Acinar Biology and Pancreatic Disease

描述（由申请人提供）：外分泌胰腺腺泡细胞制造、储存和释放大量水解酶到肠道中，这些酶对营养物质的消化和吸收至关重要。由于食物来源于细胞物质，腺泡细胞保留了保护机制，以确保消化酶在到达肠道之前不会过早激活。事实上，腺泡分泌途径的异常失调和酶的过早激活与胰腺炎和胰腺癌的发展有关，每年有超过48,000名美国人受到影响。人们普遍认为消化酶的过早激活是由腺泡菌分泌途径和溶酶体途径的异常相互作用引起的。因此，了解协调这些通路正常相互作用的基本分子机制对于理解胰腺疾病的病理生理学至关重要。本研究探讨了一种以前未被认识的重要调控途径，即腺泡细胞通过分泌和溶酶体途径调节消化酶的运输，从而调节消化酶的分泌。我们最近报道了基于胞外调节蛋白VAMP2和8的表达，腺泡细胞表达两种不同的酶原颗粒（ZGs）群体。此外，我们的数据表明，VAMP8 ZGs的成熟是由肿瘤蛋白D52（又名CRHSP-28）指导的，我们已经证明，肿瘤蛋白D52在腺泡细胞中唯一表达，并直接调节Ca2+依赖性分泌。出乎意料的是，D52定位于胞泡中一个独特的内核溶酶体和溶酶体相关的室，我们称之为内核溶酶体室。此外，在CHO-K1细胞中，D52直接调节溶酶小体膜胞吐，当D52过表达时，会引起细胞质囊泡的大量积聚。本研究旨在描述VAMP2-和VAMP8-阳性ZGs在腺泡分泌反应中的确切功能作用，并将验证d52通过内切溶酶体系统调节的运输在一个独特的溶酶体相关途径中控制VAMP8阳性ZGs的生物发生/成熟的中心假设。特异性目的1将验证vamp2阳性ZGs介导消化酶的早期即时释放，而vamp8阳性ZGs介导腺泡刺激后酶原分泌的持续平台期。特异性目标2将验证PIKfyve在腺泡中激活产生PtdIns(3,5)P2的假设，该假设可调节内溶酶体向溶酶体的运输，并抑制腺泡过度刺激时的根尖分泌。特异性Aim 3将验证D52激活“次要调节通路”的假设，即介导重要调节蛋白快速插入颗粒胞外分泌所必需的尖膜。了解腺泡杆菌如何独特地协调分泌或溶酶体室室的基本分子原理是开发治疗胰腺疾病的治疗策略的关键。

DESCRIPTION (provided by applicant): Exocrine pancreatic acinar cells manufacture, store and release large quantities of hydrolytic enzymes into the intestine that are essential for nutrient digestion and absorption. As food is derived from cellular material, acinar cells retain protective mechanisms to ensure digestive enzymes are not prematurely activated until reaching the intestine. Indeed, aberrant dysregulation of the acinar secretory pathway and premature activation of enzymes has been tied to the development of pancreatitis and pancreatic cancer, which affect over 48,000 Americans each year. It is widely believed that premature digestive enzyme activation is caused by abnormal interactions of the secretory and lysosomal pathways in acini. Thus, knowledge of the basic molecular mechanisms which orchestrate the normal interactions of these pathways is critical to understanding the pathophysiology of pancreatic disease. This proposal investigates a previously unrecognized and important regulatory pathway by which acinar cells modulate digestive enzyme trafficking within the secretory and lysosomal pathways and accordingly regulate digestive enzyme secretion. We recently reported that acinar cells express two distinct populations of zymogen granules (ZGs) based on the expression of the exocytic regulatory proteins VAMP2 and 8. Moreover, our data indicate that maturation of VAMP8 ZGs is directed by Tumor Protein D52 (aka CRHSP-28), which we have shown is uniquely expressed in acini and directly regulates Ca2+-dependent secretion. Unexpectedly, D52 was localized to a unique endosome and lysosome related compartment in acini that we term the endo-lysosomal compartment. Moreover, in CHO-K1 cells, D52 directly regulates lysosmal membrane exocytosis and when over-expressed induces a massive accumulation of cytoplasmic vesicles. This proposal aims to delineate the precise functional roles of VAMP2- and VAMP8- positive ZGs in the acinar secretory response and will test the central hypothesis that D52-regulated trafficking through the endo-lysosomal system controls the biogenesis/maturation of VAMP8-positive ZGs within a unique lysosome-related pathway. Specific Aim 1 will test the hypothesis that VAMP2-positive ZGs mediate the early immediate release of digestive enzymes, whereas VAMP8-positive ZGs mediate the sustained plateau phase of zymogen secretion following acinar stimulation. Specific Aim 2 will test the hypothesis that activation of PIKfyve to produce PtdIns(3,5)P2 in acini regulates endo-lysosomal trafficking to the lysosome and inhibits apical secretion during acinar hyperstimulation. Specific Aim 3 will test the hypothesis that D52 activation of the "minor regulated pathway" mediates the rapid insertion of important regulatory proteins into the apical membrane necessary for granule exocytosis. Understanding the basic molecular principles of how acini uniquely orchestrate the secretory or lysosomal compartments is key to the development of therapeutic strategies aimed at treating pancreatic disease.