ULTRASTRUCTURAL-LOCALIZATION OF CHARCOT-LEYDEN CRYSTAL PROTEIN (LYSOPHOSPHOLIPASE) AND PEROXIDASE IN MACROPHAGES, EOSINOPHILS, AND EXTRACELLULAR-MATRIX OF THE SKIN IN THE HYPEREOSINOPHILIC SYNDROME

  • 1 May 1990
    • journal article
    • research article
    • Vol. 62  (5) , 590-607
Abstract
Electron microscopy, ultrastructual cytochemisty, and postembedding immunogold ultrastructural immunocytochemistry were used to study a papular cutaneous lesion from a patient with the hypereosinophilic syndrome. Peroxidase activity was detected cytochemically in 40-.mu.m sections of skin utilizing the substrate diaminobenzidine; Charcot-Leyden crystal (CLC) protein was detected immunocytochemically in skin utilizing a postembedding immunogold technique; and a combined method was used where postembedding immunogold staining of CLC protein was performed on sections previously prepared to detect peroxidase activity. We describe a unique, eosinophil-rich inflammatory process in involved skin which contained extraordinary numbers of morphologically activated macrophages. Electron microscopy demonstrated (a) widespread eosinophil necrosis, (b) interstitial CLC formation, (c) macrophage activation, endocytosis, and phagocytosis, and (d) CLC formation in phagosomes of activated macrophages. Peroxidase activity was present as follows: (a) in the matrix of eosinophil specific granules in eosinophil cytoplasm, in membrane-bound specific granules released into interstitial tissues from dying eosinophils, being phagocytized by activated macrophages, and within macrophage phagosomes; (b) as amorphous interstitial deris; (c) in cytoplasm and nuclei of damaged eosinophils in the dermal tissues as well as in macrophage phagosomes; and (d) in endocytotic vesicles and vacuoles of macrophages and in CLC-containing phagosomes of macrophages. CLC protein was localized by immunocytochemistry to (a) eosinophil primary granules, (b) cytoplasm and nuclei of damaged eosinophils located in the interstitial tissues or within macrophage phagosomes, (c) CLC located in interstitial tissues adjacent to necrotic eosinophils and in macrophage phagosomes, and (d) aggregates of amorphous protein bound to macrophage surfaces; endocytotic vesicles and vacuoles of macrophages; amorphous protein aggregates in macrophage lysosomes. Combined staining for peroxidase and CLC protein demonstrated that (a) peroxidase-positive specific eosinophil granules did not stain for CLC protein; (b) macrophage endosomes generally contained CLC protein alone, whereas peroxidase was generally internalized by macrophages in association with damaged eosinophil parts or was present in CLC protein-negative vacuoles; (c) individual macrophage phagosomes contained either CLC protein alone, peroxidase-positive structures alone, or both eosinophils proteins; (d) such mixed content macrophage phagosomes showed either separately tagged domains or mixed domains; the mixed domains in phagosomes often contained recognizable eosinophil parts; (e) CLC that formed in macrophage phagosomes were generally peroxidase-negative but sometimes displayed peroxidase adsorbed to their angular and planar surfaces. These findings provide new documentation of extracellular and subcellular sites of peroxidase and CLC protein reactivity that implicate the macrophage in the biology of eosinophil-rich inflammatory processes and of these eosinophil-derived products.