FLUORESCEIN ISOTHIOCYANATE STAINING AND CHARACTERIZATION OF AVIAN HETEROPHILS

Fluorescein isothiocyanate (FITC) was found to stain cytoplasmic granu les of avian heterophil-granulocytes. In tissue sections, the fluoresc ent granulocytes were predominantly distributed adjacent to trabecular bones. The fluorescein stained granulocytes were abundant in synovial fluids of chickens with synovitis. A significant correlation was obse rved in the percent of fluorescein labeled granulocytes in blood smear s and the percent of heterophils determined using an automated countin g method, in unstained blood from normal and Escherichia coli-infected turkeys. ?The fluorescein-binding heterophils purified from chickens showed a time dependent increases in the oxidation of 2',7'-dichlorofl uorescin diacetate (DCF-DA) and the reduction of nitroblue tetrazolium (NBT) which were indicative of changes in oxidative burst in response to phorbol 12-myristate 13-acetate (PMA), Salmonella typhimurium lipo polysaccharide (LPS), and zymosan A (ZA). These heterophil-activating agents, also, caused significant degranulation at 16 h post-treatment, as indicated by the loss fluorescence. There were microscopically vis ible alterations in the cell shapes and a decrease in the density of g ranules due to treatment with LPS, PMA or ZA. In addition, these cells also showed phagocytic response which was evident at 30 min of incuba tion with fluorescent latex particles. Both chicken and turkey heterop hils produced interleukin-6 in vitro at 24 h in response to LPS but no t to PMA, FMLP or ZA. The chicken heterophils showed spontaneous produ ction of matrix metalloproteinases (MMP) which was significantly enhan ced by treatment with LPS, PMA, and ZA; however, LPS appeared to be mo st effective in inducing MR/LP production. These results demonstrate t hat the functions of heterophils can be differentially regulated by di fferent activating agents and the fluorescein binding property of thes e cells may be useful for their histochemical identification. Publishe d by Elsevier Science B.V.