【摘要】：本論文通過在斷奶犢牛日糧中添加過瘤胃谷氨酰胺(Gin),來探討Gin對斷奶犢牛生長性能、免疫能力、抗氧化能力、小腸腸道結(jié)構(gòu)、肝細(xì)胞和腸道上皮細(xì)胞自噬的影響,從細(xì)胞水平來研究Gin緩解犢牛斷奶應(yīng)激的作用機(jī)理。試驗(yàn)一Gin對斷奶前后犢牛生長性能和生理代謝指標(biāo)的影響選擇75頭出生日期、體重相近的公犢牛進(jìn)行飼喂試驗(yàn),將犢牛隨機(jī)分為5組：對照組飼喂常規(guī)日糧,4個(gè)處理組分別添加日采食量1%、2%、3%和4%的過瘤胃Gin(谷氨酰胺含量為50%,過瘤胃率為75%,小腸釋放率為87%),試驗(yàn)期為斷奶前后2周,共4周時(shí)間,研究Gin對斷奶前后犢牛生長性能和生理代謝指標(biāo)的影響。結(jié)果表明：(1)斷奶顯著了降低犢牛的日增重(P0.05),顯著增加了犢牛的干物質(zhì)采食量和飼料轉(zhuǎn)化效率(P0.05)：添加不同水平的Gin極顯著提高了斷奶犢牛的日增重和干物質(zhì)采食量(P0.01),對飼料轉(zhuǎn)化效率影響不顯著(P0.05)。添加1%Gin的處理組,犢牛的日增重、干物質(zhì)采食量和飼料轉(zhuǎn)化效率最高。(2)斷奶能顯著降低犢牛的血糖和Gin濃度,顯著升高血液中尿素氮濃度(P0.05)：添加不同水平的Gin對斷奶犢牛血糖濃度沒有顯著影響,但能顯著增加血液中Gin和尿素氮濃度(P0.05)。(3)斷奶顯著升高犢牛血液中生長激素濃度,降低血液中的胰島素濃度(P0.01)；添加不同水平Gin對斷奶犢牛血液中胰島素和生長激素濃度沒有顯著性影響(P0.05)。試驗(yàn)二Gin對斷奶前后犢牛免疫和抗氧化能力的影響試驗(yàn)牛的選擇和處理同試驗(yàn)一,結(jié)果表明：(1)犢牛斷奶顯著降低了外周血中CD3+T淋巴細(xì)胞比例和CD14+外周單核細(xì)胞比例(P0.05)。飼喂不同水平的Gin增加了犢牛外周血中CD4+、CD8+T淋巴細(xì)胞比例、CD14+外周單核細(xì)胞比例和CD4+/CD8+的比值,但沒有顯著性差異(P0.05)。(2)犢牛斷奶顯著降低了血液中IgG含量,對IgA和IgM含量沒有顯著影響。飼喂不同水平的Gin顯著增加了犢牛血液中IgG和IgM含量(P0.05),對血液中1gA含量沒有顯著性影響(P0.05)。(3)斷奶顯著降低了犢牛血液中GSH-Px活性和T-AOC (P0.05),對CAT、T-SOD.MDA活性沒有顯著影響(P0.05)。飼喂不同水平的Gin極顯著增加了犢牛血液中GSH-Px活性和T-AOC (P0.01),飼喂不同水平的Gin增加了犢牛血液中CAT和T-SOD活性,但沒有顯著性差異(P0.05)。試驗(yàn)三Gin對斷奶犢牛小腸腸道結(jié)構(gòu)的影響每組選出6頭犢牛在70日齡屠宰,取十二指腸、空腸和回腸組織樣品并在10%甲醛溶液中固定,病理切片檢測十二指腸、空腸和回腸的絨毛高度、隱窩深度及絨毛高度/隱窩深度。結(jié)果表明：飼喂不同水平的Gin對斷奶犢牛十二指腸、空腸和回腸的絨毛高度、隱窩深度及絨毛高度/隱窩深度比值沒有顯著影響(P0.05)。飼喂不同水平Gin均增加了斷奶犢牛空腸和回腸的絨毛高度和隱窩深度,但沒有顯著性差異(P0.05)。在飼喂1%Gin時(shí),斷奶犢牛十二指腸、空腸和回腸的絨毛高度,隱窩深度及絨毛高度/隱窩深度比值最大。試驗(yàn)四Gin對斷奶犢牛肝細(xì)胞和腸道上皮細(xì)胞自噬的影響每組選出6頭犢牛在70日齡屠宰,取肝臟樣品和空腸樣品用western blotting檢測肝細(xì)胞和腸道上皮細(xì)胞自噬情況。結(jié)果表明添加不同梯度的Gin均能提高犢牛肝細(xì)胞自噬體水平,其中添加3%的Gin能顯著提高犢牛肝細(xì)胞細(xì)胞自噬體水平(P0.05)。添加不同梯度的Gin均能顯著降低犢牛腸道上皮細(xì)胞自噬體水平(P0.05),其中添加1%的Gin對降低犢牛腸道上皮細(xì)胞自噬體水平的影響最顯著。添加Gin可通過PI3K/Akt信號通路、S6K1信號通路和MAPK信號通路調(diào)節(jié)肝臟和腸道上皮細(xì)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.
【學(xué)位授予單位】：中國農(nóng)業(yè)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2016
【分類號】：S823.5

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