Gastroenterology

Gastroenterology

Volume 138, Issue 2, February 2010, Pages 726-737
Gastroenterology

Basic—Liver, Pancreas, and Biliary Tract
Cathepsin L Inactivates Human Trypsinogen, Whereas Cathepsin L-Deletion Reduces the Severity of Pancreatitis in Mice

https://doi.org/10.1053/j.gastro.2009.10.048Get rights and content

Background & Aims

Acute pancreatitis is characterized by an activation cascade of digestive enzymes in the pancreas. The first of these, trypsinogen, can be converted to active trypsin by the peptidase cathepsin B (CTSB). We investigated whether cathepsin L (CTSL) can also process trypsinogen to active trypsin and has a role in pancreatitis.

Methods

In CTSL-deficient (Ctsl−/−) mice, pancreatitis was induced by injection of cerulein or infusion of taurocholate into the pancreatic duct. Human tissue, pancreatic juice, mouse pancreatitis specimens, and recombinant enzymes were studied by enzyme assay, immunoblot, N-terminal sequencing, immunocytochemistry, and electron microscopy analyses. Isolated acini from Ctsl−/− and Ctsb−/− mice were studied.

Results

CTSL was expressed in human and mouse pancreas, colocalized with trypsinogen in secretory vesicles and lysosomes, and secreted into pancreatic juice. Severity of pancreatitis was reduced in Ctsl−/− mice, whereas apoptosis and intrapancreatic trypsin activity were increased. CTSL-induced cleavage of trypsinogen occurred 3 amino acids toward the C-terminus from the CTSB activation site and resulted in a truncated, inactive form of trypsin and an elongated propeptide (trypsinogen activation peptide [TAP]). This elongated TAP was not detected by enzyme-linked immunosorbent assay (ELISA) but was effectively converted to an immunoreactive form by CTSB. Levels of TAP thus generated by CTSB were not associated with disease severity, although this is what the TAP-ELISA is used to determine in the clinic.

Conclusions

CTSL inactivates trypsinogen and counteracts the ability of CTSB to form active trypsin. In mouse models of pancreatitis, absence of CTSL induces apoptosis and reduces disease severity.

Section snippets

Materials and Methods

Please refer to Supplementary Materials and Methods for more detailed descriptions.

Subcellular Localization and Sorting of CTSL Into the Secretory Pathway

A prerequisite for a biologic relevance of CTSL in pancreatitis would be its expression in the exocrine pancreas and its colocalization with trypsinogen. All of these have previously been established for CTSB5, 8 and could also be operative for CTSL because of great similarities between the 2 lysosomal proteases. Blotting CTSL in the pancreatic juice of mice and humans resulted in a strong signal for pro-CTSL (31 kilodaltons), single chain CTSL, as well as heavy chain CTSL (Figure 1A).

Discussion

The underlying mechanism of acute pancreatitis has long been thought to involve autodigestion of the pancreas by its own digestive proteases. Under physiologic conditions, the pancreas is protected by a variety of mechanisms that include storage and processing of digestive enzymes in membrane-confined vesicles, transport of proteases to the lumen as inactive precursor zymogens, presence of protease inhibitors, and absence of the physiologic activator enterokinase from the pancreas. Several

Acknowledgments

The authors thank Dr B. Schmidt, Zentrum für Biochemie und Molekulare Zellbiologie, Göttingen, for sequencing assistance and V. Krause, C. Jechorek, K. Mülling, and N. Loth for technical assistance and D. Schwenn for writing assistance.

T.W. and J.M. contributed equally as first authors.

W.H. and M.M.L. contributed equally as senior authors.

References (31)

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Conflicts of interest The authors disclose no conflicts.

Funding Supported by grants from the Deutsche ForschungsgemeinschaftDFG HA2080/7-1, LE 625/8-1, LE 625/9-1, MA4115/1-2, DFG GRK 840 E3 and E4; Mildred Scheel Stiftung10-2031-Le I and 10-6977-Re, BMBF-NBL3 01 ZZ 0403; and Alfried-Krupp Foundation (Graduiertenkolleg Tumorbiologie) and NIHDK058088 to MST.

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