Functional defects in phagocytic cells following thermal injury. Application of flow cytometric analysis

Abstract

Defective phagocytic cell function may partially account for the morbidity and mortality associated with thermal injury. In experimental thermal injury in the rat, small circulating blood volumes increase the difficulty in obtaining significant data. Furthermore, purification and or elicitation procedures have the potential for altering the cell surface characteristics and/or the functional response of the cell in question. We have examined the circulating neutrophils and pulmonary alveolar macrophages of anesthetized rats following a 16-20% body surface area scald injury to the shaved back. The circulating neutrophils of thermally injured rats were examined by flow cytometry following stimulation with phorbol myristate acetate (PMA) (100 ng/ml) in terms of the change in fluorescence intensity of the potentiometric cyanine dye, dipentyloxocarboxyanine and the formation of the oxidized product of 2',7'-dichlorofluorescin diacetate-loaded cells. The alveolar macrophages were examined after stimulation with PMA (100 ng/ml) in terms of the change in fluorescence intensity of the potentiometric dye, dipropylthiodicarbocyanine and the generation of superoxide production, as assessed by the superoxide dismutase inhibitable reduction of cytochrome c. Both cells exhibited a profound inhibition of cell function 4 hours after the insult, with partial return toward control values at later time points. Furthermore, the plasma of thermally injured rats, 4 hours after the burn was inhibitory to normal rat neutrophils. Fluorescent compounds suggestive of in vivo lipid peroxidation were maximally detectable at this time point. Further research is needed to establish the role of these products in the induction of phagocytic cell dysfunction.