Departamento de Bioquímica Clínica, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Ciudad Universitaria, Pabellón Argentina 5000, Córdoba, Argentina
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ABSTRACT |
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The integration of innate and
adaptive immune responses is required for efficient control of
Candida albicans. The present work aimed to assess, at the
local site of the infection, the immunocompetence of macrophages in
rats infected intraperitoneally with C. albicans and exposed
simultaneously to stress during 3 days (CaS group). We studied the
1) ability to remove and kill C. albicans,
2) tumor necrosis factor- (TNF-
) release,
3) balance of the inducible enzymes NO synthase (iNOS) and
arginase, and 4) expression of interleukin (IL)-1
and
IL-1 receptor antagonist (ra) mRNA. Compared with only infected animals
(Ca group), the number of colony-forming units was significantly higher
in CaS rats (P < 0.01), and the macrophage
candidicidal activity was ~2.5-fold lower (P < 0.01). Release of TNF-
was diminished in both unstimulated and
heat-killed C. albicans restimulated macrophages of the CaS
group (Ca vs. CaS, P < 0.03 and P < 0.05, respectively). In Ca- and CaS-group rats, the rates for both the
arginase activity and the NO synthesis were significantly enhanced.
However, the stress exposure downregulated the activity of both enzymes
(CaS vs. Ca, P < 0.05). After in vitro restimulation,
the IL-1ra/IL-1
ratio was significantly diminished in CaS-group rats
(P < 0.05). Our results indicate that a correlation
exists between early impairment of macrophage function and stress exposure.
arginase; nitric oxide; tumor necrosis factor-
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INTRODUCTION |
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CELL-MEDIATED IMMUNITY IS essential for host protection against virtually all fungal pathogens, and in individuals with intact immune function, the defense mechanisms are highly efficient at preventing fungal diseases (40). The yeast Candida albicans has evolved as a successful commensal in healthy persons (31) and is also a member of the oral and gastrointestinal flora in immunocompetent humans; however, in immunocompromised hosts, the transition of C. albicans into an opportunistic pathogen is not uncommon, and disseminated candidiasis of endogenous origin may occur (22). A proper integration between the innate and the adaptive immune systems is required for efficient control of C. albicans (40, 41). Resistance to C. albicans infection is determined by phagocytic effector mechanisms enhanced by Th-1 cytokines (40, 41). Regulation of the early fungal burden, the cytokine production, and the expression of costimulatory molecules are possible pathways through which the innate immune system may condition the development of the adaptive response.
Evidence derived from experimental studies demonstrates the importance of polymorphonuclear neutrophils (PMNs) for preventing candidal dissemination. A direct correlation between the level and duration of neutropenia and the host's susceptibility to candidiasis exists (43). However, the occurrence of systemic candidiasis in hosts with normal PMN function suggests a protective role for other cells including mononuclear phagocytic cells (47). In contrast to several negative reports on the role of macrophages (40, 41, 47), many studies support the involvement of these cells in the resistance to C. albicans infection (21). Accordingly, the selective elimination of macrophages in euthymic (21, 34) or athymic (34) mice is associated with a slow clearance and an enhanced susceptibility to candidiasis.
Thus far, characterized factors that predispose one to C. albicans infection include prolonged use of broad-spectrum
antibiotics, disruption of the gastrointestinal barrier by cytotoxic
drugs, neutropenia, or T-cell dysfunction (31, 43).
Recently we developed an experimental model of candidiasis that is
suitable to study the contribution of innate effector cells in host
protection against this fungus (38, 39). We found severe
alterations in the immune response against C. albicans in
rats infected and exposed to chronic varied stress (CVS) during 10 days
(38). Interestingly, already after 3 days of stress
exposure, we observed a bigger fungal burden in kidneys and livers and
a poor inflammatory response in these organs. The number of phagocytes
(macrophages and neutrophils) recruited at the site of the infection
was significantly reduced (39), and the candidicidal
activity was diminished (38). The integrin Mac-1, which is
involved in migration and macrophage activation, was also significantly
diminished after CVS exposure (39). However, PMN oxidative
burst was not impaired (38), which is in agreement with
previous reports of the resistance of neutrophils to endogenous doses
of glucocorticoids (42). The present work aimed to assess
at the local site of the infection the immunocompetence of macrophages
from rats exposed to C. albicans infection and stress during
3 days. We studied the 1) ability to remove and kill
C. albicans, 2) tumor necrosis factor-
(TNF-
) release, 3) balance of the inducible enzymes
nitric oxide synthase (iNOS) and arginase, and 4) expression
of interleukin (IL)-1
and IL-1 receptor antagonist (ra) mRNA.
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METHODS |
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Animals. Outbred female Wistar rats (body wt, 100-150 g) were collectively housed in cages in the experimental room for at least 3 days before experiments were started. The rats were maintained at 22°C under a 12:12-h light-dark cycle with light onset beginning at 0700. Animals had continuous access to food and water except when food was removed from the stressed groups for 24 h as part of the stress procedure.
Microorganism and infection. The pathogenic C. albicans, strain no. 387, was taken from the stock culture collection of the Mycology Division, Department of Clinical Biochemistry, Faculty of Chemical Science, National University of Córdoba. Yeast cells were grown on Sabouraud glucose agar slant at 28°C, maintained by weekly subculture on the same medium, and periodically checked for assimilation pattern and virulence. For each infection, yeast cells were harvested after 48 h of culture, centrifuged at 1,000 g, washed twice in sterile 0.15 M NaCl supplemented with 0.1% gentamicin, counted, and diluted to the desired concentration. The number of viable cells was checked in triplicate by counting the colony-forming units (CFUs) on the Sabouraud agar after 48 h of incubation at room temperature (RT).
Stress procedure. Animals were exposed to different stressors between 1400 and 1600 except for food deprivation, which lasted for 24 h. In our model of candidiasis and stress (38, 39), rats are infected and exposed to a CVS paradigm that involves a different stressor each day and lasts 10 days (14). In the present work, we focused on the impairment of the immune response observed after 3 days of treatment as reported previously (38, 39). The stress procedure described (see Experimental design) is the fraction of the CVS paradigm that corresponds to the first 3 days, and includes day 0, swim (4°C for 5 min); day 1, restraint (for 2 h); and day 2, food deprivation (for 24 h). Protocols were approved by the Animal Experimentation Ethics Committee, Faculty of Chemical Science, National University of Córdoba.
Experimental design. Rats were assigned to one of four treatment groups: the uninfected and unstressed normal (N), stressed (S), C. albicans-infected (Ca), and infected and stressed (CaS) groups. Rats were infected intraperitoneally with a volume of 1 ml of inoculum (3 × 108 yeasts/ml) on day 0, and stress was imposed immediately after the infection and during the next 2 days. On day 3, animals were killed by decapitation, and peritoneal cells (PCs) were obtained by sterile lavage with 25 ml of RPMI 1640 without phenol red (Sigma) supplemented with 0.1% gentamicin and 50 U/ml heparin.
Peritoneal CFU determination. A 0.5-ml aliquot of peritoneal lavage fluid (from the 20 ml recovered) obtained as described was surface-plated in duplicate on Sabouraud glucose agar. The CFU were counted after 48 h of incubation at RT. Means and SE values were calculated from five rats per group. Experiments were performed three times. Results are expressed as CFU.
Macrophage purification. PCs were adjusted to 2 × 106 cells/ml in RPMI 1640 that contained 10% fetal calf serum and 0.1% gentamicin. Macrophages were purified by adherence during a 2-h incubation (at 37°C with 5% CO2) and depending on the experiment, 24- or 96-multiwell plates were used. Nonadherent cells were removed by washing the plates twice with cold RPMI 1640 medium. In our experimental condition, adherent cells were ~50% of the PCs plated with a purity of >90% according to morphological analysis or nonspecific esterase (Sigma) staining (35). No changes in adherence properties were observed in cells from stressed animals, which is in agreement with previous reports (36).
Killing of C. albicans by macrophages. The candidicidal activity of macrophages was assessed by a colorimetric assay (7). Briefly, C. albicans-macrophage coculture experiments were set up in 96-well, flat-bottom plates (Corning) in a final volume of 200 µl; viable C. albicans were diluted appropriately in RPMI 1640 that contained 0.1% gentamicin, and 10% of normal rat serum (NRS) was added to macrophages at an effector-target (E/T) cell ratio of 100:1. After 72 h (at 37°C with 5% CO2), plates were treated with 0.1 ml of 1% Triton X-100 in distilled water to lyse the macrophages and were washed twice with 0.1 ml of distilled water. Then 0.1 ml/well of RPMI 1640 without phenol red that contained 0.5 mg/ml of 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) (Sigma) was added, and the plates were incubated for 2 h at 37°C. After the incubation, the plates were centrifuged, the supernatants were discarded, and the pellets were resuspended in 0.2 ml of isopropanol. When the dark-blue formazan crystals had dissolved, the reaction was read at 540 nm using an automated microplate reader (TiterTek MultiScan). The color developed is proportional to the fungal mass. Wells containing only isopropanol were used as blanks. Control wells containing C. albicans or macrophages alone were included in each experiment. Each sample was assessed in triplicate. The candidicidal activity was expressed as a candidicidal index (CI) that was calculated as follows: CI = [mean optical density (OD) from N group/OD for each rat].
TNF- production.
Purified macrophages from the Ca and CaS groups were cultured with or
without opsonized, heat-killed C. albicans (E/T ratio, 100:1), and after 3 or 6 h of incubation, supernatants were
sampled to determine TNF-
quantities via an ELISA sandwich protocol. Briefly, 96-well plates were coated with primary anti-TNF-
capture antibody (10 µg/ml; Pharmingen) and blocked with PBS that contained 10% BSA. Supernatants were incubated overnight at 4°C before the diluted biotinylated anti-TNF-
-detecting monoclonal antibody (2 µg/ml) was added. The plates were developed by adding avidin peroxidase and its substrate, and absorbance was measured at 492 nm in
a microplate reader. The amount of TNF-
was extrapolated from the
standard curve, which was generated in 1:2 dilutions. Results are
expressed in micromoles.
Assessment of arginine metabolism. Purified macrophages were incubated without stimulation during 48 h (at 37°C with 5% CO2), and culture medium and cells were sampled for NO measurement and arginase assay, respectively. We determined NO as nitrite in culture supernatants via a microplate-assay method using Griess reagent (2): 100-µl aliquots were mixed with 200 µl of Griess reagent [1.5% sulfanilamide in 1 N HCl and 0.13% N-(1-naphthyl)ethylenediamide dihydrochloride in H2O]. Absorbance was measured at 540 nm in a microplate reader. Nitrite was measured by using NaNO2 as a standard. Each sample was tested in triplicate, and results are expressed in micromoles (17).
To assess arginase activity, cells were washed with PBS and treated with 0.15 ml of 0.1% Triton X-100 that contained protease inhibitors (13). After 30 min, cell lysate was mixed in a 1:1 ratio with 10 mM MnCl2, and the enzyme was activated by heating for 10 min at 56°C. Arginine hydrolysis was conducted by adding 0.5 M L-arginine at pH 9.7 to the activated lysate. Incubation was performed at 37°C for 60 min, and the reaction was stopped with 400 µl of a solution that contained 96% H2SO4, 85% H3PO4, and H2O (a 1:3:7 ratio, vol/vol/vol). The urea formed was colorimetrically quantified at 540 nm after the addition of 25 µl of 9%Immunoblot analysis. iNOS expression was assessed in macrophages that were purified in 24-well, flat-bottom plates (Corning) and cultured with or without opsonized, heat-killed C. albicans (E/T ratio, 100:1) for 24 h. Whole cell extracts were prepared as described for arginase activity. Equal amounts of protein (30 µg/lane) were fractionated in a 10% SDS-PAGE, and proteins were electrotransferred onto nitrocellulose membranes. The anti-iNOS polyclonal antibody (Santa Cruz) was diluted to 2 µg/ml and incubated overnight. Immunodetection was performed with the enhanced chemiluminiscence detection kit and was followed by exposure for 3-5 min to Amersham Hyperfilm (Uppsala, Sweden).
RNA isolation. Macrophages were purified in 100-mm plastic tissue-culture plates. For stimulation experiments, heat-killed C. albicans diluted in RPMI 1640 with 0.1% gentamicin and 10% NRS was added to macrophages (E/T ratio, 100:1) and incubated (at 37°C with 5% CO2) for 16 h. Plates were washed thoroughly with PBS, and total RNA was extracted by using the acid guanidinium method (12). Macrophages were harvested in a denaturing solution that contained 4 M guanidine thiocyanate. The suspension was mixed sequentially with 2 M sodium acetate (pH 4), phenol, and chloroform-isoamyl alcohol. The mixture was centrifuged, which yielded an upper aqueous phase that contained total RNA. After isopropanol precipitation, the RNA pellet was redissolved in the denaturing solution, reprecipitated with isopropanol, washed with 75% ethanol, and dissolved in RNase-free water.
Northern blot analysis.
Total RNA (12 µg) was subjected to electrophoresis on 1% agarose gel
that contained 0.66 M formaldehyde (8). RNA was
transferred to nylon membranes by capillary transblotting overnight,
baked for 2 h at 80°C, and prehybridized at 42°C in a solution
that contained 50% formamide, 5× Denhardt's solution, 5× sodium
chloride-sodium phosphate-EDTA (SSPE), 1% SDS, and 200 µg/ml
denatured herring testes DNA. The membranes were then hybridized
overnight at 42°C in the same solution, which contained
32P-labeled rat IL-1 cDNA, human IL-1ra, or an 18S
ribosomal RNA (rRNA) probe (20, 25). Blots were washed in
2× sodium chloride-sodium citrate (SSC) that contained 1% SDS at RT,
washed twice in 2× SSC with 1% SDS at 55°C, and exposed to Kodak
X-Omat film at
70°C. The band intensities were determined via
scanning densitometry (Shimadzu Dual-Wavelength Chromato Scanner
CS-930) at 500 nm, and the absorbance values of the IL-1
and IL-1ra
signals in arbitrary densitometric units were normalized to those of
the 18S rRNA in the same lane.
Serum corticosterone assay. To determine corticosterone levels, rats were kept undisturbed the night before the experiment. Animals from all experimental groups were killed by decapitation within <3 min of entering the room. Sampling from all rats was carried out within 30 min. Blood was allowed to clot for 45 min, and serum was obtained after centrifugation. Serum corticosterone concentration was determined as previously described (29), and results are expressed as micrograms of corticosterone per deciliter.
Statistical analysis. Differences between group means were assessed using ANOVA followed by the Student-Newman-Keuls test for multiple comparisons. A P value <0.05 was considered statistically significant.
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RESULTS |
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Increased number of C. albicans and impaired killing activity of
macrophages at infection site after stress exposure.
Rats infected intraperitoneally with C. albicans and exposed
to CVS during 10 days show an increased fungal burden in kidneys and
livers together with a poorly developed inflammatory response in these
organs (38). The effect of CVS on the immune response against the fungus is significantly pronounced after 3 days of treatment (38, 39). First, we determined the number of
viable C. albicans in peritoneal lavage aliquots of infected
(Ca) and infected and stressed (CaS) rats. As shown in Fig.
1A, the number of CFU was
significantly higher in CaS-group rats compared with the Ca group
(P < 0.01), which suggests a reduction of the lytic activity of PCs. Because the aim of our work was to evaluate the immunocompetence of macrophages exposed to C. albicans
infection and stress, we assessed the candidicidal activity of purified macrophages after restimulation in vitro with C. albicans to
define the extent of the impairment of macrophage function (Fig.
1B). The candidicidal index was ~2.5-fold lower in
CaS-group rats compared with only infected animals (P < 0.01). Clearly, the stress exposure conditioned a poor clearance of
C. albicans early after the infection.
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TNF- production.
In response to pathogen constituents, macrophages release a battery of
cytokines and other inflammatory mediators that have important local
and systemic effects (7, 40, 41). In particular, TNF-
is an inducer of a local inflammatory response that helps to contain
infections (24, 30, 33, 41). In this context, we evaluated
the production of TNF-
after 3 days of treatment in purified
macrophages from the Ca and CaS groups with or without in vitro
stimulation (Fig. 2). In 3-h
cultures, the release of TNF-
was significantly diminished in
unstimulated macrophages of the CaS group compared with the Ca group
(P < 0.03), and the same result was obtained after
restimulation with opsonized, heat-killed C. albicans (Ca
vs. CaS group, P < 0.05). A similar trend was observed
after 6 h of incubation (data not shown).
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Downregulation of iNOS-arginase activity in peritoneal macrophages
of infected rats.
In response to infection by several pathogens and inflammatory
cytokines, noticeable changes occur in macrophage arginine metabolism
(1, 28). These include, for instance, increases in NO
synthesis via iNOS and catabolism of arginine to ornithine and urea via
arginase. We evaluated the release of NO and the arginase activity in
unstimulated purified macrophages cultured in vitro for 48 h. As
shown in Fig. 3, A and
B, in infected rats (Ca and CaS groups), both the arginase
activity and the NO synthesis were significantly enhanced compared with
N and S groups (P < 0.01). However, whereas the Ca
group exhibited a 10-fold increase in arginase activity (Fig.
3A), the stress exposure downregulated significantly the
activity of this enzyme in the CaS animals (P < 0.05).
A similar pattern of response was observed for the release of NO (Fig.
3B), which suggests that although the infection with the
fungus induced the two metabolic pathways, the stress products downregulated the activity of both enzymes.
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IL-1 and IL-1ra mRNA expression.
Upon exposure to inflammatory stimuli, transcriptional activation of
genes that encode proinflammatory cytokines occurs. Therefore, the
expression of mRNA for IL-1
and IL-1ra was evaluated in macrophages after 3 days of treatment. As shown in Fig.
4, A and B, in unstimulated macrophages, an approximately twofold expression of IL-1
mRNA was
observed in infected groups (Ca and CaS vs. N group, P < 0.01), whereas the amount of IL-1ra mRNA relative to noninfected
groups showed a clear trend without reaching significant differences (Ca and CaS vs. N group, P = 0.06).
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Corticosterone levels.
In an attempt to correlate impaired killer activity and cytokine
imbalance with the continuous release of products of the hypothalamus-pituitary-adrenal (HPA) axis, we determined corticosterone levels after 3 days of treatment. As shown in Fig.
5, corticosterone levels increased in
stressed groups, but although a clear trend was observed in S-group
rats (P = 0.07), a significant increment was evident
only in rats exposed to stress and infection (P < 0.05). Interestingly, when the S and CaS groups were exposed to an
additional acute stress on day 3, a consistent increment of both ACTH and corticosterone levels was shown (data not shown). On the
other hand, an additive effect of stressors was observed after 10 days
of treatment, and the S and CaS groups exhibited increments in both
ACTH and corticosterone levels (Ref. 38; data not shown).
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DISCUSSION |
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Working with a model of candidiasis and stress, we demonstrated the impairment of the immune response against the fungus after exposure to CVS (38). Phenotypic and functional studies showed that macrophages were more affected by stress exposure than were PMNs (39). In this regard, our model is suitable to assess the biology of the macrophage under an integrated interplay of both neuroendocrine and immune systems as occurs in vivo. The present work aimed to determine how the immunocompetence of this cell is affected after stress exposure at the site of the infection.
Our first interesting observation was that in CaS-group rats, the
number of CFU recovered from the peritoneal lavage fluid was notably
larger than in animals that were only infected. Purified peritoneal
macrophages showed a significantly reduced killer capacity, which
suggests either a direct effect of the CVS treatment on the
candidicidal activity, or a modified ability to release cytokines to
respond to, or both. Phagocytosis of C. albicans by
macrophages must be accompanied by killing; otherwise, macrophages
could promote the dissemination of the pathogen and help the fungus to
avoid the immune attack. Killing of C. albicans can be
upregulated in macrophages by cytokines such as interferon (INF)- or
TNF-
(47), chemokines, or neuroendocrine products
(18, 19, 44, 51). Early during C. albicans
infection, proinflammatory cytokines such as TNF-
, IL-6, IL-1
,
and IL-18 participate both in the control of the fungus and in the
induction of protective Th-1 immunity (40, 41). TNF-
is
one of the major secretory products of macrophages (47)
with an important role in host defense against disseminated candidiasis
(6, 37, 49). TNF-
production is dose dependent, and
blood levels are directly related to the C. albicans-inoculum in both neutropenic and nonneutropenic animals (46, 48). As expected in infected rats, the infection
triggered TNF-
production. However, in infected and stressed
animals, which had a higher fungal burden at the site of the infection,
the spontaneous and stimulated release was diminished. Differences
observed between Ca- and CaS-group animals correlated with impaired
autocrine and paracrine effects of this cytokine: candidicidal activity
or cell recruitment, respectively (39).
L-Arginine can be metabolized by alternative pathways that involve the enzymes iNOS and arginase (11, 26). The competition between arginase and iNOS is more pronounced when substrate availability is compromised as occurs during sepsis and in other inflammatory sites (1). Under determined experimental conditions, the induction of one enzyme is accompanied by the suppression of another, which indicates two competitive states in murine macrophages (28). Here we demonstrated that early in the infection, during the innate immune response against C. albicans, both metabolic pathways were activated. In rats only infected (Ca group), iNOS expression and NO production were significantly increased. However, in macrophages from CaS-group animals, NO production decreased after stress exposure (38) as did the enzyme that limits the NO production. In contrast to the well-documented role of iNOS in C. albicans infection (47), no data exist about the alternative L-arginine pathway. This is the first report that demonstrates the induction of arginase activity during the course of this mycosis. After fungal infection, both enzymes exhibited increased activity. However, in macrophages from CaS-group animals that were exposed simultaneously to fungus, inflammatory cytokines, and increased glucocorticoid levels, the arginase function was clearly downmodulated. In agreement with this, induction of arginase activity by lipopolysaccharide (LPS) is largely abolished by dexamethasone with levels only twofold above control values (27).
A dual-activation paradigm has been proposed for macrophages: although
classically activated macrophages secrete proinflammatory cytokines
such as IL-1 and TNF-, alternatively activated macrophages preferentially express anti-inflammatory cytokines such as IL-1ra and
IL-10 (28). The dichotomy also involves
L-arginine metabolism and the expression of receptors
implicated in attachment of microorganisms and phagocytosis: IgG
receptor expression is associated with classic activation; and mannose
receptor (MR),
-glucan receptor, and scavenger receptor type I are
associated with the alternative activation (28, 45).
-Glucans, chitin, and mannose polymers (10) of the
C. albicans wall could interact with
-glucan receptor and
the MRs of macrophages, thereby activating the alternative pathway and
upregulating the expression of associated products. Also, the binding
of yeast to the MRs results in the elevation of mRNA of IL-1, IL-6, and
granulocyte-monocyte colony-stimulating factor (50). In
our model, the singular interaction of this opportunistic pathogen with
macrophages triggers these two metabolic pathways, and stress products
downmodulated both. The relevance of these findings deserves further study.
IL-1, IL1-
, and IL-1ra belong to the IL-1 system and share the
IL-1 receptors; by competitively binding to the receptor, IL-1ra
neutralizes the biological actions of IL-1
and IL1-
without showing any IL-1-like activity. The main source of cytokines in this
system are macrophages, and the balance between IL-1ra and IL-1 can
vary on the course of some infections (9). In candidiasis, the role of IL-1 is quite controversial (40, 41, 47) and little is known about the ratio of IL-1ra to IL-1. In our model, the
infection upregulated the IL-1
and IL-1ra transcripts in macrophages
from both Ca- and CaS-group animals with no differences associated to
stress exposure. Interestingly, after restimulation with the
fungus, lower levels of IL-1
were observed in Ca- and CaS-group
animals, whereas only CaS-group macrophages showed reduced levels of
IL-1ra transcripts. The expression of IL-1 and IL-1ra genes may be
differently regulated (3, 32). Arzt et al.
(4) reported that glucocorticoids have an inhibitory
effect on both genes and demonstrated that the ability of
glucocorticoids to suppress the synthesis of IL-1ra after stimulation
with LPS is endotoxin-dose dependent: the final effect depends on the
balance between the inflammatory stimulus and the dose of
glucocorticoids employed (4). Steroids inhibit the
synthesis of IL-1
by blocking the transcription or diminishing the
stability of the mRNA (19). The in vivo effects of other
HPA-axis hormones such as corticotropin-releasing hormone and ACTH on
IL-1 system expression are less known.
The neutralization of IL-1 biological action by IL-1ra has an important
physiological role during a sharp inflammatory process (9). The balance between endogenous IL-1 and IL-1ra in
vivo is an important determinant of the host response to infection. In
a murine myocarditis model, the increased levels of IL-1 correlated with myocardial fibrosis, whereas the increased serum levels of IL-1ra
improved survival rates and decreased myocardial inflammation and
fibrosis (23). The clinical severity of certain
inflammatory illnesses is usually associated with a decrease in the
relationship of IL-1ra to IL-1 (9). After a new contact
with the fungus, CaS-group macrophages showed a diminished
IL-1ra-to-IL-1 relationship; this suggests a compromised plasticity of
the cell to a new challenge with the pathogen, which could have
physiological relevance.
Stress can affect various facets of the immune response (5, 15). On the other hand, proinflammatory cytokines produced by activated immune cells are able to activate the HPA axis at each level: the hypothalamus, the pituitary, and the adrenal gland (16, 44). Several studies have supported a close link between the HPA axis and the adrenomedullary response during stress and have shown that this link is bidirectional in that it receives input from the nervous and immune systems (16). Serum corticosterone levels were significantly increased only in animals exposed to two associated stimuli: infection and stress. It is conceivable to speculate that both ACTH and non-ACTH-mediated input on the adrenal cortex contributed to the higher corticosterone release that was achieved in this experimental group.
Our work provides clear evidence of the in vivo relevance of macrophages during the initial control of C. albicans infection. Our findings also contribute to the understanding of how the immunocompetence of the host can be conditioned by neuroendocrine factors, thereby increasing the susceptibility for this opportunistic fungal infection.
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ACKNOWLEDGEMENTS |
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We express sincere thanks to Dr. Clelia M. Riera for discussion and critical reading of the manuscript, to Paula Icely for technical assistance, and to Luis Navarro for help.
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FOOTNOTES |
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This work was supported by grants from Consejo Nacional de Investigaciones Científicas y Tecnológicas (CONICET), Agencia Nacional de Promocion Científica y Tecnológica (05-05153), Agencia Córdoba Ciencia, and Secretaría de Ciencia y Técnica de la Universidad Nacional de Córdoba. C. Sotomayor, A. M. Cabanillas, and S. Correa are career members from CONICET.
Address for reprint requests and other correspondence: S. Correa, Inmunología, Departamento de Bioquímica Clínica, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Ciudad Universitaria, Pabellón Argentina 5000, Córdoba, Argentina (E-mail: scorrea{at}bioclin.fcq.unc.edu.ar).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
First published August 28, 2002;10.1152/ajpcell.00160.2002
Received 11 April 2002; accepted in final form 23 August 2002.
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