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Measuring the content of immunoreactive pancreatic phospholipase A2 (PLA2; EC 3.1.1.4) and the catalytic activity of PLA2 in serum samples from five patients with acute pancreatitis, we found no correlation between these two measurements overall. To test the specificity of the method for catalytic PLA2, we measured PLA2 activity in serum samples before and after immunoadsorption with an antiserum to human pancreatic PLA2. The results suggest the presence of at least two immunologically distinct PLA2 enzyme proteins in sera from these patients. One of the enzymes is pancreatic in origin and may exist in active, inactive, or inhibited form. The activity profile of the second PLA2 enzyme in serum during acute pancreatitis differs from that for other common pancreatic enzymes. In the present experiment, the catalytic activity was not removed by treatment with the anti-human pancreatic PLA2 antiserum. The source of this second PLA2 activity is unknown. Some samples contained increased activities of both PLA2 forms.

Closed duodenal loop was created in the rat by obstructing the duodenum on both sides of the common bile duct. Acute haemorrhagic pancreatitis developed in these animals within 24 hours. Increased serum amylase activity, pancreatic acinar cell necrosis, pancreatic oedema and haemorrhages, and intra-abdominal fat necroses, together with formation of ascites fluid containing high levels of amylase, were observed in the experimental animals. The findings support the view that reflux of the duodenal contents into the pancreatic duct is an important early pathogenetic mechanism in the development of acute pancreatitis. The closed-loop technique in the rat offers a simple and reliable experimental model for studies on acute pancreatitis.

The histology and ultrastructure of resected pancreas from seven patients suffering from acute haemorrhagic pancreatitis were studied. Special attention was paid to necrotic acini and zymogen granules. Acinar cells in the border of necrotic and non-necrotic parenchyma contained lipid droplets, autophagic vacuoles, bundles of intermediate filaments and degenerated cell organelles, including zymogen granules, PAS-positive material derived from secretory proteins was situated in dilated acinar lumina and in the interstitium, and proved to be fibrillar in fine structure. There were thrombosed vessels and extravasated erythrocytes at the border of the parenchymal necrosis. Bundles of intermediate filaments were often the only identifiable structures in the severely necrotic acinar cells. The amount of lipid was decreased in damaged fat cells. Older fat necroses were surrounded by myofibroblasts. It was concluded that acinar and fat cells undergo concomitant necrosis in the inflamed pancreas, zymogen granules degenerate in the acinar cells at the border of necrotic and non-necrotic areas, and secretory proteins may be displaced in the interstitium outside acinar lumina. Myofibroblasts participate in the healing of autodigestive injuries.

Within a few hours after one injection of fresh human serum by the intraperitoneal route only, mice developed pancreatic acinar cell necrosis and inflammation, fat necrosis, elevated serum amylase and a shocklike state. The extent of these lesions and mortalities were roughly dose dependent and were not noticeably modified by either different fasting cycles or pilocarpine. Acinar cell changes and necrosis usually developed first in subserosal acini. The earliest ultrastructural change detected was nonspecific swelling of cytoplasmic compartments which was reversible but also preceded the cytoplasmic degradation that developed in cells undergoing necrosis. Notably, zymogen granule dissolution neither preceded nor accompanied this swelling, but developed pari passu with cell degradation. Occasionally, intact granules were found in necrotic cells. Serum was cytotoxic for isolated acinar cells in vitro, even in the presence of soybean trypsin inhibitor. These results (1) indicate that the injury mechanism in vivo is directly initiated through contact of serum with acinar cell surfaces and is independent of zymogen secretions and trypsin activation, and (2) suggest that a rapid disturbance in cell membrane permeability results, the magnitude of which being the primary determinant of cell death. Pancreatic toxicity of human serum was abolished by aging, heating, ethylenediaminetetraacetic acid, heparin, zymosan, cobra venom factor, and absorptions with mouse red blood cells, against which fresh, unabsorbed serum was hemolytic. Pancreatic toxicity in vitro and, to a much lesser extent, in vivo was reconstituted by combining the red blood cell-absorbed serum with either heated serum, or with IgM-enriched, but not IgG serum fractions. Fresh cord serum was virtually nontoxic and could substitute for absorbed serum in such reconstitutions. These results indicate that the injury mechanism involves at least two serum components. By both circumstance and analogy, other results and a review of other examples of foreign sera toxicity suggest that they are components of a complement-dependent, cytotoxic heterophile antibody system. The relevance of this odd phenomenon is that it offers a simple model of acute pancreatitis, contributes to the debunking of traditional notions of the pivotal role of zymogens in the initiation of acute pancreatitis, and hints at a potential pathogenetic connection between pancreatitis and products of immune or related reactions.

Porcine pancreatic phospholipase A2 (PLA2) was immobilized to Sepharose 4B and porcine serum was passed through this affinity column. Bound substances were eluted by an EDTA-containing buffer and fractionated in a Sepharose 6B column. A single protein peak of the eluate from the latter column was found to inhibit PLA2 activity in a dose-dependent manner in an assay system using radioactive lecithin as a substrate and porcine pancreatic PLA2 as the enzyme source. The serum fraction containing the PLA2 inhibitory protein(s) (PIP) appeared inhomogeneous on SDS-polyacrylamide gel electrophoresis with two major bands close to each other, corresponding to a molecular weight of approximately 60,000. It was concluded that PIP might act as a protective principle against autodigestion in acute pancreatitis and other inflammatory diseases as well as playing a regulatory role in prostaglandin metabolism.

Immunoreactive phospholipase A2 (EC 3.1.1.4) was measured by a new sensitive time-resolved fluoroimmunoassay in the serum of 58 healthy subjects and 103 patients with acute pancreatitis. Patients with acute pancreatitis were grouped according to the etiology and clinical severity of the disease. The mean phospholipase A2 concentration in the reference (healthy) group was 5.5 (SD 1.9) micrograms/L. In acute pancreatitis the mean phospholipase A2 concentration was increased on the first day after hospital admission in all groups, and returned to normal somewhat more slowly than did serum amylase, especially in the patients with severe alcoholic pancreatitis. In this latter group the mean concentration of serum phospholipase A2 on the first day was 42.6 (SD 29.5) micrograms/L. In patients with pancreatic cancer, serum phospholipase A2 was 29.2 (SD 21.3) micrograms/L. The phospholipase A2 and amylase values were closely associated in all groups. The clinical sensitivities were 90.9% for severe alcoholic pancreatitis and 87.5% for pancreatic cancer. Immunochemical determination of phospholipase A2 in serum provides fast and specific detection of injury to pancreatic acinar cells. In addition to the early diagnosis of acute pancreatitis, follow-up determinations of phospholipase A2 seem to be useful in differentiating between mild and severe forms of pancreatitis.