Date of Award

2-23-2009

Document Type

Thesis

Degree Name

Medical Doctor (MD)

First Advisor

Fred Gorelick

Abstract

Acute pancreatitis (AP) is an inflammatory disease of the pancreas involving premature activation of digestive enzyme zymogens. Since energy stress and ischemia are important in the development of AP, we studied the role of a cellular energy-sensing enzyme AMP-dependent protein kinase, AMPK, in groups of pancreatic acinar cells (acini), which have phenotypic responses that match those of early pancreatitis. We used the cholecystokinin orthologue, caerulein, to stimulate acini and to induce experimental pancreatitis. Using this system, activation of AMPK with the agonist AICAR decreased zymogen activation, while inhibition with the antagonist compound C enhanced it; both AMPK modulators appeared to influence a later stage of zymogen activation. Stimulation of acini with physiologic concentrations of caerulein caused mild changes in AMPK activity. However, when exposed to hyperstimulatory doses which caused pancreatitis, caerulein caused a rapid and prolonged decrease in AMPK activity. Unexpectedly, AMPK activity varied inversely with protease activation. The hyperstimulation-induced change in AMPK activity was accompanied by modifications in phosphorylation at the enzymes activating and inhibitory sites. Further, a transient decrease in AMPK protein levels was observed with hyperstimulation. We investigated the processes involved in zymogen activation by developing a cell-free assay using zymogen-containing compartments and cytosol. We found that cytosolic factors were necessary to cause zymogen activation, and that adenosine nucleotides tended to inhibit trypsinogen, and enhance chymotrypsinogen activation. Time-course studies of secretion and cell injury in acini in the presence of AICAR and compound C revealed no effects of modulating AMPK on either process. These findings suggest that AMPK activity is linked to early pathologic protease activation in the pancreatic acinar cell. The cellular targets of AMPK will be a subject of future studies.

Comments

This thesis is restricted to Yale network users only. This thesis is permanently embargoed from public release.

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