NUTRIENT REGULATION OF SUCRASE A-DEXTRINASE MATURATION

描述(改编自调查员的摘要和/或目标)： 大鼠胃肠道经历了剧烈的成熟变化 在生命的第三周导致细胞的重大变化 动力学和酶活性。蔗糖酶-α-糊精酶，一种糖蛋白 最终吸收膳食碳水化合物所需的水解酶 在肠细胞的刷状缘膜表面，在 生命的第三周，当它突然出现并迅速上升时，如 出生后肠道成熟的主要特征。虽然 调节S-D成熟的机制尚不清楚，有明确的 充分成熟所需鲁米那营养素的证据 S-D的表达。申请人开发的独一无二的动物模型 现在已经很成熟了，将与高级 细胞和分子生物学技术，以研究 营养依赖型S-D的成熟机制。初步研究将 测定鲁米那碳水化合物对体内合成的影响， S-D在成熟肠道内的组装和降解 新生S-D多肽的腔内放射性标记及其研究 酶在其合成、组装和最终插入的不同阶段 进入灌木丛边缘。这些研究将确定动力学是否 关于细胞内转运，S-D前体的结构或 鲁米那碳水化合物改变了S-D的降解。后续 研究将确定鲁米那碳水化合物是否调节S-D 通过转录或翻译成熟通过测量特定的mRNAs 用申请人研制的聚合酶链式cDNADNA探针检测S-D 聚合酶链式反应技术及细胞内S-D结构分析 使用糖苷酶探针的组装和插入后处理， S-D的双向凝胶电泳法和氨基酸序列测定 蛋白。为了确定管腔营养素是否直接作用于 肠细胞，S-D对肠道营养物质的反应 沿隐窝绒毛轴和器官培养的吸收细胞将 学习。这些在稳态条件下进行的研究将 辅以S-D急性诱导的类比研究 申请人开发的肠道灌流技术用于 以确定是否单一的机制或 多种机制的组合参与了S-D的反应。这些研究 将产生新的信息，关于管腔内 营养物质调节S-D在出生后肠道发育过程中的成熟。

DESCRIPTION (adapted from investigator's abstract and/or aims): The gastrointestinal tract of the rat undergoes dramatic maturational changes during the third week of life resulting in major changes in cellular kinetics and enzyme activities. Sucrase-alpha-dextrinase, a glycoprotein hydrolase required for final absorption of dietary carbohydrate and located at the brush border membrane surface of the enterocyte, is absent prior to the third week of life when it abruptly appears and rises rapidly, as a major distinguishing feature of postnatal intestinal maturation. Although the mechanisms regulating S-D maturation are not known, there is clear evidence that luminal nutrients are required for full maturational expression of S-D. A unique animal model developed by the applicant which is now well established will be used in combination with advanced techniques of cellular and molecular biology in order to investigate the mechanisms of nutrient-dependent S-D maturation. Initial studies will determine the effect of luminal carbohydrate on the in-vivo synthesis, assembly and degradation of S-D in the maturing intestine using techniques of intraluminal radiolabeling of the nascent S-D peptide and investigating the enzyme at various stages of its synthesis, assembly and final insertion into the brush border. These studies will determine whether the kinetics of intracellular transport, the structure of S-D precursors or the degradation of S-D are altered by luminal carbohydrates. Subsequent studies will determine whether luminal carbohydrate regulates S-D maturation by transcription or translation by measuring specific mRNA for S-D using cDNA probes developed by the applicant using polymerase chain reaction (PCR) techniques and analyzing S-D structure, intracellular assembly and post-insertional processing using glycosidase probes, 2-dimensional gel electrophoresis and amino acid sequencing of the S-D protein. In order to determine whether luminal nutrients act directly on the enterocyte, the S-D response to luminal nutrients of the intestinal absorptive cells along the crypt-villus axis and in organ culture will be studied. These studies performed during steady-state conditions will be complemented by analogous studies of acute induction of S-D using intestinal perfusion techniques developed by the applicant for use in infant animals in order to determine whether a single mechanism or combinations of mechanisms participate in the S-D response. These studies will generate new information on the mechanisms by which intraluminal nutrients regulate S-D maturation during postnatal intestinal development.