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We have demonstrated that pancreatitis-associated ascitic fluid contributes to hepatocyte injury during acute pancreatitis; a phenomenon independent of ascites'' enzymatic content and Kupffer cell-derived cytokines. Our aim is to characterize the mechanisms of pancreatitis-associated ascitic fluid induced hepatocyte death. NIH mice were injected intraperitoneally with pathogen-free pancreatitis-associated ascitic fluid. Twenty-four hours later, serum AST, ALT, LDH, and hepatocyte apoptosis (TUNEL) were measured. Human hepatocytes (CCL-13) were treated with pancreatitis-associated ascitic fluid ±SB203580 or caspase-3 inhibitor-II. Mitochondrial membrane integrity was determined by DiOC6 staining. Apoptosis was measured by TUNEL staining and flow cytometry after dual labeling with Annexin-V/7-AAD. Data are mean ± SEM of triplicates. Pancreatitis-associated ascitic fluid increased serum AST, ALT, LDH, and apoptotic cells in the mouse liver (all P < 0.03 vs. sham). In CCL-13 cells, pancreatitis-associated ascitic fluid induced a time and dose-dependent increase in apoptosis, in addition to p38-MAPK phosphorylation (P = 0.02 vs. control), caspase-3 cleavage (P < 0.03 vs. control) and decreased DiOC6 mitochondrial staining (P < 0.01 vs. control). Both caspase-3 inhibitor-II and SB203580 decreased apoptosis, but the former had no effect on DiOC6 staining. Pancreatitis-associated ascitic fluid induces liver injury and hepatocyte apoptosis by activating p38-MAPK and caspase-3 dependent pro-apoptotic pathways. ( J Gastrointest Surg 2003;7:200–208.) [Copyright &y& Elsevier]

We have demonstrated that Kupffer cell–derived tumor necrosis factor (TNF) mediates pancreatitis-associated liver injury. The aim of this study was to determine the role of p38 mitogen-activated protein kinase (MAPK), extracellular stress-related kinase 1/2 (ERK1/2), stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK), and nuclear factor–κB (NF-κB) in TNF gene expression within Kupffer cells. TNF and TNF-mRNA were measured in rat livers perfused with elastase. TNF, TNF-mRNA, NF-κB activation, and phosphorylated p38-MAPK, SAPK/JNK, and ERK1/2 were determined in Kupffer cells treated with elastase. Elastase increased TNF and upregulated TNF-mRNA in livers (P<0.03) and Kupffer cells (P<0.001). Phosphorylated p38-MAPK, SAPK/JNK, and ERK1/2 and activated NF-κB were detected in Kupffer cells at 7 minutes; at 60 minutes, TNF-mRNA peaked and NF-κB returned to baseline, whereas all three kinases remained activated. Gadolinium inhibited elastase-induced upregulation of TNF-mRNA (P < 0.001), TNF production (P<0.001), and attenuated SAPK/JNK, as well as ERK1/2, but not p38-MAPK. Both UO126 and SB203580 significantly inhibited elastase-induced upregulation of TNF-mRNA and TNF production (P<0.001), but only UO126 inhibited activation of NF-κB. It was concluded that pretranscriptional regulation of TNF gene expression in Kupffer cells follows an orderly activation of p38-MAPK, ERK1/2, and SAPK/JNK that may not converge on NF-κB. The seemingly limited duration of NF-κB activation may be important in “switching off” the cytokine cascade during acute pancreatitis. ( J Gastrointest Surg 2003;7:20–25.) [Copyright &y& Elsevier]

Liver injury is a manifestation of the systemic inflammatory response during acute pancreatitis. We have demonstrated that elastase induces macrophage tumor necrosis factor (TNF) production in distant organs, thus mimicking pancreatitis-associated organ injury. The aim of this study was to determine the mechanism by which elastase induces hepatic cytokine production. Rat livers (n = 40) were perfused with elastase ± gadolinium (Gd) to inhibit Kupffer cells. Liver parenchymal enzymes and TNF were measured in the effluent. In vitro, rat hepatocytes or Kupffer cells were treated with elastase (1 U/ml) ± Gd (0.5 mg/ml) or pyrrolidine dithiocarbamate (PDTC; 0.5 mg/ml). TNF protein, TNF messenger RNA, and NF–κB activation were determined. In vivo, Gd blunted the elastase-induced TNF production and decreased AST, ALT, LDH, and nonviable cells (propidium iodide) (P ≤ 0.03 vs. elastase). In vitro, elastase induced TNF production from Kupffer cells (P < 0.001 vs. control) but not from hepatocytes. Gd or PDTC significantly attenuated the elastase-induced TNF production (P < 0.001). Elastase-induced overexpression of TNF messengerRNA and activation of NF-κB was attenuated by Gd. Pancreatic elastase induces a pattern of liver injury similar to that seen during acute pancreatitis by activating cytokine production and gene expression within Kupffer cells via NF-κB. Gd exhibits a protective effect against elastase-induced liver injury by inhibiting activation of NF-κB. ( J Gastrointest Surg 2002;6:474-480.) [ABSTRACT FROM AUTHOR]