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Is Required for IL-12 Responsiveness in Mice with Candida albicans Infection1
*
Microbiology Section, Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Perugia, Italy; and
Basel Institute of Immunology, Basel, Switzerland
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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in antifungal CD4+
Th-dependent immunity, 129/Sv/Ev mice deficient for IFN- receptor
(IFN-R-/-) were assessed for susceptibility to
gastrointestinal or systemic Candida albicans infection
and for parameters of innate and adaptive T helper immunity.
IFN-R-/- mice failed to mount protective Th1-mediated
acquired immunity upon mucosal immunization or in response to a live
vaccine strain of the yeast. The impaired Th1-mediated resistance
correlated with defective IL-12 responsiveness, but not IL-12
production, and occurred in the presence of an increased innate
antifungal resistance. The development of nonprotective Th2 responses
was observed in IFN-R-/- mice upon mucosal infection
and subsequent reinfection. However, under experimental conditions of
Th2 cell activation, the occurrence of Th2 cell responses was similar
in IFN-R-/- and in IFN-R+/+ mice. These
results indicate the complex immunoregulatory role of IFN- in the
induction of mucosal and nonmucosal anticandidal Th cell responses;
IFN- is not essential for the occurrence of Th2 responses but is
required for development of IL-12-dependent protective Th1-dependent
immunity. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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(4) and requires the concerted action of several cytokines
(5, 6, 7, 8), including IL-12 (9, 10, 11). Susceptibility to infection correlates
with the generation of CD4+ Th2 cells producing IL-4 and
IL-10 (4, 12, 13, 14) that have been implicated in inhibiting the
development of protective Th1 cells (15) and in opposing the
IFN--mediated activation of fungicidal macrophages (16).
Early studies in vitro indicating IFN- as a major activating factor
for fungicidal phagocytes (17) have been confirmed by recent in vivo
data (18). Analysis of cytokine gene expression in PBMC from healthy
humans stimulated in vitro with Candida Ags has revealed
appreciable levels of IFN- mRNA (19). Overproduction of IFN- may
also be involved in the acute pathology of fungal septic shock (11).
Besides its effector function, the role of IFN- in candidiasis may
encompass early regulatory effects on Th cell differentiation, leading
to the onset of protective immunity (5, 20). IFN- is rapidly
produced after infection in both resistant and susceptible mice (9).
Ag-specific secretion of IFN- protein in vitro by CD4+
cells occurs only in genetically resistant mice (10), such that its
neutralization prevents the development of protective Th1 responses
(5). However, the mechanisms underlying the Th1-promoting activity of
IFN- in candidiasis are not fully understood. High endogenous levels
of IFN- are not sufficient per se to inhibit anticandidal Th2
responses (10), thus suggesting a minimal down-regulatory effect on Th2
differentiation. IFN- might have been required for initial
production of IL-12. However, production of IL-12 by phagocytic cells
in response to Candida occurs in the absence of IFN- even
though it is potentiated by priming with IFN- (11).
IFN- exerts multiple and complex effects on the regulation of
Th induction and effector functions. It was found to be either required
(21, 22) or not required (23, 24) for IL-12-dependent Th1
differentiation of CD4+ T cells and to exert
antiproliferative effects on Th2 cells (25) due to their ability to
express the ß-chain of the IFN-R (26). The lack of this expression
would make Th1 cells resistant to IFN- by preventing transduction of
its signal (26, 27). IFN- could act also as an activator or a death
signal for T lymphocytes (28, 29), thus regulating both Th1 and Th2
responses (30). Recently, a novel and important role has been ascribed
to IFN-: its ability to maintain IL-12 responsiveness in Th1 cells
by overriding IL-4-induced inhibition of IL-12ß2 receptor expression
(31).
The availability of the inbred 129/Sv/Ev strain of mice lacking a
functional IFN- system (32) by inactivation of the 
-chain
receptor
(IFN-R-/-)3
allows us to elucidate the role of IFN- in CD4+ Th cell
development in experimental candidiasis. IFN-R-/- and
wild-type mice were infected under conditions that otherwise result in
the induction of either protective Th1 or nonprotective Th2 antifungal
immune responses. Mice were assessed for susceptibility to infection as
well as for parameters of innate and mucosal and nonmucosal Th
immunity. IFN- exerted complex effects on the regulation of immunity
to C. albicans, affecting both innate and acquired
Th1-dependent immune responses.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The mutant (129/Sv/Ev) mouse strain deficient in IFN-R
-chain (IFN-R-/-) or IL-4 (IL-4-/-)
were generated by gene targeting in murine embryonic stem cells as
previously described (32, 33, 34). IFN-R-/- mice develop a
normal immune response, possess IFN--independent macrophage and NK
cell activities, and constitutively express MHC Ag. However, they lack
a functional IFN- system (33). In these experiments we used
IFN-R-/- and wild-type 129/Sv/Ev mice of both sexes
(bred under specific pathogen-free conditions at the breeding
facilities of the University of Perugia, Perugia, Italy). For each
experiment, groups of mice were matched for sex and age as much as
possible. Procedures involving animals and their care were conducted in
conformity with national and international laws and policies.
Yeasts, infections, in vivo analysis, and treatments
The origin and characteristics of the C. albicans highly virulent strain and the low virulence live vaccine strain used in this study have been described in detail previously (4, 6). For infection, cells were washed twice in saline and diluted to the desired density to be injected i.v. via the lateral tail vein in a volume of 0.5 ml/mouse or intragastrically (108 cells) via an 18-gauge 4-cm-long plastic catheter as previously described (35, 36). The viability of the cells was >95% by trypan blue dye exclusion test and quantitative cultures. Resistance to reinfection was assessed by injecting mice with 106 virulent Candida cells i.v. 14 days after primary infection. Quantification of yeast cells in organs of infected mice (six to eight per group) was performed by a plate dilution method, using Sabouraud dextrose agar, and results (mean ± SEM) were expressed as CFU per organ.
For histology, tissues were excised and immediately fixed in formalin. Sections (3–4 mm) of paraffin-embedded tissues were stained with periodic acid-Schiff reagent and examined for histology as previously described (35, 36). Mice succumbing to yeast challenge were routinely necropsied for histopathologic confirmation of disseminated candidiasis. Soluble IL-4R (Behringwerke, Marburg/Lahn, Germany) was given i.p. at 80 µg/injection on days 1, 3, and 5 after infection as previously described (13).
Purification and culture of cells
CD4+ T splenocytes were positively selected by
sequential adherence on anti-Ig-coated plates and then on
anti-murine CD4 (mAb GK1.5); splenic macrophages (referred to as
adherent or accessory macrophages) were obtained by 2-h plastic
adherence as previously described (4, 6). CD4+ T cells were
cultured (5 x 106) in the presence of 5 x
105 accessory macrophages and 5 x 105
heat-inactivated yeast cells. Intragastric lymphocytes (IGL) were
isolated from the whole stomach after incubation in calcium and
magnesium ion-free HBSS containing DTT (0.145 µg/ml; Sigma, St.
Louis, MO) and EDTA (0.37 µg/ml) and further digestion in RPMI 1640
containing collagenase VIII (100 U/ml; Sigma) and DNase I (0.1 µg/ml,
Sigma) as described previously (8). Lymphocyte-enriched populations,
isolated at the 40/100% interface of a discontinuous Percoll gradient
(Pharmacia, Uppsala, Sweden) were used in a spot ELISA (see below) to
enumerate cytokine-producing cells. Cultures of splenic adherent
macrophages and purified peritoneal neutrophils, collected 18 h
after i.p. inoculation of aged, endotoxin-free 10% thioglycolate
solution (Difco, Detroit, MI), were performed as previously described
(37, 38) by incubating 5 x 105 cells in the presence
of 400 U/ml IFN- and 40 ng/ml LPS (Sigma). Cytokine measurement was
performed in supernatants collected after 48 h (for lymphocytes
and macrophages) and 24 h (for neutrophils) as previously
described (37, 38).
Cytokine, Ab, and ELISPOT assays
The levels of IFN-, IL-2, and IL-4 in culture supernatants
were determined by means of cytokine-specific ELISAs, using pairs of
anti-cytokine mAbs, as previously described (9, 10). The Ab pairs
used were as follows, listed by capture/biotinylated detection:
IFN-, R4–6A2/XMG1.2; IL-2, JES6-1A12/JES6-5H4; and IL-4,
BVD4-1D11/BVD6-24G2.3 (PharMingen, San Diego CA). For IL-12 p70
measurement, a modified Ab capture bioassay was used (7). Cytokine
titers were calculated by reference to standard curves constructed with
known amounts of recombinant cytokines (from PharMingen or Genetics
Institute, Cambridge, MA, for IL-12). A micro-ELISA procedure was used
to quantitate yeast-specific IgG2a, IgG1, and IgE in the sera of mice
as previously described (4). The procedure used to enumerate
cytokine-producing cells was based on an ELISPOT assay, as previously
described (7), using pairs of anti-cytokine mAbs as described
above, including the TRFK-5/TRFK-4 pair of mAbs for IL-5, and freshly
isolated IGL cultured without further stimulation.
Candidacidal assay
For the candidacidal assay, 5 x 105 splenic adherent macrophages or elicited peritoneal neutrophils were incubated with 105 C. albicans cells for 4 or 1 h, respectively, and the number of CFU was determined as previously described (16). The percentage of CFU inhibition (mean ± SE) was determined as the percentage of colony formation inhibition = 100 - (CFU experimental group/CFU control cultures) x 100.
RT-PCR
RNA extraction and amplification of synthesized cDNA from macrophages and purified CD4+ splenocytes were performed as previously described (8, 9). For hypoxanthine-guanine phosphoribosyl transferase (HPRT), IL-12p40, IL-12Rß1, and IL-12Rß2, the primers, positive controls, cycles, and temperature were previously described (8, 9). The HPRT primers were used as a control for both RT and the PCR reaction itself and also for comparing the amount of products from samples obtained with the same primer. The PCR fragments were analyzed by 1.5% agarose gel electrophoresis and visualized by ethidium bromide staining. PCR-assisted mRNA amplification was repeated at least twice for at least two separately prepared cDNA samples for each experiment. Data are representative of three different experiments.
Statistical analysis
Survival data from each group of wild-type mice were compared with those from mutant mice using the Mann-Whitney U test; significance was defined as p < 0.05. Student’s t test was used to determine statistical significance between the two groups for cytokine production and organ clearance. In vivo groups consisted of four to six animals. The data reported are from three experiments.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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R-/- mice to
gastrointestinal C. albicans infection and systemic
reinfection
To assess the functional role of IFN- in an infectious
condition that otherwise results in the development of protective Th1
responses (35, 36), IFN-R-/- and
IFN-R+/+ mice were injected intragastrically with
108 virulent C. albicans cells and monitored for
resistance to mucosal infection and systemic reinfection in terms of
survival and fungal growth in the organs (Table I
). Although survival to gastrointestinal
infection did not differ in either type of mice,
IFN-R-/- mice were more susceptible to infection;
local fungal growth in the stomach was significantly higher than that
observed in wild-type mice at 4 wk after infection. Histopathologic
examination of stomach sections from IFN-R-/- mice
(Fig. 1A) revealed the
presence of numerous intraepithelial abscesses throughout the
nonglandular portion of the stomach starting from the cardial-atrial
fold region, which is a major site of C. albicans
colonization in the murine gastrointestinal tract (35, 36). Abscesses
consisted of a thickened keratinized outer layer that enveloped a dense
aggregate of hyphae and infiltrating inflammatory cells. Conspicuous
signs of acanthosis and hypercheratosis were also visible. In wild-type
mice (Fig. 1B), fewer yeast cells were present in the
keratinized layer, with a few intraepithelial abscesses, a limited
inflammatory reaction at the mucosal level, and no signs of extensive
acanthosis and hypercheratosis. On assaying resistance to systemic
reinfection, wild-type, but not IFN-R-/-, mice showed
increased resistance, as evidenced by survival and decreased fungal
burden in the kidneys (Table I). Histopathologic examination of the
kidneys of IFN-R-/- mice (Fig. 1C) revealed
extensive fungal growth associated with the presence of numerous foci
of inflammatory reaction (mainly consisting of fungal and
polymorphonuclear cells) throughout the kidney parenchyma, as opposed
to the reduced fungal growth and few healing lesions, consisting mainly
of mononuclear cells, observed in the kidneys of
IFN-R+/+ mice (Fig. 1D). These data indicate
that in the absence of IFN- signaling, mice are highly susceptible
to mucosal C. albicans infection and are incapable of
developing resistance to systemic reinfection.
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R-/- mice to C.
albicans infection is associated with failure to mount a
protective Th1 response
To assess the type of Th cell response elicited upon
gastrointestinal infection and reinfection, mice were monitored for
patterns of cytokine and Ab production. Enumeration of
cytokine-producing cells in IGL isolated from mice with
gastrointestinal infection revealed that the numbers of IL-4- and
IL-5-producing cells were decreased in IFN-R-/- mice
compared with wild-type mice. However, the number of IFN--producing
cells was also decreased (Fig. 2). The
number of IFN-- and IL-2-producing cells was decreased in the
draining mesenteric lymph nodes (data not shown). Similarly, upon
systemic reinfection, CD4+ splenocytes from
IFN-R-/- mice produced less IFN- and IL-2 and more
IL-4 than those from IFN-R+/+ mice (Fig. 3A). In addition, serum levels
of Ag-specific IgE were higher while those of IgG2a were lower in
IFN-R-/- than wild-type mice. Similar levels of IgG1
were detected (Fig. 3B). These findings were different from
those observed in mice upon primary systemic infection, in that
production of IFN- and particularly IL-4 and levels of IgE were
equally elevated in both types of mice (data not shown). These results
clearly show the failure of IFN-R-/- mice to mount
protective anticandidal Th1 responses.
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R-/- mice with C. albicans
infection
The inability of IFN-R-/- mice to mount Th1
responses upon C. albicans infection could be due to
defective IL-12 production, defective IL-12 responsiveness, or both. To
distinguish among these possibilities, splenic macrophages and
CD4+ T cells from mice with gastrointestinal infection were
assessed for expression of IL-12 p40 and IL-12Rß1 and IL-12Rß2
chains, respectively, at 3 days after systemic reinfection. The results
show that the expression of the IL-12 p40 gene was not impaired in
IFN-R-/- compared with IFN-R+/+ mice
(Fig. 4) nor was the production of
bioactive IL-12 by splenocytes impaired upon Ag-specific stimulation in
vitro (data not shown). Interestingly, IL-12 p40 gene expression
appears to be up-regulated in IFN-R-/- mice, either
uninfected or upon gastrointestinal infection. In contrast, expression
of the ß2-chain of the IL-12R was down-regulated in
IFN-R-/- mice, with and without reinfection, but not
in wild-type mice. The results also show that the constitutive
expression of the IL-12Rß2 mRNA was different in uninfected control
mice, being apparently increased in IFN-R-/- mice. A
similar finding was observed in IL-12 p40-/- mice
(unpublished observation). No changes were observed in either type of
mouse in expression of IL-12Rß1 (Fig. 4). Both receptor subunits were
similarly expressed in the two types of mouse upon primary i.v.
infection (data not shown). These data indicate that a defective IL-12
responsiveness, rather than IL-12 production, could account for the
inability of mice with impaired IFN- signaling to mount anticandidal
Th1 responses.
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is not essential for the development of anticandidal Th2
responses
The observation that failure to activate anticandidal Th1
responses was concomitant with the expression of Th2-dependent immunity
in IFN-R-/- mice prompted us to investigate whether
IFN- could also exert an inhibitory effect on the development of
CD4+ Th2 responses. For this purpose,
IFN-R-/- and IFN-R+/+ mice were
injected i.v. with low virulence C. albicans and were
monitored for survival and cytokine production. Wild-type mice were
highly susceptible to infection, as indicated by 14-day survival (Table II) and by extensive fungal growth in the
kidneys (data not shown). On assaying cytokine production at 7 days
after infection, Ag-activated CD4+ splenocytes produced
IFN-, but relatively low levels of IL-2 and IL-4. However, IL-4
neutralization by treatment with soluble IL-4R significantly increased
resistance to infection. Similarly, 129/Sv/Ev IL-4-deficient mice were
highly resistant to infection and, importantly, to subsequent
reinfection with virulent C. albicans cells. Both IFN-
and IL-2 productions by CD4+ T cells were increased. These
data indicate that the high susceptibility of 129/Sv/Ev to acute
C. albicans infection is associated with the occurrence of
inhibitory IL-4-producing Th2-cells, such that in the absence of IL-4,
development of protective Th1 responses occurred. On infecting
IFN-R-/- mice, no significant changes in cytokine
production were observed compared with that in wild-type mice. Analysis
of IFN- and IL-4 gene expression by quantitative RT-PCR confirmed
the results obtained in terms of actual cytokine production, as the
levels of IFN- and IL-4 gene expression were not different in the
two types of mice early in the course of the infection (data not
shown). Therefore, the increased resistance to infection does not
correlate with decreased Th2 reactivity, as production of IL-4 occurs
similarly in both types of mice. We found higher innate resistance to
infection in mutant compared with wild-type mice, although they were
unable to develop resistance to reinfection. Figure 5A shows that the candidacidal
activity of macrophages and neutrophils from uninfected mice was higher
in IFN-R-/- than in IFN-R+/+ mice,
although this activity declined later in the course of infection. We
also found that production of IL-12 was not detected in macrophages and
was only minimally detected in neutrophils from IFN-R+/+
mice as opposed to the higher production observed, particularly in
neutrophils, in IFN-R-/- mice (Fig. 5B).
These data clearly point to a minimal role, if any, for IFN- in
inhibiting the development of Th2 responses in mice with C.
albicans infection and confirm its requirement for the development
of protective Th1 responses.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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in the induction of mucosal and nonmucosal anticandidal Th cell
responses. While not necessary for the induction and expression of Th2
cell responses, IFN- appeared to be an essential requirement for the
development of protective anticandidal Th1 cell responses, a finding
confirming the result obtained by IFN- neutralization in genetically
resistant mice (5).
In murine candidiasis, both resistance to mucosal infection and
subsequent systemic reinfection are dependent on the activation of Th1
cell responses (4, 35, 36). IFN-R-/- mice were highly
susceptible to gastrointestinal infection and were unable to develop
resistance to subsequent reinfection. Susceptibility to mucosal
infection correlated with a decreased frequency of intragastric T
lymphocytes producing both Th1 (IFN-) and Th2 (IL-4 and IL-5)
cytokines. Decreased production of IFN- and IL-2 was also observed
in draining lymph nodes, clearly indicating the failure of
IFN-R-/- mice to mount anticandidal Th1 responses at
the mucosal level. In systemic reinfection, susceptibility correlated
with a defective production of IFN- and IL-2 by CD4+ T
splenocytes upon Ag-specific stimulation in vitro.
Failure to mount protective anticandidal Th1 responses was not
associated with a decreased production of IL-12, which occurred in
these mice after mycobacterial (39) or viral (40) infections. This
finding is in line with our previous observation indicating that
production of IL-12 by phagocytic cells in response to
Candida occurs in the absence of IFN- even though it is
potentiated by priming with this cytokine (11). In contrast, impaired
IL-12 responsiveness was observed, as expression of the IL-12ß1
subunit receptor was unaffected but that of IL-12Rß2 subunit was
defective in IFN-R-/- mice.
That both IL-12ß1 and IL-12ß2 receptor subunits are essential
components of the functional mouse IL-12R (41) and that IFN- is
needed to override IL-4-induced inhibition of IL-12ß2 receptor
expression in Th1 cells have recently been reported (31). Our results
suggest that in murine candidiasis the Th1-promoting activity of
IFN- may rely on its ability to maintain IL-12 responsiveness rather
than to induce IL-12 production. Although IL-12 is required for Th1
development in mice with candidiasis (9, 10), we have recently shown
that IL-12 responsiveness, rather than production, correlates with the
occurrence of Th1 cell responses (8).
In the course of the present study we found that the number of
CD4+ splenocytes significantly decreased in
IFN-R-/- mice soon after primary systemic infection
(data not shown). As IFN- has been shown to act as an activator or a
death signal for T lymphocytes (28, 29), other mechanisms of
IFN--dependent regulation of Th1 induction and effector functions in
mice with candidiasis cannot be excluded.
The essential requirement for IFN- in Th1 development found in
129/Sv/Ev IFN-R-/- mice with candidiasis appears to be
in contrast with the occurrence of a Th1-type response observed in
these mice upon parasitic (42) or viral (43) infection. However, the
finding that they are resistant to Th1-mediated experimental autoimmune
encephalomyelitis (44) indicates the complex role for IFN- in
mediating Th1 induction and expression in this particular strain. This
observation would be in line with the concept of genetic differences in
default pathways for Th1 and Th2 development (45), which may include
the diverse influence of IFN- on differentiation of T cell subsets
in different mouse strains. In this regard, it has been recently
hypothesized that IFN- may have an obligate role in directing the
Th1 phenotype only under conditions where IL-12 production is limiting
(46).
Given the antiproliferative effects of IFN- on Th2 cells (25), an
increased Th2 cell activity in IFN-R-/- mice upon
C. albicans infection was expected. However, numerous
studies indicate that the production of IL-4 in the absence of IFN-
or IFN- signaling appears to be dependent on the genetic background
and the type of infection. Indeed, administration of IFN- mAb during
infection with Leishmania major (22) or Listeria
monocytogenes (47) leads to disease progression associated with
the appearance of Th2 cells. Likewise, infection of mice with a
disrupted IFN- gene with L. major (48), influenza virus
(49), or Mycobacterium tuberculosis (50) increased the
production of IL-4 and IL-5. In C57BL6 IFN-R-/- mice,
increased production of IL-4 was observed upon infection by an
intracellular pathogen (51). However, studies in 129/Sv/Ev
IFN-R-/- mice failed to demonstrate a default of
CD4+ cells to the Th2 phenotype upon viral (43), parasitic
(42), or bacterial (52) infection.
We found that production of Th2 cytokines was locally decreased in
IFN-R-/- mice with gastrointestinal infection, thus
confirming the occurrence of impaired Th2-dependent mucosal immune
response in these mice, as previously reported (53). The susceptibility
of IFN-R-/- mice to systemic reinfection was
associated with an up-regulated expression of Th2 cell responses, as
revealed by high level production of IL-4 and IgE. However, on directly
assessing the functional role of IFN- in anticandidal Th2 cell
development, we found that IFN- is not essential for the development
of anticandidal Th2 responses.
Although 129/Sv/Ev mice develop protective Th1 responses to many
parasitic, bacterial, and viral infections (52), we found that
129/Sv/Ev mice are highly susceptible to acute infection with low
virulence Candida cells and succumb to it. Although IL-4
production was barely detectable upon infection, a common finding
observed in other infections (42, 43), resistance to infection was
dramatically increased in the absence of IL-4 or upon IL-4
neutralization, clearly showing a pathogenetic role for IL-4-producing
Th2 cells in the susceptibility of mice to infection. On assaying the
pattern of Th cell responses in similarly infected
IFN-R-/- mice, a comparable level of Th2 cell
reactivity was observed, as revealed by IL-4 protein production and
cytokine gene expression by quantitative PCR. These results confirm
previous observations indicating that high levels of endogenous IFN-
are not sufficient per se to inhibit anticandidal Th2 responses (10).
They also indicate that IFN- is not essential for the occurrence of
anticandidal Th2 cell responses, a finding in line with that obtained
in mice with a disrupted IFN- gene and infected with C.
albicans under conditions of Th2 cell activation (54).
Despite the occurrence of Th2 cell responses, IFN-R-/-
mice were more resistant to primary acute systemic infection than
wild-type mice. A similar finding was observed in IFN--deficient
mice (54). This could indicate that mechanisms of innate resistance may
efficiently contribute to the early control of infection in
IFN-R-/- mice. However, other studies have indicated
an impaired macrophage activation in these mice (32, 55), which was
considered to contribute to their susceptibility to certain infections.
We found that while the number of circulating neutrophils increased
similarly in wild-type and mutant mice after infection (data not
shown), the candidacidal activity of macrophages and neutrophils was
higher in IFN-R-/- than in wild-type mice, declining
later in the course of infection. In wild-type mice no signs of
activation of these cells were observed upon infection. Production of
nitric oxide is either increased (55) or decreased (56) in 129/Sv/Ev
IFN-R-/- mice; we were unable to measure production of
nitric oxide in either type of mouse upon infection (data not shown).
Thus, a prompt and efficient antifungal effector activity of phagocytic
cells may account for the increased resistance of
IFN-R-/- mice to the acute infection.
Given the importance of cytokines acting on or secreted by macrophages
and neutrophils during infection (57), including C. albicans
infection (37, 38), it is likely that a differential production of
cytokines with activating or deactivating activity on phagocytic cells
may account for the different levels of antifungal effector function
observed in mutant and wild-type mice. We found that the production of
IL-10 was lower in macrophages and neutrophils from mutant compared
with wild-type mice (data not shown). As IL-10 is a potent inhibitor of
antifungal effector functions of IFN--activated phagocytic cells
(16) and inhibits IL-12 synthesis in these cells (57), the defective
production of IL-10 could account for the high level activation of
antifungal effector function and IL-12 production in mutant mice.
Preliminary results in wild-type mice treated with IL-10-neutralizing
mAb show an increased resistance to the primary systemic infection
associated with an increased candidacidal activity of effector
phagocytic cells, a finding confirming the suppressive role of IL-10 in
C. albicans infection (15). Given that IFN- acts as a
negative regulator of IL-10 transcription (58), it will be of interest
to investigate mechanisms by which deficiencies in IFN- signaling
lead to decreased IL-10 production in mice with C. albicans
infection.
In conclusion, this study shows that IFN- is not essential for the
occurrence of Th2 responses but is required for development of
IL-12-dependent protective Th1 responses in mice with C.
albicans infection. Our results confirm those of other
experimental models of fungal infection (59) and suggest the importance
of IFN- signaling at the levels of innate and acquired immune
responses to fungi.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Luigina Romani, Microbiology Section, Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Via del Giochetto, 06122 Perugia, Italy.
3 Abbreviations used in this paper: IFN-R-/-, IFN- receptor deficient; IGL, intragastric lymphocytes; ELISPOT, enzyme-linked immunospot; HPRT, hypoxanthine-guanine phosphoribosyl transferase.
Received for publication December 27, 1997. Accepted for publication May 27, 1998.
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) and
IFN-
) mice, uninfected (none) or
intragastrically infected with C. albicans.
Cytokine-producing intragastric T lymphocytes were enumerated by
ELISPOT assay at 4 wk after infection.
