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酶原，并包装在专门的分泌颗粒中，用于胞吐进入胰管。在胰腺炎期间，蛋白水解酶原通过与溶酶体中存在的水解酶异常混合而在腺泡细胞中过早激活。由于腺泡细胞膜运输的致病性改变，溶酶体水解酶与酶原的混合发生在大的胞质空泡中。已经确定，应激诱导的腺泡中热休克蛋白-70（HSP 70）的表达提供了对溶酶体和分泌颗粒膜室的致病性混合的天然保护作用。本研究旨在阐明HSP 70抑制腺泡细胞内酶原激活预防急性胰腺炎发病的机制。HSP 70是胞质溶胶中的分子伴侣，其需要辅伴侣蛋白来刺激ATP酶活化和底物结合。我们已经1）在腺泡细胞中鉴定了一种称为半胱氨酸串蛋白（CSP）的HSP 70共伴侣蛋白，2）将这种膜相关分子定位于整个分泌途径中，3）开发了TAT融合蛋白系统来操纵CSP在腺泡细胞中的表达，从而研究其功能。我们推测CSP通过将HSPT 0分子伴侣活性从细胞质转移到分泌途径来介导HSP 70对胰腺炎的保护作用。在特定目标1中，不再锚于分泌颗粒膜的CSP的突变形式将在腺泡细胞中过表达，以确定CSP在胰腺炎期间将HSP 70靶向这些细胞器的重要性。在特异性目的II中，将使用短干扰RNA来抑制胰腺小叶中CSP的表达，并将评估HSP 70对促分泌素诱导的腺泡细胞损伤的保护作用。这些研究将使我们能够直接评估CSP在生理和病理生理条件下腺泡分泌途径中的作用。

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.