title: Pancreatitis-associated ascitic fluid induces hepatocyte death independent of local cytokines.
abstract: BACKGROUND
  Kupffer-cell-derived cytokines mediate liver injury, yet macrophage pacification does not abolish hepatocyte injury. We undertook this study to examine the role of pancreatitis-associated ascitic fluid (PAAF) in liver injury.
  
  
  METHODS
  Pathogen-free PAAF was perfused into healthy rat livers in situ for 60 min (n = 5, sham = 5, LPS = 5). AST, ALT, LDH, and TNF were measured in the effluent. Primary cultures of rat Kupffer cells or hepatocytes were treated with PAAF; AST, ALT, LDH, and TNF were measured and cell proliferation was determined by MTT assay. A hepatocyte human cell line (CCL-13) was treated with PAAF and apoptosis was measured by flow cytometry.
  
  
  RESULTS
  Liver perfusion with PAAF induced a >15-fold increase in AST/ALT/LDH (P < 0.001 PAAF vs sham), but not in TNF. In vitro, Kupffer cell viability was sharply reduced by PAAF in a dose-dependent manner; however, 5% PAAF (50% viability) did not induce TNF production from Kupffer cells. PAAF induced a multifold increase in AST/ALT/LDH from fresh hepatocytes (P < 0.001 vs control), which was not attenuated by a protease inhibitor. The CCL-13 cell population was reduced to 15 +/- 2% of baseline by PAAF (P < 0.001 vs control), whereas elastase, trypsin, or TNF had no effect. PAAF increased the percentage of nonviable CCL-13 cells (78 +/- 4% vs 28 +/- 1%, P < 0.001 vs control). Neither protease inhibitor nor heat inactivation of PAAF altered this pattern of hepatocyte death.
  
  
  CONCLUSIONS
  PAAF induces direct hepatocyte injury and death by heat-stable factors other than pancreatic enzymes but not via local production of Kupffer-cell-derived cytokines.
authors.affiliation: Department of Surgery, James A. Haley Veterans Hospital, University of South Florida, Tampa 33620, USA.
authors.forename: Michel M
authors.initials: MM
authors.lastname: Murr
authors.forename: Jun
authors.initials: J
authors.lastname: Yang
authors.forename: Adam
authors.initials: A
authors.lastname: Fier
authors.forename: Philip R
authors.initials: PR
authors.lastname: Foulis
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Loughorn
authors.forename: P K
authors.initials: PK
authors.lastname: Epling-Burnette
authors.forename: James G
authors.initials: JG
authors.lastname: Norman
mesh_terms: D000208:Acute Disease; D000818:Animals; D017209:Apoptosis / Q000502:physiology; D001202:Ascitic Fluid / Q000503:physiopathology*; D016923:Cell Death / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D016207:Cytokines / Q000502:physiology; D004798:Enzymes / Q000502:physiology; D022781:Hepatocytes / Q000502:physiology*; D006801:Humans; D007728:Kupffer Cells / Q000502:physiology; D008099:Liver / Q000201:enzymology / Q000378:metabolism; D008297:Male; D010179:Pancreas / Q000201:enzymology; D010195:Pancreatitis / Q000378:metabolism*; D010477:Perfusion; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D016207:Cytokines; D004798:Enzymes