量1%、2%、3%和4%的过瘤胃Gin(谷氨酰胺含量为50%,过瘤胃率为75%,小肠释放率为87%),试验期为断奶前后2周,共4周时间,研究Gin对断奶前后犊牛生长性能和生理代谢指标的影响。结果表明：(1)断奶显著了降低犊牛的日增重(P0.05),显著增加了犊牛的干物质采食量和饲料转化效率(P0.05)：添加不同水平的Gin极显著提高了断奶犊牛的日增重和干物质采食量(P0.01),对饲料转化效率影响不显著(P0.05)。添加1%Gin的处理组,犊牛的日增重、干物质采食量和饲料转化效率最高。(2)断奶能显著降低犊牛的血糖和Gin浓度,显著升高血液中尿素氮浓度(P0.05)：添加不同水平的Gin对断奶犊牛血糖浓度没有显著影响,但能显著增加血液中Gin和尿素氮浓度(P0.05)。(3)断奶显著升高犊牛血液中生长激素浓度,降低血液中的胰岛素浓度(P0.01)；添加不同水平Gin对断奶犊牛血液中胰岛素和生长激素浓度没有显著性影响(P0.05)。试验二Gin对断奶前后犊牛免疫和抗氧化能力的影响试验牛的选择和处理同试验一,结果表明：(1)犊牛断奶显著降低了外周血中CD3+T淋巴细胞比例和CD14+外周单核细胞比例(P0.05)。饲喂不同水平的Gin增加了犊牛外周血中CD4+、CD8+T淋巴细胞比例、CD14+外周单核细胞比例和CD4+/CD8+的比值,但没有显著性差异(P0.05)。(2)犊牛断奶显著降低了血液中IgG含量,对IgA和IgM含量没有显著影响。饲喂不同水平的Gin显著增加了犊牛血液中IgG和IgM含量(P0.05),对血液中1gA含量没有显著性影响(P0.05)。(3)断奶显著降低了犊牛血液中GSH-Px活性和T-AOC (P0.05),对CAT、T-SOD.MDA活性没有显著影响(P0.05)。饲喂不同水平的Gin极显著增加了犊牛血液中GSH-Px活性和T-AOC (P0.01),饲喂不同水平的Gin增加了犊牛血液中CAT和T-SOD活性,但没有显著性差异(P0.05)。试验三Gin对断奶犊牛小肠肠道结构的影响每组选出6头犊牛在70日龄屠宰,取十二指肠、空肠和回肠组织样品并在10%甲醛溶液中固定,病理切片检测十二指肠、空肠和回肠的绒毛高度、隐窝深度及绒毛高度/隐窝深度。结果表明：饲喂不同水平的Gin对断奶犊牛十二指肠、空肠和回肠的绒毛高度、隐窝深度及绒毛高度/隐窝深度比值没有显著影响(P0.05)。饲喂不同水平Gin均增加了断奶犊牛空肠和回肠的绒毛高度和隐窝深度,但没有显著性差异(P0.05)。在饲喂1%Gin时,断奶犊牛十二指肠、空肠和回肠的绒毛高度,隐窝深度及绒毛高度/隐窝深度比值最大。试验四Gin对断奶犊牛肝细胞和肠道上皮细胞自噬的影响每组选出6头犊牛在70日龄屠宰,取肝脏样品和空肠样品用western blotting检测肝细胞和肠道上皮细胞自噬情况。结果表明添加不同梯度的Gin均能提高犊牛肝细胞自噬体水平,其中添加3%的Gin能显著提高犊牛肝细胞细胞自噬体水平(P0.05)。添加不同梯度的Gin均能显著降低犊牛肠道上皮细胞自噬体水平(P0.05),其中添加1%的Gin对降低犊牛肠道上皮细胞自噬体水平的影响最显著。添加Gin可通过PI3K/Akt信号通路、S6K1信号通路和MAPK信号通路调节肝脏和肠道上皮细胞的自噬水平。
[Abstract]:In this paper, the effects of Gin on growth performance, immunity, antioxidant capacity, intestinal structure, hepatocyte and intestinal epithelial autophagy of weaned calves were studied by adding rumen glutamine (Gin) to the diet of weaned calves. The effects of growth performance and physiological and metabolic indexes on the growth performance of post-calves were studied by feeding 75 male calves with similar birth dates and weights. The calves were randomly divided into 5 groups: control group was fed with conventional diet, and 4 groups were fed with 1%, 2%, 3% and 4% perruminal Gin (50% glutamine content, 75% transruminal rate, small intestinal release rate) respectively. The results showed that: (1) Weaning significantly decreased daily gain (P 0.05), significantly increased dry matter intake and feed conversion efficiency (P 0.05). Gin supplementation at different levels significantly increased feed conversion efficiency (P The daily gain, dry matter intake and feed conversion efficiency of weaned calves were the highest in the treatment group with 1% Gin. (2) Weaning significantly reduced blood sugar and Gin concentration, significantly increased blood urea nitrogen concentration (P 0.05): Gin at the same level had no significant effect on blood glucose concentration of weaned calves, but could significantly increase the concentrations of Gin and urea nitrogen in blood (P 0.05). (3) Weaning significantly increased the concentration of growth hormone in blood and decreased the concentration of insulin in blood (P 0.01); Gin at different levels had no significant effect on the concentration of insulin and growth hormone in blood of weaned calves. The results showed that: (1) Weaning significantly reduced the proportion of CD3 + T lymphocytes in peripheral blood and the ratio of CD14 + peripheral mononuclear cells (P 0.05). Feeding different levels of Gin increased the percentage of CD in peripheral blood of calves. 4 +, CD8 + T lymphocyte ratio, CD14 + peripheral mononuclear cell ratio and CD4 + / CD8 + ratio, but there was no significant difference (P 0.05). (2) Weaning significantly reduced the blood IgG content, but had no significant effect on IgA and IgM content. Gin at different levels significantly increased the blood IgG and IgM content of calves (P 0.05), but did not affect the blood 1gA content. (3) Weaning significantly decreased GSH-Px activity and T-AOC (P 0.05), but had no significant effect on CAT, T-SOD. MDA activity (P 0.05). Gin at different levels significantly increased GSH-Px activity and T-AOC (P 0.01), and Gin at different levels increased CAT and T-SOD activity in blood of calves, but not at different levels. There was no significant difference (P 0.05). Six calves were slaughtered at 70 days of age in each group. Duodenum, jejunum and ileum tissue samples were fixed in 10% formaldehyde solution. The villus height, crypt depth and villus height / crypt depth of duodenum, jejunum and ileum were detected by pathological section. The results showed that different levels of Gin had no significant effect on the villus height, crypt depth and the ratio of villus height to crypt depth in duodenum, jejunum and ileum of weaned calves (P 0.05). Gin at different levels increased the villus height and crypt depth of jejunum and ileum of weaned calves, but there was no significant difference (P 0.05). The highest ratio of villus height, crypt depth and villus height/crypt depth was found in duodenum, jejunum and ileum of weaned calves fed with 1% Gin. Experiment 4 Gin affected the autophagy of liver cells and intestinal epithelial cells in weaned calves. Six calves were slaughtered at 70 days of age. Liver and jejunum samples were taken for detection of liver fineness by Western blotting. The results showed that different gradients of Gin could increase the autophagy level of calf hepatocytes, and the addition of 3% Gin could significantly increase the autophagy level of calf hepatocytes (P 0.05). The addition of different gradients of Gin could significantly reduce the autophagy level of calf intestinal epithelial cells (P 0.05), and the addition of 1% Gin could significantly increase the autophagy level of calf hepatocytes (P 0.05). Gin supplementation could regulate the autophagy level of intestinal epithelial cells in calves through PI3K/Akt signaling pathway, S6K1 signaling pathway and MAPK signaling pathway.
【学位授予单位】：中国农业大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S823.5

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