Pancreatic ascites as a powerful inducer of inflammatory cytokines. The role of known vs unknown factors.
OBJECTIVE
To determine if pancreatic ascites will induce interleukin 1 beta (IL-1 beta) or tumor necrosis factor alpha (TNF-alpha) production outside the pancreas and examine the possible components responsible.


METHODS
Severe pancreatitis was induced in rats (n = 30) by pancreatic duct infusion with 4% glycodeoxycholic acid; pancreatic ascites was collected 18 hours later. In vitro studies used quiescent murine splenic or pulmonary macrophages (10(5)/mL) which were exposed to media alone (control), trypsin, chymotrypsin, cathepsin-B, 20% ascites (vol/vol), 50% ascites, or endotoxin (lipopolysaccharide, 10 micrograms/mL, positive control) for 4 hours. Subsequently, pancreatic ascites was cultured for bacteria and assayed for endotoxin and cytokines (interleukin 1, interleukin 6, interleukin 8, TNF-alpha, or interferon gamma). The experiments were then repeated using 20% and 50% ascites that was sterile and cytokine-free (SCF ascites). In vivo studies used 100% (n = 8) or 50% (n = 12) SCF ascites or normal rat serum (control, n = 12) for a 10-second pulmonary lavage (100 microL) in adult mice, with lungs collected at 6 hours for cytokine gene analysis.


METHODS
Surgical basic science research laboratory.


METHODS
Interleukin 1 beta and TNF-alpha gene induction was assessed by quantitative competitive reverse-transcription polymerase chain reaction and cytokine protein production was determined by enzyme-linked immunosorbent assay.


RESULTS
Macrophages responded to untested and SCF ascites in a dose-dependent fashion, with a multifold increase in both IL-1 beta and TNF-alpha messenger RNA (mRNA) and protein, which was often more potent than lipopolysaccharide. Expression of IL-1 beta and TNF-alpha mRNA could not be induced by trypsin, chymotrypsin, or cathepsin-B. All animals undergoing lavage with 100% SCF ascites died within 2 hours, while those undergoing lavage with 50% SCF ascites showed a multifold increase in pulmonary IL-1 beta and TNF-alpha mRNA.


CONCLUSIONS
Pancreatic ascites contains factors that are capable of inducing IL-1 beta and TNF-alpha production in vitro and in vivo. This effect cannot be reproduced by activated digestive enzymes and is propagated despite the absence of known inducers of cytokines such as bacteria, endotoxin, or other inflammatory cytokines.
Denham
W
W
Department of Surgery, University of South Florida, Tampa, USA.
Yang
J
J
Fink
G
G
Zervos
E E
EE
Carter
G
G
Norman
J
J
D000818:Animals; D001201:Ascites / Q000209:etiology / Q000276:immunology*; D007375:Interleukin-1 / Q000096:biosynthesis / Q000276:immunology*; D008264:Macrophages / Q000276:immunology / Q000378:metabolism; D051379:Mice; D010195:Pancreatitis / Q000150:complications / Q000276:immunology*; D051381:Rats; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000276:immunology*
D007375:Interleukin-1; D014409:Tumor Necrosis Factor-alpha