Early trypsin activation develops independently of autophagy in caerulein-induced pancreatitis in mice.

Abstract

Premature intrapancreatic trypsinogen activation is widely regarded as an initiating event for acute pancreatitis. Previous studies have alternatively implicated secretory vesicles, endosomes, lysosomes, or autophagosomes/autophagolysosomes as the primary site of trypsinogen activation, from which a cell-damaging proteolytic cascade originates. To identify the subcellular compartment of initial trypsinogen activation we performed a time-resolution analysis of the first 12 h of caerulein-induced pancreatitis in transgenic light chain 3 (LC3)-GFP autophagy reporter mice. Intrapancreatic trypsin activity increased within 60 min and serum amylase within 2 h, but fluorescent autophagosome formation only by 4 h of pancreatitis in parallel with a shift from cytosolic LC3-I to membranous LC3-II on Western blots. At 60 min, activated trypsin in heavier subcellular fractions was co-distributed with cathepsin B, but not with the autophagy markers LC3 or autophagy protein 16 (ATG16). Supramaximal caerulein stimulation of primary pancreatic acini derived from LC3-GFP mice revealed that trypsinogen activation is independent of autophagolysosome formation already during the first 15 min of exposure to caerulein. Co-localization studies (with GFP-LC3 autophagosomes versus Ile-Pro-Arg-AMC trypsin activity and immunogold-labelling of lysosomal-associated membrane protein 2 [LAMP-2] versus trypsinogen activation peptide [TAP]) indicated active trypsin in autophagolysosomes only at the later timepoints. In conclusion, during the initiating phase of caerulein-induced pancreatitis, premature protease activation develops independently of autophagolysosome formation and in vesicles arising from the secretory pathway. However, autophagy is likely to regulate overall intracellular trypsin activity during the later stages of this disease.

• Authors: Sudarshan R Malla, Burkhard Krueger, Thomas Wartmann, Matthias Sendler, Ujjwal M Mahajan, F Ulrich Weiss, Franziska G Thiel, Carina De Boni, Fred S Gorelick, Walter Halangk, Ali A Aghdassi, Thomas Reinheckel, Anna S Gukovskaya, Markus M Lerch, Julia Mayerle
• Journal: Cellular and molecular life sciences : CMLS (Cell Mol Life Sci) Vol. 77 Issue 9 Pg. 1811-1825 (May 2020) ISSN: 1420-9071 [Electronic] Switzerland
• PMID: 31363815 (Publication Type: Journal Article)

Chemical References

• Ceruletide
• Trypsinogen
• Trypsin

Topics

• Animals
• Autophagosomes (metabolism)
• Autophagy
• Ceruletide (toxicity)
• Endosomes (metabolism)
• Male
• Mice
• Mice, Inbred C57BL
• Pancreatitis (chemically induced, metabolism, pathology)
• Secretory Vesicles (metabolism)
• Trypsin (metabolism)
• Trypsinogen (metabolism)