By
From the * Microbiology Section, Department of Experimental Medicine and Biochemical Sciences,
University of Perugia, 06122 Perugia, Italy; and The Basel Institute for Immunology, CH-L005
Basel, Switzerland
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Abstract |
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Interleukin (IL)-4-deficient mice were used to assess susceptibility to systemic or gastrointestinal Candida albicans infections, as well as parameters of innate and elicited T helper immunity. In the early stage of systemic infection with virulent C. albicans, an unopposed interferon
(IFN)- response renders IL-4-deficient mice more resistant than wild-type mice to infection.
Yet, IL-4-deficient mice failed to efficiently control infection in the late stage and succumbed
to it. Defective IFN-
and IL-12 production, but not IL-12 responsiveness, was observed in
IL-4-deficient mice that failed to mount protective T helper type 1 cell (Th1)-mediated acquired immunity in response to a live vaccine strain of the yeast or upon mucosal immunization in vivo. In vitro, IL-4 primed neutrophils for cytokine release, including IL-12. However,
late treatment with exogenous IL-4, while improving the outcome of infection, potentiated
CD4+ Th1 responses even in the absence of neutrophils. These findings indicate that endogenous IL-4 is required for the induction of CD4+ Th1 protective antifungal responses, possibly
through the combined activity on cells of the innate and adaptive immune systems.
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Introduction |
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Thelper cells play a central role in regulating immune
responses to the fungus Candida albicans by secreting
cytokines that modulate the development and activity of
immune effectors. The dominance of either one of the two
Th subsets (Th1 and Th2) directly correlates with the outcome and severity of infection (1, 2). In experimental
models of candidiasis, protection correlates with the occurrence of Th1 responses (3), whereas Th2 responses are
associated with disease exacerbation and pathology (6). A
variety of factors control CD4+ Th subset development
and regulation in murine candidiasis (1, 2). Cytokines are
among these, acting not only as modulators of antifungal
effector functions but also as key regulators in the development of the different Th subsets from precursor Ths. Early
in infection, neutralization of Th1 cytokines (IFN- and IL-12; references 6, 9) and administration of Th2 cytokines (IL-4 or IL-10; reference 12) lead to the onset of
Th2 rather than Th1 responses, whereas Th2 cytokine
neutralization allows the development of Th1 rather than
Th2 responses (4, 5, 8).
The finding that IL-4 fails to convert an already established Th1 response into a Th2 response (12) and that it is
indeed required for the expression of protective immunity
in mice with candidiasis (13), suggests complex levels of cytokine-mediated regulation of Th development and effector function, which were previously unappreciated. In particular, recent studies in IL-4-deficient mice have revealed
important novel roles for IL-4, which would not have been
predicted from data on IL-4 neutralization in vivo (14).
Thus, in genetically susceptible IL-4-deficient mice with toxoplasmosis, IL-4 appears to have different roles, depending on the phase of infection (16). The infection is also exacerbated in genetically resistant IL-4-deficient mice, IL-4
being required for protection in the late stage of infection
(15). The protective effect of IL-4 appears to be due to its
ability to sustain IFN- production by activated CD4+ T
cells (15), a finding that, although previously described (17,
18), is not easy to accommodate in the paradigm of CD4+
Th regulation by IL-4 (19). However, recent lineage ablation studies by Flavell et al. provide convincing evidence
on the requirement of IL-4 in the differentiation of both
Th2 and Th1 effector cells from precursor CD4+ Ths
(20).
In this study, we assessed susceptibility and parameters of innate and adaptive Th immunity in IL-4-deficient mice infected with C. albicans. The results show the ability of these mice to successfully control the infection in the early stage but not the late, IL-4 being required for induction of protective CD4+ Th1 anticandidal responses.
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Materials and Methods |
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Mice.
Breeding pairs of homozygous IL-4-deficient (IL-4Yeasts, Infections, In Vivo Analyses, and Treatments.
The origin and characteristics of the C. albicans highly virulent CA-6 strain and the live vaccine strain PCA-2 used in this study have already been described in detail (4). For infection, cells were washed twice in saline, and diluted to the desired density to be injected intravenously via the lateral tail vein in a volume of 0.5 ml/mouse or intragastrically via an 18-gauge 4-cm-long plastic catheter, as described (24). The viability of the cells was >95% by trypan blue exclusion and quantitative cultures. Quantification of yeast cells in organs of infected mice (6-8/group) was performed by a plate-dilution method, using Sabouraud dextrose agar, and results (mean ± SEM) were expressed as CFU per organ. Resistance to reinfection was assessed by injecting mice with 106 CA-6 cells intravenously, 14 d after primary infection. Mice succumbing to yeast challenge were routinely necropsied for histopathological confirmation of disseminated candidiasis. Recombinant murine IL-4 (provided by Dr. Robert Coffman, DNAX Research Institute, Palo Alto, CA) was given intraperitoneally together with the anti-IL-4 mAb 11B11 hybridoma (American Type Culture Collection, Rockville, MD; 3 µg IL-4 + 30 µg mAb, each injection) as described (12, 13). Anti-IL-4 mAb was injected intraperitoneally at the dose of 0.5 mg affinity-purified mAb/injection, as described (4, 13). Control groups were injected with saline or isotype-matched Ab (Zymed Laboratories, Inc., South San Francisco, CA). Both treatments were performed on days 6, 8, 10, and 12 after C. albicans infection. Long-lasting neutrophil depletion was obtained as described (25), by administering RB6-8C5 (anti-Ly6G) mAb intraperitoneally at the dose of 100 µg/mouse 2 d before reinfection. Endotoxin was removed from all solutions with Detoxi-gel (Pierce Chemical Co., Rockford, IL).Purification and Culture of Cells.
CD4+ T lymphocytes were positively selected from pools of spleen cells by means of a panning procedure using purified anti-murine CD4 mAb (GK1.5 hybridoma from American Type Culture Collection), which resulted in a >90% pure population on flow cytometric analysis (3). CD4+ cells (5 × 106) were cultured in the presence of 5 × 105 accessory macrophages and 5 × 105 heat-inactivated yeast cells. Unfractionated splenocytes (5 × 106/ml) were cultured in the presence of 10 µg/ml of Concanavalin A (Sigma Chemical Co., St. Louis, MO). Cultures of purified peritoneal neutrophils and macrophages, collected 18 or 72 h, respectively, after intraperitoneal inoculation of aged, endotoxin-free 10% thioglycollate solution (Difco, Detroit, MI), were done as described (25, 26) by incubating 5 × 105 cells in the presence of IFN-Cytokine Assays.
The levels of IFN-Candidacidal Assay and Nitrite Determination.
For the candidacidal assay, 5 × 105 splenic macrophages (obtained by 2-h plastic adherence, >95% pure on esterase staining) or elicited peritoneal neutrophils were plated (0.1 ml/well) in 96-well flat-bottomed microtiter plates (Costar, Cambridge, MA) and incubated with 105 PCA-2 cells for 4 or 1 h, respectively, as described (27). Triton X-100 (final concentration: 0.1%) was then added to the wells, and serial dilutions from each well were made in distilled water. Pour plates (four to six replicate samples) were made by spreading each sample on Sabouraud glucose agar. The number of CFU was determined after incubation at 37°C and the percentage of CFU inhibition (means ± SE) was determined as follows: percentage of colony formation inhibition = 100RNA Preparation and Reverse Transcriptase PCR.
Splenic macrophages, peritoneal neutrophils, and CD4+ splenocytes were subjected to RNA extraction by the guanidium thiocyanate-phenol-chloroform procedure, as described (28). In brief, 3 µg of total RNA was incubated with 0.5 µg of oligo(dT) (Pharmacia Biotech AB, Uppsala, Sweden) for 3 min at 65°C and chilled on ice for 5 min. Each sample was then incubated for 2 h at 42°C after adding 20 U RNase inhibitors (Boehringer Mannheim, Milan, Italy), 1.5 mM deoxynucleoside triphosphates, 25 U avian myeloblastosis virus reverse transcriptase (RT) (Boehringer Mannheim), and RT buffer (50 mM Tris-HCl, pH 8.3, 8 mM MgCl2, 30 mM KCl, and 10 mM dithiothreitol, final concentrations) in a final volume of 20 µl. cDNA was diluted to a total volume of 500 µl with TE buffer (10 mM Tris-HCl, 1 mM EDTA, pH 8.0) and frozen atCompetitive RT-PCR.
The semiquantitative competitive PCR developed by Reiner et al. (30), was performed using the competitor construct containing sequences for multiple cytokines, the primers for HPRT and IFN-Statistical Analysis.
Survival and organ clearance data from each group of wild-type mice were compared with those from IL-4-deficient mice using the Mann-Whitney U test; P < 0.05 was considered significant. Student's t test was used to determine statistical significance between cytokine production of the two groups. In vivo groups consisted of four to six animals. The data reported are pooled from three experiments. ![]() |
Results |
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IL-4/
and IL-4+/+ mice were injected intravenously with different doses (106, 5 × 105 and 2 × 105) of highly virulent
CA-6, with 106 cells of the live vaccine strain PCA-2, or
intragastrically with 108 CA-6. Mice were monitored for
resistance to primary and secondary infections (Table 1)
and for fungal growth in the organs (Fig. 1). The results
show that, although the median survival time (MST) of
mice injected with the highest inoculum of CA-6 did not
differ between the two types of mice, IL-4-deficient mice
were more resistant than IL-4+/+ to the lower inocula of
the yeast, as observed by the increased survival. IL-4
/
mice were as resistant as wild-type mice to infection with
PCA-2 or to intragastric infection with CA-6. However,
on assaying the susceptibility of survivors to a subsequent
lethal CA-6 challenge, IL-4+/+ mice either survived or
showed a remarkable resistance to reinfection, whereas
IL-4-deficient mice did not.
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Quantification of yeast cells recovered from infected
mice revealed that the number of C. albicans cells decreased
in the kidneys of IL-4/
mice throughout the first week
of infection with either 5 × 105 or 2 × 105 CA-6 cells.
However, at 14 d after infection, when no yeast cells were
recovered in mice surviving the lowest inoculum, an extensive fungal growth was observed in the kidneys of mice
succumbing to infection. In contrast, the fungal load in the
kidneys of wild-type mice was always higher and increased
continuously until death. In mice with gastrointestinal infection, the number of yeast cells recovered from the stomach was significantly lower in IL-4
/
than in IL-4+/+
mice, even though both types of mice eventually survived
the infection. Resistance to CA-6 intravenous reinfection
was lower in IL-4
/
than IL-4+/+ mice, as revealed by the
high number of yeast cells recovered from the kidneys of
IL-4
/
reinfected mice. Histopathological examination of
the kidneys of systemically infected IL-4-deficient mice revealed patterns of lesions similar to those observed in resistant or susceptible strains of mice infected intravenously (3-
5), with signs of extensive fungal growth and numerous
foci of inflammatory reaction (mainly consisting of polymorphonuclear cells) throughout the kidney parenchyma of mice succumbing to infection and absence of pathological lesions and fungal growth in kidneys of mice resistant to
infection (data not shown). Therefore, these data clearly
show a two-stage control of C. albicans infection in mice.
IL-4-deficient mice were highly resistant in the early stage
of infection, but highly susceptible in the late stage.
To evaluate the contribution of cells of the innate immune
system on the ability of IL-4-deficient mice to efficiently
oppose infectivity in the early stage of infection, the antifungal effector functions of macrophages and neutrophils
were assessed. Splenic macrophages and peritoneal neutrophils were obtained from IL-4/
and +/+ mice at 3 d after
intravenous infection with 5 × 105 CA-6 cells and assessed
for candidacidal activity and production of NO. The results
(Table 2) showed that the candidacidal activity of both
types of cells was severely depressed in IL-4+/+ mice upon
infection, as opposed to the unimpaired activity of those
from IL-4
/
mice. Interestingly, the candidacidal activity
of macrophages, and lesser that of neutrophils, appeared to
be higher in uninfected IL-4
/
mice as opposed to wild-type mice. Likewise, production of NO occurred successfully in IL-4-deficient mice, and to a lesser extent in wild-type mice. Moreover, the number of peripheral white blood cells did not differ between IL-4
/
and IL-4+/+ uninfected mice and increased comparably upon infection
(data not shown). Therefore, these data suggest that a successful innate antifungal immune response occurs in IL-4
/
mice upon infection.
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IFN- is a potent activator of antifungal effector functions
of phagocytic cells (31, 32) and is produced by a variety of
cells, including NK and CD4+ Th1s (33). Therefore, we
assessed the level of IFN-
production in cultures of unfractionated or purified CD4+ T splenocytes obtained from
mice infected with 5 × 105 CA-6 cells, soon after infection. We also extended the analysis to IL-2, IL-4, and IL-10 production, as resistance and susceptibility to C. albicans
is associated with preferential expansion of cells producing
Th1 and Th2 cytokines, respectively (1, 2). We found that
production of IFN-
by mitogen-stimulated splenocytes was higher in IL-4
/
than +/+ mice, either uninfected or
at 3 d after infection, at a time when no IL-4 or minimal
IL-10 were detected in the former mice compared to wild-type mice (Fig. 2). On assaying cytokine levels produced
by antigen-stimulated CD4+ splenocytes, a similar pattern
of Th1 cytokine production was observed in IL-4
/
and
+/+ mice, in that minimal or no IFN-
and IL-2 were produced. As expected, IL-4 and IL-10 productions were increased in infected wild-type mice, as opposed to no (IL-4)
or minimal (IL-10) detection in mutant mice (Fig. 2).
Therefore, these data indicated that the early sustained production of IFN-
in IL-4
/
mice was derived from cells
other than CD4+ Th1 lymphocytes.
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To investigate whether CD4+ Th1 development may
occur in IL-4-deficient mice, we measured cytokine production in culture supernatants of purified CD4+ splenocytes from IL-4-deficient mice infected under conditions that would otherwise result in the activation of protective
CD4+ Th1s. For this purpose, cytokine production by
splenic CD4+ T cells was assessed in mice reinfected with
virulent CA-6, 14 d after the primary intravenous challenge with PCA-2 or after the primary intragastic challenge
with CA-6. The results showed (Fig. 3) that production of
IFN- and IL-2 were elevated, as expected, in IL-4+/+
mice either surviving reinfection or showing increased resistance to it (Table 1). In contrast, minimal production of
both cytokines was observed in IL-4 mutant mice, succumbing to reinfection. We also assessed levels of IFN-
gene expression in CD4+ T cells from both types of mice
by competitive PCR. IL-4
/
and IL-4+/+ mice were injected with PCA-2 cells and assessed for IFN-
gene expression 3 d after reinfection with CA-6 cells. The results showed a three- to fourfold increase in the IFN-
message
in CD4+ cells from wild-type as compared to mutant mice
(Fig. 4). Interestingly, treatment with exogenous IL-4 restored IFN-
gene expression to a level comparable to that
observed in wild-type-resistant mice.
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Because IL-12 is
both required and prognostic for Th1 development in mice
with candidiasis (9, 10), we evaluated whether the failure of
IL-4-deficient mice to default to the Th1 pathway was associated with a defective production of IL-12. We have recently shown that neutrophils, more than macrophages, are
important sources of IL-12 in vivo in C. albicans infection
(25, 26). Therefore, we looked for IL-12 gene expression
in both neutrophils and macrophages from either type of
mouse after PCA-2 infection. We found that IL-12p40
message, while present in both neutrophils and macrophages from either type of mouse soon after infection (data
not shown), could not be detected in IL-4-deficient mice
later in infection unless mice were treated with IL-4 (Fig.
5). The message also disappeared in IL-4+/+ mice upon IL-4
neutralization, a finding in line with previous data (9). We
also measured IL-12 responsiveness in both types of mice,
by assessing levels of IL-12R1 and IL-12R
2 expression, as
it has recently been demonstrated that loss of IL-12 responsiveness, due to a selective loss of IL-12R
2 subunit expression, represents an early step in the commitment of T
cells to the Th2 pathway (34, 35). We found that expression of the IL-12R
1 subunit gene was similar in either
type of uninfected mice and was not modified upon C. albicans infection. For the IL-12R
2 subunit, expression was
induced by infection and was continuously present in either type of mouse (Fig. 5). Therefore, these data suggest that IL-12 production, but not IL-12 responsiveness, was
impaired in IL-4-deficient mice infected with C. albicans.
To verify whether IL-12 deficiency could be responsible
for the inability of IL-4-deficient mice to mount a protective CD4+ Th1 response, an obvious approach would have
been to supply mice with exogenous IL-12. However, due
to an adverse effect on neutrophils (11), administration of
exogenous IL-12 to mice with candidiasis does not have a
beneficial effect (9, 11). Nevertheless, the finding that in
vitro exposure to IL-12 greatly increased production of
IFN-
by CD4+ T cells from IL-4
/
mice cultured with
Candida Ag and accessory cells (data not shown), indicated
that the defective production of IL-12 may be a limiting factor in the induction of CD4+ Th1 responses in IL-4
/
deficient mice.
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To assess whether IL-4 would directly induce production of IL-12 in mice with candidiasis, we evaluated the effect of IL-4 priming on the ability of neutrophils and macrophages to release immunomodulating cytokines, including IL-12. Elicited peritoneal neutrophils and macrophages from IL-4-deficient mice were exposed in vitro to IL-4 before assessing cytokine levels produced in response to yeast cells. The results showed that priming with IL-4 greatly increased the ability of neutrophils, but not of macrophages, to release IL-12, with the levels of production being similar to those observed in IL-4+/+ mice (Fig. 6 A). Interestingly, priming with IL-4 increased IL-6 production from both types of cells. The potent stimulatory effect on neutrophils appeared to be due to a direct effect of IL-4 on these cells. In fact, expression of IL-4R in response to Candida cells was induced by priming with IL-4 as opposed to what observed in macrophages, whose IL-4R expression appeared to be decreased upon IL-4 priming (Fig. 6 B). The same potentiating effect was observed by adding IL-4 and yeast cells simultaneously (data not shown). These data suggest that IL-4 may be required for an optimal production of IL-12 by neutrophils in mice with candidiasis.
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We have already shown that IL-4 production late in infection is associated with the detection of protective CD4+ Th1s and is
positively regulated by IL-12 (13). Neutralization, but not
administration, of IL-4 late in the course of infection alters
an already established CD4+ Th1 response to C. albicans
(13). Here we show that treatment with IL-4 early in the
course of infection greatly exacerbated the infection in
both IL-4/
and +/+ mice (data not shown), as expected
(12). Late treatment with IL-4 greatly increased resistance
of PCA-2-infected mutant mice to reinfection, as revealed
by increased survival, decreased fungal growth in the kidneys, and increased production of IFN-
and IL-2 by
CD4+ T cells. In contrast, IL-4 neutralization reduced resistance of wild-type mice (Table 3). To assess whether
mechanisms other than regulation of IL-12 produced by
neutrophils may contribute to the ability of IL-4 to sustain
Th1-dependent anticandidal immunity, IL-4 was administered to IL-4
/
and IL-4+/+ mice depleted of neutrophils.
To this purpose, groups of mice with primary infection
were injected with the anti-Ly6G mAb 2 d before reinfection and treated with exogenous IL-4. Depletion of neutrophils decreased resistance to reinfection in both types of
mice, as expected (25). However, exogenous IL-4 partially restored resistance of neutrophil-depleted mice, as indicated by the increased survival, decreased fungal growth,
and increased production of IFN-
and IL-2. Interestingly, the
effects of IL-4 administration were similar to those observed in
mice with primary infection and treated with IL-12-neutralizing Ab before reinfection (data not shown). All together these data indicate that exogenous IL-4 can increase
Th1-mediated anticandidal resistance in IL-4-deficient mice,
and that mechanisms other than those regulating IL-12
production by neutrophils may significantly contribute to
this effect.
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Discussion |
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The results of the present study reveal a novel, previously
unappreciated, role for IL-4 in vivo in mice with C. albicans infection, in which endogenous IL-4 is required for
the induction of protective antifungal CD4+ Th1 responses. Previous studies demonstrating the beneficial (4, 5)
or detrimental (12) effect of early IL-4 neutralization or administration, respectively, in mice with candidiasis led us to
conclude that the occurrence of Th1 or Th2 responses positively correlated with the presence or the absence of IL-4.
However, the subsequent finding that peak production of
IL-4 occurred in mice with candidiasis concomitantly with
the expression of protective, Th1-mediated resistance, and
that depletion but not administration of IL-4 late in infection decreased resistance and production of IFN- and IL-12, suggested that IL-4 may have a positive effect in C. albicans infection, at least in the late stage (13). In the present
study, we found that IL-4-deficient mice, while having an
impaired Th2 response, did not default to the Th1 pathway, thus becoming highly susceptible in the late stage of
C. albicans infection. Susceptibility to infection was not associated with signs of Th2 activation, such as production of
IL-10 and high levels of circulating specific IgE (data not
shown). Also, the message of IL-13 was only transiently induced in IL-4
/
and IL-4+/+ mice upon infection (data
not shown), thus ruling out the possibility that IL-13,
which signals through the IL-4R
chain (36), may compensate for IL-4 deficiency. Therefore, IL-4 is the most
important cytokine in the induction of CD4+ Th2 responses in candidiasis. A similar finding was observed in IL-4-
deficient mice infected with a variety of pathogens, which
have impaired Th2 responses but enhanced (23, 37) or not (14, 42) Th1 responses. We found that production of
IFN-
by CD4+ Th splenocytes was reduced in IL-4
/
mice compared to wild-type mice in response to virulent
C. albicans, but also in experimental conditions of infection
that otherwise result in the induction of CD4+ Th1. Defective activation of CD4+ Th1 was not associated with defective IL-12 responsiveness, as the expression of IL-12R
2s
on these cells was not different from that observed in wild-type mice mounting a CD4+ Th1 response. Therefore,
these data indicate that the unimpaired expression of IL-12
2
on CD4+ cells, due to the lack of inhibitory IL-4 (34, 35),
may not correlate with the functional activation of CD4+
Th1s in murine candidiasis.
The results obtained in the present study clearly evidence
a two-stage control of infection in mice with C. albicans infection. In the early stage, an innate immune response, if
unopposed, may successfully control infectivity in the absence of a supportive CD4+ Th1 response, as observed in
IL-4-deficient mice exposed to low or moderate yeast inocula or with gastrointestinal infection. In this condition,
IFN- derived from a non-T cell compartment, presumably NK cells (33), represents one possible activator of antifungal effector cells (31, 32). However, in the late stage of
infection, IL-12 and CD4+ Th1s producing IFN-
are
required to cope successfully with the pathogen. Indeed,
IL-4
/
mice failed to develop protective CD4+ Th1 responses, as observed upon intravenous or mucosal immunization, thus becoming susceptible to infection at the late
stage. Although IL-4-deficient mice appeared to be resistant to mucosal infection, the data of the present paper do
not seem to suggest a possible late exacerbation of the infection, a finding compatible with the notion that IL-4 may
mediate both protection (43) and pathology (44) at the
mucosal level.
IL-4 appears to be required for the optimal occurrence
of both innate and adaptive immune responses. Fungal
elimination in IL-4/
BALB/c mice was not as efficient as
in genetically resistant similarly infected mice (our unpublished observation), thus suggesting that IL-4 may exert a
positive effect on the antifungal effector function of phagocytic cells. In this regard, IL-4 has been reported to enhance murine macrophage mannose receptor activity (45) and to stimulate phagocytosis and killing of yeast cells by
macrophages (46) and neutrophils (47). Both types of cells
express surface receptors for IL-4 (48, 49). We have previously reported that IL-4 inhibits candidacidal activity and
NO release by macrophages (27), and in the present study
we found that both activities were unimpaired in IL-4-
deficient mice. It is likely that IL-4 may both positively and
negatively affect the antifungal effector functions of phagocytic cells, the net result being dependent on the dose and
time of infection, as observed in leishmaniasis (50).
Whatever the effect of IL-4 on the antifungal effector functions of phagocytic cells, in this study we found that IL-4 efficiently primed neutrophils for IL-12 production in response to the fungus. The effect was associated with the induction of IL-4R on these cells. That IL-4 can prime for IL-12 production has already been observed (51, 52). In human mononuclear cells (51) the positive effect of IL-4 priming on IL-12 production appeared to be due to IL-10 inhibition. Because neutrophils produce IL-12 and IL-10 in response to C. albicans (25, 26), this mechanism could be at work in our system. Interestingly, priming with IL-4 also resulted in the release of high levels of IL-6, which is known to regulate IL-4 receptors on murine myeloid progenitor cells (53). Therefore, it appears that a positive amplification loop exists between IL-6 and IL-4 at the level of neutrophil response, which may be one possible mechanism underlying the beneficial effect of IL-6 in mice with candidiasis (54).
The defective production of IL-12 may likely contribute
to the impairment of CD4+ Th1 development in IL-4-deficient mice with C. albicans infection, as it has been shown
that exposure to IL-12 restores IFN- production in CD4+
T cells from IL-4
/
mice (22). However, the ability of IL-4
to increase IFN-
and IL-2 production in the relative absence of neutrophils also suggests a possible direct effect of
IL-4 on effector Th1s. Elegant studies by Flavell et al. have
recently shown that developing Th1 lineage cells produce
low levels of IL-4 as they differentiate into Th1 effectors
(20, 21), thus implying that endogenous IL-4 could play an
as yet not completely defined physiologic role in modulating Th1 effectors (20). Further studies will elucidate the
important role of endogenous IL-4 in CD4+ Th1 differentiation and maintenance in C. albicans infection.
One interesting and still partially unresolved issue raised
by our studies is where does IL-4 come from and which
yeast/host factors regulate its production in mice with C. albicans infection. Evidence indicates that TCR-/
+ cells,
with an activated phenotype and a biased V
receptor expression, are the early producers of IL-4 in infected mice
(55). The interaction between Candida cells and/or fungal
products with cells producing IL-4 appears to occur
through a superantigen-like mediated mechanism (55, 56).
Whether the late peak production of IL-4 occurring in
mice with candidiasis also occurs in recognition of superantigen-like molecules produced by fungal processing or fungal growth polymorphism remains an interesting possibility. Late peak production of IL-4 has also been observed in
resistant mice infected with Leishmania major, and appears
to be an essential component of the immune response to
this parasite (57, 58).
In conclusion, this study provides several novel features of the immune response to C. albicans, including a physiologic role for IL-4 in the induction and maintenance of IL-12-dependent protective cell-mediated immunity.
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Footnotes |
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Address correspondence to Luigina Romani, Microbiology Section, Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Via del Giochetto, 06122 Perugia, Italy. Phone: 39-75-585-3411; Fax: 39-75-585-3400; E-mail: lromani{at}unipg.it
Received for publication 11 July 1997 and in revised form 15 October 1997.
1 Abbreviations used in this paper: HPRT, hypoxanthine-guanine phosphoribosyl transferase; mRNA, messenger RNA: MST, median survival time; NO, nitric oxide; RT, reverse transcriptase.The authors are grateful to Eileen Mahoney Zannetti for dedicated secretarial and editorial assistance.
This study was supported by AIDS Project 50A.0.28, Italy. The Basel Institute of Immunology has been founded and is supported by Hoffman LaRoche.
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