Post-Transcriptional Processing of the Small Intestinal Alkaline Phosphatase in the Postnatal Developing Pig

猪的全身能量和营养物质利用效率在出生时和哺乳期间较高，然后在断奶过渡期间急剧下降，并在断奶后生长期间变得较低。断奶相关的肠道疾病和生长迟缓是当前全球养猪业面临的主要挑战。目前使用饲料抗生素来控制断奶仔猪营养中的发病率和死亡率，导致与抗生素耐药性相关的全球公共卫生威胁不断上升。我的NSERC发现计划的长期目标是了解小肠水解酶和营养转运蛋白的表达和调节因子，它们影响沿着隐窝-绒毛轴和猪肠道发育期间的肠道功能。断奶标志着哺乳动物肠道从哺乳到固体饮食喂养的主要适应。肠碱性磷酸酶（IAP）是一种丰富的顶端膜蛋白，其主要生理功能是解毒细菌内毒素脂多糖。因此，IAP的表达在保护肠道免受炎症和感染以及维持全身健康状态方面至关重要。早期断奶通过降低IAP基因在转录和转录后水平的表达来抑制IAP最大酶活性，从而显著降低仔猪中顶端IAP的丰度和亲和力。肠内氨基酸（AA）的利用率和膳食益生元可以调节IAP的表达和功能。我假设，由于营养AA的饮食供应增加，低蛋白饮食喂养的断奶仔猪中IAP表达、功能和肠道健康状态可以通过IAP生物合成和IAP糖基化的转录后调节而显著增强，即，结晶谷氨酸盐、谷氨酰胺、亮氨酸和选择的可发酵的非粘性可溶性纤维作为益生元。拟议的研究的具体目标是调查的作用，增加这些营养膳食AA的可用性和改变选定的非粘性可溶性纤维组合物在调节空肠IAP分数合成率和IAP功能，并支持两个研究生进行拟议的研究。主要的研究方法包括IAP动力学测定显色和纯化的生理底物，在体内IAP分数合成率同位素标记，IAP糖基化免疫沉淀和化学分析，和肠道蛋白质翻译途径的启动和延伸因子丰度的蛋白质印迹。拟议的研究将有助于提高对上调肠道内源性碱性磷酸酶表达和功能的饮食因素的理解，并有助于开发改善断奶仔猪肠道健康的替代策略。

Efficiency of whole body energy and nutrient utilization is high at birth and during suckling, then drops dramatically during the weanling transition and becomes low during the post-weaning growth in pigs. Weaning-associated enteric diseases and growth retardation are major challenges facing the current global swine production. Current use of feed antibiotics for curbing morbidity and mortality in weanling swine nutrition contributes to the rising global public health threat associated with antibiotic resistance. Long-term goals of my NSERC Discovery program aim to understand the expression and regulatory factors of the small intestinal hydrolases and nutrient transporters in affecting gut functions along the crypt-villus axis and during the gut development in pigs. Weaning marks a major intestinal adaptation from suckling to feeding on solid diets in mammals. Intestinal alkaline phosphatase (IAP) is an abundant apical membrane protein with a primary physiological function of detoxification of bacterial endotoxin lipopolysaccharides. Thus, expression of IAP is pivotal in protecting the gut from inflammation and infection and in maintaining whole body health status. Early weaning dampens the IAP maximal enzyme activity by reducing the IAP gene expression at the transcriptional and post-transcriptional levels with considerable decreases in the apical IAP abundance and affinity in piglets. Enteral amino acid (AA) availability and dietary prebiotics could modulate IAP expression and functionality. I hypothesize that IAP expression, functionality and enteral health status in weanling piglets fed a low-protein diet could be substantially enhanced by post-transcriptional regulation of IAP biosynthesis and IAP glycosylation because of increases in dietary supply of trophic AA, i.e., crystalline glutamate, glutamine, leucine and selected fermentable non-viscous soluble fibres serving as prebiotics. Specific objectives for the proposed research are to investigate roles of increasing these trophic dietary AA availability and changing the selected non-viscous soluble fibre composition in regulating the jejunal IAP fractional synthesis rate and IAP functionality, and in supporting two graduate students to conduct the proposed research. Major methodology for the proposed research includes IAP kinetics measured with chromogenic and purified physiological substrates, in vivo IAP fractional synthesis rate by isotopic labeling, IAP glycosylation by immune-precipitation and chemical profiling, and intestinal protein translational pathway initiation and elongation factor abundances by Western blotting. The proposed research will contribute to improved understanding of dietary factors in up-regulating intestinal endogenous alkaline phosphatase expression and functionality and the development of alternative strategies to improve gut health in weanling pigs.