Acute-phase hepatocytes regulate liver sinusoidal cell mediator production.
BACKGROUND
Overproduction of liver sinusoidal cell (LCS) mediators in response to endotoxemia or gram-negative infection that follows tissue injury may contribute to hepatic dysfunction.


OBJECTIVE
To better define the role of hepatocyte-derived acute-phase reactants in the regulation of sinusoidal cell mediator production following sequential insults, we tested the hypothesis that interleukin-6 (IL-6) prestimulation alters hepatocyte regulation of lipopolysaccharide (LPS)-stimulated sinusoidal cell tumor necrosis factor (TNF), IL-6, and nitric oxide production.


METHODS
Hepatocytes and LSCs were isolated from Wistar rats, and in vitro responses were compared between LSCs alone and hepatocyte-LSC cocultures. Cocultures and LSCs alone were sequentially stimulated with IL-6 (5000 U/mL) then LPS (dose-response), and culture supernatants were analyzed for TNF (L929 cytolysis), IL-6 (7TD1 proliferation), and nitric oxide (Griess reaction). Induction of acute-phase protein synthesis by the stimulation of hepatocytes with IL-6 and dexamethasone (0.1 mumol/L) was assayed by methionine radiolabeling and SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis). Coculture levels of messenger RNA for TNF-alpha and IL-6 were examined by RNA extraction and reverse transcriptase polymerase chain reaction with specific primers.


RESULTS
Interleukin-6 and dexamethasone signal hepatocyte acute-phase protein synthesis. Prestimulation of cocultures, but not of LSCs alone, with IL-6 inhibits LPS-stimulated IL-6 and nitric oxide production significantly. Bioactivity of TNF is reduced to a lesser extent. Polymerase chain reaction analysis demonstrated similar levels of TNF and IL-6 message following sequential stimulation.


CONCLUSIONS
Interleukin-6-stimulated acute-phase hepatocytes limit LPS-stimulated coculture cytokine bioactivity and nitric oxide production. This hepatocyte response may provide a local counterregulatory mechanism to limit LSC-mediated injury.
Bankey
P
P
Department of Surgery, University of Texas, Southwestern Medical Center, Dallas.
Prager
M
M
Geldon
D
D
Taylor
S
S
McIntyre
K
K
D000209:Acute-Phase Proteins / Q000096:biosynthesis; D000210:Acute-Phase Reaction / Q000378:metabolism*; D000704:Analysis of Variance; D000818:Animals; D066298:In Vitro Techniques; D018836:Inflammation Mediators / Q000378:metabolism*; D015850:Interleukin-6 / Q000096:biosynthesis / Q000502:physiology; D008099:Liver / Q000098:blood supply / Q000166:cytology / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis; D016133:Polymerase Chain Reaction; D051381:Rats; D017208:Rats, Wistar; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
D000209:Acute-Phase Proteins; D018836:Inflammation Mediators; D015850:Interleukin-6; D014409:Tumor Necrosis Factor-alpha; D009569:Nitric Oxide