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

猪的全身能量和养分利用效率在出生时和哺乳期间都很高，然后在断奶过渡阶段急剧下降，在断奶后生长阶段变得很低。断奶相关的肠道疾病和生长迟缓是当前全球生猪生产面临的主要挑战。目前使用饲料抗生素来控制断奶仔猪营养方面的发病率和死亡率，加剧了与抗生素耐药性相关的全球公共卫生威胁。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在猪隐窝绒毛轴和肠道发育过程中影响肠道功能的表达和调控因素。断奶标志着哺乳动物从哺乳到以固体饲料喂养的主要肠道适应。肠道碱性磷酸酶(IAP)是一种含量丰富的顶膜蛋白，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起着关键作用。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的最大酶活性，并显著降低仔猪顶端IAP的丰度和亲和力。肠道氨基酸(AA)利用率和日粮益生素可以调节IAP的表达和功能。我推测，饲喂低蛋白日粮的断奶仔猪的IAP表达、功能和肠道健康状况可以通过IAP生物合成和IAP糖基化的转录后调控而显著提高，因为日粮中营养氨基酸的供应增加，即结晶谷氨酸、谷氨酰胺、亮氨酸和精选的可发酵非粘性可溶性纤维作为益生元。建议研究的具体目标是探讨增加这些营养饲料AA的利用率和改变选定的非粘性可溶性纤维成分在调节空肠IAP组分合成速率和IAP功能方面的作用，以及支持两名研究生进行建议研究的作用。研究的主要方法包括：发色和纯化生理底物测定IAP动力学，同位素标记测定体内IAP合成速率，免疫沉淀和化学图谱测定IAP糖基化，Western blotting测定肠道蛋白翻译途径启动和延伸因子的丰度。这项拟议的研究将有助于更好地了解日粮因素上调肠道内源性碱性磷酸酶的表达和功能，并开发替代策略来改善断奶仔猪的肠道健康。

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.