(Journal of Leukocyte Biology. 2000;68:194-200.)
© 2000
by Society for Leukocyte Biology
Effects of theophylline on human eosinophil functions: comparative study with neutrophil functions
Kozo Yasui,
Kazunaga Agematsu,
Koji Shinozaki,
Sho Hokibara,
Haruo Nagumo,
Shinji Yamada,
Norimoto Kobayashi and
Atsushi Komiyama
Department of Pediatrics, Shinshu University School of Medicine, Matsumoto, Japan
Correspondence: Kozo Yasui, Department of Pediatrics, Shinshu University School of Medicine, Asahi 3-1-1, Matsumoto 390-8621, Japan. E-mail: koyasui{at}gipac.shinshu-u.ac.jp
 |
ABSTRACT
|
|---|
The understanding of theophylline as a bronchodilator has been
reconsidered in recent years. We undertook to determine its
immunomodulatory actions in granulocytes and elucidate their mechanism.
Preincubation of neutrophils with theophylline (10-5 to
5 x 10-3 M) had a biphasic effect on
O2- production stimulated with
N-formyl-methionyl-leucyl-phenylalanine or C5a.
Theophylline potentiates O2- production via
adenosine A2A receptor antagonism induced by
receptor-linked agonists from neutrophils, but not from eosinophils.
The addition of theophylline caused a significant decline in neutrophil
chemotaxis at lower concentrations than those for eosinophil motility.
Theophylline reduces neutrophil chemotaxis via adenosine A1
receptor antagonism. At high concentrations, with an intracellular cAMP
accumulation as a result of phosphodiesterase (PDE) inhibition,
theophylline also exerts an inhibitory effect on the
O2- production and chemotaxis of both types of
cells. The difference in theophyllines effect on neutrophils and
eosinophils appears to depend on the existence of specific adenosine
receptors. Theophylline thus modulates granulocyte functions in
association with specific adenosine receptor antagonism and cAMP-PDE
inhibition.
Key Words: superoxide anion production chemotaxis adenosine receptor antagonism airway inflammation
 |
INTRODUCTION
|
|---|
Theophylline has been widely used in the treatment of patients
with bronchial asthma. The primary pharmacological mechanism of
theophylline has long been considered to be bronchodilation due to the
inhibition of cyclic nucleotide phosphodiesterase (PDE) isoenzymes,
leading to accumulation of adenosine 3,5-cyclic monophosphate
(cAMP) in the airway to relax smooth muscle. However, this view has
come under review in recent years. The hypothesis was challenged on the
basis of the observation that the concentrations of theophylline
required to induce airway smooth muscle relaxation in vitro
are higher than the therapeutic concentrations usually achieved in
plasma (520 µg/mL; 3 to 11 x 10-5 M), so that
therapeutic concentrations of theophylline can produce a slight
inhibition of PDE activity [1
2
3
]. These findings
suggest that theophylline has a beneficial effect on asthma control
that is not related to PDE inhibition or mild bronchodilator action.
This effect is thought to involve the suppression of the inflammatory
processes underlying the development and exacerbation of asthma.
There is increasing evidence that theophylline has anti-asthma
properties other than bronchodilation, which include anti-inflammatory
actions [4
, 5
]. The inhibition of a late
asthmatic response by theophylline has been demonstrated in conjunction
with significant reduction of inflammatory cells in the airway
[6
, 7
], and these effects were seen at
plasma concentrations below those traditionally regarded as
therapeutically useful [6
]. Bronchial
hyper-responsiveness is an important feature in asthma, while
inflammation associated with granulocyte (neutrophil as well as
eosinophil) infiltration is involved in its development. Granulocytes
are thought to damage the normal airway tissue through releasing toxic
substances such as superoxide and harmful proteins.
Because neutrophil and eosinophil activation is associated with
late-phase asthmatic reactions [3
, 6
,
7
], to elucidate the effects of theophylline on these
cells may be of importance.
Several studies have been published on the effects of theophylline on
neutrophil functions such as generation of oxygen metabolites and
chemotactic movement [8
9
10
11
12
]. It has been established
that the physiological mechanism for the effect of theophylline on the
modulation of neutrophil functions may be dependent on adenosine
receptor antagonism and/or on PDE inhibition. Adenosine, which is
normally present in plasma at concentrations up to 3 x
10-7 M [13
, 14
], is one of the
pharmacological and hormonal agents that modulate neutrophil functions
in vitro [10
, 14
15
16
17
18
].
Neutrophils release a certain amount of adenosine [13
],
and adenosine A1 and A2A but not
A2B receptors are known to be strongly expressed on
neutrophils [19
, 20
]. Theophylline (an
A1, A2A, and A2B antagonist) is
believed to modulate neutrophil functions at least in part through
selective adenosine receptor antagonism. However, very limited
information on the effects and mechanism of theophylline on eosinophil
functions is available compared with those on neutrophils, and little
confused [21
22
23
24
].
This study was undertaken to determine the involvement of adenosine
receptor antagonism and PDE inhibition in the effect of theophylline on
eosinophil functions and to compare it with that on neutrophils to
reach a consolidated view of the functions and mechanisms of
theophylline.
 |
MATERIALS AND METHODS
|
|---|
Reagents
Dextran T500 was purchased from Pharmacia (Uppsala, Sweden).
N-formyl-methionyl-leucyl-phenylalanine (fMLP), complement
C5a, phorbol 12-myristate 13-acetate (PMA), cytochrome c
(type VI), N-ethylmalemide, superoxide dismutase,
theophylline, dibutyryl-adenosine 3,5-monophosphate (dbcAMP),
adenosine deaminase (ADA; type V),
N6-cyclohexyladenosine (CHA; A1 >>
A2 agonist), 5-N-ethylcarboxamidoadenosine
(NECA; A2A and A2B > A1),
8-cyclopentyl-1,3-dipropylxanthine (DPCPX; A1 and
A2B receptor antagonist), dimethyl sulfoxide (DMSO),
Histopaque, HEPES, and propidium iodide (PI) were purchased from Sigma
Chemical (St. Louis, MO).
1-Deoxyl-1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-N-methyl-ß-D-ribofuranuronamide
(IB-MECA; A3 agonist),
8-p-sulfophenyltheophylline (8-SPT) and
3-n-propylxanthine (enprofylline) were obtained from RBI
(Natick, MA). 2-p-(2-carboxyethyl)
phenethylamino-5-N-ethylcarboxamidoadenosine (CGS21680; a
selective adenosine A2A receptor agonist)
[20
] was acquired from Tocris Cookson (Bristol, UK).
(E)-1,3-dipropyl-8-(3,4-dimethoxystyryl)-7-methylxanthine
(KF-17837; a selective adenosine A2A receptor antagonist
with a low affinity for A2B receptors) [25
]
was provided by Kyowa Hakko (Tokyo, Japan). Anti-CD16 mAb-coated
immunologic magnetic beads and the magnetic cell sorter system were
obtained from Miltenyi Biotec (Bergisch Gladbach, Germany).
Preparation of cells
Heparinized venous blood was obtained from healthy volunteers.
Neutrophils were isolated by using dextran sedimentation and
centrifugation on a Histopaque gradient (without endotoxin) as
previously described [26
]. Contaminating red blood cells
were removed by means of cold hypotonic water lysis. Cells were then
subjected to another density cut using centrifugation over a Percoll
gradient in order to isolate neutrophils from contaminating
eosinophils. Eosinophils were purified from the peripheral blood of
slightly allergic donors (mild rhinitis) with the aid of gradient
centrifugation and negative selection with anti-CD16 mAb-coated
immunologic magnetic beads and a magnetic cell sorter. The purity of
the isolate was assessed by preparing cytocentrifuged smears and
staining with May-Grünwald-Giemsa (Merck, Darmstadt, Germany)
stain. The purity of the neutrophils was >97% and that of the
eosinophil preparations was >98%, whereas the granulocyte viability
was >99% as determined by trypan blue dye exclusion (Sigma). Purified
cells were suspended in Hanks balanced salt solution (HBSS) without
calcium chloride or magnesium sulfate.
Superoxide anion (O2-) production
Superoxide production was determined at 37°C, with the
modified method of Cohen and Chovaniec [26
,
27
]. The amount of superoxide released was determined
with a Hitachi spectrophotometer U 2000 (Tokyo, Japan) as the change in
absorbance at 550 nm resulting from superoxide dismutase
(SOD)-inhibitable cytochrome c reduction. The reaction was
carried out for 5 min with fMLP (10-6 M), C5a
(10-7 M), and PMA (5 ng/mL), after which the reaction was
stopped by the addition of 0.5 mM N-ethylmalemide. The
generation of superoxide was calculated by subtracting the change in
absorbance in the presence of SOD (1 mM) from that in its absence, and
then dividing this value by 21.1 x 103/M/cm for the
molar extinction coefficient.
Cell motility (chemotaxis)
Cell motility was measured with the agarose method
[26
, 28
]. Briefly, 5 mL of 1.2% agarose
dissolved in HBSS containing Ca2+ and Mg2+ and
supplemented with 10% fetal calf serum (FCS) was placed in 60 x
15-mm Petri dishes. A 10-µL aliquot of cell suspension, containing
5 x 105 cells, was placed in the center well of the
plate, and equal volumes of the chemoattractant (2 x
10-7 M fMLP) and HBSS were placed in the outer and inner
wells, respectively. The wells were 3 mm in diameter, and the outer and
inner wells were located at a distance of 7 mm from the center one.
After incubation at 37°C in 5% CO2 for 2 h, the
plates were fixed with ethanol and formalin and the cells stained with
Wrights stain (Sigma). Cell motility was defined as the linear
distance the cells moved from the center well in the direction of the
well containing the chemoattractant (chemotaxis), and the migration
distance was compared to that of controls. All results represent the
averages of duplicate measurements.
Statistical analysis
All data are presented as means ± SD.
Comparisons between paired conditions were made with the aid of paired
t tests. One-way factorial analysis of variance and multiple
comparison tests (Fishers PLSD) were used for comparative analyses of
data for several groups. P < 0.05 was considered
significant in all cases.
 |
RESULTS
|
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Effects of theophylline on superoxide anion production
Isolated neutrophils (1 x 107 cells/mL) and
eosinophils (2 x 106 cells/mL) in HBSS were
preincubated with various concentrations of theophylline at 37°C for
15 min. Control O2- production stimulated with
1 x 10-6 M fMLP ranged from 60 to 100 nmol cytochrome
c reduced/107 cells for 5 min (86 ± 14
nmol; mean ± SD of five individual donors) in
neutrophils, and from 49 to 63 nmol cytochrome c
reduced/2 x 106 cells for 5 min (55 ± 6 nmol;
mean ± SD of four individual donors) in eosinophils.
This concentration of fMLP caused the strongest activation for
O2- production. As shown in Figure 1
, preincubation of neutrophils with 10-5 to 5 x
10-3 M theophylline had a biphasic, concentration-related
effect on O2- production stimulated with fMLP.
Theophylline potentiated O2- production by
2050% at a clinically relevant concentration range of
10-5 to 10-4 M, and maximal potentiation was
observed at a concentration of 5 x 10-5 M
(P < 0.002), whereas theophylline at higher
concentrations (10-3 to 5 x 10-3 M)
inhibited O2- production from 25 to 60% of
control value (5 x 10-3 M; P <
0.0001). Theophylline induced similar degrees of potentiation and
biphasic influence on O2- production
stimulated with C5a (1 x 10-7 M) in neutrophils
(control value of 103 ± 18 nmol; mean ± SD,
n = 4), but no significant change in
O2- production was in PMA (5 ng/mL)-stimulated
neutrophils (control value of 83 ± 12 nmol, n =
4).

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Figure 1. Effects of theophylline on superoxide anion
(O2-) production from neutrophils
(107 cells/mL) and eosinophils (2 x 106
cells/mL) stimulated with several agents. Cells were preincubated with
buffer or the indicated concentrations of theophylline for 15 min at
37°C before addition of stimulants, after which
O2- production was determined. The reaction
was conducted for 5 min. Results from five donors were normalized to
obtain percentages of control values and are shown as the mean ±
SD. *Significant differences from control
values: P < 0.05.
|
|
The effect of theophylline on O2- production
in eosinophils was substantially different from that in neutrophils
because no potentiation was observed at any concentration of
theophylline. Preincubation of eosinophils with 10-4 to
5 x 10-3 M theophylline significantly inhibited
O2- production stimulated with fMLP and C5a
(P < 0.01, n = 3) but not with PMA.
Effects of theophylline on chemotaxis
The maximum linear distance that cells had migrated from the
margin of the center well was measured and compared with that in
control experiments (Fig. 2
). The control migration distance for four individual donors was
3120 ± 160 µm in neutrophils, and 2250 ± 150 µm in
eosinophils. As shown, theophylline caused a concentration-dependent
reduction in the chemotactic response of both cells. The addition of
theophylline to neutrophils caused a significant decline in chemotaxis
at lower concentrations than those for eosinophils.

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Figure 2. Effects of theophylline on fMLP (2 x 10-7 M)-induced
chemotaxis of neutrophils and eosinophils. Chemotaxis was defined as
the maximum distance cells migrated under agarose (2 h) from the edge
of the center well. Results are shown as means ± SD
from three separate experiments and normalized as control percentage.
*Significant differences from control values:
P < 0.05.
|
|
Effects of 8-p-sulfophenyltheophylline and enprofylline
on superoxide anion production
Theophylline had a biphasic influence on fMLP-stimulated
generation of superoxide anions in neutrophils (Fig. 1)
. In contrast to
theophylline, 8-SPT produced only potentiation of
O2- production (Fig. 3
). Major potentiation of more than 100% was observed at a
concentration of 5 x 10-5 M (P <
0.0001, n = 3) when stimulated with fMLP. The same
effect of 8-SPT on O2- production was observed
in neutrophils stimulated with C5a (data not shown) but not with
enprofylline. Incubation with 0.1 U/mL of adenosine deaminase (ADA) or
1 µM of KF-17837 (10 min, 37°C) also augmented superoxide
production more than twice as much as in control experiments, but DPCPX
had no effect (data not shown). In contrast, none of the effects seen
in neutrophils, i.e., potentiation by 8-SPT, ADA, or KF-17837, was
observed in eosinophils. Incubation of neutrophils at various
concentrations of enprofylline (5 x 10-6 to
10-3 M) caused a concentration-dependent inhibition of
O2- production. A similar inhibition was seen
in eosinophils treated with enprofylline.

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Figure 3. Effects of 8-sulfophenyltheophylline (8-SPT) or enprofylline on
O2- production from neutrophils (A) and
eosinophils (B) stimulated with fMLP (10-6 M). Cells were
preincubated with buffer or the indicated concentrations of drugs for
15 min at 37°C before addition of the stimulant, after which
O2- production was determined. The reaction
was conducted for 5 min. The cells were also incubated with 0.1 U/mL
ADA (filled triangles) or 1 µM KF-17837 (open triangles). Results are
shown as means ± SD from three separate experiments
and normalized as control percentage. *Significant
differences from control values: P < 0.05.
|
|
Effects of 8-p-sulfophenyltheophylline and enprofylline
on chemotaxis
As shown in Figure 4
, the addition of 8-SPT or enprofylline caused a
concentration-dependent reduction in the chemotactic response in
neutrophils (Fig. 4A)
, but significant reduction was not observed at
lower concentrations of enprofylline (
10-4 M).
Preincubation with 0.1 U/mL of ADA also significantly inhibited
neutrophil chemotaxis, whereas treatment of eosinophils with 8-SPT or
ADA (Fig. 4B)
had no effect on eosinophil chemotaxis. Inhibitory effect
was observed in eosinophils treated with enprofylline at higher
concentrations.

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Figure 4. Effects of 8-SPT or enprofylline on fMLP (2 x
10-7 M)-induced chemotaxis of neutrophils (A) and
eosinophils (B). Chemotaxis was defined as the maximum distance cells
migrated under agarose (2 h) from the edge of the center well. The
cells were also treated with 0.1 U/mL ADA (filled triangles) or 1 µM
KF-17837 (open triangles). Results are shown as means ±
SD from three separate experiments and normalized as
percentages of control values. *Significant differences
from control values: P < 0.05.
|
|
Effects of adenosine analogs on superoxide anion production
To determine what class of adenosine receptor exists on
neutrophils or eosinophils and affects their functions, we studied the
effects of several adenosine analogs on O2-
production in these granulocytes.
All of the adenosine analogs inhibited O2-
production from neutrophils stimulated with fMLP (Fig. 5A
), with a rank order for potency obtained with selective adenosine
receptor agonists of CGS-21680 (A2A) > NECA
(A1 and A2A, A2B) > CHA
(A1) > IB-MECA (A3). CGS-21680 was most
potent in adenosine analogs, suggesting that an
A2A-receptor was involved in neutrophil events. Exposure to
5 x 10-7 M CGS-21680 for 15 min resulted in a 50%
inhibition of O2- production from neutrophils.

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Figure 5. Effects of adenosine analogs on
O2- production from neutrophils (A) or
eosinophils (B) stimulated with fMLP (10-6 M). Neutrophils
(107 cells/mL) and eosinophils (2 x 106
cells/mL) were preincubated with adenosine analogs (final DMSO
concentration < 0.1%) for 15 min at 37°C. Representative data
are shown. Essentially the same results were obtained in three
similarly designed experiments.
|
|
IB-MECA was the most potent inhibitor of superoxide production in
eosinophils. Exposure to 1 x 10-6 M IB-MECA resulted
in a 50% inhibition of O2- production from
eosinophils.
Comparative study between theophylline and 8-SPT in
O2- production from neutrophils
A likely explanation of the inhibitory effects of
O2- production at high concentrations of
theophylline (>10-3 M) can be found in the intracellular
cAMP accumulation as a result of PDE inhibition. On the basis of the
finding that 8-SPT does not inhibit PDE activity [10
,
29
], we used 8-SPT and an excess of dbcAMP to reproduce
the biphasic effect of theophylline on O2-
production from neutrophils stimulated with fMLP (10-6 M).
dbcAMP was used as a cell-permeant cAMP analog that can modulate
intracellular signaling and cellular function. The results are shown in
Figure 6
. The potentiation of O2- production from
neutrophils by 8-SPT was reversed by the addition of dbcAMP, and an
excess of dbcAMP (5 x 10-4 M) significantly inhibited
O2- production from neutrophils in comparison
to that seen in controls (ANOVA: F3,9 = 60.642,
P < 0.0001). In other words, the biphasic effect of
theophylline could be reproduced with these chemicals.

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Figure 6. Effects of 8-SPT and dbcAMP on O2- production
from neutrophils. Neutrophils (107 cells/mL) were
preincubated for 15 min at 37°C with buffer and the indicated
concentrations of 8-SPT alone or in the presence of dbcAMP.
O2- production was conducted for 5 min under
stimulation with fMLP (10-6 M). Results are shown as
means ± SD from three separate experiments and
normalized as percentages of control.
|
|
DISCUSSION
Our results confirm that theophylline has a biphasic effect on
O2- production from neutrophils to
receptor-linked agonists such as fMLP and C5a as previously reported
[10
, 11
, 17
]. Superoxide anion
production was potentiated by 50% at concentrations of
10-5 to 10-4 M of theophylline, which are
within the therapeutic range. We also observed that a specific
adenosine A2A receptor agonist, CGS-21680, has the
strongest inhibitory effect on O2- production
in neutrophils, and that this potentiation can also be induced with the
A2A receptor-selective antagonist KF-17837 or adenosine
deaminase but not with DPCPX (an A1 and A2B
receptor antagonist) or enprofylline (A1 and
A2B antagonist [11
, 30
]). These
results strongly suggest that A2A receptors are involved in
the modulation of superoxide production in neutrophils. A recent study
has shown that a selective A2A agonist reduces superoxide
release from neutrophils and inhibits inflammation in a rat model of
meningitis [31
]. These findings are in agreement with
the previous observation that adenosine acts on A2
receptors and inhibits some neutrophil functions [32
].
A2A receptor activation is linked to stimulation of
adenylate cyclase, and adenosine increases intracellular cAMP levels in
neutrophils [25
]. Consequently, the elevated cAMP levels
have been postulated as representing the second messenger mediating
neutrophil functions with adenosine A2A agonists.
Meanwhile, a serine/threonine protein phosphatase has been suggested as
an alternative candidate [33
], but more detailed
information is needed to reach a definitive conclusion.
It should be noted that a different effect of theophylline was observed
in eosinophils, where neither theophylline nor A2A-specific
adenosine analog had any potentiating effect on
O2- production. This appears to be consistent
with the results of receptor binding studies that few adenosine
A2A receptors occur in eosinophils [20
,
34
]. The inhibitory effect of theophylline on
O2- production from eosinophils was previously
reported by several investigators [21
, 23
,
24
]. With regard to the effect of theophylline,
contradictory findings to ours have been reported by different parties
[21
, 24
]. They reported that lower
concentrations of theophylline may potentiate
O2- production from eosinophils. Yukawa and
colleagues [25
] reported that eosinophil activation is
caused by competing with circulating adenosine for eosinophil
A2 receptors. One explanation for this clear discrepancy
could be the insufficient purity of eosinophils. Although high-purity
eosinophils should have been obtained for their functional study, the
centrifugation over Percoll density gradients was used, resulting in
low-purity eosinophils (80%). In contrast, the magnetic bead method is
recommended for the isolation of eosinophils with a high purity
(>98%) without functional priming [35
]. It can
therefore be assumed that the eosinophils used in the study contained
20% neutrophils; the potentiation of superoxide production by
theophylline could therefore be the result of neutrophil contamination.
In a recent study, however, Ezeamuzie and Al-Hage [24
]
again reported a biphasic effect of theophylline on the activation of
highly purified eosinophils. Furthermore, they observed significant
inhibition of O2- production from eosinophils
at much lower concentrations (10-6 and 10-5 M).
These findings are contrary to other studies [21
,
23
], but so far the causes of these discrepancies are not
fully understood.
Higher concentrations (
10-3 M) of theophylline caused
significant inhibition of O2- production from
both neutrophils and eosinophils. Our findings showed that theophylline
analog 8-SPT, more potent A1 and A2A receptor
antagonists, which lacks PDE inhibitory activity [10
,
29
], had only a potentiating effect on
O2- production in neutrophils. On the other
hand, enprofylline, which is known as an adenosine A1 and
A2B antagonist [11
, 30
] with
PDE inhibitory activity, inhibited O2-
production from both cells. This suggests that the difference in
effects between theophylline and enprofylline might be due to the
presence or absence of A2A antagonism and that the
inhibitory effects of these xanthine derivatives on
O2- production may be due to PDE inhibition.
The inhibitory effects of theophylline and enprofylline on cAMP-PDE are
already well known [11
, 36
]. In addition,
we were able to reproduce the biphasic effect of theophylline on
O2- production from neutrophils by using 8-SPT
and an excess of dbcAMP. The addition of dbcAMP reversed the
potentiation of O2- production from
neutrophils by 8-SPT. Selective PDE IV inhibitors have been found to be
very effective for the inhibition of superoxide production from
activated eosinophils [14
, 23
,
37
]. These findings support our contention that the
inhibitory effects of O2- production at a high
concentration of theophylline can be explained by intracellular cAMP
accumulation as a result of PDE inhibition.
The elevation of intracellular cAMP inhibits activation of several
functions of human granulocytes including O2-
production induced by receptor-linked agonists [38
]. In
contrast, when PMA was used to activate cells, theophylline had no
effect on O2- production from either
neutrophils or eosinophils. cAMP PDE inhibitor in combination with
prostaglandin E1 markedly inhibits superoxide production
from neutrophils stimulated with fMLP, but not from those stimulated
with phorbol ester [39
], whereas inactive C5b67 inhibits
C5a- and fMLP-, but not PMA-induced superoxide production
[40
]. These results indicate that such inhibitions occur
independent of protein kinase C activation, and that cAMP levels
represent the second messenger interfering with intracellular signaling
stimulated with receptor-linked agonists and not with PMA in
granulocytes.
The addition of theophylline to neutrophils caused a significant
decline in chemotaxis at lower concentrations than those used for
eosinophils. The effect thus seems to depend on the existence of a
specific receptor for adenosine. A1 receptor activation in
neutrophils is known to be associated with augmentation of chemotaxis
[15
]. Theophyllines antagonism does not discriminate
between A1 and A2A receptor subtypes
[20
], and neutrophils express both A1 and
A2A receptors [15
, 19
]. The
difference in effects of 8-SPT and enprofylline on neutrophil
chemotaxis was according to their affinities for adenosine
A1 receptor [20
]. On the other hand,
eosinophils possess fewer binding sites for adenosine A1
and A2A agonists [34
] and cAMP is known as
an inhibitory modulator of leukocyte chemotaxis [41
];
the inhibitory effects on eosinophil chemotaxis at a high concentration
of enprofylline and theophylline can be explained by intracellular cAMP
accumulation as a result of PDE inhibition.
Recently, it was found that human eosinophils express higher levels of
adenosine A3 receptors [34
], and that
A3 receptor stimulation modulates the functions of
eosinophils [30
, 42
, 43
]. We
also confirmed that a selective A3 receptor agonist
(IB-MECA) reduces O2- production in
eosinophils most effectively [43
]. Furthermore,
theophylline did not influence the effect of IB-MECA (data not shown),
which may be due to the lack of A3 receptor antagonism.
The primary aim of this study was to organize the currently available
information on the effects of theophylline on granulocyte functions,
and to understand the mechanism that mediates cellular functions. We
can conclude that theophylline modulates granulocyte functions in
conjunction with adenosine receptor antagonism (A1 and
A2A) and cAMP-PDE inhibition. Furthermore, the difference
in effect on neutrophils and eosinophils appears to depend on the
existence of specific adenosine receptors. In connection with the
anti-inflammatory actions of theophylline, the results presented here
may raise fears that neutrophils subjected to theophylline therapy may
exhibit enhanced superoxide anion production and cause normal tissue
damage. However, the priming effect of theophylline on
O2- production is very small compared to that
of inflammatory cytokines and is normalized by the inhibitory action of
PDE at therapeutic concentrations. In addition, accelerated neutrophil
apoptosis by theophylline [44
] may well prevent any
prolonged airway inflammation. In conclusion, the anti-inflammatory
effects of theophylline on granulocyte functions are partly the result
of inhibition of the release of toxic oxygen metabolites from
eosinophils and/or suppression of chemotactic migration by both
neutrophils and eosinophils.
 |
ACKNOWLEDGEMENTS
|
|---|
This work was supported in part by a Grant-in-Aid from the Ministry
of Health and Welfare and a grant from the Ministry of Culture
(12670738) of Japan.
Received November 22, 1999;
revised February 29, 2000;
accepted March 1, 2000.
 |
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