Published in Cancer Detection and Prevention 2000; 24(Supplement 1).

Effect of dibutyryl cyclic AMP on production of tumor necrosis factor alpha in rat Kupffer cells

T Goto MD 1, M Komatsu MD 1, K Mikami MD 1, K Yoneyama MD 1, K Miura MD 1, K Nakane MD 1, JG Lin MD 1, S Watanabe MD 1

1 First Dept Internal Medicine, Akita University of Medicine, Akita, Japan,

AIMS: Dibutyryl cyclic AMP (DBcAMP) is an analogues of cAMP. DBcAMP has many effects on hepatocellular carcinoma, liver cirrhosis, ischemic liver failure and endotoxin-induced inflammatory liver injury. However, little is known about the mechanism of DBcAMP action in Kupffer cells. We examined the effects of DBcAMP on phagocytic activity and production of tumor necrosis factor(TNF)-alpha in cultured rat Kupffer cells treated with lipopolysaccharide (LPS). METHODS: Kupffer cells were obtained from the livers of Sprague-Dawley male rats by a method employing collagenase. The phagocytic activity was measured according to the number of latex particles incorporated into the cytoplasm of Kupffer cells. TNF-alpha levels was measured with ELISA. Immunofluorescent staining for nuclear factor-kappaB (NF-kappaB) was visualized using an anti-NF-kappaB p65 antibody. RESULTS: DBcAMP premedication had no effect on LPS-stimulated phagocytic activity of Kupffer cells but inhibited TNF-alpha production dose-dependent manner. Genistein, an inhibitor of protein tyrosin kinases, did not block; but H-89, an inhibitor of cAMP-dependent protein kinase, did block the inhibitory effect of DBcAMP on LPS-stimulated TNF-alpha production in Kupffer cells. Using immunofluorescent staining for NF-kappaB, we demonstrate that the effect of DBcAMP dose not involve signal transduction through NF-kappaB. CONCLUSIONS: We conclude that DBcAMP inhibits LPS-stimulated TNF-alpha production by activating cyclic AMP-dependent protein kinase.

KEY WORDS: cytokine, cAMP.

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Paper presented at the International Symposium on Impact of Biotechnology on Cancer Diagnostic & Prognostic Indicators; Geneva, Switzerland; October 28 - 31, 2000; in the section on novel therapies.