Calcium-dependent enzyme activation and vacuole formation in the apical granular region of pancreatic acinar cells
Calcium-dependent enzyme activation and vacuole formation in the apical granular region of pancreatic acinar cells
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DOI：
10.1073/pnas.97.24.13126
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发表时间：
2000-11-21
影响因子：
11.1
通讯作者：
Petersen, OH
中科院分区：
文献类型：
--
作者：
Raraty, M;Ward, J;Petersen, OH
胰腺腺泡细胞产生强大的消化酶，包装在顶端的酶原颗粒中。由乙酰胆碱或胆囊收缩素（CCK）引起的Ca 2+信号通过顶膜的胞吐作用启动酶分泌。细胞内酶的激活通常保持在最低限度，但在经常致命的人类疾病急性胰腺炎中，会发生自动消化。酶是如何被不适当地激活的尚不清楚。我们监测细胞内Ca 2+浓度（[Ca 2 +]（i）），细胞内胰蛋白酶的激活，并与特定的荧光探针在分离的活细胞中的定位和研究细胞内空泡形成的电子显微镜以及定量图像分析（光学显微镜）。生理CCK水平（10 pM）引发定期Ca 2+尖峰没有引起细胞内胰蛋白酶激活或空泡形成。然而，用10 nM CCK刺激。引起[Ca ~（2+）]（i）的持续升高，诱导明显的胰蛋白酶活化和广泛的空泡形成，两者都位于顶极。这两个过程被取消，通过防止异常[Ca 2 +]（i）升高，无论是通过预孵育与特定的Ca 2+螯合剂1，2-双（O-氨基苯氧基）乙烷-N，N-N '，N'-四乙酸（BAPTA）或通过去除外部Ca 2+。CCK过度刺激引起细胞内胰蛋白酶激活和顶端颗粒极空泡形成。这两个过程都是由[Ca 2 +]（i）的异常持续升高介导的。
The pancreatic acinar cell produces powerful digestive enzymes packaged in zymogen granules in the apical pole. Ca2+ signals elicited by acetylcholine or cholecystokinin (CCK) initiate enzyme secretion by exocytosis through the apical membrane. Intracellular enzyme activation is normally kept to a minimum, but in the often-fatal human disease acute pancreatitis, autodigestion occurs. How the enzymes become inappropriately activated is unknown. We monitored the cytosolic Ca2+ concentration ([Ca2+](i)), intracellular trypsin activation, and its localization in isolated living cells with specific fluorescent probes and studied intracellular vacuole formation by electron microscopy as well as quantitative image analysis (light microscopy). A physiological CCK level (10 pM) eliciting regular Ca2+ spiking did not evoke intracellular trypsin activation or vacuole formation. However, stimulation with 10 nM CCK. evoking a sustained rise in [Ca2+](i), induced pronounced trypsin activation and extensive vacuole formation, both localized in the apical pole. Both processes were abolished by preventing abnormal [Ca2+](i) elevation, either by preincubation with the specific Ca2+ chelator 1,2-bis(O-aminophenoxy)ethane-N,N-N',N'-tetraacetic acid (BAPTA) or by removal of external Ca2+. CCK hyperstimulation evokes intracellular trypsin activation and vacuole formation in the apical granular pole. Both of these processes are mediated by an abnormal sustained rise in [Ca2+](i).