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Infection and Immunity, October 2000, p. 6005-6011, Vol. 68, No. 10
0019-9567/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Expression of Interleukin-9 Leads to Th2
Cytokine-Dominated Responses and Fatal Enteropathy in Mice with Chronic
Schistosoma mansoni Infections
Padraic G.
Fallon,1,*
Philip
Smith,1
Emma J.
Richardson,1
Frances J.
Jones,1
Helen C.
Faulkner,2,
Jacques
Van
Snick,3
Jean-Christophe
Renauld,3
Richard K.
Grencis,2 and
David W.
Dunne1
Department of Pathology, University of
Cambridge, Cambridge,1 and School of
Biological Sciences, University of Manchester,
Manchester,2 United Kingdom, and
Experimental Medicine Unit, Ludwig Institute for Cancer
Research, University of Louvain, Brussels,
Belgium3
Received 11 May 2000/Returned for modification 14 June
2000/Accepted 16 July 2000
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ABSTRACT |
Mice infected with Schistosoma mansoni develop Th2
cytokine-mediated granulomatous pathology that is focused on the liver and intestines. In this study, transgenic mice constitutively expressing IL-9 were infected with S. mansoni and the
outcome of infection was determined. Eight weeks after infection,
transgenic mice with acute infections had a moderate increase in Th2
cytokine production but were overtly normal with respect to parasite
infection and pathological responses. Transgenic mice with chronic
infections died 10 weeks after infection, with 86% of transgenic mice
dead by week 12 of infection, compared to 7% mortality in infected wild-type mice. Stimulation of mesenteric lymph node cells from infected transgenic mice with parasite antigen elicited elevated interleukin-4 (IL-4) and IL-5 production and reduced gamma interferon and tumor necrosis factor alpha production compared to the responses in
wild-type mice. Morbid transgenic mice had substantial enlargement of
the ileum, which was associated with muscular hypertrophy, mastocytosis, eosinophilia, goblet cell hyperplasia, and increased mucin expression. We also observed that uninfected transgenic mice
exhibited alterations in their intestines. Although there was hepatic
mastocytosis and eosinophilia in infected transgenic mice, there was no
hepatocyte damage. Death of transgenic mice expressing IL-9 during
schistosome infection was primarily associated with enteropathy. This
study highlights the pleiotropic in vivo activity of IL-9 and
demonstrates that an elevated Th2 cytokine phenotype leads to death
during murine schistosome infection.
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INTRODUCTION |
Interleukin-9 (IL-9) is a Th2
cytokine initially described as a T-cell and mast cell growth factor
(29). Studies of transgenic mice that constitutively express
IL-9 have shown that a spectrum of immunological responses are
potentially mediated by IL-9, including lymphomagenesis
(30), intestinal mastocytosis (13), bronchial hyperresponsiveness (4, 23), pulmonary eosinophilia (4, 23), expansion of B-1 lymphocytes (38), and resistance
to intestinal nematode infection (10, 11). In addition,
transgenic mice with lung-specific expression of IL-9 develop airway
inflammation, mast cell hyperplasia, eosinophilia, and increased airway
hyperresponsiveness (37). These studies have implicated IL-9
as an important cytokine in a number of Th2 cytokine-mediated
pathologies, in particular asthma.
Infection of mice with the helminth parasite Schistosoma
mansoni elicits a dynamic pathological process that is associated with both Th1 and Th2 responses (5). In mice with
schistosome infections, the liver and intestine are the major organs
affected. The tissue damage in the liver is primarily caused by
granulomatous inflammation surrounding parasite eggs trapped in hepatic
parenchyma, whereas the intestine is subject to inflammation elicited
by parasite eggs being translocated through the intestinal wall. In
this study, a transgenic mouse strain that constitutively expresses
IL-9 (30) was infected with S. mansoni to address
the influence of expression of IL-9 in an in vivo Th2-mediated
pathological process. Transgenic mice with chronic infections developed
a marked Th2 cytokine-dominated response that was associated with high
mortality and enteropathy. This study highlights the pleiotropic in
vivo activity of IL-9.
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MATERIALS AND METHODS |
Mice and parasite.
A Puerto Rican strain of S. mansoni was used in all experiments. Treatment of mice expressing
IL-9 constitutively has been described; the Tg54 transgene-positive
line and wild-type FVB/N mice were used for all experiments
(30). Mice were maintained under specific-pathogen-free conditions.
Parasitological and pathological techniques.
Infection and
portal perfusions were done as described previously (34).
The livers and intestines were removed, stored at 
20°C, and used
for tissue homogenates (see below) or tissue egg counts. Tissues were
digested in 4% KOH, and eggs were counted as described previously
(2). Fecal samples were collected on the day the mice were
terminated and eggs were counted (2). In accordance with
United Kingdom Home Office regulations, any infected mice that became
morbid were humanely killed. Analysis of pathology was performed as
described in previous studies (6, 7). Liver and ileums were
fixed in Formal-saline or Carnoy's fixative. Three 4-µm-thick
sections of liver or 10 to 15 sections of intestine were cut at
150-µm intervals. Sections were stained with hematoxylin and eosin
(granuloma measurements), toluidine blue (mast cells), Martius scarlet
blue (collagen), periodic acid-Schiff (PAS) (goblet cells), and Giemsa
(eosinophils) stain. The diameter of the muscularis propria (serosa to
submucosa) was measured at eight different angles on transverse
sections of ileum with an ocular micrometer. The mean diameter of the
muscularis propria in individual mice was calculated from measurements
on at least 10 sections. All pathological parameters were assayed in a
double-blind manner.
Cytokine analysis.
Spleen or mesenteric lymph nodes (MLN)
were aseptically removed and cultured in vitro, and cytokine was
assayed in culture supernatant as described previously (6,
8). Cells were restimulated in vitro with soluble egg antigens
(20 µg/ml). Anti-cytokine monoclonal antibodies and recombinant
cytokine standards to detect IL-4, IL-5, IL-9, gamma interferon
(IFN-
), and tumor necrosis factor alpha (TNF-
) in an
enzyme-linked immunosorbent assay (ELISA) were purchased from
PharMingen (San Diego, Calif.) or Genzyme (Kent, United Kingdom).
Preparation of tissue homogenates.
Liver or intestine tissue
was weighed and chopped with scissors. Tissue was processed for
eosinophil peroxidase (EPO) assays essentially as described previously
(1). In brief, approximately 5% (wt/vol) tissue suspensions
in Hanks balanced salt solution (HBSS) (without phenol red, 10 mM
HEPES) (pH 7.4) (Sigma, Dorset, United Kingdom) were homogenized on ice
using a T8 Ultra-Turrax homogenizer (Janke and Kunkel GmbH, Staufen,
Germany). The supernatant was centrifuged at 1,952 × g
for 10 min (4°C). The pellet was retained, and red blood cells were
lysed by hypotonic shock. Homogenization and centrifugation were
repeated two more times. The cell pellet was resuspended in HBSS
containing 0.5% hexadecyltrimethylammonium bromide (HBSS-HTAB),
rehomogenized, freeze-thawed three times in liquid nitrogen, and stored
at 20°C until assayed. Intestinal tissue was processed for murine
mast cell proteinase (mMCP) analysis as described previously
(16). Chopped intestinal tissue was homogenized in 1 ml of
20 mM Tris-1 M NaCl (pH 7.5). Following centrifugation
(9,000 × g for 30 min), the supernatant was stored at
20°C until assayed. Protein content was assayed in all intestinal homogenates.
mMCP-1 analysis.
Serum or intestinal homogenates were
assayed for mMCP-1 activity using an antibody capture ELISA
(26). The assay was performed with a commercial kit
following the manufacturer's instructions (Moredun Scientific Ltd.,
Midlothian, Scotland).
EPO assay for tissue eosinophilia.
Tissue eosinophilia was
determined using the EPO assay (35) as described previously
(1). Intestine or liver tissue was processed as described
above. The numbers of eosinophils (expressed as 106 or
107 cells per gram of tissue) were interpolated from a
standard curve prepared from murine eosinophils. Blood from
Nippostrongylus brasiliensis-infected mice (10 to 20% of
circulating leukocytes were eosinophils) were used as a source of
eosinophils. Murine eosinophils were isolated from mouse blood as
described previously (36). In brief, eosinophils were
purified from blood by dextran T-500 (Amersham Pharmacia Biotech AB,
Uppsala, Sweden) sedimentation. Red blood cells were removed from the
leukocyte-rich supernatant, and following washing, eosinophils were
counted on eosin-stained cytospins. Eosinophils were adjusted to a
stock dilution of 106 eosinophils per ml in HBSS-HTAB,
freeze-thawed, and stored at 20°C.
Intestinal mucin detection.
Mucous expression was analyzed
by Western blots of intestinal homogenates (prepared as described for
EPO assays) using a monoclonal antibody against human colonic mucin
(R35.3.3) as described previously (17, 27). Protein level
estimation was performed on supernatants from the homogenates of
intestines from individual mice. Supernatants were resolved on a 4 to
12% polyacrylamide gel under nonreducing conditions; 50 µg of
protein was loaded per lane. Gels were stained with silver to ensure
equal protein loading in wells. Following electrotransfer to
nitrocellulose paper, strips were probed with R35.3.3. Mucin was
detected at 79 kDa, and relative expression of this band was performed
using Kodak Digital Science 1D Image Analysis Software (Rochester,
N.Y.).
Statistical analysis.
Statistical differences between the
values for different groups were determined by Student's t
test. P values of <0.05 were considered statistically significant.
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RESULTS AND DISCUSSION |
IL-9 production during S. mansoni infection of
mice.
During infection of mice with S. mansoni, there
are dynamic temporal changes in the relative production of Th1 and Th2
cytokines (5). Characteristically, during the first 4 weeks
of infection, there is an early Th1 cytokine response that is then
superseded by a marked Th2 cytokine phenotype (14, 28). The
Th2 cytokine response peaks approximately 8 weeks after infection,
after which there is a progressive decline in Th2 cytokine production.
To date, there have not been extensive studies on the role of IL-9 in
schistosome infection. Following polyclonal (mitogen) stimulation of
spleen cells recovered from infected mice at various stages of
infection, we observed that IL-9 production followed the general temporal pattern of Th2 cytokine production during schistosome infection (data not shown). The increased production of IL-9 during the
first 8 weeks of murine schistosome infection is consistent with the
results of a previous study (18). However, while we observed
reduced IL-9 production from weeks 8 to 16 after infection, these
researchers observed a plateauing or marginal reduction in IL-9
production from weeks 8 to 14 after infection. These data demonstrate
that IL-9 production is elevated in the Th2 cytokine-dominated acute
stages of schistosome infection of mice and is reduced in the chronic
stages of infection.
Increased Th2 cytokine responses in an acute S. mansoni
infection of transgenic mice that constitutively express IL-9.
Transgenic and wild-type mice were exposed to an acute (150-cercaria)
schistosome infection to determine if expression of IL-9 altered the
induction of Th2 cytokines or modified the course of infection. Mice
were killed 8 weeks after infection, coincident with the peak in Th2
cytokine responses in mice. No deaths or differences in overt morbidity
were observed for the groups. Parasitologically, expression of IL-9 did
not influence parasite infectivity (number of worms recovered) or
fecundity (egg production per worm pair) (Table
1). Infected wild-type and transgenic
mice had similar-sized granulomas surrounding eggs in the liver and had
comparable levels of hepatic fibrosis (Table
2). There was, however, a slight increase in the numbers of eosinophils within the egg granulomas in the livers
of transgenic mice over those of wild-type mice (Table 2). Measurement
of the levels of aspartate aminotransferase, a marker for hepatocyte
damage, in plasma demonstrated that the infected transgenic mice and
wild-type mice had comparable levels of hepatic damage (Table 2). There
was no major difference in intestinal pathology in the infected
wild-type and transgenic mice, with the exception of an increase in the
size of the ileum of two of the eight transgenic mice. Both groups of
mice had comparable levels of excretion of parasite eggs in the feces
(Table 1).
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TABLE 1.
Parasitological data obtained from wild-type and
IL-9-expressing transgenic mice with acute or chronic S. mansoni infectionsa
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TABLE 2.
Hepatic alterations in wild-type and IL-9-expressing
transgenic mice with received acute or chronic
S. mansoni infectionsa
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MLN cells recovered from infected mice were restimulated with parasite
egg antigens. MLN cells from infected transgenic mice
had significant
greater production of IL-9 after restimulation
with egg antigens
relative to IL-9 release from cells from infected
wild-type mice (Fig.
1A). Infected transgenic mice had a
nonsignificant
increase in production of Th2 cytokines (IL-4 and IL-5)
and reduced
Th1 cytokine (IFN-
and TNF-
) responses compared to
those of
wild-type mice (Fig.
1A). ELISAs for the levels of parasite
antigen-specific
antibody in serum demonstrated there were elevated
immunoglobulin
E (IgE) and IgG1 responses in the transgenic mice than
in the
infected wild-type mice (data not shown). Collectively, the data
indicate that the expression of IL-9 during an 8-week acute schistosome
infection caused a moderate increase in the production of Th2
cytokines
but did not cause any major parasitological or pathological
alterations.
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FIG. 1.
Th1 and Th2 cytokine profiles in wild-type mice and
transgenic (Tg) mice that constitutively express IL-9 (Tg) with acute
S. mansoni infections. Mice were terminated on day 58 after
infection. MLN from two to four mice were pooled and stimulated with
parasite egg antigens (20 µg/ml). Data presented are means ± standard deviations from three cultures.
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Th2 cytokine-dominated responses and high mortality during chronic
schistosome infection of transgenic mice that constitutively express
IL-9.
In mice with chronic schistosome infections, the production
of IL-9 and other Th2 cytokines is down modulated from approximately 8 to 12 weeks after infection. To determine whether the expression of
IL-9 influenced the ability of mice to down modulate Th2 cytokine responses, transgenic mice were infected with a light (25-cercaria) chronic infection. Unexpectedly, for mice with chronic infections, there was high mortality of transgenic mice, with transgenic animals dying from day 74 after infection (Fig.
2). By 83 days after infection, 86% (12 of 14 mice infected) of the transgenic mice had died, whereas 7% (1 of
15 mice infected) of wild-type mice had succumbed. This IL-9-expressing
transgenic mouse strain is predisposed to develop thymic lymphomas from
3 to 9 months of age (31). We therefore checked if death of
transgenic mice was associated with thymus enlargement. No infected
transgenic mice had evidence of alterations in thymus size. As
uninfected transgenic mice of the same age that were housed in the same
room as that of infected mice did not die, we attribute the deaths of
the transgenic mice to schistosome infection.
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FIG. 2.
Survival of mice with chronic S. mansoni
infections. The mortality for IL-9-expressing transgenic (Tg) mice was
much higher than that of the wild-type (Wt) mice.
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In a second chronic infection, mice were terminated on day 72 after
infection when the infected transgenic mice had started
to develop
morbidity. There were no parasitological differences
between
schistosome-infected transgenic and wild-type mice (Table
1). Cells
from the MLN were restimulated with egg antigens, and
cytokine
production was measured by ELISA. Infected transgenic
mice had a marked
elevation in the production of IL-4 and IL-5
relative to wild-type mice
and a reciprocal reduced production
of IFN-
and TNF-
(Fig.
3). Other Th2 cytokines (IL-10 and IL-13)
were also elevated in the infected transgenic mice than in the
wild-type mice. Transgenic mice also had increased levels of parasite
antigen-specific IgE (data not shown). With respect to the induction
of
cytokine production following a chronic schistosome infection,
the
transgenic mice have a Th2 cytokine-dominated response relative
to
wild-type mice. It has been previously observed that Th2 cytokine
responses are required in schistosome infection to prevent exacerbated
Th1 responses evoking pathology (
5,
7,
33). The data
presented
here illustrate that a Th2-polarized cytokine response can
lead
to death during chronic schistosome infection of mice. In
agreement
with this observation, a recent study has also demonstrated
that
a Th2 cytokine-dominated response causes lethal pathology during
murine schistosome infection (
15).
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FIG. 3.
Th1 and Th2 cytokine profiles in wild-type and
IL-9-expressing transgenic (Tg) mice with chronic S. mansoni
infections. Mice were terminated on day 72 after infection. MLN from
two to four mice were pooled and stimulated with parasite egg antigens
(20 µg/ml). Data presented are means ± standard deviations from
three cultures.
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Enteropathy in infected transgenic mice.
Gross examination
during autopsy of chronically infected mice demonstrated marked
distension of the small intestines of all infected mice relative to
those of uninfected mice (Fig. 4A). There
was striking enlargement of the ileums of all infected transgenic mice
compared to infected wild-type animals, with the size of the ileum in
transgenic mice that had overt morbidity at termination being increased
three- to sixfold (Fig. 4A). The enlargement in size of the intestine
of infected transgenic mice was not associated with alterations in the
submucosa or serosa but was due to a profound increase in the size of
the muscularis propria (Fig. 4B). Measurements of the diameter of the
muscularis propria demonstrated the significant (P < 0.001) increase in its size in the infected transgenic mice over
that of infected wild-type mice (Fig.
5A). The intestinal enlargement observed
in wild-type mice is comparable to a previous study in
schistosome-infected mice (3), and similar intestinal changes have been observed during infection of other animals (9, 20). It was also observed that uninfected transgenic mice had enlarged small intestines relative to the size of the intestines in
wild-type mice (Fig. 4A), with a significant (P < 0.05) increase in the diameter of the muscularis propria (Fig.
5A). However, the intestinal enlargement in uninfected transgenic mice
was extremely variable between individual mice, with approximately 50%
of transgenic mice having intestines of comparable size to those of
wild-type mice.
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FIG. 4.
Intestinal and hepatic alterations in IL-9-expressing
transgenic (Tg) and wild-type (Wt) mice. (A) Uninfected () or
chronically infected (+) Tg mice had a marked increase in the size of
the ileum relative to that of uninfected or infected wild-type mice
(hematoxylin and eosin stained). (B) Intestinal enlargement in the
infected Tg mice was associated with marked distension of the
muscularis propria compared to that of infected wild-type mice
(hematoxylin and eosin stained). Bar, 45 µm. (C and D) Within the
muscularis propria of infected Tg mice, there was marked infiltration
of mast cells (C) and eosinophils (D). Toluidine blue (C) and
hematoxylin and eosin (D) stain was used. Bars, 40 µm (C) and 50 µm
(D). There was marked goblet cell hyperplasia in the ileums of Tg mice
(PAS stained). Bar, 30 µm. (F) In the livers of infected Tg mice,
there was a pronounced infiltration of eosinophils within the granuloma
surrounding the egg compared to wild-type mice (hematoxylin and eosin
stained). Bar, 30 µm.
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FIG. 5.
Enteropathy in uninfected and chronically infected
IL-9-expressing transgenic (Tg) mice. (A) The diameter of the
muscularis propria was measured on histological sections with an ocular
micrometer. (B and C) To quantify the extent of eosinophilia and
mastocytosis in the ileums of mice, tissue homogenates were assayed for
EPO (B) and mMCP-1 activity (C). Results are from 4 to 12 mice per
group, and data are presented as means ± standard errors.
Statistical differences between groups were determined by Student's
t test.
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Histological analysis demonstrated that the increase in the size of the
muscularis propria of infected transgenic mice was
due to muscular
hypertrophy (Fig.
4B) and infiltration of mast
cells (Fig.
4C) and
eosinophils (Fig.
4D). In the infected transgenic
mice, eosinophils
were detected within the egg granuloma and throughout
the muscularis
propria (Fig.
4D), whereas in infected wild-type
mice, eosinophils were
primarily associated with the granuloma
surrounding the egg (not
shown). To quantify the levels of intestinal
eosinophilia, we measured
EPO activity in tissue homogenates.
Uninfected transgenic mice had
slightly elevated numbers of intestinal
eosinophils than wild-type mice
(Fig.
5B). Schistosome infection
elicited a marked intestinal
eosinophilia in both wild-type and
transgenic mice, with infected
transgenic mice having two- to
threefold-more intestinal eosinophils
than infected wild-type
mice (
P < 0.001) (Fig.
4B).
Infiltration of the intestine by eosinophils
is a normal process during
schistosome infection of mice, with
eosinophils within the intestine
proposed to reduce intestinal
inflammation (
5) and aid the
translocation of the egg through
the intestinal wall (
19).
However, IL-9-expressing transgenic
mice and wild-type mice with acute
or chronic schistosome infections
had comparable numbers of eggs
excreted, even though the transgenic
mice had intestinal eosinophilia
(Table
1).
Intestinal mastocytosis during schistosome infection of mice has been
reported previously, with increased intraepithelial
intestinal
mast cells and intestinal mast cell protease (mMCP-1)
activity in
infected mice (
24). Initially, as performed previously
in
the same mouse strain following gastrointestinal nematode challenge
(
10,
11), we counted the numbers of intraepithelial mast
cells
on toluidine blue-stained sections of intestine. However, as we
had observed marked mast cell infiltration of the muscularis propria
(Fig.
4C), we measured the levels of mMCP-1 in intestinal homogenates
to quantify total intestinal mast cells. In intestinal homogenates
of
uninfected wild-type mice, there was limited mMCP-1 activity
but
significantly elevated protease activity in transgenic mice
(
P < 0.001) (Fig.
5C). The mastocytosis induced during
schistosome
infection was reflected in elevated mMCP-1 activity in
infected
wild-type mice and fourfold-greater protease activity in
infected
transgenic mice (Fig.
5C). The intestinal mastocytosis in
uninfected
IL-9-expressing transgenic mice has been reported previously
(
11,
13). Our data on elevated intestinal mast cells and
mMCP-1 activity
in schistosome-infected transgenic mice is consistent
with previous
studies in the same mouse strain following challenge with
gastrointestinal
nematodes (
10,
11).
PAS-stained sections of ileum demonstrated marked goblet cell
hyperplasia in the infected transgenic mice (Fig.
5E). Numeration
of
PAS-positive goblet cells per villous crypt unit demonstrated
the
elevation in goblet cells in the transgenic mice, with naive
transgenic
mice having approximately twice the numbers of goblet
cells as
wild-type mice (Fig.
6A). To more
accurately analyze
intestinal mucous expression in individual animals,
a monoclonal
anti-human colonic mucin (R35.3.3) antibody was used to
detect
mucin in Western blots of intestinal homogenates (
17,
27).
Using this method, the relative expression of mucin, a band
at
79 kDa, in different animals was evident (Fig.
6B). Densitometry
was
used to quantify intestinal mucin expression in individual
mice (Fig.
6C). There was greater intestinal expression of mucin
in uninfected
transgenic mice than in wild-type mice (
P < 0.05).
Schistosome infection elicited an increase in intestinal mucin
expression (
P < 0.001) in both wild-type and
transgenic animals
compared to that in uninfected mice (Fig.
6B and C).
Mucin expression
was significantly greater in infected transgenic mice
than wild-type
mice. The observation that IL-9 may elicit mucin
expression in
this study is in agreement with recent studies that
demonstrate
this cytokine may directly stimulate mucin production
(
21,
22).
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FIG. 6.
Goblet cell hyperplasia and mucin expression in
transgenic (Tg) mice that constitutively express IL-9. (A) The numbers
of goblet cells per villous crypt unit (vcu) were counted on
PAS-stained ileum sections from uninfected or infected Tg and wild-type
mice. (B and C). Mucin expression in the ileum was determined by
Western blotting of sodium dodecyl sulfate-polyacrylamide gel
electrophoresis-resolved ileum homogenates using an anti-mucin
monoclonal antibody. (B) A representative Western blot showing mucin
expression (79 kDa) in uninfected and infected wild-type (Wt) and
transgenic (Tg) mice is shown. Densitometry of the intensities of the
mucin band on a series of Western blots was used to quantify the levels
of mucin expression in groups of mice. (C) Data are presented as the
increase in mucin expression in different groups of mice relative to
the mucin levels in uninfected wild-type mice, which was set at 1. Data
are from 3 to 10 mice per group and are presented as means ± standard errors. Statistical differences between groups were determined
by Student's t test.
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With respect to the livers of chronically infected animals, there was
no difference between transgenic and wild-type mice
with respect to the
size of the granuloma or the degree of hepatic
fibrosis (Table
2).
However, there was a striking predominance
of eosinophils within the
granuloma of infected transgenic mice
relative to that in wild-type
mice (Fig.
4E). We also detected
sporadic eosinophil infiltrates
throughout the hepatic parenchyma
and surrounding portal tracts of the
infected transgenic mice
(not shown). Assays for EPO activity in
hepatic tissue confirmed
there was two- to threefold more eosinophils
within the livers
of chronically infected transgenic mice than in
infected wild-type
animals (Table
2). On toluidine blue-stained liver
sections,
there was increased mast cells in transgenic mice, with
assays
for mMCP-1 activity on liver homogenates demonstrating
significant
mastocytosis in the infected transgenic mice relative to
wild-type
mice (Table
2). The marked eosinophil infiltration of the
livers
of infected transgenic mice is potentially significant, as
eosinophils
have been causally associated with a number of liver
diseases
(
25).
With respect to murine schistosomiasis, it has been suggested that
various mediators that are released by degranulation of
eosinophils may
stimulate hepatic damage in schistosome-infected
mice (
12).
However, despite the increased hepatic eosinophilia
and mastocytosis in
the infected transgenic mice, these mice and
wild-type mice had
comparable plasma transaminase levels (Table
2), implying there was no
increase in hepatocyte
damage.
In conclusion, infection of mice expressing IL-9 with a helminth
infection that elicits elevated Th2 cytokine caused fatalities
in
transgenic mice. A range of Th2 cytokine-mediated responses
that are
elicited in wild-type mice during schistosome infection
were
exacerbated in IL-9-expressing transgenic mice including
the following:
(i) a marked elevation in Th2 cytokine production
and reduction in Th1
production, (ii) increased tissue (intestine
and liver) eosinophilia
and mastocytosis, and (iii) intestinal
goblet cell hyperplasia and
mucin expression. In addition, transgenic
mice had marked hypertrophy
of the muscularis in the intestine,
implicating a potential effect of
IL-9 on smooth muscle. In contrast,
a number of other responses were
not affected, including parasite
infectivity or fecundity and egg
excretion. We cannot attribute
the various responses that we have
observed in schistosome-infected
IL-9-expressing transgenic mice as
solely due to IL-9, since expression
of IL-9 elicited a marked
elevation in Th2 cytokine production
following infection. However,
certain Th2 cytokine-mediated responses
(hepatic granuloma formation
and fibrosis) were not affected by
expression of IL-9. With respect to
the elevated mucin expression
in transgenic mice, there is emerging
evidence that IL-9 may have
a direct role in this response; e.g., IL-9
has been shown to directly
induce mucin production in respiratory
epithelial cells (
21),
and the IL-9-expressing transgenic
mouse strain used in this study
has been shown to have elevated mucin
expression in the lung (
22).
Similarly, tissue eosinophilia
and mastocytosis have been reported
in previous studies in
IL-9-expressing transgenic mice (
4,
37), and with respect to
eosinophilia, in vivo neutralization
of IL-9 activity by antibody
treatment blocks parasite-induced
blood eosinophilia (
32).
The data presented in this study enlarge
the spectrum of responses
elicited in mice that constitutively
express IL-9. This study
illustrates the pleiotropic activities
of IL-9 and its potential role
in Th2 cytokine-mediated
pathologies.
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ACKNOWLEDGMENTS |
We are grateful to Fiona Culley for advice on eosinophil
peroxidase assays, Hugh Miller and Elisabeth Thornton for assistance with mMCP-1 assays, and Daniel Podolsky for kindly providing the anti-mucin monoclonal antibody. We are grateful to Barry Potter for
performing histology.
This work was supported by the Medical Research Council and the
Wellcome Trust. P.G.F. is supported by a Wellcome Trust Career Development Award.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: Department of
Pathology, University of Cambridge, Tennis Court Rd., Cambridge CB2
1QP, United Kingdom. Phone: 44 1223 339768. Fax: 44 1223 333741. E-mail: pgf20{at}cam.ac.uk.
Present address: Department of Biological Sciences, University of
Salford, Salford, United Kingdom.
Editor:
W. A. Petri Jr.
| |
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Infection and Immunity, October 2000, p. 6005-6011, Vol. 68, No. 10
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Copyright © 2000, American Society for Microbiology. All rights reserved.
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Top
Abstract
Introduction
