Specific pancreatic enzymes activate macrophages to produce tumor necrosis factor-alpha: Role of nuclear factor kappa B and inhibitory kappa B proteins

The triggering events by which mononuclear cells throughout the body are induced to produce large amounts of cytokines during acute pancreatitis are unclear. However, recent work in our laboratory demonstrated that three specific pancreatic enzymes (elastase, carboxypeptidase A, and lipase) induced dramatic tumor necrosis factor-alpha (TNF-α) protein production from macrophages, whereas all others could not. This series of experiments was designed to examine the second messenger system by which this occurs. The rat macrophage cell line NR8383 was incubated for 3 hours with elastase, carboxypeptidase A, lipase, trypsin, or lipopolysaccharide (positive control). Activation of nuclear factor kappa B (NF-KB) was demonstrated by electrophoretic mobility shift assay, presence of inhibitory kappa B alpha and beta (IKB-a and IKB-13) by Western ~lot analysis, and TNF-α protein production by enzyme-linKed immunosorbent assay. Elastase, carboxypeptidase A, and lipase induced degradation of IKB-13 (but not IKB-a), activation of NF-KB, and production of TNF-α protein, whereas inhibition of IKB with pyrrolidine dithiocarbamate attenuated this response. Trypsin was unable to elicit any of these responses. Macrophages can be induced by specific activated pancreatic enzymes-elastase, carboxypeptidase A, and lipase--to produce TNF-α. This process is dependent on IKB-13 degradation and NF-KB activation, suggesting that these enzymes trigger this second messenger system through specific membrane-bound receptors.