Suppression by citrus auraptene of phorbol ester- and endotoxin-induced inflammatory responses: Role of attenuation of leukocyte activation

Auraptene (AUR), a citrus coumarin derivative, is one of the promising chemopreventive agents against skin, tongue, esophagus and colon carcinogenesis in rodents. We reported previously that AUR suppresses superoxide anion (O₂^-) generation from inflammatory leukocytes in in vitro experiments. In the present study, we investigated the anti-inflammatory activities of AUR using a 12-O-tetradecanoylphorbol-13-acetate-treated mouse skin model, and compared them with those of umbelliferone (UMB), a structural analog of AUR that is virtually inactive toward O₂- generation inhibition. Double pre-treatments of mouse skin with AUR, but not UMB, markedly suppressed edema formation, hydrogen peroxide production, leukocyte infiltration, and the rate of proliferating cell nuclear antigen-stained cells. These inhibitory effects by AUR are attributable to its selective blockade of the activation stage, as revealed by single pre-treatment experiments. In a murine macrophage line, RAW 264.7, AUR significantly attenuated the lipopolysaccharide-induced protein expression of inducible isoforms of both nitric oxide synthase and cyclooxygenase, with decreased production of nitrite anion and prostaglandin E₂, and yet suppressed the release of tumor necrosis factor-α. Conversely, UMB did not show any inhibitory effect. This contrasting activity profile between AUR and UMB was rationalized to be a result of their distinct differences in cellular uptake efficiencies, i.e. the geranyloxyl group in AUR was found to play an essential role in incorporation. Thus, our findings indicate that AUR is an effective agent to attenuate the biochemical responsiveness of inflammatory leukocytes, which may be essential for a greater understanding of the action mechanism that underlies its inhibition of inflammation-associated carcinogenesis.