Abstract
Zymogen secretory granules in pancreatic acinar cells express two vesicle-associated membrane proteins (VAMP), VAMP2 and -8, each controlling 50% of stimulated secretion. Analysis of secretion kinetics identified a first phase (0-2 min) mediated by VAMP2 and second (2-10 min) and third phases (10-30 min) mediated by VAMP8. Induction of acinar pancreatitis by supramaximal cholecystokinin (CCK-8) stimulation inhibits VAMP8-mediated mid- and late-phase but not VAMP2-mediated early-phase secretion. Elevation of cAMP during supramaximal CCK-8 mitigates third-phase secretory inhibition and acinar damage caused by the accumulation of prematurely activated trypsin. VAMP8-/- acini are resistant to secretory inhibition by supramaximal CCK-8, and despite a 4.5-fold increase in total cellular trypsinogen levels, are fully protected from intracellular trypsin accumulation and acinar damage. VAMP8-mediated secretion is dependent on expression of the early endosomal proteins Rab5, D52, and EEA1. Supramaximal CCK-8 (60 min) caused a 60% reduction in the expression of D52 followed by Rab5 and EEA1 in isolated acini and in in vivo The loss of D52 occurred as a consequence of its entry into autophagic vacuoles and was blocked by lysosomal cathepsin B and L inhibition. Accordingly, adenoviral overexpression of Rab5 or D52 enhanced secretion in response to supramaximal CCK-8 and prevented accumulation of activated trypsin. These data support that acute inhibition of VAMP8-mediated secretion during pancreatitis triggers intracellular trypsin accumulation and loss of the early endosomal compartment. Maintaining anterograde endosomal trafficking during pancreatitis maintains VAMP8-dependent secretion, thereby preventing accumulation of activated trypsin.
Highlights
Zymogen secretory granules in pancreatic acinar cells express two vesicle-associated membrane proteins (VAMP), VAMP2 and -8, each controlling 50% of stimulated secretion
Noting that 1) acini prepared from VAMP8Ϫ/Ϫ mice show significantly elevated basal secretion and a diminished secretory inhibition in response to supramaximal CCK-8, 2) VAMP8-mediated acinar secretion is acutely dependent on endosomal trafficking in WT acini, and 3) VAMP8 is a ubiquitously expressed endosomal SNARE, we investigated the impact of acinar pancreatitis on the VAMP8secretory pathway and its potential role in intracellular activated zymogen accumulation
We recently reported that the early phase is mediated by the zymogen granules (ZGs) SNARE protein VAMP2 and the later phase by VAMP8 [1]
Summary
Zymogen secretory granules in pancreatic acinar cells express two vesicle-associated membrane proteins (VAMP), VAMP2 and -8, each controlling 50% of stimulated secretion. Adenoviral overexpression of Rab or D52 enhanced secretion in response to supramaximal CCK-8 and prevented accumulation of activated trypsin These data support that acute inhibition of VAMP8-mediated secretion during pancreatitis triggers intracellular trypsin accumulation and loss of the early endosomal compartment. We recently reported that zymogen activation in response to supramaximal CCK-8, bile salts, or cigarette smoke toxin occurs as a consequence of enhanced EE to LE/lysosome trafficking when acinar secretion is inhibited. Noting that 1) acini prepared from VAMP8Ϫ/Ϫ mice show significantly elevated basal secretion and a diminished secretory inhibition in response to supramaximal CCK-8, 2) VAMP8-mediated acinar secretion is acutely dependent on endosomal trafficking in WT acini, and 3) VAMP8 is a ubiquitously expressed endosomal SNARE, we investigated the impact of acinar pancreatitis on the VAMP8secretory pathway and its potential role in intracellular activated zymogen accumulation
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