Cysteine String Protein and Acute Pancreatitis

描述（由申请人提供）：急性胰腺炎是一种炎症性疾病，由腺泡细胞中蛋白水解酶原的过早激活引发。作为一种保护机制，酶原被合成为无活性的前酶，并被包装在专门的分泌颗粒中，最终分泌到胰管中。在胰腺炎期间，由于溶酶体中的水解酶与蛋白水解酶的异常混合，在腺泡细胞中蛋白水解酶被过早激活。溶酶体水解酶与酶原的混合发生在大细胞质空泡中，这是由于腺泡细胞膜运输的致病性改变。已经证实，应激诱导的热休克蛋白70 （HSP70）在腺泡中表达对溶酶体和分泌颗粒膜区室的致病性混合具有天然的保护作用。本研究旨在阐明HSP70抑制腺泡内细胞酶原激活以预防急性胰腺炎发作的机制。HSP70是细胞质溶胶中的分子伴侣，需要一种共伴侣蛋白来刺激atp酶激活和底物结合。我们在腺泡细胞中发现了一种HSP70共伴侣蛋白，称为半胱氨酸串蛋白（CSP）， 2)在整个分泌途径中定位了这种膜相关分子，3)开发了tat融合蛋白系统来控制CSP在腺泡细胞中的表达，从而研究其功能。我们假设CSP通过从细胞质到分泌途径靶向hsp0伴侣活性来介导HSP70对胰腺炎的保护作用。在Specific Aim 1中，不再锚定在分泌颗粒膜上的CSP突变形式将被腺泡细胞过度表达，以确定CSP在胰腺炎期间靶向HSP70到这些细胞器中的重要性。在Specific Aim II中，将使用短干扰rna抑制胰腺小泡中CSP的表达，并评估HSP70对分泌剂诱导的腺泡细胞损伤的保护作用。这些研究将使我们能够在生理和病理生理条件下直接评价CSP在腺泡细胞分泌途径中的作用。

DESCRIPTION (provided by applicant): Acute pancreatitis is an inflammatory disease that is triggered by the premature activation of proteolytic zymogens in acinar cells. As a protective mechanism, zymogens are synthesized as inactive pro-enzymes and packaged in specialized secretory granules destined for exocytosis into the pancreatic duct. During pancreatitis, proteolytic zymogens are prematurely activated in acinar cells by aberrantly mixing with hydrolases present in lysosomes. The mixing of lysosomal hydrolases with zymogens occurs in large cytoplasmic vacuoles as a result of pathogenic alterations in acinar cell membrane trafficking. It is well established that the stress-induced expression of heat shock protein-70 (HSP70) in acini provides a natural protective effect against the pathogenic mixing of lysosomal and secretory granule membrane compartments. This proposal is designed to elucidate the mechanism by which HSP70 inhibits the intra-acinar cell activation of zymogens in preventing the on-set of acute pancreatitis. HSP70 is a molecular chaperone in the cytosol that requires a co-chaperone protein to stimulate ATPase activation and substrate binding. We have 1) identified an HSP70 co-chaperone protein called cysteine string protein (CSP) in acinar cells, 2) localized this membrane associated molecule throughout the secretory pathway and 3) developed the TAT-fusion protein system to manipulate CSP expression in acinar cells and thereby study its function. We hypothesize that CSP mediates the protective effects of HSP70 against pancreatitis by targeting HSPT0 chaperone activity from the cytosol to the secretory pathway. In Specific Aim 1, mutant forms of CSP that no longer anchor to secretory granule membranes will be overexpressed acinar cells to determine the importance of CSP in targeting HSP70 to these organelles during pancreatitis. In Specific Aim II, short interference RNAs will be used to inhibit CSP expression in pancreatic Iobules and the protective effects of HSP70 on secretagogue-induced acinar cell damage will be evaluated. These studies will allow us to directly evaluate the role of CSP in the secretory pathway of acini under physiological and pathophysiological conditions.