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Defensin-rich dense granules of human neutrophils
WG Rice, T Ganz, JM Kinkade , ME Selsted, RI Lehrer and RT Parmley
Defensins are a newly recognized class of small, cationic polypeptides that
have in vitro microbicidal activity toward certain bacteria, fungi, and
viruses. Human neutrophil granules were separated into 13 density fractions
by using a high-resolution Percoll gradient centrifugation procedure, and
the distribution of the three defensin polypeptides in these fractions was
determined. Levels of defensins and several granule marker proteins were
estimated in each fraction from relative staining intensities of bands
following acid-urea and sodium dodecyl sulfate-polyacrylamide gel
electrophoresis (SDS-PAGE) of total acid-extractable proteins. These
results were confirmed by enzyme immunoassay measurements of defensins and
quantitative determinations of the typical azurophil granule components,
myeloperoxidase, beta- glucuronidase, lysozyme, and elastase. The five
higher density granule fractions (H1 through H5) contained fourfold higher
relative amounts of defensins as compared with the eight lower density
fractions (L1 through L8), accounting for approximately 50% of the total
protein. In particular, fraction H5 was especially enriched in defensins
but was relatively deficient in myeloperoxidase, beta-glucuronidase,
lysozyme, and elastase. Ultrastructural morphology showed that fraction H5
contained the largest granules. Seventy percent of these granules exhibited
electron-dense rims and electron-lucent central regions when stained with
methanolic uranyl acetate-lead citrate, and 70% showed this same
characteristic rim-staining pattern after limited reaction (30 minutes) for
peroxidase with diaminobenzidine. These distinctively large, rim-stained
granules were identified in intact, mature peripheral blood neutrophils as
well as in human bone marrow promyelocytes, indicating that their synthesis
occurs during early myeloid development. This unusual granule type may play
a specialized role in the microbicidal functions of the neutrophil,
distinct from that of typical azurophil granules.
Volume 70,
Issue 3,
pp. 757-765,
09/01/1987
Copyright © 1987 by The American Society of Hematology
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