Morphological and functional analysis of PTA granules in human NK cells |
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Authors: | S A Kolb and Peter Groscurth |
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Institution: | (1) Institute of Anatomy, Division of Cell Biology, University of Zurich-Irchel, Winterthurerstrasse 190, CH-8057 Zurich, Switzerland Tel.: +41-1-2575311; Fax: +41-1-2575702, CH |
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Abstract: | Human natural killer (NK) cells contain unique granules with parallel tubular arrays (PTA granules) of approximately 30 nm
diameter that can be seen only by electron microscopy. In order to clarify the role of PTA granules in NK cell-mediated cytolysis
we examined these structures with regard to frequency and expression of lytic proteins (perforin, granzymes). NK cells (CD3−, CD16+, CD56+) were obtained from heparinized blood of healthy donors and enriched by double-step negative selection using mAb coupled
to magnetic beads. PTA granules were found in 31.3% of freshly separated NK cells. When NK cells were cultivated, even in
the presence of various stimulating agents (rhIL-2, rhIL-4, rhIL-6, rhIL-12, GM-CSF, rhIFNα, anti-CD16 mAb, dexamethasone),
PTA granules disappeared and transformed into conventional primary lysosomes. By immune electron microscopy using antibodies
directed against perforin and granzyme B we observed distinct immuno-reactivity in the tubules and in the tubule-associated
faintly electron-dense matrix of PTA granules. Immuno-labelling for perforin and granzyme B was also found in the fine granular
matrix of primary lysosomes. Finally, we tested the distribution of chondroitin 4-sulfate which is suggested to inactivate
lytic proteins. Immuno-reactivity for chondroitin 4-sulfate was detected only in the moderately electron-dense matrix but
not in the tubules of PTA granules. These observations indicate that perforin and granzyme B are stored in an inactive form
in PTA tubules due to highly ordered paracrystalline protein folding. As soon as the tubules decay the lytic proteins are
kept in an environment of chondroitin 4-sulfate for inactivation.
Accepted: 24 March 1997 |
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Keywords: | Natural killer cells Ultrastructure Parallel tubular arrays Perforin Granzyme B Chondroitin sulfate |
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