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  • Title: An ultrastructural analysis of tumor-promoting phorbol diester-induced degranulation of human basophils.
    Author: Dvorak AM, Warner JA, Morgan E, Kissell-Rainville S, Lichtenstein LM, MacGlashan DW.
    Journal: Am J Pathol; 1992 Dec; 141(6):1309-22. PubMed ID: 1466396.
    Abstract:
    Release reactions stimulated in human basophils by a variety of secretagogues show biochemical and morphologic differences as well as similarities. Biochemical differences include those of rate, amount, and order of mediator release, as well as mediator type released or generated. Morphologic diversity of release reactions includes prototypic anaphylactic degranulation (AND), or piecemeal degranulation (PMD), and a continuum of anatomic release comprised of PMD followed by AND that is seen when human basophils are stimulated by the bacterial peptide, formyl methionyl leucyl phenylalanine (FMLP). AND is characterized by extrusion of membrane-free granules through multiple plasma membrane pores; PMD is characterized by partially to completely empty, nonfused granule containers in the cytoplasm of basophils. AND is further characterized by diminished-to-absent granules and reduced cytoplasmic vesicles at peak histamine release intervals; PMD does not show decreases in numbers of granules, and cytoplasmic vesicles are plentiful. Smooth membrane-bound vesicles with granule particles and vesicles that appear empty comprise this organelle population. PMD is the single most evident activation change present in basophils that traffic into tissues in multiple diseases in vivo. In this study, we examined the ultrastructural kinetic morphology associated with stimulation of human basophils with tetradecanoyl phorbol acetate (TPA)--a tumor-promoting phorbol diester known to elicit histamine (but not LTC4) release. Partially purified human basophils were prepared for electron microscopy and examined either after control incubations in buffer alone or at 0 time, 1, 2, 5, 10, 30, and 45 minutes after TPA stimulation. Standard morphology and ultrastructural quantitation of vesicles and granules and contents of vesicles or alteration of granules was done and compared with previous ultrastructural kinetic analyses of human basophil release reactions stimulated by different triggers. Like biochemical studies that have determined that TPA is a unique secretogogue for human basophils, the morphology stimulated by TPA and associated with histamine release was also unique. For example, very minor images of AND were evident. Far greater amounts of PMD were imaged. PMD was associated with approximately 50% alteration of cytoplasmic granules by 45 minutes after TPA stimulation. This evidence of empty granules was associated with, and preceded by, a rapid, extensive, and sustained increase in particle-containing cytoplasmic vesicles, as compared with buffer controls (P < 0.001 for each TPA stimulation time compared with unstimulated basophils). In addition, previously undescribed interactions of releasing granules and their overlying plasma membranes characterized TPA-stimulated cells.(ABSTRACT TRUNCATED AT 400 WORDS)
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