Effects of reactive oxygen and nitrogen metabolites on
MCP-1-induced monocyte chemotactic activity in vitro
Etsuro
Sato1,
Keith L.
Simpson1,
Matthew B.
Grisham2,
Sekiya
Koyama3, and
Richard A.
Robbins1
1 Research Services, Tucson and
Overton Brooks Veterans Affairs Medical Centers, and Department of
Medicine, University of Arizona, Tucson, Arizona 85723;
2 Department of Molecular and
Cellular Physiology, Louisiana State University Medical Center,
Shreveport, Louisiana 71131; and
3 The First Department of Internal
Medicine, Shinshu University School of Medicine, Matsumoto 390-8621, Japan
 |
ABSTRACT |
Peroxynitrite, an
oxidant generated by the interaction between superoxide and nitric
oxide (NO), can nitrate tyrosine residues, resulting in compromised
protein function. Monocyte chemoattractant protein-1 (MCP-1) is a
chemokine that attracts monocytes and has a tyrosine residue critical
for function. We hypothesized that peroxynitrite would alter MCP-1
activity. Peroxynitrite attenuated MCP-1-induced monocyte chemotactic
activity (MCA) in a dose-dependent manner
(P < 0.05) but did not attenuate
leukotriene B4 or
complement-activated serum MCA. The reducing agents dithionite,
deferoxamine, and dithiothreitol reversed the MCA inhibition by
peroxynitrite, and exogenous L-tyrosine abrogated the
inhibition by peroxynitrite. PAPA-NONOate, an NO donor, or superoxide
generated by xanthine and xanthine oxidase did not show an inhibitory
effect on MCA induced by MCP-1. The peroxynitrite generator
3-morpholinosydnonimine caused a concentration-dependent inhibition of
MCA by MCP-1. Peroxynitrite reduced MCP-1 binding to monocytes and
resulted in nitrotyrosine formation. These findings are consistent with
nitration of tyrosine by peroxynitrite, with subsequent inhibition of
MCP-1 binding to monocytes, and suggest that peroxynitrite may play a
role in regulation of MCP-1-induced monocyte chemotaxis.
nitric oxide; monocyte migration; superoxide; nitrotyrosine; monocyte chemoattractant protein-1
 |
INTRODUCTION |
MONOCYTE CHEMOATTRACTANT protein-1 (MCP-1) belongs to a
family of low-molecular-weight proteins collectively called chemokines, which mediate the recruitment of specific subsets of leukocytes including monocytes and lymphocytes (11, 18, 29). The activity of MCP-1
in inducing monocyte infiltration implies its role in several
inflammatory diseases such as rheumatoid arthritis (19, 26),
atherosclerosis (33, 42, 49), and inflammatory bowel disease (17).
Peroxynitrite, the reaction product of superoxide with nitric oxide
(NO) (5, 22), is a strong oxidant (28) that oxidizes proteins (32),
lipids (36), DNA (25), and low-molecular-weight biomolecules such as
glutathione (35), methionine (34), and ascorbate (3). In addition to
its role in oxidative reactions, peroxynitrite also nitrates free or
protein-associated tyrosine residues to form the stable product
nitrotyrosine by addition of a nitro group to the three-position
adjacent to the hydroxyl group of tyrosine (22, 25). Peroxynitrite is
known to alter the function of some proteins. For example,
peroxynitrite inactivates manganese superoxide dismutase activity;
3-nitrotyrosine is increased in a tissue homogenate of transplanted
renal allografts during chronic rejection (30); and peroxynitrite
inhibited protein phosphorylation by tyrosine kinases, thus interfering
with signal transduction mechanisms (16).
Recently, the tyrosine residue at position 13 of MCP-1 has been shown
to be important in monocyte chemotaxis. Substitution of an alanine
results in a dramatic loss in MCP-1 function (41). We hypothesized that
peroxynitrite might regulate monocyte chemotactic activity (MCA) by
nitrating tyrosine and modulating MCP-1-induced MCA. To test this
hypothesis, the chemotactic responses of human monocytes to MCP-1
incubated with peroxynitrite or the peroxynitrite generator
3-morpholinosydnonimine (SIN-1) were evaluated in vitro. The results
suggest that peroxynitrite may play a regulatory role in inflammation
by regulating MCA.
 |
METHODS |
Measurement of MCA. Mononuclear cells
for the chemotaxis assay were obtained from nonsmoking healthy
volunteers by Ficoll-Hypaque density centrifugation (Histopaque 1077;
Sigma Chemical, St. Louis, MO) to separate the red blood cells and
neutrophils from the mononuclear cells (8). The mononuclear cells were
harvested at the interface, centrifuged at 400 g for 5 min, and washed three times
with Hanks' balanced salt solution (HBSS; Biofluids, Rockville, MD).
The resulting cell pellet routinely consisted of ~30% monocytes and
70% lymphocytes by morphology and esterase staining (Sigma) with
>98% viable cells. The cells were suspended in Gey's balanced salt
solution (Life Technologies, Grand Island, NY) containing 2% BSA
(Sigma) at a final concentration of 5 × 106 cells/ml.
To ensure that lymphocytes were not playing a major role in regulating
human MCA, some experiments were performed using highly purified
monocytes. Mononuclear cells obtained by Ficoll-Hypaque density
centrifugation were suspended at a concentration of 2 × 106 cells/ml. Two milliliters of
the cell suspension were placed in tissue culture flasks (Corning) for
90 min at 37°C. The supernatant fluids containing the nonadherent
lymphocytes were removed, and the monocytes were detached by adding 0.2 ml of 0.05% trypsin and 0.53 mM EDTA.
MCA was assayed in 48-well microchemotaxis chambers (Neuro Probe, Cabin
John, MD) as previously described (20). The bottom wells of the chamber
were filled with 25 µl of the chemotactic stimulus or medium in
duplicate. A 10-µm-thick polyvinylpyrrolidone-free polycarbonate
filter with a pore size of 5 µm was placed over the samples. The
silicon gasket and the upper pieces of the chamber were applied, and 50 µl of the cell suspension were placed in the upper wells. The
chambers were incubated in humidified air in 5%
CO2 at 37°C for 90 min.
Nonmigrated cells were wiped away from the filter. The filter was
immersed in methanol for 5 min, stained with a modified Wright's
stain, and mounted on a glass slide. Cells that had completely migrated
through the filter were counted using light microscopy. MCA is
expressed as the mean number of migrated cells per high-power field
from duplicate wells.
Effect of peroxynitrite on MCP-1-induced
MCA. Peroxynitrite was evaluated for its capacity to
modulate MCP-1-induced MCA in vitro. Recombinant human MCP-1 (R&D
Systems, Minneapolis, MN) was incubated for 2 h at 37°C with each
concentration of peroxynitrite (Calbiochem, La Jolla, CA) before the
MCA assay. In control experiments, MCP-1 was incubated with medium alone.
Effect of peroxynitrite on leukotriene
B4- and activated serum-induced MCA.
The capacity of peroxynitrite to modulate leukotriene
B4
(LTB4)- and activated
serum-induced MCA was similarly evaluated and compared with MCP-1.
LTB4
(10
6 M; Sigma) or
complement-activated serum (1:10 dilution; see Ref. 40) was incubated
with peroxynitrite (10
4 M)
for 2 h at 37°C before the MCA assay was performed.
Effect of PAPA-NONOate on MCP-1-induced
MCA. To evaluate the effect of NO alone on
MCP-1-induced MCA, PAPA-NONOate (Alexis, San Diego, CA) was used as an
NO donor (47). MCP-1 (10
7
g/ml) was incubated with PAPA-NONOate
(10
3 to
10
6 M) for 2 h at 37°C
before the MCA assay was performed. The samples were dialyzed overnight
at 4°C against HBSS using tubing with a molecular-mass cutoff of 3 kDa to remove NO because NO has been reported to effect cell migration.
The half-life of PAPA-NONOate is 15 min in physiological buffer at
37°C, and two moles of NO are released per mole of PAPA-NONOate
(24).
Effect of xanthine/xanthine oxidase on MCP-1
MCA. To evaluate the effect of superoxide on
MCP-1-induced MCA, xanthine
(10
6 to
10
3 M; Sigma) and xanthine
oxidase (3.4 × 10
6,
3.4 × 10
5, 3.4 × 10
4, and 3.4 × 10
3 U/ml; Sigma)
were combined to produce superoxide (14). MCP-1 (10
7 g/ml) was incubated
with xanthine alone, xanthine oxidase alone, or xanthine and xanthine
oxidase for 2 h at 37°C before the MCA assay was performed.
Effect of SIN-1 on MCP-1-induced MCA.
To confirm the effect of peroxynitrite on MCP-1, SIN-1 (Alexis), a
peroxynitrite generator (21), was evaluated by incubating MCP-1
(10
7 g/ml) with SIN-1
(10
3 to
10
8 M) for 2 h at 37°C
before the MCA assay was performed. At pH 7.4 and 37°C, SIN-1
generates peroxynitrite at a rate roughly equivalent to 1% of the
SIN-1 concentration per minute, e.g., 1 mM SIN-1 produces ~10 µM
peroxynitrite/min. SIN-1 decomposition is essentially complete after 3 h (12).
Effect of reducing agents on peroxynitrite-induced
attenuation of MCA by MCP-1. The capacity of the
reducing agents dithiothreitol, deferoxamine, or dithionite to
attenuate the effect of peroxynitrite on MCP-1-induced MCA by
peroxynitrite was assessed. Dithiothreitol (1 mM; Sigma), deferoxamine
(50 µM; Sigma), dithionite (1 mM; Fluka, St. Louis, MO), and
peroxynitrite (10
4 M) were
added to MCP-1 (10
7 g/ml)
and incubated for 2 h at 37°C before evaluation of MCA.
Effect of L-tyrosine on
peroxynitrite-induced attenuation of MCA by MCP-1. The
capacity of L-tyrosine to reserve the attenuation of MCA
induced by peroxynitrite was assessed by addition of
L-tyrosine (10
3
and 10
4 M; Sigma) to MCP-1
(10
7 g/ml) before exposure
to peroxynitrite (10
4 M).
Detection of nitrotyrosine on MCP-1 incubated with
peroxynitrite. Nitrotyrosine on MCP-1 incubated with
peroxynitrite was evaluated using modifications of a previously
described ELISA (23). MCP-1 (100 ng/ml) was incubated with
peroxynitrite (100 µM) as above and frozen until assayed. Goat
anti-human MCP-1 IgG (R&D Systems) was dissolved in Voller buffer (1.59 g sodium carbonate, 2.93 g sodium bicarbonate, and 0.2 g sodium azide
in 1 liter of distilled water, pH = 9.6) at a final concentration 200 ng/ml. Two hundred milliliters were added to flat-bottomed 96-well
plates (Costar, Cambridge, MA) and were allowed to adsorb to the
plastic overnight at 4°C. After the flat-bottomed plate was washed
three times with PBS-Tween (0.075% Tween 20; Sigma), 200 µl of MCP-1 were added to the MCP-1 antibody-coated plate and incubated for 60 min
at room temperature. After three washes with PBS-Tween, 200 µl of a
1:400 dilution of rabbit polyclonal anti-nitrotyrosine (Calbiochem)
were added to the wells and incubated for 90 min. After another three
washes with PBS-Tween, 200 µl of a 1:500 dilution of
peroxidase-conjugated anti-rabbit IgG were added to the wells and
incubated for 90 min. Two hundred microliters of
o-phenylenediamine (100 µg/ml;
Sigma) in 0.003%
H2O2
were added and visually monitored. The reaction was terminated by
addition of 25 µl of 8 N
H2SO4, and the absorbance was read at 490 nm.
In separate experiments, plates were coated with MCP-1 that had been
either exposed or not exposed to peroxynitrite. Subsequently, anti-nitrotyrosine, peroxidase-conjugated goat anti-rabbit IgG, and
o-phenylenediamine were added
sequentially, with washing between each addition. The reaction was
terminated with
H2SO4,
and the absorbance was read at 490 nm.
Effect of peroxynitrite on MCP-1 binding to
monocytes. To investigate the peroxynitrite effect on
MCP-1 binding to monocytes, MCP-1 was incubated with 100 µM of
peroxynitrite for 2 h at 37°C. In control experiments, MCP-1 was
incubated with medium alone. Subsequently, MCP-1 with or without
peroxynitrite was incubated with monocytes
(106 cells) at 4°C for 30 min.
Next, supernatants were removed, and monocytes were washed three times
by HBSS. Monocytes were suspended in 1 ml PBS-Tween, sonicated for 20 s
(MSE Soniprep, Crawley, UK), and then centrifuged at 20,000 g for 30 min in a refrigerated microcentrifuge to obtain a supernatant (soluble) and particulate fraction. MCP-1 was measured using a commercially available MCP-1 ELISA
(R&D Systems).
Statistics. Data were analyzed by
one-way ANOVA. In all cases, a P value
of <0.05 was considered significant. The data are expressed as means ± SE.
 |
RESULTS |
Effect of peroxynitrite on MCA by
MCP-1. Differing concentrations of MCP-1 were incubated
with peroxynitrite (100 µM). At each concentration, exposure to
peroxynitrite caused a reduction in MCA (Fig.
1; n = 4 experiments, P < 0.05). Incubation
of MCP-1 (100 ng/ml) with various amounts of peroxynitrite induced a
significant, concentration-dependent attenuation of MCA (Fig.
2; n = 4, P < 0.05). The lowest dose of
peroxynitrite tested, 10
5
M, significantly inhibited MCA (P < 0.05). Peroxynitrite itself was not chemotactic for monocytes (data not
shown). Similarly, incubation of peroxynitrite (100 µM) with the
monocytes before the chemotaxis assay did not inhibit MCA to MCP-1
(data not shown).

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Fig. 1.
Inhibition of monocyte chemotactic activity (MCA) by peroxynitrite.
Monocyte chemoattractant protein-1 (MCP-1) was incubated for 2 h in the
presence of medium alone or 100 µM peroxynitrite;
n = 4 experiments for each condition.
, MCA by MCP-1 with 100 µM peroxynitrite; , MCA by MCP-1
without peroxynitrite incubation.
* P < 0.05 compared with MCA
by MCP-1 incubated with medium alone.
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Fig. 2.
Dose-responsive inhibition of MCA by peroxynitrite. MCP-1 (100 ng/ml)
was incubated for 2 h in the presence of medium alone or peroxynitrite
at the concentration indicated; n = 4 experiments for each condition.
* P < 0.05 compared with MCP-1
incubated with medium alone.
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Effect of peroxynitrite on LTB4- and
activated serum-induced MCA.
To ensure that the effect of peroxynitrite was not a nonspecific effect
on monocyte chemotaxis, the effect of peroxynitrite on MCA induced by
MCP-1 (Fig.
3A),
complement-activated serum (Fig.
3B), and
LTB4 (Fig.
3C) was assessed. Peroxynitrite did not significantly inhibit the MCA of
LTB4 or complement-activated serum.

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Fig. 3.
Effect of peroxynitrite on MCA induced by MCP-1
(A), activated serum
(B), and leukotriene
B4
(LTB4;
C). MCP-1, activated serum, and
LTB4 were incubated for 2 h in the
presence of medium alone or 100 µM peroxynitrite;
n = 4 experiments for each condition.
* P < 0.05 compared with MCP-1
incubated with medium alone.
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Effect of PAPA-NONOate on MCP-1-induced
MCA. To investigate the capacity of NO to modulate
MCP-1-induced MCA, the effect of the NO donor PAPA-NONOate was
evaluated. PAPA-NONOate
(10
3 to
10
6 M) did not
significantly change MCA induced by MCP-1
(P > 0.05, all comparisons).
Effect of xanthine/xanthine oxidase on MCP-1-induced
MCA. To evaluate the effect of superoxide on MCA
induced by MCP-1, MCP-1 was incubated with xanthine
(10
6 to
10
3 M), xanthine oxidase
(3.4 × 10
6, 3.4 × 10
5, 3.4 × 10
4, and 3.4 × 10
3 U/ml), or xanthine and
xanthine oxidase. None significantly altered MCA to MCP-1
(P > 0.05, all comparisons).
Effect of SIN-1 on MCP-1-induced MCA.
SIN-1 spontaneously decomposes under aqueous conditions, generating
first O
2 and then NO at comparable rates.
SIN-1 induced a significant, concentration-dependent attenuation of MCA
by MCP-1 (Fig. 4;
n = 6, P < 0.05). The lowest dose of SIN-1
to inhibit MCA was 10
6 M
(P < 0.05). One hundred micromolar
SIN-1 induced ~80% inhibition of MCA by MCP-1. SIN-1 itself was not
chemotactic for monocytes (data not shown).

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Fig. 4.
Effect of 3-morpholinosydnonimine (SIN-1) on MCA induced by MCP-1.
MCP-1 was incubated for 2 h in the presence of medium alone or SIN-1 at
the concentration indicated; n = 4 experiments for each condition.
* P < 0.05 compared with MCP-1
incubated with medium alone.
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Effect of dithionite, dithiothreitol, and deferoxamine
on peroxynitrite-induced attenuation of MCA by MCP-1.
The reducing agents dithiothreitol, deferoxamine, and dithionite were
added to MCP-1 before incubation with peroxynitrite. Each inhibited the
peroxynitrite effect on MCA (Fig. 5;
n = 4, P < 0.05). Dithionite, dithiothreitol, and deferoxamine alone were not chemotactic for monocytes (data not shown).

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Fig. 5.
Effect of dithionite, dithiothreitol, and deferoxamine on peroxynitrite
(ONOO )-induced
attenuation of MCA by MCP-1. MCP-1 was cultured with peroxynitrite for
2 h in the presence of dithionite, dithiothreitol, and deferoxamine at
the concentration indicated; n = 4 experiments for each condition. HBSS, Hanks' balanced salt solution.
* P < 0.05 compared with MCP-1
incubated with peroxynitrite.
** P < 0.05 compared with
MCP-1 incubated with medium alone.
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Effect of L-tyrosine on
peroxynitrite-induced attenuation of MCA by MCP-1. One
mechanism of peroxynitrite inhibition may be through nitrating tyrosine
residues. Therefore, the effect of L-tyrosine addition to
MCP-1 before incubation with peroxynitrite was investigated. Addition
of L-tyrosine to MCP-1 abrogated the attenuation of MCA
induced by peroxynitrite (Fig. 6;
n = 4, P < 0.05). The addition of 100 µM
L-tyrosine prevented the inhibition of MCA induced by 100 µM of peroxynitrite. L-Tyrosine itself was not
chemotactic for monocytes (data not shown).

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Fig. 6.
Effect of L-tyrosine on peroxynitrite-induced attenuation
of MCA by MCP-1. MCP-1 was cultured with 100 µM of peroxynitrite for
2 h in the presence of medium alone or L-tyrosine at the
concentration indicated; n = 4 experiments for each condition.
* P < 0.05 compared with MCP-1
incubated with peroxynitrite.
** P < 0.05 compared with
MCP-1 incubated with medium alone.
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Detection of nitrotyrosine on MCP-1 incubated with
peroxynitrite. Optical density of MCP-1 with
peroxynitrite incubation was significantly higher than that of MCP-1
without peroxynitrite incubation. Peroxynitrite resulted in
nitrotyrosine formation on MCP-1 (Fig. 7;
n = 6, P < 0.05).

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Fig. 7.
Nitrotyrosine detection of MCP-1 incubated with peroxynitrite. MCP-1
was incubated with 100 µM of peroxynitrite for 2 h. MCP-1 with and
without peroxynitrite incubation was allowed to adsorb to an uncoated
plate (A) or was added to the MCP-1
antibody-coated plate (B).
* P < 0.05 compared with MCP-1
incubated without peroxynitrite.
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Effect of peroxynitrite on MCP-1 binding to
monocytes. MCP-1 induces chemotactic activity by
binding to monocytes. Addition of peroxynitrite to MCP-1 resulted in an
inhibition of MCP-1 binding to monocytes (Fig.
8; n = 4, P < 0.05).

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Fig. 8.
Effects of peroxynitrite on MCP-1 binding to monocytes. MCP-1 was
incubated with and without 100 µM of peroxynitrite for 2 h at
37°C, and after that, MCP-1 was incubated with monocytes
(106 cells) at 37°C for 30 min. MCP-1 binding to monocytes was measured by ELISA.
* P < 0.05 compared with MCP-1
incubated with medium alone.
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 |
DISCUSSION |
The results of this study show that peroxynitrite significantly
attenuated MCP-1-induced MCA in vitro. Sodium dithionite, deferoxamine,
dithiothreitol, or tyrosine attenuated the inhibition. NO or superoxide
did not cause a reduction in MCP-1-induced MCA because PAPA-NONOate and
xanthine/xanthine oxidase did not show an inhibitory effect. The
peroxynitrite donor SIN-1 induced a significant,
concentration-dependent inhibition of MCA by MCP-1. Nitrotyrosine was
detected in the MCP-1 incubated with peroxynitrite by ELISA. These data
suggest that peroxynitrite may play an important role in regulating
human monocyte locomotion in response to MCP-1.
NO has been reported to inhibit monocyte chemotaxis (4) and to inhibit
MCP-1 expression (44). Other studies have reported that NO synthase
inhibitors attenuate MCA to a variety of stimuli (6). However, in this
study, we examined the effect of an end product of NO production,
peroxynitrite, on a chemotactic factor itself.
The concentration of peroxynitrite in vivo is unknown. Peroxynitrite is
a transient intermediate in free radical chemistry and is highly
reactive at physiological pH. Thom et al. (43) reported that
nitrotyrosine concentrations in lung homogenates were 30-60 ng/mg
protein. If peroxynitrite concentrations were of the same order, the
peroxynitrite concentration in this experiment should be sufficient to
modulate MCP-1 function.
MCP-1 is a prototypical C-C chemokine that was isolated on the basis of
its ability to attract monocytes in Boyden chambers in vitro (31, 45,
50). MCP-1 expression has been detected in a variety of pathological
conditions including atherosclerosis and rheumatoid arthritis (19, 26,
33, 42, 49). Various inflammatory mediators are reported to enhance
production of NO and superoxide, and they may lead to the formation of
peroxynitrite during inflammation. Evidence of peroxynitrite reaction
products, specifically immunoreactive 3-nitrotyrosine residues, occurs
in lung tissue sections from patients with bronchial asthma (38) and
adult respiratory distress syndrome (27) and in atherosclerotic blood
vessels (10). Chemotactic factors, including MCP-1, are likely to be
exposed to high local concentrations of NO, superoxide, and
peroxynitrite at inflammatory sites.
Coincubation of MCP-1 with several peroxynitrite scavengers ameliorated
the peroxynitrite-induced MCA inhibition. The protective effect of
dithiothreitol on MCP-1 has been suggested to be by attenuating the
effects of nitrotyrosine on cysteine residues; however, dithiothreitol
has also been reported to prevent peroxynitrite-mediated nitration of
tyrosine (37). The iron chelator deferoxamine also inhibited
peroxynitrite-induced inhibition of MCP-1 chemotactic activity but is
also a scavenger of peroxynitrite reaction independent of iron
chelation (13). Dithionite, which has been proposed to modify
3-nitrotyrosine by substitution of an amine group (2), ameliorated the
peroxynitrite inhibition. In addition, L-tyrosine abrogated
the peroxynitrite MCA inhibition. These results are consistent with
tyrosine nitration by peroxynitrite as a mechanism for MCP-1 inhibition.
MCP-1 is a 76-amino acid chemokine, and it has two tyrosine residues.
Several studies reported that tyrosine nitration is related to
inactivation of protein and enzymatic activity (30, 48). Consistent
with the concept that tyrosine is important in binding of MCP-1 to its
receptor, Zhang et al. (51) reported that changing tyrosine-28 to
aspartate or arginine-30 to leucine produced proteins with essentially
no monocyte chemoattractant activity. Steitz et al. (41) reported that
point mutations of tyrosine-13 greatly lowered MCP-1 receptor binding
and activity. Our findings of nitrotyrosine formation on MCP-1 after
peroxynitrite are consistent with these observations and suggest that
tyrosine nitration by peroxynitrite on MCP-1 receptor is a likely
mechanism altering MCP-1 binding and chemotactic function. However,
deletion or mutational analysis of MCP-1 shows that other regions of
the molecule contribute to its activity. Peroxynitrite may potentially affect protein function by other mechanisms, including methionine (46), tryptophan (1), or formation of S-nitrosothiol groups on
cysteine residues (15).
Although NO and peroxynitrite are physiological regulators, they have
been shown to alter respiration (7, 39) and induce cell death (9). To
estimate the effect of peroxynitrite on monocytes, we incubated
monocytes with peroxynitrite for 90 min at 37°C before chemotaxis
experiments. It induced no significant cytotoxicity, as assessed by
trypan blue exclusion, compared with medium alone, and it had no
significant effect on MCA by MCP-1.
In summary, we found that peroxynitrite modulates MCP-1-induced MCA in
vitro. The results are consistent with the mechanism of inhibition
being nitration of a tyrosine residue. These data demonstrate that
peroxynitrite attenuates MCP-1 chemotactic activity and suggest a role
for peroxynitrite in regulating human monocyte locomotion during inflammation.
 |
ACKNOWLEDGEMENTS |
This work was supported by a Merit Review grant from the Department
of Veterans Affairs and a grant from Rotary International.
 |
FOOTNOTES |
The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement"
in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Address for reprint requests and other correspondence: R. A. Robbins,
Research Health Care Group, Tucson VA Medical Center, 3601 S. 6th Ave.,
Tucson, AZ 85723 (E-mail:
Richard.Robbins2{at}med.va.gov).
Received 17 December 1998; accepted in final form 16 April 1999.
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