Impaired Autolysosome Formation Correlates With Lamp-2 Depletion: Role of Apoptosis, Autophagy, and Necrosis in Pancreatitis

Background & Aims

Acute pancreatitis constitutes a life-threatening condition in which pancreatic acinar cells undergo massive cell death. We investigated the incidence of apoptosis, autophagy, and necrosis affecting acinar cells in the early onset of acute pancreatitis induced by chronic alcohol feeding and acute endotoxemia.

Methods

Rats were fed either an ethanol-containing or a control diet over 14 weeks and killed 3 or 24 hours after a single lipopolysaccharide injection. Apoptosis, necrosis, and autophagy of pancreatic acinar cells were assessed by histology, electron microscopy, immunofluorescence, and biochemical methods.

Results

The combination of alcohol exposure and endotoxemia resulted in the depletion of several lysosomal proteins including lysosomal-associated membrane protein-2 (Lamp-2), a protein that is required for the proper fusion of autophagosomes with lysosomes. Accordingly, Lamp-2 depletion correlated with the accumulation of autophagosomes and a relative paucity of autolysosomes, reduced adenosine-5′-triphosphate levels, and a switch from apoptotic to necrotic cell death. This switch to necrosis was accompanied by reduced caspase activation and the nuclear release of the proinflammatory factor high mobility group box 1. Importantly, human patients with alcoholic pancreatitis also exhibited local Lamp-2 depletion, which points to a crucial role for Lamp-2 and autophagy in pancreatic acinar cell death.

Conclusions

Our data suggest that acinar cell vacuolization in pancreatitis is mediated by an endotoxemia-induced inhibition of the late stage of autophagy. The combination of alcohol and endotoxemia attenuated apoptosis response yet enhanced acinar cell necrosis. The depletion of lysosomal proteins plays a critical role in the early onset of acute pancreatitis.

Abbreviations used in this paper: EtOH, alcohol-fed, Gramp-92, granule membrane protein 92, HMGB1, high mobility group box 1, Lamp-2/-1, lysosomal-associated membrane protein-2/-1, LC3-I, microtubule-associated protein 1 light chain 3-I, LC3-II, autophagosomal form, LPS, lipopolysaccharide, TUNEL, terminal deoxynucleotidyl transferase-mediated digoxigenin-dNTP nick-end labeling