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Levels of Schistosoma mansoni-induced interleukin (IL)-4 and IL-5 and posttreatment levels of immunoglobulin E recognizing the parasite's tegument (Teg) correlate with human resistance to subsequent reinfection after treatment. We measured changes in whole-blood cytokine production in response to soluble egg antigen (SEA), soluble worm antigen (SWA), or Teg after treatment with praziquantel (PZQ) in a cohort of 187 individuals living near Lake Albert, Uganda. Levels of SWA-induced IL-4, IL-5, IL-10, and IL-13 increased after treatment with PZQ, and the greatest relative increases were seen in the responses to Teg. Mean levels of Teg-specific IL- 5 and IL-10 increased ∼10—15-fold, and mean levels of IL-13 increased ∼5-fold. Correlations between the changes in cytokines suggested that their production was positively coregulated by tegumentally derived antigens. Levels of SEA-, SWA-, and Teg-induced interferon-g were not significantly changed by treatment, and, with the exception of IL-10, which increased slightly, responses to SEA also remained largely unchanged. The changes in cytokines were not strongly influenced by age or intensity of infection and were not accompanied by corresponding increases in the numbers of circulating eosinophils or lymphocytes.
Treatment with S. mansoni Increases Th2 Responses JID 2004:190 (15 August) 835
Increases in Human T Helper 2 Cytokine Responses
to Schistosoma mansoni Worm and Worm-
Tegument Antigens Are Induced by Treatment
with Praziquantel
Sarah Joseph,
Frances M. Jones,
Klaudia Walter,
Anthony J. Fulford,
Gachuhi Kimani,
Joseph. K. Mwatha,
Timothy Kamau,
Henry C. Kariuki,
Francis Kazibwe,
Edridah Tukahebwa,
Narcis B. Kabatereine,
John H. Ouma,
Birgitte J. Vennervald,
and David W. Dunne
Department of Pathology, University of Cambridge, Cambridge, United Kingdom;
Kenya Medical Research Institute and
Division of Vector-Borne
Diseases, Nairobi, Kenya;
Vector Control Division, Kampala, Uganda;
Danish Bilharziasis Laboratory, Charlottenlund, Denmark
Levels of Schistosoma mansoni–induced interleukin (IL)–4 and IL-5 and posttreatment levels of immuno-
globulin E recognizing the parasite’s tegument (Teg) correlate with human resistance to subsequent reinfection
after treatment. We measured changes in whole-blood cytokine production in response to soluble egg antigen
(SEA), soluble worm antigen (SWA), or Teg after treatment with praziquantel (PZQ) in a cohort of 187 individuals
living near Lake Albert, Uganda. Levels of SWA-induced IL-4, IL-5, IL-10, and IL-13 increased after treatment
with PZQ, and the greatest relative increases were seen in the responses to Teg. Mean levels of Teg-specific IL-
5 and IL-10 increased 10–15-fold, and mean levels of IL-13 increased 5-fold. Correlations between the changes
in cytokines suggested that their production was positively coregulated by tegumentally derived antigens. Levels
of SEA-, SWA-, and Teg-induced interferon-g were not significantly changed by treatment, and, with the exception
of IL-10, which increased slightly, responses to SEA also remained largely unchanged. The changes in cytokines
were not strongly influenced by age or intensity of infection and were not accompanied by corresponding increases
in the numbers of circulating eosinophils or lymphocytes.
Schistosomiasis affects 200 million people in the de-
veloping world and causes chronic morbidity in pop-
ulations living in areas where the disease is endemic
[1]. The disease is caused by infection with blood-dwell-
ing worms of the genus Schistosoma, which infect hu-
mans after contact with water containing skin-pene-
trating parasite larvae. Epidemiological studies in areas
of endemicity have shown that, in general, adults have
lower intensities of infection than do children, sug-
Received 29 October 2003; accepted 28 February 2004; electronically published
12 July 2004.
Financial support: Medical Research Council and the Commission of the
European Community’s Science and Technology for DevelopmentProgramme(INCO-
DC contract IC18 CT97-0237 and INCO-DEV contract ICA4-CT-1999-10003).
Present affiliation: London School of Hygiene and Tropical Medicine, London,
United Kingdom.
Reprints or correspondence: Dr. Sarah Joseph, Dept. of Pathology, University
of Cambridge, Tennis Court Rd., Cambridge CB2 1QP, UK (
The Journal of Infectious Diseases 2004;190:835–42
2004 by the Infectious Diseases Society of America. All rights reserved.
gesting that resistance to infection develops with age,
manifesting around the age of puberty [2]. Reinfection
studies, in which the acquisition of new infections is
monitored after treatment, have shown that adults be-
come reinfected far less often than children, even in
fishing communities where adults have the greatest ex-
posure to the parasite [3]. The measurement of parasite-
specific immune responses near the time of treatment
has identified correlates of low intensities of reinfection.
These include eosinophilia (Schistosoma haematobium
[4]), parasite-specific IgE (S. haematobium [5], Schisto-
soma mansoni [6, 7], and Schistosoma japonicum [8]),
and the production of interleukin (IL)–4 and IL-5 by
peripheral blood mononuclear cells stimulated in vitro
with parasite antigens (Ags) (S. haematobium [9] and S.
mansoni [10]). Such reinfection studies have led to the
consensus that schistosome-specific T helper 2 (Th2)
responses are associated with, and perhaps directly in-
volved in, human age-dependent immunity to reinfec-
tion after treatment.
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836 JID 2004:190 (15 August) Joseph et al.
In areas of Kenya where transmission of S. mansoni is sea-
sonal, immunological correlates of resistance to reinfection
were strongest when parasite-specific immune responses were
measured after treatment with praziquantel (PZQ) but before
exposure to reinfection had occurred [7]. In that Kenyan study,
posttreatment levels of adult worm–specific IgE correlated neg-
atively with the intensity of reinfection [7], providing evidence
for the hypothesis that drug-dependent killing of worms in situ
might boost protective immunity against reinfection by im-
munizing with Ags released from dying worms [11, 12]. Adult
S. mansoni worms in infected Kenyan populations have been
estimated to have a mean life span of 6–10 years [13]; thus, in
such areas of endemicity, exposure to Ags from naturally dying
worms might be expected to be age dependent in untreated
infected populations. It follows that the slow development of
age-dependent immunity in such areas where schistosomiasis
is endemic might arise as the result of the cumulative exposure
to host-protective Ags derived from naturally dying worms.
The tissues of schistosome-infected individuals are simul-
taneously and chronically exposed to 2 distinct developmental
stages of the parasite: the egg and the adult worm. Hundreds
of eggs are released into the bloodstream of the host daily by
each female worm. In studies in Kenya, posttreatment levels of
soluble worm Ag (SWA)–specific IgE, but not soluble egg Ag
(SEA)–specific IgE, were found to be associated with resistance
to subsequent reinfection [7]. The interface between the adult
worm and its environment occurs at its outer tegument (Teg),
a highly specialized organelle with a syncytial structure that
forms a highly responsive outer layer over the entire surface of
the worm. The Teg can be isolated from live intact worms, and
high posttreatment IgE responses against this worm substruc-
ture are also predictive of low levels of reinfection [7]. Western
blots of parasite extracts, by use of serum samples obtained from
chronically infected Kenyans, identified an 22-kDa Ag located
in the Teg of S. mansoni as the predominant target of S. mansoni
specific IgE responses in humans [7]. This tegumental Ag has
been identified as a homologue of cytoplasmic dynein light chain
[14], and IgEs recognizing either the purified native Ag (Sm22)
[15] or a bacterially expressed recombinant form (rSm22.6) [16]
were significantly correlated with resistance to reinfection after
treatment. In addition, immunization with preparations of the
Teg has been shown to induce some protection against experi-
mental S. mansoni infections in mice [17, 18].
It is known that PZQ rapidly and specifically disrupts the
schistosome Teg, exposing hitherto sequestered parasite Ags to
the host immune system [19, 20]. In fact, the killing of schis-
tosomes by PZQ in vivo depends, at least in part, on the in-
tegrity of the host immune system, and efficacy of PZQ may
depend on host immune responses against newly exposed teg-
umental Ags [21, 22]. The Teg is crucial to the survival of
schistosomes in the bloodstream, but, despite important in-
teractions between the Teg, the host immune system, and PZQ
in vivo, human cellular responses with specificity for this struc-
ture have not been previously described. We therefore examined
the effects of treatment with PZQ on the cytokine responses to
the Teg of S. mansoni in a cohort of 187 Ugandans living in a
fishing community who were exposed to high levels of parasite
transmission. We assessed changes in the levels of IL-4, IL-5, IL-
10, IL-13, and interferon (IFN)–g in whole-blood cultures in
response to stimulation in vitro with S. mansoni Teg before and
7 weeks after treatment and compared them with changes in
responses to either homogenized SEA or SWA. We observed
significant increases in several Th2 cytokines in response to Teg
and SWA in this area where S. mansoni is endemic.
Study population. A study cohort was selected from the
fishing village of Booma (in the parish of Butiaba), which is
adjacent to Lake Albert, Masindi district, in northwestern
Uganda. One hundred eighty-seven individuals (in equal num-
bers from all age groups and of both sexes) were selected on
the following criteria: (1) they lived close to the lake shore, (2)
they were 7–50 years old, (3) they had resided in the village
for at least 10 years or since birth, and (4) they agreed to
participate and to donate blood samples before and 7 weeks
after treatment. Informed consent was obtained from all par-
ticipants in this study, in accordance with the national guide-
lines of the Ugandan Ministry of Health, whose ethical review
committees approved all the protocols used. Before treatment,
3 stool samples were obtained on consecutive days from each
member of the study cohort. For each stool sample, 2 Kato
slides (50 mg) were prepared and examined, and S. mansoni
eggs were counted. The intensity of S. mansoni infection for
the cohort members varied from 0 to 8100 eggs/gram of feces,
with an estimated prevalence of infection of 94%. After do-
nating a blood sample, each cohort member was treated im-
mediately with PZQ (40 mg/kg of body weight) and was treated
again 2 weeks later; an additional blood sample was obtained
7 weeks after the first treatment. At the parasitological exam-
inations 7 weeks after treatment, 20% of the cohort members
were excreting at least 1 egg. Overall, the burden of infection
was reduced by 99.7%.
Whole-blood cultures. Venous blood was collected into
heparin, at a concentration of 10 U/mL (Sigma), and was di-
luted immediately 1:4 in RPMI 1640 medium (Sigma) supple-
mented with penicillin (50 U/mL), streptomycin (50 mg/mL),
and l-glutamine (2 mmol/L) (Sigma). Whole-blood cultures
were set up as described elsewhere [23], and parasite-derived
Ags were added to the appropriate wells, to a final concentration
of 10 mg/mL. Cultures were stimulated with SEA, SWA, or Teg
or were left unstimulated. Levels of IL-4 were measured in
supernatants from 48-h cultures, and levels of IL-5, IL-10, IL-
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Treatment with S. mansoni Increases Th2 Responses JID 2004:190 (15 August) 837
13, and IFN-g were measured in supernatants from 120-h cul-
tures. The supernatants were immediately frozen at 20C and
were transported frozen to the United Kingdom. Subsequently,
the culture supernatants were thawed and treated with inactive
enveloped viruses, by incubation for 2 h in the presence of
0.3% tri-butyl phosphate (Sigma) and 1% Tween 80 (Sigma).
Subsequently, the supernatants were either assayed for cytokines
immediately or were stored at 70C. None of the cytokine
assays was affected by the viral inactivation procedure (data
not shown). No supernatant sample went through
12 cycles of
freezing and thawing before being assayed for cytokine content.
Samples were diluted as required to ensure that the assay values
were within the linear part of the standard curve for that assay.
Parasite Ags. The PBS-soluble fractions of S. mansoni
SWA and SEA homogenates were prepared according to estab-
lished protocols [16]. The schistosome outer Teg was stripped
from freshly isolated worms by incubation in PBS for 1 h at
room temperature and then was collected from the supernatant
[23]. In the whole-blood cultures, all Ags were used at a final
concentration of 10 mg/mL. Before use, the Ags were titrated
in vitro by use of murine splenocytes from infected mice. En-
dotoxin content was measured by use of a limulus amebocyte
lysate kit (QCL-1000; BioWhittaker). The levels of endotoxin
in the native Ag used in these studies were 1.44 ng of endotoxin/
mg of SEA, 1.23 ng of endotoxin/mg of SWA, and 1.09 ng of
endotoxin/mg of Teg. This level of lipopolysaccharide did not
induce detectable levels of any of the tested cytokines when
added to our whole-blood cultures (data not shown).
Cytokine assays. Cytokines were measured using capture
ELISAs and commercially available anticytokine antibodies (Abs)
and standards from BD Biosciences, as described elsewhere [23].
All Abs were purchased from BD Biosciences and were titrated
for optimal concentration before use. For the cytokine assays,
the following Ab pairs were coated on microtiter plates to capture
target cytokines or were used to detect captured cytokines (IL-
4, coating 8D4-8 and detection MP4-25D2; IL-5, coating TRFK5
and detection JRS1-5A10; IL-10, coating JES3-9D7 and detection
JES3-12G8; IL-13, coating JES10-5A2 and detection B69-2; and
IFN-g, coating NIB42 and detection 45B3).
Statistical analyses. All the analyses and graphs were per-
formed in R, an open-source language and environment for
statistical computing and graphics (available at: http://www Paired t tests were used to assess whether re-
sponses had changed significantly between the 2 blood samples
and were performed using nontransformed data. Correlations
between variables were determined by calculating Spearman’s
rank correlation coefficient for each pair of immune responses,
again using nontransformed data. The approximate percentage
increase in responses after treatment with PZQ for all study
subjects was determined using the following calculation: 100
(exp{mean[ln(B+1)ln(A+1)]}1). Effects of sex, age, and in-
tensity of infection on changes in cytokine responses were ex-
amined by analysis of variance on log-transformed cytokine re-
sponses, log(+0.1). The adequacy of the models was checked using
variance ratios, which under the null hypothesis, follow an F dis-
tribution using appropriate df. The model fit was also checked by
examining appropriate plots, and results were confirmed by Spear-
man’s rank correlation coefficients on the raw data.
Effects of treatment with PZQ on production of cytokines.
Figure 1 shows the mean level of production of cytokines by
diluted whole blood stimulated with SEA, SWA, or Teg or un-
stimulated, before and then again 7 weeks after treatment with
PZQ. Treatment with PZQ had no significant effect on the mean
levels of IL-4, IL-5, IL-10, IL-13, or IFN-g produced in culture
in the absence of Ag stimulation (figure 1A–1E). The mean
levels of IL-4, IL-5, and IL-13 in response to SWA and Teg
increased significantly after treatment, whereas the mean levels
of these cytokines in response to SEA did not change signifi-
cantly (figure 1A, 1B, and 1D). By contrast, the mean levels of
IL-10 seen in response to all S. mansoni–derived Ags (SEA,
SWA, and Teg) increased significantly during the 7 weeks after
treatment (figure 1C). The mean levels of production of IFN-
g in response to SEA, SWA, and Teg were not significantly
altered by treatment (figure 1E). The highest posttreatment
levels of IL-5, IL-10, and IL-13 were seen in response to SWA,
and the levels were significantly higher than the mean levels
measured in response to Teg (figure 1B, 1C, and 1D). There
was no significant difference in the mean posttreatment levels
of IL-4 detected in whole-blood cultures in response to SWA
and Teg (figure 1A), and these levels were higher than the
corresponding responses to SEA.
Relative changes in cytokine response to SWA and Teg.
Of the responses that changed significantly after treatment, the
mean response to Teg increased more (as a proportion of pre-
treatment levels) than did the response to SWA, with mean
levels of IL-5 and IL-10 increasing 10–15-fold and mean levels
of IL-13 increasing 5-fold in response to Teg, whereas mean
levels of the same cytokines in response to SWA increased only
2–4.5-fold after treatment. The production of IL-4 did not
show the same pattern, with similar relative increases seen (2–
4-fold) in response to both SWA and Teg (table 1).
Correlations between the changes in specific cytokine re-
sponses. The strongest correlations were between changes
in SWA- and Teg-specific responses for each cytokine, and all
such correlation coefficients were
10.675 and statistically sig-
nificant (table 2). Changes in all SWA-specific responses were
positively correlated with each other, as were all changes in Teg-
specific responses (table 2). With the exception of Teg-specific
IL-5 and IL-13, pairs of SWA-specific responses were always
more strongly correlated than were the same responses to Teg
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838 JID 2004:190 (15 August) Joseph et al.
Figure 1. Effect of treatment with praziquantel (PZQ) on production of cytokines. production of cytokines in the whole cohort wasMean SEM
determined by use of whole-blood culture before (Bleed A) and 7 weeks after (Bleed B) treatment with PZQ. The cultures were left unstimulated
(MED) or were stimulated with Schistosoma mansoni antigen (Ag), soluble egg Ag (SEA), adult worm outer tegument (TEG), or soluble adult worm
Ag (SWA). A, Interleukin (IL)–4 ( ); B, IL-5 ( ); C, IL-10 ( ); D, IL-13 ( ); and E, interferon (IFN)–g ( ). All cytokinesn p 164 n p 186 n p 187 n p 186 n p 187
are shown in picograms per milliliter, except IFN-g, which is shown in units per milliliter.
, for comparison of cytokine production beforeP ! .001
and after treatment for each individual (paired t test of the raw data).
(table 2). Changes in the levels of SEA-specific IL-10 were most
strongly correlated with the changes in SWA- and Teg-specific
IL-10. They were also weakly correlated with changes in SWA-
specific IL-13, IL-5, and IL-4, but not with any Teg-specific
cytokines other than IL-10. There were no responses that were
negatively correlated.
Influence of age on the changes in cytokine production.
There was a very weak negative correlation between subject age
and the change in production of IFN-g in response to SEA
( and [ ]), SWA ( andr p 0.13 P p .074 n p 187 r p 0.18
[ ]), and Teg ( and [P p .013 n p 187 r p 0.20 P p .006 n p
]). The effect was stronger when nonresponders were re-187
moved from the analysis ( and [ ],r p 0.18 P p .039 n p 137
and [ ], and andr p 0.24 P p .003 n p 156 r p 0.23 P p
[ ], respectively). Age was not significantlycorrelated.003 n p 159
with changes in the production of any other cytokines.
Influence of intensity of infection on the changes in cytokine
production. The changes in SEA IL-5 ( and Ppr p 0.193
.008 [ ]), SEA IL-10 ( and [npn p 186 r p +0.248 P p .001
187]), and Teg IL-10 ( and [ ]) werer p +0.17 P p .018 n p 187
positively correlated with pretreatment intensity of infection. In-
tensity of infection was not significantly correlated with changes
in the production of any other cytokines.
Changes in cell populations after treatment. The mean
number of total white blood cells decreased significantly after
treatment, from cells/mL to cells/mL (P
6.72 10 5.46 10
! .000 [ ]), as did the mean number of circulating eo-n p 165
sinophils, which decreased from cells/mL to 0.44-
0.73 10
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Treatment with S. mansoni Increases Th2 Responses JID 2004:190 (15 August) 839
Table 1. Changes in cytokine responses to Schistosoma mansoni soluble egg antigen
(SEA), soluble worm antigen (SWA), and worm-tegument (Teg) as a percentage of pretreat-
ment values.
boost (95% CI) P
boost (95% CI) P
boost (95% CI) P
IL-4 46 (6 to 99) .271 238 (179–29)
!.001 401 (327–476)
IL-5 133 (7–259) .667 460 (344–576)
!.001 1518 (1290–1747)
IL-10 305 (202–408)
.007 444 (355–533)
!.001 1032 (901–1162)
IL-13 84 (20–148) .564 231 (166–296)
!.001 478 (349–607)
IFN-g 68 (26–109) .228 43 (10 to 96) .816 142 (84–199) .051
NOTE. Percentage boost is the percentage change in cytokine responses averaged for all study subjects.
Data were calculated as described in Subjects, Materials, and Methods. CI, confidence interval; IFN, interferon;
IL, interleukin.
A paired t test was performed on raw cytokine data for pretreatment cytokine levels vs. posttreatment
cytokine levels.
.P .01
cells/mL ( [ ]). The mean number of lym-P ! .000 n p 151
phocytes was not significantly altered by treatment (3.7110
cells/mL before treatment and cells/mL 7 weeks after
3.5 10
treatment [ ]).n p 155
The present study has described the effects of treatment with
PZQ on the production of cytokines in response to SEA, SWA,
and, for the first time, the Teg, in a cross-sectional cohort from
an area where S. mansoni is endemic. Both the quantities and
types of cytokines produced after treatment were influenced by
the developmental stage of the parasite from which the Ags
were derived. Before treatment, Th2 cytokines, including IL-4,
IL-5, IL-10, and IL-13, were produced primarily in response
to SWA and Teg, albeit at lower levels in response to the latter,
whereas IL-10 was the only cytokine produced in comparable
amounts in response to SEA. The effects of treatment were also
polarized and Ag specific, with significant increases seen largely
in Th2 cytokines in response to worm-derived Ags, with only
small (and nonsignificant) increases seen in responses to SEA.
After treatment, the greatest increases in IL-4, IL-5, IL-10, and
IL-13 were seen in response to worm-derived Ags and, in par-
ticular, to Teg.
In contrast to what has been described for schistosome-specific
Ab responses, particularly those that have been associated with
resistance to reinfection, such as SWA- or Teg-specific IgE [24],
age exerted little effect on the posttreatment levels of parasite-
specific cytokines. Little association was seen between any cy-
tokine response or change in such response and the pretreatment
intensity of infection. Of all responses measured, only the changes
in the mean SEA- or Teg-specific IL-10 were significantly posi-
tively associated with intensity of infection (albeit weakly).
Our data suggest that treatment with PZQ results in the
release of Ags from adult worms in vivo and that the differential
effects of treatment on the responses to Teg and SWA perhaps
reflect the fact that, before treatment, tegumental Ags are se-
questered from the host immune system. According to this
argument, Teg-specific responses might be specifically boosted
when parasite Ags are synchronously released into the host
circulation after treatment with drugs. It has been suggested
elsewhere that infection with schistosomes causes the down-
regulation of certain cellular immune responses [25–26] and
that the increases in production of cytokines seen after treat-
ment occur as a result of the removal of this immunosup-
pression. This down-regulation has been ascribed to IL-10, pos-
sibly via its direct action on Ag-presenting cells [27, 28]. IL-10
has been shown to play a role in the down-regulation of po-
tentially harmful Th1 parasite-specific cytokines, such as IFN-
g and tumor necrosis factor–a, by cells from individuals chron-
ically infected with S. haematobium [29, 30], and has been
shown to be elicited by the onset of egg laying during murine
infections [31], suggesting that egg-derived Ags might stimulate
its production. The striking specificity of the increases in worm
Ag–induced IL-4, IL-5, and IL-13 that we have described argues
against the notion that such increases are the result of the
removal of general immunosuppression. In the present study,
the mean levels of SEA- and SWA-specific IL-10 increased sig-
nificantly after treatment, suggesting that the specific increases
in worm Ag–induced IL-4, IL-5, and IL-13 seen after treatment
were not dependent on the removal of IL-10–mediated im-
munosuppression. Although IL-10 may be important in con-
trolling responses, there does not appear to be a simple negative
relationship between the levels of IL-10 and any of the other
responses we measured.
The greater increases in responses to worm Ags, compared
with egg Ags, seen after treatment may reflect variations in the
exposure of the host to these Ags during the course of a chronic
infection. Adult worms live for many years in the bloodstream
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840 JID 2004:190 (15 August) Joseph et al.
Table 2. Correlations between the cytokine responses to Schistosoma mansoni soluble egg antigen (SEA), soluble worm antigen
(SWA), and worm-tegument (Teg) that changed significantly after treatment.
IL-4 SWA IL-4 Teg IL-5 SWA IL-5 Teg IL-10 SEA IL-10 SWA IL-10 Teg IL-13 SWA IL-13 Teg
IL-4 SWA 0.765 (164)
0.476 (162)
0.478 (162)
0.213 (163)
0.371 (163)
0.483 (163)
0.397 (163)
0.438 (163)
IL-4 Teg 0.377 (162)
0.385 (162)
NS (163) 0.184 (163)
0.329 (163)
0.234 (163)
0.325 (163)
IL-5 SWA 0.720 (185)
NS (185) 0.380 (185)
0.339 (185)
0.524 (184)
0.463 (184)
IL-5 Teg NS (187) 0.220 (185)
0.360 (185)
0.351 (184)
0.569 (185)
IL-10 SEA 0.529 (186)
0.464 (186)
0.205 (185)
NS (185)
IL-10 SWA 0.678 (186)
0.372 (185)
0.294 (185)
IL-10 Teg 0.287 (186)
0.290 (186)
IL-13 SWA 0.730 (186)
NOTE. Data are Spearman’s rank correlations between the specific cytokine responses that were significantly changed by treatment with praziquantel (no.
of samples included in each correlation). Specific cytokine levels were calculated by subtracting the level of each cytokine produced in the absence of antigen
from that produced in the presence of either SEA, SWA, or Teg for each individual. The change in specific responses was determined by subtracting these
pretreatment from posttreatment levels for each individual. IL, interleukin.
.P ! .001
..001 ! P ! .01
..01 ! P ! .05
and continuously produce large numbers of relatively short-
lived eggs each day, many of which become trapped and die
in host tissues. Thus, the human host is continuously exposed
to the complete repertoire of egg-derived Ags, whereas worm-
derived Ags are less likely to present a constant and chronic
stimulus to the immune system, especially since adult worms
exploit many strategies of immune evasion to preserve the im-
munological invisibility of crucial functional molecules [32].
As a result, even though eggs and worms are targets for PZQ,
many worm Ags are only likely to be exposed to the host when
worms die [33–35]. Posttreatment increases in cytokine pro-
duction could not be explained simply by increases in the num-
bers of circulating lymphocytes or eosinophils, although it re-
mains possible that particular subsets of cells proliferated in
vivo after treatment.
Immune responses to schistosomes in chronically infected
individuals are typically polarized toward Th2 cytokines and
are characterized by eosinophilia, high levels of parasite-specific
IgE, and the predominance of Th2 cytokines, such as IL-4 and
IL-5, in vitro [2]. Previous studies have not specifically ad-
dressed the question of the differential effects of treatment on
cytokine responses to SEA and SWA [10, 36, 37], although some
have shown increases in certain Th2 cytokines in vitro after
treatment for S. haematobium [36, 37]. The significant increases
we observed were polarized toward Th2 cytokines and were
largely specific for worm and, more particularly, Teg Ags. Reg-
ulation of cytokine production by S. mansoni Ags was de-
termined by examining the extent to which the changes in
different pairs of responses were correlated. The strongest cor-
relations were seen between the changes seen in SWA- and Teg-
specific responses for each cytokine, and these correlations were
stronger than those between any pairs of different SWA- or
Teg-specific cytokines. This result suggests that Teg and SWA
Ags might coregulate the production of Th2 cytokines from a
common cellular source and that treatment with PZQ results
in the release of worm-derived and, more particularly, tegu-
mentally derived Ags that initiate this increased production.
The changes in SWA- and Teg-specific IL-5, IL-13, IL-4, and
(to a lesser extent) IL-10 were all positively correlated with each
other, with the strongest correlations generally seen between
the changes in pairs of SWA-specific cytokines. Such positive
correlations again suggest that there is coregulation of these
Th2 cytokines by S. mansoni worm Ags and argue against a
negative regulatory role for IL-10. That there was no significant
change in lymphocyte counts and that eosinophil counts de-
creased significantly after treatment suggest that either (1) the
number of S. mansoni–specific cells had increased but was un-
detectable by the methods used or (2) the cellular production
of SWA- and Teg-specific IL-5, IL-13, IL-4, and IL-10 and SEA-
specific IL-10 had been up-regulated by treatment, which, con-
sidering the magnitude of the increases observed, we consider
to be the most likely explanation.
Immune evasion, partially mediated via the maintenance of
the integrity of the worm Teg, is vital to the long-term survi-
val of the parasite in the bloodstream of the host [38]. Mice can
be successfully immunized against schistosome infection with
homogenized tegumental membranes from adult worms, and
several of the targets of the protective humoral responses are
located in the Teg [39], with levels of specific Abs correlating
with the degree of protective immunity [17, 18]. In Western blot
analysis, serum of infected humans detects a much more re-
stricted set of Ags in preparations of tegumental membranes,
compared with those detected in homogenized whole worms. In
spite of this, the predominant human IgE response seen during
both S. mansoni [7] and S. japonicum [40] infections is directed
against a 22.6-kDa Ag located in the outer Teg. Teg-specific IgE
[7] and parasite-specific IL-4 and IL-5 [9, 10] have been asso-
ciated with resistance to reinfection with schistosomes, suggest-
by guest on February 2, 2016 from
Treatment with S. mansoni Increases Th2 Responses JID 2004:190 (15 August) 841
ing that a Th2 immune response to parasite tegumental Ags may
contribute either directly or indirectly to immunity in humans.
We speculate that the responses that we have observed are driven
by the synchronized release of hitherto sequestered tegumentally
derived Ags into the circulation. The polarized responses that
follow may play a role in the generation and/or maintenance of
the equally polarized Ab responses that have been shown to
correlate with immunity to infection and reinfection in human
populations. It is also interesting to note that the severe hepa-
tosplenic form of human schistosomiasis, which is seen in a
minority of infected individuals, has been attributed in experi-
mental murine studies to parasite egg–specific IL-4 and IL-13
responses that give rise to granulomatous and fibrotic liverlesions
[41]. If this notion is correct, the subdued responses to SEA and
vigorous anti-SWA responses observed in the present study may
be the combination of antiparasite responses that allow infected
human populations in areas of endemicity to maintain protective
responses to the threat of additional infection, while minimizing
the risk of egg-induced, chronic, severe immunopathology.
We thank the people of Booma for their participation in this study and
Colin Fitzsimmons for critical reading of the manuscript.
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... The interaction of PZQ with schistosomes results in a quantitative and qualitative alteration of host-parasitespecific immune responses. Early studies reported modifications in the cell proliferative responses (4) and in the levels and types of antibody (5,6) and cytokine responses (7) following treatment. While this infection and treatment approach to inducing immunologic resistance to reinfection has been well articulated (8), the antigen targets of protective immune responses in human subjects is poorly understood. ...
... 6 significance of difference in mean signal intensity of DIV cohort 18 months after PZQ treatment compared to DIV cohort before treatment. 7 proteins for array inclusion selected from bioinformatic analysis of second-generation S. mansoni array (16) or analysis of S. haematobium proteomes (13,14). T, S. haematobium adult tegument, AES, S. haematobium adult excretory/secretory products, SEA, S. haematobium soluble egg antigen. ...
... PZQ is widely used to treat human schistosome infections and has two main effects on adult flukes -paralysis and tegument damage. Experimental work in mice has demonstrated that PZQ treatment exposes tegumental antigens (22,23), and studies in disease-endemic areas have shown that repeated rounds of PZQ therapy results in enhanced Th2 cytokine and antibody responses, notably IgE against surface membrane proteins (7,24). In addition, adult worms and eggs suppress schistosome-specific immune responses (25) so that worm death results in increased responsiveness to schistosome antigens. ...
Full-text available
Despite the enormous morbidity attributed to schistosomiasis, there is still no vaccine to combat the disease for the hundreds of millions of infected people. The anthelmintic drug, praziquantel, is the mainstay treatment option, although its molecular mechanism of action remains poorly defined. Praziquantel treatment damages the outermost surface of the parasite, the tegument, liberating surface antigens from dying worms that invoke a robust immune response which in some subjects results in immunologic resistance to reinfection. Herein we term this phenomenon Drug-Induced Vaccination (DIV). To identify the antigenic targets of DIV antibodies in urogenital schistosomiasis, we constructed a recombinant proteome array consisting of approximately 1,000 proteins informed by various secretome datasets including validated proteomes and bioinformatic predictions. Arrays were screened with sera from human subjects treated with praziquantel and shown 18 months later to be either reinfected (chronically infected subjects, CI) or resistant to reinfection (DIV). IgG responses to numerous antigens were significantly elevated in DIV compared to CI subjects, and indeed IgG responses to some antigens were completely undetectable in CI subjects but robustly recognized by DIV subjects. One antigen in particular, a cystatin cysteine protease inhibitor stood out as a unique target of DIV IgG, so recombinant cystatin was produced, and its vaccine efficacy assessed in a heterologous Schistosoma mansoni mouse challenge model. While there was no significant impact of vaccination with adjuvanted cystatin on adult worm numbers, highly significant reductions in liver egg burdens (45-55%, P<0.0001) and intestinal egg burdens (50-54%, P<0.0003) were achieved in mice vaccinated with cystatin in two independent trials. This study has revealed numerous antigens that are targets of DIV antibodies in urogenital schistosomiasis and offer promise as subunit vaccine targets for a drug-linked vaccination approach to controlling schistosomiasis.
... Indeed, PZQ treatment markedly alters both the polarization and magnitude of schistosome specific cytokine responses with longer-term immunological impact than just transient clearance of infection. In the human host, Th2 biased response in form of increased parasite-specific IgE, eosinophil numbers, solublehigh and low affinity IgE receptors on eosinophils and CD23+B cells following PZQ administration is observed (23,171,172), which has been related to resistance to reinfection (172)(173)(174). In addition, PZQ chemotherapy alleviates the immune suppression induced by schistosomes by downregulating the immune dampening T-reg, Th17 cell numbers and IL-10 cytokine levels (23,24,175,176). ...
... Indeed, PZQ treatment markedly alters both the polarization and magnitude of schistosome specific cytokine responses with longer-term immunological impact than just transient clearance of infection. In the human host, Th2 biased response in form of increased parasite-specific IgE, eosinophil numbers, solublehigh and low affinity IgE receptors on eosinophils and CD23+B cells following PZQ administration is observed (23,171,172), which has been related to resistance to reinfection (172)(173)(174). In addition, PZQ chemotherapy alleviates the immune suppression induced by schistosomes by downregulating the immune dampening T-reg, Th17 cell numbers and IL-10 cytokine levels (23,24,175,176). ...
Full-text available
Despite mass drug administration programmes with praziquantel, the prevalence of schistosomiasis remains high. A vaccine is urgently needed to control transmission of this debilitating disease. As some promising schistosomiasis vaccine candidates are moving through pre-clinical and clinical testing, we review the immunological challenges that these vaccine candidates may encounter in transitioning through the clinical trial phases in endemic settings. Prior exposure of the target population to schistosomes and other infections may impact vaccine response and efficacy and therefore requires considerable attention. Schistosomes are known for their potential to induce T-reg/IL-10 mediated immune suppression in populations which are chronically infected. Moreover, endemicity of schistosomiasis is focal whereby target and trial populations may exhibit several degrees of prior exposure as well as in utero exposure which may increase heterogeneity of vaccine responses. The age dependent distribution of exposure and development of acquired immunity, and general differences in the baseline immunological profile, adds to the complexity of selecting suitable trial populations. Similarly, prior or concurrent infections with other parasitic helminths, viral and bacterial infections, may alter immunological responses. Consequently, treatment of co-infections may benefit the immunogenicity of vaccines and may be considered despite logistical challenges. On the other hand, viral infections leave a life-long immunological imprint on the human host. Screening for serostatus may be needed to facilitate interpretation of vaccine responses. Co-delivery of schistosome vaccines with PZQ is attractive from a perspective of implementation but may complicate the immunogenicity of schistosomiasis vaccines. Several studies have reported PZQ treatment to induce both transient and long-term immuno-modulatory effects as a result of tegument destruction, worm killing and subsequent exposure of worm antigens to the host immune system. These in turn may augment or antagonize vaccine immunogenicity. Understanding the complex immunological interactions between vaccine, co-infections or prior exposure is essential in early stages of clinical development to facilitate phase 3 clinical trial design and implementation policies. Besides well-designed studies in different target populations using schistosome candidate vaccines or other vaccines as models, controlled human infections could also help identify markers of immune protection in populations with different disease and immunological backgrounds.
... IL-10 is a cytokine with multiple functions in immunoregulation and inflammation. It downregulates the expression of 1 cytokines such as IFN-c and TNF-α, MHC Class II antigens, and costimulatory molecules on macrophages [25]. It also improves B cell survival, proliferation, and antibody production. ...
Full-text available
Schistosomiasis is a parasitic disease that affects millions of people in 78 countries globally. Children under the age of 14, who have the chronic disease may suffer from anemia and malnutrition that contribute to lost days at school and pervasive learning disabilities. The infection is prevalent in Kenya, especially in endemic areas, contributing to significant morbidity. The cellular response pattern is associated with both the acute and chronic phases of the disease, in which cytokines play a critical role. The objective of this study was to evaluate the cytokine profiles of IL-4, IL-2, IL-10, IL-5, IFN-γ, and TNF in serum samples of infected school-aged children by using flow cytometry before and after treatment. The analysis indicated a shift in the expression of the cytokines after treatment with all the cytokines being downregulated, except TNF. There was a general trend of decrease in the expression of the cytokines at six and twelve weeks after treatment as compared to the pretreatment levels. There were statistically significant differences in the expression in IL-2 (P=0.001∗∗), IL-4 (P=0.033∗), IL-10 (P=0.001∗∗∗), IFN-γ (P=0.023∗), and IL-5 (P=0.0001∗∗∗), except in TNF (P=0.095). The reduction in the cytokine levels can be directly related to the influence of the drug praziquantel, modulating the cytokine response by elimination of adult worms, decline in parasitic load, and reduction of morbidity. Therefore, cytokine response is directly related with the influence of treatment in the variation of the immune response.
... However, the cytokine expression profile of PBMCs following PZQ therapy shows a mixed Th1/Th2/Th17 response with nearly all of the cytokines measured were upregulated compared to the cytokine profile at week 13 with the exceptions of TGFβ being downregulated and TNF-α and IL-6 showing no significant change. Destruction of the adult schistosome worm releases antigens that may be concealed from the host immune response, such as antigens in the tegument, thus causing a strong cellular immune response, although marked variability of cytokine expression between individuals can exist (Joseph et al., 2004a;Martins-Leite et al., 2008;Castro et al., 2018;Supplementary Figure 3). ...
Full-text available
For decades, mass drug treatment with praziquantel (PZQ) has been utilized to treat schistosomiasis, yet reinfection and the risk of drug resistance are among the various factors precluding successful elimination of schistosomiasis. Tractable models that replicate “real world” field conditions are crucial to effectively evaluate putative schistosomiasis vaccines. Herein, we describe the cellular immune responses and cytokine expression profiles under field conditions that include prior infection with schistosomes followed by treatment with PZQ. Baboons were exposed to Schistosoma mansoni cercariae through trickle infection over 5 weeks, allowed for chronic disease to develop, and then treated with PZQ. Peripheral blood mononuclear cells (PBMCs) were monitored for cellular immune response(s) at each disease stage and PZQ therapy. After initial infection and during chronic disease, there was an increase in non-classical monocytes, NK and NKT cells while the CD4:CD8 T cell ratio inverted from a 2:1 to 1:2.5. The cytokine expressions of PBMCs after trickle infections were polarized more toward a Th2 response with a gradual increase in Th1 cytokine expression at chronic disease stage. Following PZQ treatment, with the exception of an increase in B cells, immune cell populations reverted back toward naïve levels; however, expression of almost all Th1, Th2, and Th17 cytokines was significantly increased. This preliminary study is the first to follow the cellular immune response and cytokine expression profiles in a non-human primate model simulating field conditions of schistosomiasis and PZQ therapy, providing a promising reference in predicting the immune response to future vaccines for schistosomiasis.
... Chronic infections with schistosomes are associated with immune hyporesponsiveness characterized by reduced in vitro proliferation of T lymphocytes and decreased cytokine production in response to not only schistosomal antigens [2] but also to third-party antigens such as Mycobacterium tuberculosis purified protein derivative [3,4]. Treatment with the antischistosomal drug praziquantel (PZQ) leads to clearance of infection [5], elevated antigen-specific proliferation [6], and cytokine production [7,8]. The mechanisms of T-cell hyporesponsiveness during chronic schistosomiasis are not well understood, but several recent reports have shown that in addition to impaired dendritic cell (DC) activity during schistosome infection [9], regulatory T-cells (Tregs) [10] and regulatory B cells [11] are expanded in schistosome-infected compared with uninfected individuals. ...
Full-text available
Background Although Schistosoma haematobium infection has been reported to be associated with alterations in immune function, in particular immune hyporesponsiveness, there have been only few studies that have used the approach of removing infection by drug treatment to establish this and to understand the underlying molecular mechanisms. Methods Schistosoma haematobium-infected schoolchildren were studied before and after praziquantel treatment and compared with uninfected controls. Cellular responses were characterized by cytokine production and flow cytometry, and in a subset of children RNA sequencing (RNA-Seq) transcriptome profiling was performed. Results Removal of S haematobium infection resulted in increased schistosome-specific cytokine responses that were negatively associated with CD4+CD25+FOXP3+ T-cells and accompanied by increased frequency of effector memory T-cells. Innate responses to Toll like receptor (TLR) ligation decreased with treatment and showed positive association with CD4+CD25+FOXP3+ T-cells. At the transcriptome level, schistosome infection was associated with enrichment in cell adhesion, whereas parasite removal was associated with a more quiescent profile. Further analysis indicated that alteration in cellular energy metabolism was associated with S haematobium infection and that the early growth response genes 2 and 3 (EGR 2 and EGR3), transcription factors that negatively regulate T-cell activation, may play a role in adaptive immune hyporesponsiveness. Conclusions Using a longitudinal study design, we found contrasting effects of schistosome infection on innate and adaptive immune responses. Whereas the innate immune system appears more activated, the adaptive immunity is in a hyporesponsive state reflected in alterations in CD4+CD25+FOXP3+ T-cells, cellular metabolism, and transcription factors involved in anergy.
... In this study, praziquantel treatment did not significantly alter the trajectory of the concentration of any of the cytokines examined. Our results differ from previous studies among non-pregnant individuals infected with S. mansoni that have reported increases in Th2 cytokines following praziquantel treatment [50][51][52]. It, however, remains possible that schistosomiasis infection alters the inflammatory milieu at the MFI, but the prolonged immune response to treatment does not allow modification of this milieu during gestation, suggesting active treatment of all women of reproductive age as recently recommended [53]. There are limitations to this study that should be addressed. ...
Full-text available
Background: The objectives of this study were to 1) evaluate the influence of treatment with praziquantel on the inflammatory milieu in maternal, placental, and cord blood, 2) assess the extent to which proinflammatory signatures in placental and cord blood impacts birth outcomes, and 3) evaluate the impact of other helminths on the inflammatory micro environment. Methods/findings: This was a secondary analysis of samples from 369 mother-infant pairs participating in a randomized controlled trial of praziquantel given at 12-16 weeks' gestation. We performed regression analysis to address our study objectives. In maternal peripheral blood, the concentrations of CXCL8, and TNF receptor I and II decreased from 12 to 32 weeks' gestation, while IL-13 increased. Praziquantel treatment did not significantly alter the trajectory of the concentration of any of the cytokines examined. Hookworm infection was associated with elevated placental IL-1, CXCL8 and IFN-γ. The risk of small-for-gestational age increased with elevated IL-6, IL-10, and CXCL8 in cord blood. The risk of prematurity was increased when cord blood sTNFRI and placental IL-5 were elevated. Conclusions: Our study suggests that fetal cytokines, which may be related to infectious disease exposures, contribute to poor intrauterine growth. Additionally, hookworm infection influences cytokine concentrations at the maternal-fetal interface. Clinical trial registry number and website: (NCT00486863).
... This boost in IgE to adult worm-derived antigens, and antigens cross-reactive with worm-derived antigens, is accompanied by an increase in histamine release upon antigen stimulation of basophils, indicating that the increased IgE is biologically active [50]. The elevated specific IgE levels are also accompanied by an increase in immune cell proliferation [51] and production of type-2 immune cytokines, such as IL-4, 5 and 13 by peripheral blood mononuclear cell and whole blood cultures [52,53]. IL-5, an eosinophil maturation factor, in particular, is elevated in post-treatment supernatants of SWA stimulated immune cells [54,55]. ...
Full-text available
Mass drug administration (MDA) for control of schistosomiasis is likely to affect transmission dynamics through a combination of passive vaccination and reduction of local transmission intensity. This is indicated in phenomenological models of immunity and the impact of MDA, yet immunity parameters in these models are not validated by empirical data that reflects protective immunity to reinfection. There is significant empirical evidence supporting the role of IgE in acquired protective immunity. This is proposed to be a form of delayed concomitant immunity, driven at least in part by protective IgE responses to the tegument allergen-like (TAL) family of proteins. Specific questions have arisen from modeling studies regarding the strength and duration of the protective immune response. At present, field studies have not been specifically designed to address these questions. There is therefore a need for field studies that are explicitly designed to capture epidemiological effects of acquired immunity to elucidate these immunological interactions. In doing so, it is important to address the discourse between theoretical modelers and immuno-epidemiologists and develop mechanistic models that empirically define immunity parameters. This is of increasing significance in a climate of potential changing transmission dynamics following long-term implementation of MDA.
... Most importantly, the egg that induces the immunopathology associated with chronic schistosomiasis had minimal levels of transcripts. Because treatment with PZQ alters immune responses to adult Schistosoma antigens, 58 it could indirectly modify host immune responses to the schistosomula. 59 This implies that cross-reactive antibodies against larvae could be boosted by killing the adults and releasing antigens. ...
Full-text available
While antigens from Schistosoma schistosomula have been suggested as potential vaccine candidates, the association between antibody responses to schistosomula antigens and infection intensity at reinfection is not well‐known. Schistosoma mansoni infected individuals were recruited from a schistosomiasis endemic area in Uganda (n=372), treated with 40mg/kg praziquantel (PZQ) and followed up at five weeks and at one‐year post‐treatment. Pre‐treatment and five‐week post‐treatment immunoglobulin (Ig) E, IgG1 and IgG4 levels against recombinant schistosomula antigens rSmKK7, rSmLy6A, rSmLy6B and rSmTSP7 were measured using ELISA. Factors associated with detectable pre‐treatment or post‐treatment antibody response against the schistosomula antigens and the association between five‐week antibody responses and one‐year post‐treatment reinfection intensity among antibody responders were examined. Being male was associated with higher pre‐treatment IgG1 to rSmKK7, rSmLy6a and AWA. Five‐week post‐treatment antibody responses against schistosomula antigens were not associated with one‐year post‐treatment reinfection intensity among antibody responders’ antibody levels against rSmKK7, rSmLy6B and rSmTSP7 dropped, but increased against rSmLy6A, AWA and SEA at five weeks post‐treatment among antibody responders. S. mansoni infected individuals exhibit detectable antibody responses to schistosomula antigens that are affected by treatment. These findings indicate that schistosomula antigens induce highly varied antibody responses and could have implications for vaccine development. This article is protected by copyright. All rights reserved.
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Background Among the different faces of immune reconstitution inflammatory syndrome (IRIS) developing in HIV-patients, no clinical definition has been reported for Schistosomiasis-IRIS (Schisto-IRIS). Although Schisto-IRIS remains largely uninvestigated, matrix metalloproteinases (MMP) and tissue inhibitors of metalloproteinases (TIMP) have previously been associated with S. mansoni infection and tuberculosis-IRIS. Here, we aimed to investigate the relevance of these markers in Schisto-IRIS. Methodology Patients were diagnosed with IRIS related to S. mansoni within a cohort of patients with Schistosomiasis-HIV co-infection, using a clinical working definition of Schisto-IRIS. We compared 9 patients who developed Schisto-IRIS to 9 Schisto⁺HIV⁺ controls who did not, and 9 Schisto⁻HIV⁺ controls. Plasma levels of MMP-1, MMP-7, MMP-10, TIMP-1, TIMP-2, sCD14, intestinal fatty-acid binding protein, C-reactive protein, and 8 anti-nuclear antibodies (ANA) were analyzed prior to and during 3 months of ART. Principal findings Although no differences were observed for MMP-1 and -7, MMP-10 levels decreased significantly in Schisto⁺HIV⁺ controls during 3 months of ART (p = 0.005) while persisting in Schisto-IRIS patients at significantly higher levels compared to Schisto⁻HIV⁺ controls (p≤0.030). In contrast TIMP-1 levels only decreased significantly in Schisto-IRIS patients (p = 0.012), while TIMP-2 levels were lower compared to Schisto⁺HIV⁺ controls at 2 weeks (p = 0.007), 1 month (p = 0.005) and 3 months (p = 0.031) of ART. Five out of 8 ANAs studied decreased significantly in Schisto-IRIS patients after 1 month of ART(p≤0.039), whereas only 1 ANA decreased for Schisto⁺HIV⁺ controls (p = 0.027). Conclusions/Significance In this study, we propose a working definition for the diagnosis of Schisto-IRIS in resource limited settings. We report persistent plasma levels of MMP-10, along with a more pronounced decrease in TIMP-1 and ANA-levels, and low levels of TIMP-2 during 3 months of ART. Corresponding to the clinical symptoms, these data suggest that Schisto-IRIS is marked by unbalanced MMP/TIMP dynamics which favor inflammation.
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Immunoregulation is considered a common feature of Schistosoma mansoni infections, and elevated levels of T regulatory (Treg) lymphocytes have been reported during chronic human schistosomiasis. We now report that the removal of Treg (CD4+/CD25hi/CD127low lymphocytes) from peripheral blood mononuclear cells (PBMCs) of S. mansoni-infected individuals leads to increased levels of phytohemagglutinin (PHA)-stimulated interferon gamma (IFNγ) production and decreased interleukin-10 (IL-10) responses. Exposure to schistosome antigens did not result in measurable IFNγ by either PBMC or Treg-depleted populations. Interleukin-10 responses to soluble egg antigens (SEA) by PBMC were unchanged by Treg depletion, but the depletion of Treg greatly the decreased IL-10 production to soluble worm antigenic preparation (SWAP). Proliferative responses to PHA increased upon Treg removal, but responses to SEA or SWAP did not, unless only initially low responders were evaluated. Addition of anti-IL-10 increased PBMC proliferative responses to either SEA or SWAP, but did not alter responses by Treg-depleted cells. Blockade by anti-TGF-β increased SEA but not SWAP proliferative responses by PBMC, whereas anti-TGF-β increased both SEA- and SWAP-stimulated responses by Treg-depleted cultures. Addition of both anti-IL-10 and anti-TGF-β to PBMC or Treg-depleted populations increased proliferation of both populations to either SEA or SWAP. These studies demonstrate that Treg appear to produce much of the antigen-stimulated IL-10, but other cell types or subsets of Treg may produce much of the TGF-β. The elevated levels of Treg seen in chronic schistosomiasis appear functional and involve IL-10 and TGF-β in antigen-specific immunoregulation perhaps leading to regulation of immunopathology and/or possibly decreased immunoprotective responses.
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This study examined the hypothesis that the nature of the host cellular immune response to schistosome ova is a risk factor for urinary tract morbidity in areas in which Schistosoma haematobium is endemic. S. haematobium–infected children and adolescents with bladder pathology assessed by ultrasonography had 54-fold greater tumor necrosis factor (TNF)–α production and a 120-fold greater ratio of TNF-α to interleukin (IL)–10 release by peripheral blood mononuclear cells in response to egg antigens, in comparison with control children and adolescents matched by age, sex, and infection severity. Mycobacterial antigens also stimulated 7-fold more TNF-α among subjects with bladder morbidity than in control subjects, which suggests an innate predisposition to enhanced TNF-α production. Levels of egg antigen–induced IL-4 and -5 and interferon-γ were equivalent in subjects with and without bladder pathology. Thus, children and adolescents predisposed to increased TNF-α production to S. haematobium infection are more likely to develop an exaggerated granulomatous response to ova trapped in the bladder wall, with associated urinary tract pathology
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Schistosomamansoni-infectedindividualswhohavelowintensitiesofreinfectionfollowingtreatmentproduce immunoglobulin E (IgE) antibodies against a range ofS. mansoniadult-worm antigens. One of the targets of the IgE response is an adult-worm sodium dodecyl sulfate-polyacrylamide gel electrophoresis band of 22 kDa (Sm22), which contains an antigen(s) located within the tegument and gut lining of adult worms and relatively late schistosomula life cycle stages only. A significant negative correlation between the level of anti-Sm22 IgE and the intensity of reinfection following treatment suggests that IgE responses against this antigen(s) are characteristic of individuals who are resistant to reinfection. To identify the antigen(s) in the Sm22 band that are associated with these IgE responses, we have cloned and characterized a recombinant 22-kDa protein (rSm22) that cross-reacts immunologically with Sm22. There was a high correlation between native and recombinant Sm22 isotype responses, indicating that the correct antigen had been cloned and that responses against rSm22 made up the majority of the responses against Sm22. By analyzing human isotype responses to rSm22 with human sera from a longitudinal treatment and reinfection study and correlating the anti-rSm22 isotype responses, retrospectively, with the intensity of reinfection following treatment for each individual, we observed a negative correlation between the IgE response to rSm22 and the intensity of reinfection. This relationship remained significant after allowing for age and other isotype responses to rSm22, in particular IgG4.
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The susceptibility of two Venezuelan (YT and SM) and one Brazilian (BH) strain of Schistosoma mansoni to single oral doses of praziquantel (Pz; 250 or 500 mg/kg), oxamniquine (Ox; 40, 60, or 100 mg/kg) or to low-dose combinations of both drugs (33 mg/kg Pz and 25 mg/kg Ox; 66 mg/kg Pz and 12.5 mg/kg Ox; 250 mg/kg Pz and 40 mg/kg Ox) was experimentally evaluated in mice. At lower doses of either drug, adult worms of the SM isolate were less susceptible than those of the BH and YT isolates. However, no difference in liver or intestinal egg counts (IECs) could be detected among the isolates after this treatment. At such doses, Pz was better than Ox at reducing IECs. In spite of lowered IECs, eggs continued to accumulate in the liver after Ox treatment. At higher individual doses or following treatment with low-dose combinations of both drugs, no difference in susceptibility could be detected among the parasite isolates. Under such conditions, oviposition was drastically reduced in all three isolates. We confirm that Ox preferentially kills male parasites and present for the first time evidence for the preferential killing of female worms by Pz. We propose that the synergistic effect obtained in the present study and in other investigations using low-dose combinations of both drugs may be due to the preferential cytotoxicity of each drug against a different parasite sex.
Previous studies in school children have demonstrated the slow development with age of resistance to reinfection after chemotherapy of Schistosoma mansoni infections, and have indicated that inappropriate (“blocking”) antibody responses prevent the expression of immunity in young children. The present study was designed to investigate further the nature of the protective responses, by serological studies on a group of 151 S. mansoni-infected individuals resident in an endemic areain Machakos District, Kenya. Antibody levels against various antigens in blood samples before treatment were related to intensity of previous infections; antibodies in blood samples taken 6 months after treatment were related to cumulative reinfection rates over the following 30 months. IgE against an adult-worm antigen preparation correlated positively with age andnegatively with reinfection. In contrast, IgE antibodies against other life-cycle stages showed either no relationship or the reverse correlation. Furthermore, antibodies of other isotypes against adult-worm antigens showed no correlations withreinfection. The correlation with IgE could be demonstrated for different preparations of adult worms, including a periodate-treated preparation presumptively depleted of carbohydrate epitopes. For both the intact and the periodate-treated preparations, multiple regression analysis of the results for children ≤16 years old demonstrated an IgEeffect after allowing for age, although this effect was not observed in a previouslystudied group of school children. Western blot analysis of the adult-worm preparation revealed a limited set of antigens recognized by IgE, among which an antigen of 22 kDa was prominent. The qualitative presence of IgE against this antigen could also beshown to be related to a lack of subsequent reinfection.
Some structural, biophysical and molecular features of the tegument are shown, highlighting their changes during maturation as important adaptative mechanisms for the survival of the parasite and resistance to immune attack. On the other hand, many antigens, targets of the immune response against the schistosome, are located on the tegument. Some of the features of these molecules are presented and the possibilities of using them in the immunological control of the disease are discussed.
The efficacy of praziquantel treatment was significantly enhanced (P < 0.01) in CBA/Ca mice that had been immunized prior to Schistosoma mansoni infection with a crude extract of worm membrane antigens. In Western immunoblots sera from the worm antigen-immunized animals had a poly specific antibody response, with a 25–27 kDa antigen being reacted against with particular intensity. A molecule of similar size was also recognized by rabbit antisera raised against an antigen with esterase activity that has been previously identified as a sensitive target for drug-antibody synergy. The increase in efficacy of subcurative doses of praziquantel in immunized animals is attributed to drug-induced tegumental damage causing antigens to become exposed on the worm surface. Thus, specific antigens, including the 25–27kDa antigen, become accessible to circulating schistosomicidal antibodies. The role of antibodies that can synergize with praziquantel to kill schistosome worms is discussed.,
Host immune responses have been shown to enhance the efficacy of several schistosomicidal drugs. The evidence derives mainly from experiments on Schistosoma mansoni infections in the mouse with their immune status variously modulated; this review emphasises praziquantel (PZQ), which is now the main drug used for treatment of human schistosomiasis. Electron microscopy and indirect immunofluorescence indicate that PZQ disrupts the integrity of the surface membranes of S. mansoni, particularly those covering the dorsal tubercles of adult male worms, and this causes antigens which are the targets of antibody attack to be revealed. We review the evidence that two S. mansoni antigens in particular are implicated in the immune-dependent action of PZQ: a 200 kDa glycoprotein and a 27 kDa antigen with non-specific esterase activity. Consistent with the involvement of the latter antigen, increased non-specific esterase activity was demonstrated histochemically on the surface of intact PZQ-treated male worms, and we describe a chromogenic substrate assay for quantifying the amount of esterase activity that is exposed after drug treatment. The potential relevance of these observations for enhancing the efficacy of drugs currently used to treat human schistosomiasis, and for devising novel therapeutic strategies, is discussed.
To characterize Th cell populations induced by helminth infection, spleen cells from mice infected with Schistosoma mansoni were stimulated with parasite (worm or egg Ag) or mitogen (Con A) and the supernatants assayed for the Th1-specific cytokines IFN-gamma and IL-2 and the Th2-specific cytokines IL-4 and IL-5. Th2 cytokine production was not detected in substantial quantity until the 6 to 8th wk of infection and after reaching peak levels at 8 to 12 wk declined slowly thereafter. The time courses of IL-4 and IL-5 production, whereas differing from each other, closely resembled corresponding published data on IgE and peripheral blood eosinophil levels during murine schistosome infection. In contrast, Th1 cytokine responses occurred only during the first 6 wk of infection and were virtually absent during the peak period of Th2 production. To assess the role of egg deposition in the observed pattern of Th response, cytokine production was assayed in mice carrying unisexual schistosome infections in which parasite eggs are absent. Splenocytes from these animals displayed only marginal Th2 cytokine synthesis but greater Th1 cytokine responses than the corresponding cells from mice with bisexual infections. Moreover, cultures of liver tissue or isolated granulomas from infected mice constitutively produced high levels of IL-4 and IL-5 but failed to synthesize significant amounts of IL-2 and IFN-gamma even when stimulated with egg Ag or mitogen. Taken together the data indicate that egg deposition is the major stimulus of Th2 cytokine response in S. mansoni-infected mice and suggest that T cells belonging to this subset must play a major role in egg granuloma formation.