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Papers in Microbiology Papers in the Biological Sciences

8-1-2007

Exogenous Farnesol Interferes with the Normal Progression of

Cytokine Expression during Candidiasis in a Mouse Model
Dhammika H. M. L. P. Navarathna
University of Nebraska-Lincoln

Kenneth W. Nickerson
University of Nebraska-Lincoln, [email protected]

Gerald E. Duhamel
University of Nebraska-Lincoln, [email protected]

Thomas R. Jerrels
University of Nebraska Medical Center, Omaha, Nebraska

Thomas M. Petro
University of Nebraska-Lincoln, [email protected]

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Navarathna, Dhammika H. M. L. P.; Nickerson, Kenneth W.; Duhamel, Gerald E.; Jerrels, Thomas R.; and
Petro, Thomas M., "Exogenous Farnesol Interferes with the Normal Progression of Cytokine Expression
during Candidiasis in a Mouse Model" (2007). Papers in Microbiology. 79.
https://digitalcommons.unl.edu/bioscimicro/79

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INFECTION AND IMMUNITY, Aug. 2007, p. 4006–4011 Vol. 75, No. 8
0019-9567/07/$08.00⫹0 doi:10.1128/IAI.00397-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Exogenous Farnesol Interferes with the Normal Progression of

Cytokine Expression during Candidiasis in a Mouse Model䌤
Dhammika H. M. L. P. Navarathna,1,2 Kenneth W. Nickerson,1 Gerald E. Duhamel,2
Thomas R. Jerrels,3 and Thomas M. Petro4*
School of Biological Sciences1 and Department of Veterinary and Biomedical Sciences,2 University of Nebraska, Lincoln, Nebraska;
Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska3; and Department of
Oral Biology, University of Nebraska Medical Center, Lincoln, Nebraska4
Received 15 March 2007/Returned for modification 25 April 2007/Accepted 7 May 2007

Candida albicans, a dimorphic fungus composed of yeast and mycelial forms, is the most common human fungal
pathogen. Th1 cytokines such as interleukin-2 (IL-2), gamma interferon (IFN-␥), and tumor necrosis factor alpha
(TNF-␣), which are induced by macrophage IL-12, are critical to resistance against systemic candidiasis, while Th2
cytokines such as IL-4 and IL-5 are less critical. Farnesol is a quorum-sensing molecule produced by C. albicans that
controls the formation of mycelia but is also a virulence factor. To determine whether farnesol enhances the
virulence of C. albicans by modulating the production of Th1 and Th2 cytokines, mice were pretreated with farnesol
prior to intravenous infection with a sublethal dose of farnesol-producing C. albicans. Production of IL-2, IL-4, IL-5,
TNF-␣, IFN-␥, and IL-12 was evaluated by bead-array flow cytometry and enzyme-linked immunosorbent assay.

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Mice exhibited an elevation in serum TNF-␣ levels at 48 h and an elevation in IFN-␥ and IL-12 levels at 6 to 12 h
after infection with C. albicans. Pretreatment with farnesol significantly reduced the elevation of both IFN-␥ and
IL-12 but not TNF-␣. In contrast, mice pretreated with farnesol exhibited an unexpected elevation in IL-5 levels. To
determine whether farnesol has a direct effect on macrophage production of IL-12, peritoneal macrophages were
pretreated with farnesol prior to stimulation with IFN-␥ plus lipopolysaccharide (LPS). Farnesol inhibited pro-
duction of both IL-12 p40 and p70 from IFN-␥/LPS-stimulated macrophages. Therefore, the role of farnesol in
systemic candidiasis is likely due to its ability to inhibit the critical Th1 cytokines IFN-␥ and IL-12 and perhaps to
enhance a Th2 cytokine, IL-5.

Candida albicans is a dimorphic commensal fungus and a This indication that farnesol might be a virulence factor was
medically important opportunistic pathogen, particularly in explored more fully in our recent work, which showed that
immunocompromised individuals (37). Previous work showed farnesol promoted virulence by C. albicans (30). This increased
that yeast-mycelium dimorphism in C. albicans is regulated in virulence was observed when mice received farnesol either
part by secretion of a lipophilic molecule identified as farnesol intraperitoneally or orally in their drinking water. It was also
(15). Farnesol prevents mycelial development in a quorum- observed for a C. albicans mutant with a knockout of DPP3,
sensing or cell density-dependent manner, and it was the first encoding a phosphatase that converts farnesyl pyrophosphate
quorum-sensing molecule (QSM) discovered in a eukaryotic to farnesol. This mutant (KWN2) produced 6 times less far-
organism. This work was recently reviewed by Nickerson et al. nesol and was 4.2 times less pathogenic to mice than its parent.
(33). However, all of the work on farnesol as a QSM has been The strain with DPP3 reconstituted (KWN4) regained both its
done in vitro, leaving open the question of what role farnesol farnesol production levels and pathogenicity.
or farnesol analogs have in vivo. In this regard, Hornby et al. These data suggest that farnesol excretion should be in-
(15) proposed two competing hypotheses. The first was that cluded among a list of virulence factors for C. albicans, but they
the shift from yeast to mycelium was a critical step in patho- say nothing about what the mode of action of farnesol might
genesis and that exogenous farnesol could block this transition, be. At that time we suggested four possibilities: farnesol might
thus acting in a therapeutic manner. The alternate hypothesis (i) promote invasiveness by altering the membrane fluidity of
was that the lipophilic farnesol excreted during infection would host cells; (ii) lyse red blood cells, making iron available for
interact with host cells in a manner that promoted virulence. fungal growth; (iii) protect C. albicans from phagocytes by
The latter idea was supported by our work on the pathogenicity countering their oxidative burst (51); or (iv) interfere with
of C. albicans cells pretreated with subinhibitory concentra- aspects of host defense necessary to survive fungal infection.
tions (0.5 to 1 ␮M) of fluconazole (29). These cells secreted 10 The present work shows that the last possibility is a correct one
times more farnesol than did untreated cells, and they were 4.2 but possibly not the only correct one. Farnesol interferes with
to 8.5 times more lethal (P ⬍ 0.001) than untreated cells in a the normal progression of cytokine induction in mice infected
mouse model of intravenously disseminated candidiasis. with C. albicans.
The pathophysiologies of disseminated candidiasis in mice
and humans are very similar. During early innate immune
* Corresponding author. Mailing address: Department of Oral Bi- responses against systemic candidiasis, phagocytosis and both
ology, University of Nebraska Medical Center, Lincoln, NE 68583.
Phone: (402) 472-1327. Fax: (402) 472-2551. E-mail: tpetro@unmc
oxidative and nonoxidative mechanisms (8) of monocytes and
.edu. macrophages (4) destroy both yeast and hyphae of C. albicans.
䌤
Published ahead of print on 21 May 2007. During the adaptive immune response, cell-mediated immu-

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VOL. 75, 2007 FARNESOL AND CYTOKINE PRODUCTION DURING CANDIDIASIS 4007

nity, with macrophages, T cells, and their cytokines, is required housed at five animals per cage and cared for according to approved protocols of
for complete resistance against disseminated candidiasis. In the Institutional Animal Care and Use Committee of the University of Nebraska.
The institutional standards are in accordance with all federal, state, and univer-
this regard, macrophage interleukin-12 (IL-12) induces the sity rules and regulations. Forty-eight mice were inoculated with 1.3 ⫻ 106 C.
differentiation of CD4 T cells to a Th1 phenotype. The cyto- albicans cells. Of those, 24 animals were also given farnesol i.p. Other control
kines produced by Th1 cells, gamma interferon (IFN-␥) and groups consisted of 24 animals given farnesol administered i.p. and 24 animals
tumor necrosis factor alpha (TNF-␣), signal an increase in the given 0.1% Tween 80 (the solvent for the farnesol stock) administered i.p. These
control groups received no fungal challenge. Therefore, the experimental and
anticandidal activities of macrophages and reinforce expres-
control groups were as follows: C. albicans infected and also given an i.p. injec-
sion of IL-12 (2). Therefore, IL-12 is a critical macrophage tion of Tween 80, C albicans infected and also given an i.p. injection of farnesol
cytokine in immunity to systemic candidiasis. in Tween 80, uninfected with an i.p. injection of farnesol in Tween 80, and
On the other hand, IL-4 plays the pivotal role in develop- uninfected with an i.p. injection of Tween 80. Three animals from each group
ment of an another subset of CD4⫹ T cells, Th2, which secrete were sacrificed at 1, 3, 6, 12, 18, 24, and 48 h postinoculation (p.i.). Twelve
control animals, i.e., untreated and uninfected, were sacrificed, and the serum
IL-4, IL-5, IL-10, and IL-13 (28, 45). These cytokines are was collected. The mean results for these 12 control animals are shown as time
necessary for antibody responses but antagonize the develop- zero values in Fig. 1 through 3. Sera separated from the blood collected from
ment and activity of Th1 cells. Therefore, cytokines from Th1 individual mice were stored at ⫺80°C until the day of analysis. The same exper-
cells have been associated with resistance to C. albicans infec- iment was repeated to examine the IL-12 response by enzyme-linked immu-
nosorbent assay (ELISA) but did not include the 18-h p.i. groups.
tion, while Th2 appears to increase susceptibility to infection.
Macrophages. Macrophages were harvested from the peritoneal cavity 3 days
This hypothesis is supported by a study which showed that after injection of 3% thioglycolate medium (Difco, Detroit, MI). Macrophages
patients with chronic mucocutaneous candidiasis exhibit Th2 were maintained in Dulbecco modified Eagle medium (Invitrogen, Carlsbad,
cytokine production during the immune response to C. albi- CA) containing 10% fetal bovine serum (FBS) (Invitrogen) and 0.05 mg/ml
cans (20). Furthermore, TNF-␣, released by both hematopoi- gentamicin (Invitrogen) and incubated at 37°C with 5% CO2. Peritoneal macro-
phages were cultured overnight, nonadherent cells were removed, and adherent
etic and nonhematopoietic cells, has been shown to play a vital
cells were either left untreated or were pretreated with 100 ␮M farnesol and then

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role in host defense in systemic candidiasis (5). Therefore, were either left unstimulated or were stimulated with 10 ng/ml recombinant
TNF-␣, IFN-␥, and IL-12 are associated with effective resis- mouse IFN-␥ (BD Pharmingen, San Diego, CA) plus 500 ng/ml Escherichia coli
tance to C. albicans infection in mice. A comparative study of O111:B4 lipopolysaccharide (LPS) (Invivogen, San Diego, CA). After 24 h,
C. glabrata and C. albicans revealed that delayed induction or supernatants were collected for IL-12 ELISAs.
Cytokine assays. (i) Bead array. Murine serum was collected from sacrificed
absence of these cytokines during early infection increases mice at various time points following infection with C. albicans with and without
susceptibility to C. albicans (5). Therefore, it is likely that if farnesol pretreatment. The BD Cytometric Bead Array (CBA), the mouse Th1/
innate macrophage and adaptive cell-mediated immune re- Th2 cytokine kit (catalog no. 551287) of BD Biosciences, San Diego, CA, was
sponses are compromised, then the containment of commensal used for the detection IL-2, IL-4, IL-5, TNF-␣, and IFN-␥, according to the
manufacturer’s specifications. Briefly, microbead populations with distinct fluo-
C. albicans at mucosal surfaces is impaired, leading to systemic
rescence intensities were coated with capture antibody specific for each cytokine.
candidiasis (2, 39). The capture beads, phycoerythrin-conjugated detection antibodies, and recom-
C. albicans may have a number of virulence factors that binant standards or test samples were incubated at room temperature. The
delay T-cell and macrophage cytokine expression. We have samples are then washed with 1 ml of wash buffer and recovered by centrifuga-
shown that C. albicans produces farnesol, which acts as a QSM tion (200 ⫻ g for 5 min). After careful aspiration of the supernatant, the beads
were resuspended in 300 ␮l of wash buffer, vortexed briefly, and analyzed with a
to control mycelial development in vitro. In this study, we BD FACScan and BD CBA analysis software. Each assay had a sensitivity range
examined a possible role of farnesol in modulating resistance of 20 to 5,000 pg/ml.
against systemic candidiasis, as well as in modulating T-cell and (ii) ELISA. Ninety-six-well polyvinylchloride plates were coated with 2.0 ␮g/ml
macrophage cytokine expression. The results show that farne- purified unconjugated anti-IL-12 p70 (clone 9A5) or anti-IL-12 p40 (clone
C15.6) monoclonal antibody (BD Pharmingen, San Diego, CA) in bicarbonate
sol increases the susceptibility of mice to systemic candidiasis.
buffer (pH 9.6) at 4°C overnight. After three washes with phosphate-buffered
In addition, farnesol decreases expression of the Th1 cytokine saline (PBS)–0.05% Tween 20 (Sigma, St. Louis, MO), each plate was blocked
IFN-␥ and the Th1-inducing cytokine IL-12 and increases ex- with PBS–10% FBS. For standard curves, serial dilutions of recombinant IL-12
pression of the Th2 cytokine IL-5. p70 and IL-12 p40 (BD Pharmingen) were added to additional wells coated with
antibody. The plates were incubated at room temperature for 2 h. After four
washes with PBS-Tween 20, each plate was incubated with 2.5 ␮g/ml biotinylated
MATERIALS AND METHODS anti-mouse IL-12 p40/p70 (clone C17.8) in PBS–10% FBS at room temperature
Preparation of farnesol. Commercially available E,E-farnesol (Sigma, St. for 30 min. After five washes with PBS-Tween 20, the plates were incubated with
Louis, MO) was used in all experiments. Tween 80 (0.5%) in sterile nonpyro- diluted (1:1,000) avidin-peroxidase (BD Pharmingen) at room temperature for
genic normal saline was used to dilute the farnesol to the required concentra- 30 min. The plates were washed five times with PBS-Tween 20 and incubated
tions. The solubility of farnesol in water is 1.2 mM; therefore, 0.5% Tween 80 was with 3,3⬘,5,5⬘-tetramethylbenzidine substrate/hydrogen peroxide solution (BD
used to increase the concentrations at which farnesol could be resuspended. Pharmingen). IL-12 p40 and p70 were measured by determining the optical
Farnesol in 0.5% Tween 80 or Tween 80 alone (control) was delivered by densities at 450 nm with an ELISA plate reader.
intraperitoneal (i.p.) injection immediately before inoculation. Statistics. The mixed procedure of the SAS system was used to analyze serum
C. albicans and animal inoculation. Stock cultures of C. albicans A72 cells cytokine expression patterns among all treatment groups at various time points.
were kept at 4°C and passaged once a month to maintain viability. Before each Time trends of treatments were compared for significant differences by estimat-
experiment, C. albicans A72 cells were grown for 24 h in 50 ml of modified ing and comparing means of each triplicate reading at various time points.
glucose salts biotin medium (15) at 30°C with aeration. Cells were harvested by
centrifugation at 4,750 ⫻ g for 10 min. Cells were washed three times with 50 ml
of sterile nonpyrogenic saline, counted with a hemacytometer, and resuspended
RESULTS
in saline to the required concentrations. Mice were inoculated intravenously
(i.v.) in the lateral tail vein. For those mice receiving both C. albicans (i.v.) and Farnesol reduces serum IFN-␥ levels but increases serum
farnesol/Tween 80 or Tween 80 alone, i.p. treatment occurred within 5 min of the
i.v. inoculation.
IL-5 levels during systemic candidiasis. IFN-␥ and TNF-␣ are
Mice. Female CF-1 mice (9 to 11 weeks old, 20 to 25 g) (Charles River critical for immunity against systemic candidiasis. To deter-
Laboratories, Wilmington, MA) were used for all experiments. Mice were mine whether farnesol modulates these cytokines during sys-
4008 NAVARATHNA ET AL. INFECT. IMMUN.

FIG. 3. Expression of IL-5 in mouse sera at 1 to 48 h p.i. Experi-

mental groups are represented as described in the legend to Fig. 1.
Data are means ⫾ standard deviations for 3 mice except where “a”
indicates 12 mice per group; “b” indicates means that are significantly
FIG. 1. Concentrations of TNF-␣ in mouse sera at 1 to 48 h p.i. different from control values.
Values at time zero are mean values for control sera from 12 mice,
open bars represent mean values for mice administered i.v. C. albicans
and i.p. Tween 80, closed bars represent mean values for mice admin-
istered i.v. C. albicans and i.p. farnesol in Tween 80, vertical-lined bars levels after 12 and 18 h (P, ⬍0.03 and 0.01, respectively), and,
represent mean values for mice administered i.p. farnesol in Tween 80, in contrast to its effect on TNF-␣, farnesol significantly de-
and horizontal-lined bars represent mice administered i.p. Tween 80. creased IFN-␥ levels (Fig. 2) following C. albicans infection at
Data are means ⫾ standard deviations for 3 mice except where “a” 12 h (P ⬍ 0.04).
indicates 12 mice per group; “b” indicates means that are significantly
Unlike Th1 cytokines, Th2 cytokines may antagonize effec-
different from control values.

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tive immunity against systemic candidiasis. C. albicans infec-
tion alone did not increase serum IL-5 levels compared to
control mice. However, farnesol significantly elevated serum
temic candidiasis, the following experimental design was em- IL-5 levels in C. albicans-infected mice at 3 and 6 h p.i. (P,
ployed. Sera from 12 control mice were compared with a 1- to ⬍0.05 and 0.0001, respectively). In addition, farnesol itself
48-h time sequence for four categories of three test mice each: elevated IL-5 levels at 3, 6, and 12 h p.i. (P, 0.03, 0.0001, and
C. albicans infected and also given an i.p. injection of Tween ⬍0.0001, respectively). As another control, the Tween 80 used
80, C. albicans infected and also given an i.p. injection of to dissolve farnesol did not significantly affect serum IL-5 levels
farnesol in Tween 80, uninfected with an i.p. injection of far- (Fig. 3).
nesol in Tween 80, and uninfected with an i.p. injection of Farnesol reduces serum IL-12 levels during systemic candi-
Tween 80. Serum TNF-␣, IFN-␥, IL-5, and IL-12 concentra- diasis. IL-12 is the determining macrophage cytokine that
tions are shown in Fig. 1 through 4, respectively. For conve- stimulates both development of the Th1 subset and IFN-␥
nience, the control sera (uninfected and untreated) are shown production from CD4 T cells. Therefore, we examined serum
at time zero. C. albicans infection significantly elevated TNF-␣ IL-12 levels in untreated or farnesol-treated mice infected with
levels at 48 h p.i. (P ⬍ 0.01), but farnesol alone did not stim- C. albicans. Significantly different levels of IL-12 were observed
ulate TNF-␣ (Fig. 1) nor did it modulate TNF-␣ production only at 12 h after C. albicans infection. At this time, farnesol
during systemic candidiasis. Similarly, the serum IL-2 and IL-4 pretreatment significantly reduced the levels of serum IL-12
levels were present and detectable but not affected by C. albi- (P ⬍ 0.04) in mice infected with C. albicans (Fig. 4).
cans infection or by farnesol administration either by itself or Farnesol reduces in vitro macrophage production of IL-12
in combination with C. albicans infection (data not shown). p70 and p40 expressed in response to IFN-␥ plus LPS. The
However, C. albicans infection significantly increased IFN-␥ data so far indicate that farnesol suppresses IFN-␥ production

FIG. 2. Concentrations of IFN-␥ in mouse sera at 1 to 48 h p.i. FIG. 4. Serum IL-12 levels at 12 h p.i. Treatments were C. albicans
Experimental groups are represented as described in the legend to Fig. (CA)/Tween 80 (Tw80), C. albicans plus farnesol (CA/F), farnesol (F),
1. Data are means ⫾ standard deviations for 3 mice except where “a” and Tween 80 (Tw80) only. Data are means ⫾ standard deviations for
indicates 12 mice per group; “b” indicates means that are significantly three mice. “a” indicates means that are significantly lower than those
different from control values. for the C. albicans only and Tween 80 only groups.
VOL. 75, 2007 FARNESOL AND CYTOKINE PRODUCTION DURING CANDIDIASIS 4009

TABLE 1. Effects of farnesol on IL-12 expression in macrophages Romani et al. (40) similarly reported that C. albicans-infected
Mean IL-12 production ⫾ SD (pg/ml)
mice did not exhibit increased IL-2 and IL-4 levels at 2 days p.i.
Treatment but that they were elevated at 3 days p.i. Therefore, time after
p40 p70
C. albicans infection is likely critical to IL-2 and IL-4 expres-
None 1.5 ⫾ 2 4.25 ⫾ 8 sion. Interestingly, Scaringi et al. (43) reported that as early as
Farnesola 0.33 ⫾ 0.5 2 ⫾ 3.4 2 h after i.p. infection with C. albicans, and persisting for 5
IFN-␥ and LPSb 3,215 ⫾ 2,300 40.8 ⫾ 24 days, IL-2 expression was detected in cells of the peritoneal
Farnesol/IFN-␥ and LPSc 1.7 ⫾ 1.4 7.1 ⫾ 14
cavity, while TNF-␣ and IL-5 expression was not detected. In
a
Macrophages pretreated with 100 ␮M farnesol. contrast, we clearly showed that after i.v. inoculation with C.
b
Macrophage IL-12 induced with 10 ng/ml IFN-␥ and 500 ng/ml LPS.
c
Macrophages pretreated with 100 ␮M farnesol; IL-12 induced with 10 ng/ml
albicans, TNF-␣ and IL-5 were detected in serum. This sug-
IFN-␥ and 500 ng/ml LPS. gests that the route of C. albicans inoculation may dictate the
T-cell cytokine phenotype.
In view of the robust expression of IFN-␥ by 48 h p.i., and
in response to C. albicans infection through its impact on since IL-4 expression is repressed by IFN-␥, it was not surpris-
macrophage IL-12 p70, a cytokine composed of p35 and p40 ing that IL-4 expression was not induced further at 48 h p.i. It
subunits. In addition, p40 can form homodimers that have was, however, somewhat surprising that we detected IL-5, an-
antagonistic and agonistic effects. However, IL-12 p70-induced other Th2 cytokine, in response to farnesol. Since IL-4 is the
IFN-␥ feeds back and reinforces IL-12 production from mac- prototypical Th2 cytokine, IL-5 is secreted by activated Th2
rophages, while p40 homodimers antagonize this effect (22). following IL-4-induced differentiation (45, 46). While it is pos-
Macrophages produce the maximum amount of IL-12 p70 in sible that farnesol directly stimulates expression of IL-5 from
response to IFN-␥ plus LPS (49). Thus, to determine whether CD4 T cells, the source of IL-5 is more likely to be mast cells

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farnesol directly suppresses IL-12 p70 production compared (35) or eosinophils (9) within the time frame that it was in-
with IL-12 p40, primary peritoneal macrophages were pre- duced in the present study. IL-5, together with IL-4 and IL-2,
treated with 100 ␮M farnesol prior to stimulation with IFN-␥ contributes to activation and proliferation of B cells, while IL-5
plus LPS. This concentration of farnesol is considered relevant by itself is also the terminal eosinophil differentiation factor
for cells cultured in medium with 10% FBS (27). The data (53). Therefore, it is likely that farnesol-treated mice exhibit a
indicate that the combination of IFN-␥ and LPS potently stim- surge in eosinophil production. IL-5 is also a significant con-
ulates production of both IL-12 p40 and p70, 2,100- and 10- tributing cytokine to the development of asthma (47), due to its
fold, respectively (Table 1), while farnesol significantly sup- stimulation of eosinophil development (12). Others have re-
presses production of IL-12 p40 (1,900-fold) and p70 (5-fold) ported IL-5 expression during C. albicans infection (6), and it
from IFN-␥/LPS-stimulated macrophages. Therefore, farnesol is thought that C. albicans may play a role in the development
suppression of immunity against systemic candidiasis likely oc- of certain allergies, including asthma (26, 34). Therefore, for
curs through suppression of both IFN-␥ production by T cells the first time, we show the mechanism by which C. albicans
and IL-12 production by macrophages. induction of IL-5 could contribute to this crucial event during
the development of asthma (10, 17, 31, 38). It remains to be
determined whether the elevated levels of IL-5 produced in
DISCUSSION
response to farnesol play a significant role in inhibiting Th1
The results of the present investigation show that farnesol, a cytokine expression or in suppressing immunity to systemic
QSM of C. albicans, suppresses production of IFN-␥ and IL- candidiasis.
12, but not TNF-␣, induced during the host response to C. In contrast to IL-2, we found that serum IFN-␥ levels in-
albicans. It is clear that TNF-␣, IFN-␥, and IL-12 are required creased in mice infected with C. albicans. Numerous studies
for resistance against systemic candidiasis. TNF-␣ is required with a mouse model noted significant IFN-␥ responses to C.
for resistance to a number of pathogens, including Mycobacte- albicans infection (2, 40). However, in the present study, ex-
rium tuberculosis (42), Listeria monocytogenes (19), and Neisse- perimental groups administered farnesol and then challenged
ria meningitidis (50). It is reported to be secreted by hemato- with C. albicans exhibited a significant decrease in IFN-␥ levels
poietic and nonhematopoietic cells during fungal infections compared with groups challenged with C. albicans but not
(21). Even though low levels of TNF-␣ play a protective role by administered farnesol. Therefore, our finding of depressed el-
inducing macrophages to produce microbicidal reactive inter- evation of IFN-␥ levels in Candida-infected mice treated with
mediates (11) and natural killer cells to produce IFN-␥ (23), farnesol may explain why farnesol decreases resistance to sys-
high levels of TNF-␣ are associated with organ failure and temic C. albicans infection (30). It is not surprising that farne-
septic shock in experimental lethal infections (24). Our find- sol had no effect on IFN-␥ levels after 12 h, since our recent
ings showing increased TNF-␣ at 48 h p.i. in C. albicans- report (30) shows that administered farnesol is cleared from
infected mice agree with previous findings (18, 36, 39, 41). the system after 12 h. Similar to IL-5, the source of IFN-␥
TNF-␣ is produced by a number of cell types following expo- during C. albicans infection at 12 h is not likely to be CD4 T
sure to C. albicans. This cytokine recruits inflammatory cells to cells of the adaptive immune response. It is more likely that
the site of infection, promotes growth and differentiation of T NK cells are a source of early IFN-␥ secretion (1). However,
cells, and activates endothelia (2). Therefore, increased TNF-␣ systemic candidiasis usually involves chronic infection of the
has been associated with Candida infection. kidney. Spellburg et al. (44) reported that the basal concentra-
It was somewhat surprising that serum IL-2 and IL-4 were tion of IFN-␥ is markedly higher in the kidneys than in spleen.
present but not elevated at 48 h p.i. with C. albicans. However, In addition, this group describes cells other than CD4⫹ lym-
4010 NAVARATHNA ET AL. INFECT. IMMUN.

phocytes in the kidney that produce IFN-␥. Therefore, C. 10. Faergemann, J. 2002. Atopic dermatitis and fungi. Clin. Microbiol. Rev.
15:545–563.
albicans may have a virulence factor, such as farnesol, that 11. Farah, C. S., Y. Hu, S. Riminton, and R. B. Ashman. 2006. Distinct roles for
suppresses IFN-␥ production in the kidney. It is likely that the interleukin-12p40 and tumour necrosis factor in resistance to oral candidiasis
significant source of serum IFN-␥ is the kidney, and we pro- defined by gene-targeting. Oral Microbiol. Immunol. 21:252–255.
12. Frigas, E., and G. J. Gleich. 1986. The eosinophil and the pathophysiology of
pose that farnesol produced locally in the kidney by C. albicans asthma. J. Allergy Clin. Immunol. 77:527–537.
has a direct effect on IFN-␥ production there. 13. Goriely, S., B. Vincart, P. Stordeur, J. Vekemans, F. Willems, M. Goldman,
While the mechanism by which farnesol induces IL-5 is un- and D. De Wit. 2001. Deficient IL-12(p35) gene expression by dendritic cells
derived from neonatal monocytes. J. Immunol. 166:2141–2146.
clear, the data presented here suggest that farnesol likely re- 14. Hilliard, B. A., N. Mason, L. Xu, J. Sun, S. E. Lamhamedi-Cherradi, H. C.
duces expression of IFN-␥ through a concomitant reduction in Liou, C. Hunter, and Y. H. Chen. 2002. Critical roles of c-Rel in autoimmune
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Our next goal is to examine what C. albicans genetically cytokines in response to Candida albicans infection in patients with chronic
programmed to produce more farnesol would do to cytokine mucocutaneous candidiasis. Infect. Immun. 71:5690–5699.
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Induction of interferon-␥ from natural killer cells by immunostimulatory
CpG DNA is mediated through plasmacytoid-dendritic-cell-produced inter-
ACKNOWLEDGMENTS feron-␣ and tumour necrosis factor-␣. Immunology 117:38–46.
This work was supported by the University of Nebraska Tobacco 24. Mencacci, A., E. Cenci, G. Del Sero, C. Fe d’Ostiani, P. Mosci, C. Montagnoli,
A. Bacci, F. Bistoni, V. F. Quesniaux, B. Ryffel, and L. Romani. 1998. Defective
Settlement Biomedical Research Enhancement Fund, the John C. and co-stimulation and impaired Th1 development in tumor necrosis factor/lympho-
Nettie V. David Memorial Trust Fund, and the Farnesol and Candida toxin-alpha double-deficient mice infected with Candida albicans. Int. Immunol.
albicans Research Fund of the University of Nebraska Foundation. 10:37–48.
25. Mirani, M., I. Elenkov, S. Volpi, N. Hiroi, G. P. Chrousos, and T. Kino. 2002.
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Editor: A. Casadevall