THE FIRST APPLICATION OF A CHEMILUMINESCENCE PROBE, 2-METHYL-6-[P-METHOXYPHENYL]-3,7-DIHYDROIMIDAZO[1,2-A- ]PYRAZIN-3-ONE (MCLA), FOR DETECTING O-2(-) PRODUCTION, IN-VITRO, FROM KUPFFER CELLS STIMULATED BY PHORBOL-MYRISTATE ACETATE

The purpose of this study was to investigate the generation of superoxide anion radical, O-2(-) by Kupffer cells in vivo in rat liver. Phorbol myristate acetate (PMA) was infused into perfused rat liver which had been continuously infused with 2-methyl-6-[p-methoxyphenyl]-3,7-dihydroimidazo-[1,2-a]pyrazin-3-one (MCLA, a highly sensitive luminescence reagent for O-2(-)) and the MCLA luminescence from the liver surface was detected with a sensitive photon counter. Under identical conditions, but without MCLA infusion, PMA and nitro blue tetrazolium were infused to verify generation of O-2(-) by formation of formazan deposition. Based on MCLA luminescence and formazan deposition, both dependent on the O-2(-) reaction, we have concluded that liver Kupffer cells generate O-2(-) in vivo in response to stimuli such as PMA. Further, the generation of O-2(-) by Kupffer cells may be by the same mechanism as displayed by macrophages.