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Electron Spin Resonance Spectroscopy Does Not Reveal Hydroxyl Radical Production in Activated Natural Killer Lymphocytes
Author(s) -
Gibboney James J.,
Haak Richard A.,
Kleinhans Frederick W.,
Brahmi Zacharie
Publication year - 1988
Publication title -
journal of leukocyte biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.819
H-Index - 191
eISSN - 1938-3673
pISSN - 0741-5400
DOI - 10.1002/jlb.44.6.545
Subject(s) - hydroxyl radical , radical , natural killer cell , electron paramagnetic resonance , chemistry , cytotoxicity , spin trapping , biochemistry , microbiology and biotechnology , biology , in vitro , physics , nuclear magnetic resonance
We investigated hydroxyl radical (OH) production by human natural killer (NK) cells, using electron spin resonance (ESR) spectroscopy and 5,5 dimethyl‐1‐pyrroline‐N‐oxide (DMPO), a spin trap specific for OH production. We confirmed that hydroxyl radical scavengers, n‐propyl gallate and catechin, inhibited NK cell‐mediated cytotoxicity (NK‐CMC) in a dose‐dependent manner and demonstrated that DMPO also inhibited NK‐CMC. Polymorphonuclear leukocytes (PMNL) activated by opsonized zymosan (2.4 mg/ml) and mixed with DMPO (0.14 M) showed an early increase in hydroxyl radical production, leading to a net production of free radical of almost 400 pMol/10 6 cells. We then mixed NK cells with K562, an NK‐sensitive tumor cell, at a 1:1 ratio and added DMPO (0.14 M). We pelleted the cells to increase EC to TC binding before taking the sample readings. Activated NK cells showed no increase in OH production, leading to a net production of free radicals less than 1% that of activated PMNL. These data strongly suggest that hydroxyl radical production does not play a role in the early events of NK cell activation; they indicate a need to reevaluate the mechanism of inhibition of NK‐CMC by OH scavengers.