Full text of DOI:10.1211/0022357001777414

J. Pharm. Pharmacol. 2000, 52: 1411±1416
Received March 30, 2000
Accepted June 26, 2000
# 2000 J. Pharm. Pharmacol.
Protective Effect of Erdosteine Against Hypochlorous Acid-
induced Acute Lung Injury and Lipopolysaccharide-induced
Neutrophilic Lung In¯ammation in Mice
KEN-ICHI HAYASHI, HISASHI HOSOE, TOSHIHIKO KAISE AND KENJI OHMORI
Drug Development Research Laboratories, Pharmaceutical Research Institute, Kyowa Hakko Kogyo Co.,
Ltd, Japan
Abstract
The effect of erdosteine, a mucoactive drug, on hypochlorous acid (HOCl)-induced lung
injury, and the lipopolysaccharide (LPS)-induced increase in tumour necrosis factor-a
(TNF-a) production and neutrophil recruitment into the airway, was investigated.
Male BALB=c mice were orally administered erdosteine (3±100 mg kg 1), ambroxol
1
hydrochloride (ambroxol) (3±30 mg kg ), S-carboxymethyl-L-cysteine (S-CMC) (100±
1
1
600 mg kg ) or prednisolone (10 mg kg ), 1 h before intratracheal injection of HOCl or
LPS. In the HOCl-injected mice, erdosteine markedly suppressed increases in the ratios of
lung wet weight to bodyweight and lung dry weight to bodyweight, whereas the other
mucoactive drugs ambroxol and S-CMC had little effect. Erdosteine also inhibited the
LPS-induced neutrophil in¯ux, although it did not affect the increased level of TNF-a in
the bronchoalveolar lavage ¯uid.
The results suggest that attenuation of reactive oxygen species and neutrophil recruit-
ment is involved in the clinical ef®cacy of erdosteine in the treatment of chronic bronchitis.
During acute exacerbations in chronic bronchitis,
large numbers of neutrophils accumulate in the
airways, carrying a repository of potentially injur-
ious substances including reactive oxygen species
(ROS), arachidonic acid metabolites and protei-
nases (McCusker & Hoidal 1988). The secretory
products of neutrophils increase mucus production
by epithelial cells and impair cilial function
(McCusker & Hoidal 1988; Repine et al 1997).
These observations suggest that neutrophils and
their secretions are closely related to abnormal
mucus accumulation in the airway in¯ammation
including chronic bronchitis. Drugs which can
attenuate neutrophil recruitment and products such
as ROS may be bene®cial in the treatment of
chronic bronchitis.
tive pulmonary disease including chronic bronchitis
(Dechant & Noble 1996; Miyata et al 1998).
Erdosteine facilitates sputum expectoration by
reducing sputum viscosity and=or by increasing
mucociliary clearance (Dechant & Noble 1996). In
addition, erdosteine can protect against cigarette
smoke-induced loss of a1-antitrypsin activity in the
lungs of rats and man (Dechant & Noble 1996). a1-
Antitrypsin is known to decrease the activity of
neutrophil elastase and to prevent a variety of tis-
sues from injury (Carp & Janoff 1980). Moreover,
it has been reported that erdosteine attenuates
paraquat toxicity in mice, characterized by pro-
gressive and irreversible respiratory failure, leading
to death due to the formation of ROS (Kaise et al
1993; Inglesi et al 1994; Dechant & Noble 1996).
We have previously shown that erdosteine sup-
presses the pulmonary oedema and the increase in
the alveolar±arterial oxygen partial pressure dif-
ference induced by bleomycin in rats (Kaise et al
1993), suggesting that erdosteine exerts an anti-
in¯ammatory action via an antioxidative effect in
addition to its direct mucoregulatory activity.
Clinically, both the antioxidative effect and the
mucoregulatory effect of erdosteine may contribute
Erdosteine, (Æ )-((((tetrahydro-2-oxo-3-thienyl)
carbamoyl)methyl)thio)acetic acid, a mucoactive
drug which enhances respiratory ventilation, was
developed for the treatment of chronic obstruc-
Correspondence: Ken-ichi Hayashi, Drug Development
Research Laboratories, Pharmaceutical Research Institute,
Kyowa Hakko Kogyo Co., Ltd, 1188 Shimotogari, Nagai-
zumi-cho, Sunto-gun, Shizuoka 411-8731, Japan.

1412
KEN-ICHI HAYASHI ET AL
to ameliorating the decrease in respiratory function
due to the airway obstruction accompanying airway
in¯ammation and abnormal mucus production.
In this study we assessed the effects of erdosteine
on the acute lung injury induced by hypochlorous
acid (HOCl), a ROS produced by neutrophils
(Weiss et al 1982), and on lipopolysaccharide
(LPS)-induced neutrophil recruitment into the
airway.
wet lung weight to body weight (lung wet wt=BW)
and that of lung dry weight to body weight (lung
dry wt=BW) were calculated.
LPS-induced airway in¯ammation
BALB=c mice were administered orally with either
drug or distilled water and 1 h later, either 0Á1 mL
LPS (E. coli 055 : B5, Difco Laboratories, Detroit,
1
MI) (10 ng mL in saline), or saline alone was
injected intratracheally under anaesthesia with
ether. At the appropriate time, mice were killed by
an intraperitoneal injection of pentobarbital sodium
Materials and Methods
1
(120 mg kg ). The trachea was cannulated and
then a total of 4 mL (four 1-mL portions) of saline
was immediately injected into the lung and the
intrapulmonary contents were aspirated. The
bronchoalveolar lavage (BAL) ¯uid was cen-
trifuged at 200 g for 10 min (4ꢀC), and the super-
natant was stored at 80ꢀC until the determination
of tumour necrosis factor-a (TNF-a), and the pre-
cipitate was suspended in 0Á4 mL saline for cell
counts.
Animals
Male BALB=c mice, 20±30 g, were purchased
from Japan Charles Liver (Hino, Japan), and were
allowed to acclimatize in an animal room main-
tained at a room temperature of 22±24ꢀC and a
relative humidity of 50±60%, with a 12-h light±
dark cycle (light between 07 00 and 19 00 h) for 1
week. Healthy mice were chosen for use in the
experiment which was approved by the Animal
Ethical Committee of Kyowa Hakko Kogyo Co.,
Ltd (Shizuoka, Japan).
The total cell count was determined on a cell
counter (Celltac a MEK-6158; Nihon Kohden,
Tokyo, Japan). For cell differentiation, a cytocen-
trifuge preparation was made by centrifuging the
1
cells at 600 rev min for 3 min in a Cytospin 3
Drugs
(Shandon Inc., Pittsburgh, PA); the specimen was
then stained with Giemsa stain, and differential cell
counts were performed for 500 cells.
On the day of the experiments, erdosteine (Edmond
Pharma, Milan, Italy) was dissolved with an
equivalent molar quantity of sodium bicarbonate in
distilled water; ambroxol hydrochloride (ambroxol)
(Sigma Chemical Co., St Louis, MO) was dissolved
in distilled water; S-carboxymethyl-L-cysteine (S-
CMC) (Sigma) was dissolved in 1 M NaOH and
then neutralized with 1 M HCl (adjusted to pH 6±
7); prednisolone (Sigma) was suspended in 0Á5%
methylcellulose. All drugs were prepared in the
appropriate concentrations and administered in a
The level of TNF-a was measured with an
enzyme-linked immunosolvent assay kit (mouse
TNF-a ELISA kit; Endogen Inc., Woburn, MA). If
a measurement value was lower than the quantita-
1
tion limit of 50 pg mL , the limit value was
regarded as the level of TNF-a.
Statistical analysis
1
volume of 10 mL kg .
Statistical signi®cance was analysed using the SAS
system for Windows ver. 6.12 (SAS Institute Inc.,
Cary, NC). The Wilcoxon rank sum test was used
for comparisons between the two groups. Multiple
comparisons were made ®rst by the Kruskal±
Wallis test and, when appropriate, followed by the
Steel test. All data were expressed as meanÆ s.e.m.
Differences were considered signi®cant at P < 0Á05.
HOCl-induced lung injury
At 1 h after oral administration of either drug or
distilled water, BALB=c mice were injected intra-
tracheally with either 0Á1 mL of the desired con-
centrations of NaOCl (Wako Pure Chemical
Industries, Ltd, Osaka, Japan) in saline (buffered to
pH 7), or with saline alone under anaesthesia with
ether. At various intervals, mice were anaesthetized
1
Results
(pentobarbital sodium; 120 mg kg , i.p.) and
exsanguinated. The lungs were surgically removed,
the extrapulmonary bronchi were cut away and the
wet weight was recorded. The lungs were placed in
a drying oven at 120ꢀC for 24 h and the dry weight
was determined. As an index of injury, the ratio of
HOCl-induced lung injury
Intratracheal instillation of HOCl (0Á3, 1, 3 and
1
10 mmol L ) produced concentration-dependent
increases in lung wet wt=BW and lung dry wt=BW

ANTIINFLAMMATORY EFFECT OF ERDOSTEINE
1413
1 h after the instillation. Both ratios in mice
exposed to HOCl at a concentration of 1 mmol L
maximum inhibition (61 and 100%, respectively)
at 100 mg kg
1
1
(Table 3). Ambroxol tended to
suppress the increase in lung wet wt=BW at
or higher, were signi®cantly greater than those
in the saline-treated control group. However,
10 mmol L HOCl caused death in 8 out of 12
1
10 mg kg
and signi®cantly inhibited it at
1
1
30 mg kg , although the inhibition was just 29 and
23%, respectively. Ambroxol even at a concentra-
tion of 30 mg kg failed to inhibit the increase in
mice within 1 h of the instillation (Table 1).
The lung wet wt=BW at 0Á5, 1 and 3 h, and the
lung dry wt=BW at 1 and 3 h, after the instillation
1
lung dry wt=BW (Table 3). S-CMC (100±
600 mg kg ) did not alter the increases in lung wet
1
1
of 1 mmol L HOCl were signi®cantly elevated
compared with the saline-treated control group. In
particular, at 1 h later, these increases were highly
signi®cant. The lung wet wt=BW at 6 h, and the
lung dry wt=BW at 0Á5 and 6 h, after the instillation
was not affected by HOCl (Table 2). Exposure to
wt=BW and lung dry wt=BW at any of the doses
tested (Table 3).
LPS-induced airway in¯ammation
1
At 3 h after intratracheal instillation of LPS (1 ng),
the number of total cells in the BAL ¯uid did not
differ from that in the saline-treated control group,
however, neutrophil counts signi®cantly increased
and macrophage counts signi®cantly decreased
(Table 4). Under the same experimental conditions,
the level of TNF-a in the BAL ¯uid was sig-
ni®cantly increased by LPS (Table 4). Erdosteine
1 mmol L HOCl for 1 h, which was used in the
subsequent experiments, produced suf®cient injury
to assess the effects of drugs.
Erdosteine signi®cantly prevented HOCl-induced
increases in lung wet wt=BW and lung dry wt=BW
1
at concentrations of 10 mg kg or higher, and
30 mg kg or higher, respectively, and produced a
1
1
(10±100 mg kg ) did not signi®cantly alter these
events, whereas prednisolone (10 mg kg ) inhib-
Table 1. Effects of increasing concentrations of hypochlor-
ous acid (HOCl) on the ratios of lung wet weight to body-
weight (lung wet wt=BW) and lung dry weight to bodyweight
(lung dry wt=BW) 1 h after the intratracheal instillation.
1
ited the increase in neutrophil counts and TNF-a
level (Table 4). At 24 h after the instillation of LPS,
total cell counts and neutrophil counts in the BAL
Treatment Concentration
(mmol L
Lung wet
Lung dry
3
1
)
wt=BW (Â10 3) wt=BW (Â10
)
Table 3. Effects of erdosteine, ambroxol hydrochloride
(ambroxol) and S-carboxymethyl-L-cysteine (S-CMC) on
hypochlorous acid (HOCl)-induced increases in the ratios of
lung wet weight to bodyweight (lung wet wt=BW) and lung
dry weight to bodyweight (lung dry wt=BW).
Saline
HOCl
6Á09Æ 0Á06
1Á24Æ 0Á01
0Á3
1
3
10
6Á27Æ 0Á13
1Á26Æ 0Á02
7Á05Æ 0Á14*** 1Á32Æ 0Á02**
10Á57Æ 0Á17*** 1Á44Æ 0Á02***
13Á43Æ 0Á33** 1Á72Æ 0Á03**
Drug
Dose
(mg kg
Lung wet
wt=BW
Lung dry
wt=BW
Each value represents the meanÆ s.e.m. (saline, 0Á3, 1 and
1
)
1
1
3 mmol L
HOCl; n ˆ 12, 10 mmol L
value in the saline-treated control group.
HOCl; n ˆ 4).
3
3
(Â10
)
(Â10
)
**P < 0Á01, ***P < 0Á001 compared with the corresponding
Negative control
Positive control
Erdosteine
5Á94Æ 0Á09
1Á21Æ 0Á01
7Á28Æ 0Á14{{{ 1Á32Æ 0Á02{{
3
10
30
100
6Á98Æ 0Á12
6Á73Æ 0Á09*
6Á79Æ 0Á11*
1Á27Æ 0Á02
1Á27Æ 0Á02
1Á25Æ 0Á01*
Table 2. Time-course of changes in the ratios of lung wet
weight to bodyweight (lung wet wt=BW) and lung dry weight
6Á46Æ 0Á09*** 1Á21Æ 0Á02**
to bodyweight (lung dry wt=BW) after the intratracheal instil-
lation of saline or 1 mmol L hypochlorous acid (HOCl).
1
Negative control
Positive control
Ambroxol
5Á73Æ 0Á10
1Á19Æ 0Á02
7Á04Æ 0Á09{{{ 1Á28Æ 0Á02{{
3
10
30
6Á85Æ 0Á10
6Á66Æ 0Á13
6Á74Æ 0Á10*
6Á08Æ 0Á09
1Á25Æ 0Á01
1Á25Æ 0Á02
1Á24Æ 0Á01
1Á24Æ 0Á01
Treatment Time after
Lung wet
Lung dry
wt=BW (Â10
3
3
injection (h) wt=BW (Â10
)
)
Negative control
Positive control
S-CMC
Untreated
Saline
±
5Á73Æ 0Á09
7Á37Æ 0Á09
6Á03Æ 0Á07
5Á55Æ 0Á08
5Á66Æ 0Á12
7Á76Æ 0Á12*
1Á24Æ 0Á02
1Á28Æ 0Á02
1Á20Æ 0Á01
1Á24Æ 0Á02
1Á25Æ 0Á02
1Á29Æ 0Á02
7Á31Æ 0Á16{{{ 1Á31Æ 0Á02{
0Á5
100
300
600
7Á16Æ 0Á12
7Á07Æ 0Á12
7Á01Æ 0Á14
1Á29Æ 0Á02
1Á30Æ 0Á02
1Á27Æ 0Á02
1
3
6
HOCl
0Á5
Each mouse was administered orally with distilled water
(negative control and positive control), erdosteine, ambroxol or
S-CMC 1 h before intratracheal instillation of saline or
1
7Á23Æ 0Á13***
6Á06Æ 0Á06***
5Á74Æ 0Á10
1Á33Æ 0Á01***
1Á30Æ 0Á01*
1Á27Æ 0Á02
3
6
1
1 mmol L HOCl. Each value represents the meanÆ s.e.m.,
n ˆ 12. {P < 0Á05, {{P < 0Á01, and {{{P < 0Á001, compared
with the corresponding value in the negative control group.
*P < 0Á05, **P < 0Á01, and ***P < 0Á001, compared with the
corresponding value in the positive control group.
Each value represents the meanÆ s.e.m., n ˆ 12. *P < 0Á05,
***P < 0Á001 compared with the corresponding value in the
saline-treated control group.

1414
KEN-ICHI HAYASHI ET AL
¯uid signi®cantly increased, whereas no changes in
macrophage counts were observed compared with
the saline-treated control group (Table 5). At this
time, the release of TNF-a in the BAL ¯uid of LPS-
injected mice was at a normal level (data not
shown). Erdosteine signi®cantly inhibited the
known to cause disruption of various plasma
membranes, including the epithelium and the
È
endothelium, by oxidation (Schraufstatter et al
1990; Guo et al 1995; Ochoa et al 1997). We
assume that, in this study, the increased perme-
ability of the airway epithelium and blood vessels
resulted in the plasma leakage into the bronch-
oalveolar space, leading to the formation of pul-
monary oedema. Moreover, the increased lung dry
wt=BW suggests that the increase in lung wet
wt=BW resulted not only from excess water but also
from the increased unknown factors in the lung, and
that it may involve pulmonary congestion.
increase in total cell counts and neutrophil counts
at concentrations of 30 mg kg
1
or higher and
1
100 mg kg , respectively. Erdosteine did not affect
the macrophage counts at any of the doses tested
(Table 5).
We assessed the effects of three mucoactive
drugs, erdosteine, ambroxol and S-CMC, on the
HOCl-induced lung injury. Erdosteine signi®cantly
inhibited the increase in lung wet wt=BW and lung
Discussion
Activated neutrophils, which play a major role in
chronic bronchitis (McCusker & Hoidal 1988;
Repine et al 1997), produce HOCl (Weiss et al
1982). A previous report showed that intratracheal
injection of HOCl evoked airway in¯ammation, as
demonstrated by marked increases in protein con-
centration and the percentage of polymorpho-
nuclear cells in the BAL ¯uid of rats (Ghio et al
1996). In this study, we found that instillation of
HOCl into the airway increased lung wet wt=BW
and lung dry wt=BW. In addition, signi®cant
increases in wet=dry and accumulation of Evans
blue dye in the BAL ¯uid were observed after
treatment with HOCl (data not shown). HOCl is
1
dry wt=BW at a concentration of 10 mg kg or
1
higher and 30 mg kg
or higher, respectively.
However, the increase in lung wet wt=lung dry wt
was not signi®cantly in¯uenced by erdosteine as a
result of inhibited increases in both lung wet
wt=BW and lung dry wt=BW (data not shown). A
previous study showed that erdosteine protected
against cigarette smoke-induced loss of a1-anti-
trypsin activity, suggesting that this drug scavenges
ROS (Dechant & Noble 1996). Erdosteine contains
two blocked sulphhydryl groups, which are
unmasked after metabolic transformation in the
Table 4. Effects of erdosteine and prednisolone on changes in the number of leukocytes and the TNF-a level in BAL ¯uid 3 h
after the intratracheal instillation of LPS.
1
)
1
Drug
Dose (mg kg
Total cells (Â105)
Neutrophils (Â105)
Macrophages (Â105)
TNF-a (pg mL
51Á7Æ 1Á7
)
Negative control
Positive control
Erdosteine
1Á5Æ 0Á2
1Á6Æ 0Á2
1Á4Æ 0Á4
1Á5Æ 0Á2
1Á5Æ 0Á2
1Á3Æ 0Á3
0Á0Æ 0Á0
0Á8Æ 0Á2{{
0Á7Æ 0Á2
0Á5Æ 0Á2
0Á3Æ 0Á1
0Á1Æ 0Á0**
1Á5Æ 0Á2
0Á7Æ 0Á0{{
0Á7Æ 0Á2
0Á9Æ 0Á2
1Á1Æ 0Á2
1Á2Æ 0Á3
1360Æ 104Á2{{
1782Æ 319Á3
998Á0Æ 292Á9
880Á0Æ 246Á4
441Á2Æ 74Á9**
10
30
100
10
Prednisolone
Each mouse was administered orally with distilled water (negative control and positive control), erdosteine or prednisolone 1 h
before intratracheal instillation of saline or 1 ng LPS. Each value represents the meanÆ s.e.m., n ˆ 5±6. {{P < 0Á01 compared with
the corresponding value in the negative control group. **P < 0Á01 compared with the corresponding value in the positive control
group.
Table 5. Effects of erdosteine on changes in the number of leukocytes in BAL ¯uid 24 h after the intratracheal instillation of LPS.
1
Drug
Dose (mg kg
)
Total cells (Â105)
Neutrophils (Â105)
Macrophages (Â105)
Negative control
Positive control
Erdosteine
1Á7Æ 0Á2
0Á1Æ 0Á0
4Á8Æ 0Á7{{
3Á0Æ 0Á6
1Á9Æ 0Á6
1Á4Æ 0Á6*
1Á6Æ 0Á2
1Á8Æ 0Á3
1Á3Æ 0Á2
1Á2Æ 0Á2
1Á5Æ 0Á2
6Á6Æ 0Á7{{
4Á3Æ 0Á6
10
30
100
3Á1Æ 0Á8*
3Á0Æ 0Á7*
Each mouse was administered orally with distilled water (negative control and positive control) or erdosteine 1 h before the
intratracheal instillation of saline or 1 ng LPS. Each value represents the meanÆ s.e.m., n ˆ 5±6. {{P < 0Á01 compared with the
corresponding value in the negative control group. *P < 0Á05 compared with the corresponding value in the positive control group.

ANTIINFLAMMATORY EFFECT OF ERDOSTEINE
1415
liver. The liberated sulphhydryl groups of this
metabolite are suggested to display free radical
scavenging activity (Dechant & Noble 1996). N-
Acetylcysteine, which has an oxidizable thiol
group, is a scavenger of HOCl (Gillissen et al
1997). Similarly, erdosteine seems to have a
scavenging activity against HOCl, which could
contribute to the inhibition of HOCl-induced lung
injury. S-CMC did not suppress the increase in lung
wet wt=BW and lung dry wt=BW at any of the
doses tested. Although the effect of S-CMC on
HOCl is unknown, the lack of free sulphhydryl
groups may be the reason for its weak activity.
Ambroxol tended to suppress the increase in lung
ni®cantly reduced the increased neutrophil counts
at a dose of 100 mg kg . These results indicate that
erdosteine exerts an inhibitory effect on LPS-
induced in¯ammation in a later phase rather than in
the TNF-a-mediated early phase.
1
This study showed that erdosteine inhibited neu-
trophil in®ltration without affecting TNF-a produc-
tion in mice intratracheally treated with LPS.
Although erdosteine did not signi®cantly inhibit the
TNF-a generation, it may have in¯uenced the
expression of adhesion molecules and=or the related
chemotactic factors in the process between TNF-a
and neutrophil recruitment. The recruited neu-
trophils release a variety of products, such as elastase
and ROS (McCusker & Hoidal 1988), the latter of
which can damage various cells to reinforce the
activity of elastase by inactivating a1-antitrypsin
(Carp & Janoff 1980). Moreover, neutrophil elastase
is shown to cause detachment and deformation of
human bronchial epithelial cells and to induce gene
expression and protein production of interleukin-8
(Shibata et al 1996). Thus, erdosteine may have
inhibited the production of neutrophil chemotactic
factors via prevention of airway epithelium injury
because of its scavenging action against ROS from
recruited neutrophils. N-Acetylcysteine is reported
to suppress nuclear factor-kB activation, cytokine-
induced neutrophil chemo-attractant messenger
RNA expression, and neutrophilic lung in¯amma-
tion in rats (Blackwell et al 1996). As regards the
exact mechanism for the anti-in¯ammatory effect of
erdosteine without affecting TNF-a production,
further investigation is needed.
1
wet wt=BW at a concentration of 10 mg kg and
signi®cantly inhibited it at 30 mg kg , although
1
the inhibition was just 29 and 23%, respectively.
Chemically, ambroxol and N-acetylcysteine are
completely different molecules. Ambroxol has
an aromatic ring with an amine group and a
hydroxycyclohexyl ring. Nevertheless, an HOCl
scavenging action by ambroxol has been reported
(Lapenna et al 1994; Gillissen et al 1997). In this
study, however, ambroxol did not signi®cantly
inhibit lung injury. Considering that erdosteine but
not ambroxol is a prodrug-type compound, the
possibility remains that ambroxol is almost oxi-
dized before arriving at airway tissues.
We previously reported that congestion, in®ltra-
tion of in¯ammatory cells, oedema and epithelium
disintegration were observed in the lung of mice 3
days after paraquat treatment, and that erdosteine
reduced these changes, delayed the onset of the
lethal event, and ®nally prevented the mortality
(Kaise et al 1993). The toxicity in acute paraquat
poisoning is believed to result from free radical
injury (Bus et al 1974). These ®ndings support the
notion that erdosteine suppresses ROS in-vivo, and
thus that the prevention of HOCl-induced lung
injury by erdosteine in this study was mediated via
its ROS scavenging action.
The effect of erdosteine on LPS-induced airway
in¯ammation was tested. We observed increases in
neutrophil counts and TNF-a level in BAL ¯uid 3 h
after intratracheal instillation of LPS. de Moraes
et al (1996) have previously reported increases in
TNF-a level and neutrophil counts in BAL ¯uid 3 h
after LPS inhalation in mice. In addition, they
demonstrated that TNF-a was produced before
neutrophil recruitment and that anti-TNF-a anti-
bodies blocked neutrophil recruitment. In this
study, erdosteine did not signi®cantly inhibit the
neutrophil in¯ux at 3 h after the LPS challenge,
presumably because of the inability of the drug
to prevent the production of TNF-a. In contrast,
at 24 h after the LPS instillation, erdosteine sig-
In conclusion, this study demonstrated that
erdosteine inhibited HOCl-induced acute lung
injury in mice, suggesting that this drug exerts a
scavenging effect on ROS in-vivo. The anti-
oxidative effect may be involved in the prevention
by erdosteine of LPS-induced neutrophil recruit-
ment into the airway. Our results suggest that the
attenuation of ROS and neutrophil in¯ux may
contribute to the clinical ef®cacy of erdosteine in
the therapy of chronic bronchitis.
References
Blackwell, T. S., Blackwell, T. R., Holden, E. P., Christman,
B. W., Christman, J. W. (1996) In vivo antioxidant treatment
suppresses nuclear factor-kappa B activation and neutro-
philic lung in¯ammation. J. Immunol. 157: 1630±1637
Bus, J. S., Aust, S. D., Gibson, J. E. (1974) Superoxide- and
singlet oxygen-catalyzed lipid peroxidation as a possible
mechanism for paraquat (methyl viologen) toxicity. Bio-
chem. Biophys. Res. Commun. 58: 749±755
Carp, H., Janoff, A. (1980) Potential mediators of in¯amma-
tion: phagocyte-derived oxidants suppress the elastase-
inhibitory capacity of alpha1-proteinase inhibitor in vitro.
J. Clin. Invest. 66: 987±995

1416
KEN-ICHI HAYASHI ET AL
Dechant, K. L., Noble, S. (1996) Erdosteine. Drugs 52: 875±
881
Lapenna, D., De Gioia, S., Ciofani, G., Cuccurullo, F. (1994)
Ambroxol is a scavenger of hypochlorous acid and mono-
chloramine. Pharmacology 49: 132±135
McCusker, K. T., Hoidal, J. (1988) Leukocyte function and
chronic bronchitis. Semin. Respir. Infect. 3: 5±13
Miyata, T., Kai, H., Isohama, Y., Takahama, K. (1998) Current
opinion of muco-active drug research: strategies and prob-
lems. Eur. Respir. J. 11: 480±491
Ochoa, L., Waypa, G., Mahoney, J. R., Rodriguez, L., Min-
near, F. L. (1997) Contrasting effects of hypochlorous acid
and hydrogen peroxide on endothelial permeability: preven-
tion with cAMP drugs. Am. J. Respir. Crit. Care. Med. 156:
1247±1255
de Moraes, V. L. G., Vargaftig, B. B., Lefort, J., Meager, A.,
Chignard, M. (1996) Effect of cyclo-oxygenase inhibitors
and modulators of cyclic AMP formation on lipopolysac-
charide-induced neutrophil in®ltration in mouse lung. Br. J.
Pharmacol. 117: 1792±1796
Ghio, A. J., Marshall, B. C., Diaz, J. L., Hasegawa, T.,
Samuelson, W., Povia, D., Kennedy, T. P., Piantadosi, A.
(1996) Tyloxapol inhibits NF-kappa B and cytokine release,
scavenges HOCl, and reduces viscosity of cystic ®brosis
sputum. Am. J. Respir. Crit. Care. Med. 154: 783±788
È
Gillissen, A., Scharling, B., Jaworska, M., Bartling, A.,
Rasche, K., Schultze-Werninghaus, G. (1997) Oxidant sca-
venger function of ambroxol in vitro: a comparison with N-
acetylcysteine. Res. Exp. Med. 196: 386±398
Repine, J. E., Bast, A., Lankhorst, I. (1997) Oxidative stress in
chronic obstructive pulmonary disease. Oxidative Stress
Study Group. Am. J. Respir. Crit. Care. Med. 156: 341±357
È
Guo, Y., Schneider, L. A., Wangensteen, O. D. (1995) HOCl
effects on tracheal epithelium: conductance and permeabil-
ity measurements. J. Appl. Physiol. 78: 1330±1338
Inglesi, M., Nicola, M., Fregnan, G. B., Bradamante, S.,
Pagani, G. (1994) Synthesis and free radical scavenging
properties of the enantiomers of erdosteine. II Farmaco 49:
703±708
Kaise, T., Hosoe, H., Sano, J., Ohmori, K. (1993) Effects of
KW-9144 on paraquat toxicity in mice and bleomycin-
induced pulmonary injury in rats. Jpn. J. Pharmacol. 61
(Suppl. 1): 81P
Schraufstatter, I. U., Browne, K., Harris, A., Hyslop, P. A.,
Jackson, J. H., Quehenberger, O., Cochrane, G. (1990)
Mechanisms of hypochlorite injury of target cells. J. Clin.
Invest. 85: 554±562
Shibata, Y., Nakamura, H., Kato, S., Tomoike, H. (1996)
Cellular detachment and deformation induce IL-8 gene
expression in human bronchial epithelial cells. J. Immunol.
156: 772±777
Weiss, S. J., Klein, R., Slivka, A., Wei, M. (1982) Chlorination
of taurine by human neutrophils: evidence for hypochlorous
acid generation. J. Clin. Invest. 70: 598±607