Studies on the locomotory, phagocytic and metabolic activities of neutrophils from the plaice (Pleuronectes platessa L.)
The aim of this project was to study the mechanisms of disease resistance in plaice (Pleuronectes platessa L.). Within this broad area, the work concentrated on investigating the migration, phagocytic activity and luminol-dependent chemiluminescence (CL) of plaice phagocytic leucocytes in vitro. In these studies two sources of leucocytes were used: (i) glycogen-elicited, peritoneal exudate cells and (ii) kidney leucocytes that had been purified on a discontinuous Percoll gradient. Both cell isolates comprised of at least 70% neutrophils. Several factors were found to chemoattractants for both peritoneal and kidney neutrophils. Endogenous attractants included leukotriene B4 and two neutrophil-derived factors, which differed in their solubilities in organic solvents. Exogenous attractants included the formyl-peptide, N-formylmethionyl-leucyl-phenylalanine, and an organic solvent soluble factor released by viable Vibrio alginolyticus. The interactions of these attractants and their in vivo activities were also investigated. The migration of neutrophils in vitro was significantly reduced in the presence of cortisol at levels equivalent to those found in stressed plaice (> 400 ng/ml). The opsonic activity of plaice serum was investigated by analysing the phagocytosis and CL of plaice neutrophils. Sheep erthrocytes were only opsonized by the presence of specific antibody and an activatable factor(s) found in normal serum. The bacterium, V. alginolyticus, was opsonized by a factor(s) in normal serum, the activity of which was not further enhanced by the presence of specific antibody. In fact, two different opsonins were identified in normal plaice serum: (i) a very heat-stable activity (stable at 70oC for 30 min) recognised by kidney neutrophils and (ii) a heat-labile activity (50oC for 30 min) recognised by peritoneal neutrophils. Plaice neutrophils were shown to produce several reactive oxygen species: superoxide anion (O-2), hydrogen peroxide (H2O2) and their derivatives, possibly including hydroxyl radical (OHbullet).