Activation peptide of carboxypeptidase B in serum and urine in acute pancreatitis

Abstract

Background: The pathophysiology of acute pancreatitis involves activation of the pancreatic proenzymes. Levels of the trypsinogen activation peptide in urine in acute pancreatitis has been shown to correlate with the severity of disease. However, this peptide is unstable in urine and, because of its low molecular mass, difficult to measure. Procarboxypeptidase B has a larger activation peptide which could be more suitable for analysis in serum and urine.

Aims: To study the presence of the activation peptide from procarboxypeptidase B (CAPAP) in serum and urine in acute pancreatitis.

Patients: Urine and serum samples were obtained within 48 hours of admittance from 40 patients with acute pancreatitis. Severity was classified retrospectively according to levels of C-reactive protein and clinical course. Thirty four patients with abdominal pain from other causes were studied as controls.

Methods: CAPAP was purified from human pancreatic juice. Specific antibodies were obtained and a radioimmunoassay was developed.

Results: Levels of CAPAP in serum and urine in acute pancreatitis correlate with the severity of the attack. CAPAP is very stable, and urine contains only CAPAP whereas, in serum, cross reacting procarboxypeptidase B is found together with CAPAP.

Conclusions: CAPAP could be a valuable tool in the diagnosis and early determination of severity in acute pancreatitis.

Figure 1

Agarose gel electrophoresis at pH 8.6 of pure procarboxypeptidase B (lane 2), pure active carboxypeptidase B (lane 3) and pure carboxypeptidase B activation peptide (lane 4). The electrophoretic pattern of human serum is shown as a reference in lane 1. A indicates the application slit and B the albumin band in serum.

Figure 2

Sodium dodecyl sulphate/polyacrylamide gel electrophoresis (12% gel) of procarboxypeptidase B (proCPB) before activation (lane 2), active CPB together with the activation peptide (CAPAP) (after activation of proCPB with trypsin) (lanes 3 and 4), and purified CAPAP (lanes 5 and 6). Molecular mass standards are shown in lanes 1 and 7.

Figure 3

Electrospray mass spectroscopic analysis of the purified activation peptide.

Figure 4

Assay for immunoreactive carboxypeptidase B activation peptide (CAPAP). Dilution curve for pure CAPAP (CAPAP standard), together with dilution curves for procarboxypeptidase B (proCPB) before and after activation with trypsin (activated proCPB).

Figure 5

Immunoreactive carboxypeptidase B activation peptide (CAPAP) in serum from patients with acute pancreatitis divided up according to severity of attack. Samples from patients with acute abdominal pain for known reasons other than acute pancreatitis (no pancreatitis) and from patients with newly diagnosed pancreatic cancer are also shown.

Figure 6

Characterisation of carboxypeptidase B activation peptide (CAPAP)-like immunoreactivity by gel filtration (Sephadex G-50 column) of serum and urine samples from a patient with acute pancreatitis.

Figure 7

Immunoreactive carboxypeptidase B activation peptide (CAPAP) in urine from patients with acute pancreatitis divided up according to severity of attack. Samples from patients with acute abdominal pain for known reasons other than acute pancreatitis (no pancreatitis) and from patients with newly diagnosed pancreatic cancer are also shown.

Figure 8

Characterisation of immunoreactive carboxypeptidase B activation peptide (CAPAP), in urine from a patient with acute pancreatitis, by preparative gel electrophoresis. Application slit at zero. Anode to the right.