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Department of Internal Medicine, Division of Infectious Diseases, University of Kentucky and Veterans Affairs Medical Center, Lexington, KY 40506
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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and IFN-
mRNA and chemokines
lymphotactin, RANTES, and macrophage inflammatory protein-1
mRNA.
Phagocytic cells from neonatal mice were significantly less efficient
than adult cells at migrating to the draining lymph nodes after
phagocytosing fluorescent beads. There were fewer dendritic cells and
Ia+ myeloid cells in the lungs of P.
carinii-infected neonatal mice compared with adults. These data
indicate that the lung environment of neonatal mice is insufficient for
migration of T cells, due at least in part to inefficient phagocytosis
and migration of APCs to the lymph nodes as well as delayed chemokine
and TNF- mRNA expression. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Normal, healthy neonatal rabbits and mice can harbor subclinical
P. carinii infections until the age of weaning, when an
inflammatory response takes place and clears the organisms (3, 4). Additionally, Vargas and colleagues recently reported a high
incidence of P. carinii infection in autopsy lung specimens
obtained from infants who had died of sudden infant death syndrome,
confirming that human infants also harbor subclinical P.
carinii infections (5). We have reported that there
is a delay in the inflammatory response in the lungs of mice infected
with P. carinii as neonates compared with that of infected
adult mice (3). Adult mice challenged with an intranasal
(i.n.) inoculation of P. carinii develop a T cell response
within 7 days and resolve the infection within 3 wk. In contrast,
neonatal mice challenged with a comparable dose of P.
carinii do not have detectable T cells in the alveolar spaces
until 
3 wk of age and do not resolve the infection until 6 wk
(3). This delayed inflammatory response in neonates could
be due either to inadequate number or function of specific
CD4+ T cells that are required for resolution of
the infection, or, alternatively, the lung environment may not be
conducive to processing and presenting Ag or attracting effector
lymphocytes.
It has been known for some time that resolution of PCP is dependent on the presence of competent CD4+ T cells (6, 7). More recently, it has been determined that B cells are also critical for host defense against primary P. carinii infection (8, 9). The function of neonatal lymphocytes has generally been considered to be immature. However, more recent studies have suggested that there is no intrinsic defect in neonatal T cells, but the dose and mode of Ag challenge can affect the data obtained (10, 11, 12). Furthermore, stimulating neonatal T cells with costimulatory molecules or adult dendritic cells results in responses similar to those of adults (13, 14). These data suggest that the environment surrounding neonatal T cells is critical to their functional capacity. In this regard it has been shown that infant rats do not express adult level staining patterns for MHC class II-positive airway epithelial and alveolar dendritic cells until the age of weaning (15, 16). This is consistent with the timing of the inflammatory response to P. carinii seen in mice infected as neonates, suggesting that the lung environment may not be conducive to mounting an inflammatory response to P. carinii until the age of weaning (3).
We have used an adoptive transfer strategy to distinguish between the
effects of the lung environment and the function of
lymphocytes in the host response to P. carinii in neonatal
mice. Neonatal or adult SCID mice were given P. carinii
infections and 5 days later were reconstituted with splenocytes from
adult BALB/c donors. We then examined the P. carinii lung
burden, inflammatory response, and lung cytokine and chemokine mRNA
expression and found that the delay in inflammatory response and
clearance of P. carinii in mice is due to deficiencies in
the lung environment and can be attributed to, at least in part,
delayed TNF- and chemokine production. This may be due to
ineffective phagocytosis of P. carinii in the lungs and Ag
presentation in the draining lymph nodes.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Pregnant BALB/c or BALB/c SCID mice in midgestation and adult female BALB/c or BALB/c SCID mice (8 wk old) were purchased from the National Cancer Institute (Bethesda, MD). Donor 6- to 8-wk-old female BALB/c mice were purchased from the National Cancer Institute or from Harlan Sprague Dawley (Indianapolis, IN). P. carinii were obtained from Dr. Allen Harmsen at Trudeau Institute (Saranac Lake, NY) and maintained in C.B17 SCID mice originally obtained from The Jackson Laboratory (Bar Harbor, ME) and bred in the Lexington Veterans Affairs Medical Center veterinary medical unit.
Inoculation of P. carinii
Lungs were excised from P. carinii-infected mice and pushed through stainless steel mesh in HBSS. Cell debris was removed by centrifugation at 100 x g for 2 min. Aliquots of lung homogenates were spun onto glass slides, fixed in methanol, and stained with Diff-Quik (Dade International, Miami, FL). P. carinii nuclei were enumerated by microscopy as previously described (3). Mice were anesthetized lightly with halothane gas to inhibit the diver’s reflex. The P. carinii inoculum was placed over both nares. The entire volume of 10 µl for neonates and 50 µl for adults was aspirated because mice are obligate nose breathers. Neonatal mice (<72 h old) were routinely inoculated with 2 x 106 P. carinii organisms, and adults (8 wk old) were injected with 107 organisms.
Adoptive transfer of immunocompetent splenocytes
Spleens were obtained from adult (8-wk-old) female BALB/c mice and pushed through mesh screens in HBSS to form single cell suspensions. Erythrocytes were removed using a hypotonic lysing buffer, and cells were washed and resuspended at 107/ml in PBS. Four days after infection with P. carinii, adult and neonatal mice were given i.p. injections of 5 x 107 and 107 cells, respectively.
Isolation of cells from alveolar spaces, lungs, and lymph nodes
Mice were killed by exsanguination under deep halothane anesthesia. The lungs were lavaged with HBSS containing 3 mM EDTA. Right lung lobes were excised, minced, and digested in RPMI supplemented with 3% FCS, 1 mg/ml collagenase A, and 50 U/ml DNase for 1 h at 37°C. Digested lung fragments were pushed through mesh screens, and aliquots were removed for enumeration of P. carinii. Erythrocytes were removed from digests using a hypotonic lysing buffer, cells were washed, and single-cell suspensions were enumerated. Tracheobronchial lymph nodes (TBLN) were pushed through mesh screens in HBSS and enumerated.
Enumeration of P. carinii in the lungs of neonatal and adult mice
Aliquots of lung digests were diluted, and 100-µl aliquots were spun onto glass slides, fixed in methanol, and stained with Diff-Quik (Dade International). P. carinii nuclei were enumerated microscopically as previously described (3). Lung burden is expressed as log10 P. carinii nuclei per right lung lobes. The limit of detection ranged from 2.93–3.23 in pups and was 3.23 in adults.
Flow cytometric analysis of lung and lymph node lymphocytes
Lung lavage, lung digest, and TBLN cells were washed in PBS with 0.1% BSA and 0.02% NaN3 (PBA) and stained with the appropriate concentrations of fluorochrome-conjugated Abs specific for murine Iad, CD4, CD8, CD11c, CD44, CD62 ligand, CD19, CD80, and CD86. Abs were purchased from PharMingen (San Diego, CA). Expression of these molecules on the surface of lymphocytes was determined by multiparameter flow cytometry using a FACScalibur cytofluorometer (Becton Dickinson, Mountain View, CA).
For intracellular cytokine staining, lung digest cells were stimulated
for 4 h with 50 ng/ml PMA and 1 µg/ml ionomycin at 37°C under
an atmosphere of 5% CO2. Brefeldin A (10
µg/ml) was added for the final 2 h of incubation to inhibit
secretion of cytokines. Cells were surface stained with anti-CD4
and anti-CD8 as described above, fixed in 5% formalin, and
permeabilized with PBA containing 0.5% saponin. Nonspecific binding
sites were blocked with anti-CD16/CD32 (Fc block, PharMingen). For
detection of cytokines, cells were incubated with
fluorochrome-conjugated Abs specific for IFN- and IL-10.
Quantitation of the number of CD4+ or
CD8+ cells expressing IFN- or IL-10 was
performed using a FACScalibur cytofluorometer.
Migration of fluorescent beads from the lungs to the TBLN
Neonatal and adult mice were given i.n. inoculations of 1.2 x 107 1-µm fluorescent (NYOcarboxylate, Polysciences, Warrington, PA) microspheres/g of body weight. Two days later lungs were lavaged with HBSS/EDTA, and TBLN were excised and pushed through mesh screens in HBSS. Cells associated with fluorescent microspheres were enumerated using a Nikon Eclipse microscope (Nikon, Melville, NY) with a mercury arc lamp. Using this method it was not possible to determine definitively that the particles were phagocytosed and not bound to the surface of the cells. However, experiments performed by Harmsen et al. (17) in which beads were placed into different lung lobes of dogs indicate that it is unlikely that unphagocytosed beads migrated to the draining lymph nodes.
RNase protection assays (RPAs)
Total RNA was prepared from frozen lung tissue using TRIzol
(Life Technologies, Gaithersburg, MD) according to manufacturer’s
instructions. Briefly, 50–100 mg of lung tissue was homogenized in
TRIzol reagents followed by chloroform extraction and ethanol
precipitation. Air-dried RNA pellets were dissolved in
diethylpyrocarbonate-treated water. Multiprobe DNA templates for
cytokines (IFN-, TNF-, IL-2, IL-4, IL-5, IL-6, IL-9, IL-13, and
IL-15) and chemokines (lymphotactin, RANTES, eotaxin, macrophage
inflammatory protein (MIP)-1, MIP-1, MIP-2, monocyte
chemoattractant protein (MCP)-1, and TCA-3) were purchased from
PharMingen along with the RiboQuant in vitro transcription kit and
RiboQuant RPA kit. RPA was performed according to manufacturer’s
instructions. DNA templates were used to synthesize antisense
riboprobes labeled with [-32P]UTP (Amersham
Pharmacia Biotech, Piscataway, NJ) using T7 RNA polymerase. Labeled
probes were hybridized with 10 µg of total RNA and samples were
digested with RNase A and T1 and treated with proteinase K. Labeled,
protected RNA was extracted with phenol/chloroform/isoamyl alcohol and
resolved on 5% denaturing polyacrylamide gels. Dried gels were exposed
to storage PhosphorImager screens, and images were developed using a
Storm 860 imaging system (Molecular Dynamics, Sunnyvale, CA). The
intensity of each specific band was measured using ImageQuant software
(Molecular Dynamics) and was standardized for mRNA levels by dividing
by the intensity signal of the housekeeping gene L32 for each
sample.
Statistical analysis
Differences between experimental groups were determined using Student’s t tests or ANOVA, followed by Student-Newman-Keuls post-hoc test where appropriate. Differences were considered statistically significant when p < 0.05. SigmaStat statistical software (SPSS, Chicago, IL) was used for all analyses.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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To examine the possibility that delayed resolution of P.
carinii in mice challenged as neonates is due to the lung
environment, SCID pups or adults were infected with P.
carinii and reconstituted 4 days later with splenocytes from adult
immunocompetent BALB/c mice. Because neonatal mice (pups) weighed 5- to
10-fold less than adult female mice, pups were routinely inoculated
with 2 x 106 organisms and
107 cells, and adults were given
107 organisms and 5 x
107 cells. The normal kinetics of clearance of
P. carinii from BALB/c mice infected as neonates is delayed
compared with that from adults as shown in Fig. 1
A. SCID pups reconstituted
with adult splenocytes also had delayed clearance of P.
carinii compared with reconstituted SCID adults (Fig. 1B). Adult SCID mice had resolved PCP by about 5 wk
postreconstitution, whereas only two of five SCID pups had cleared
P. carinii from the lungs by almost 7 wk postreconstitution
(Fig. 1B). These data suggest that the delayed clearance of
P. carinii in neonatal mice is due to the inability of the
lung environment to support effector function of lymphocytes.
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We have previously reported that there is a delay in the lung
inflammatory response to P. carinii in immunocompetent mice
infected as neonates compared with that in adults (3).
Consistent with these data, there was a delayed inflammatory response
in P. carinii-infected SCID pups reconstituted with adult
lymphocytes compared with that in reconstituted adult SCID mice.
P. carinii-infected adult SCID mice had a significant number
of CD4+ cells in the draining lymph nodes, lung
digest, and lung lavages when examined on day 13 postreconstitution
using flow cytometry (Fig. 2). In
contrast, CD4+ cells were nearly undetectable in
SCID pups until day 24 postreconstitution. CD4+ T
cells appeared in the TBLN and lung digest of the pups on day 24;
however, very few cells were found in the alveolar spaces until day 37
postreconstitution (Fig. 2C). The appearance of B and
CD8+ T cells in neonatal tissues was also delayed
compared with that in adults (data not shown). Notably, increasing the
inoculation dose of P. carinii in the neonatal mice to equal
that in adult mice did not result in an earlier inflammatory response
or clearance of PCP (data not shown). Because the appearance of
CD4+ cells in the alveolar spaces precedes
clearance of the organisms, these results suggest that failure of
CD4+ cells to appear in the lungs contributes to
the delayed clearance of P. carinii.
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and IL-10 was delayed in the
lungs of reconstituted SCID pups compared with that in adults (data not
shown). This was due to the overall low numbers of cells that migrated
into the lungs of the SCID pups. However, the proportion of
CD4+ cells that produced IFN- (>40%) or
IL-10 (5–10%), as determined by flow cytometry, was similar in pups
and adult mice (data not shown). This suggested that the small numbers
of cells that were able to migrate into the lungs of reconstituted SCID
pups were activated and producing cytokines. Migration of APCs from the lungs to the draining lymph nodes is impaired in neonatal mice
One possible reason why there is a delay in the migration of
CD4+ cells to the lungs of neonatal mice might be
that Ag presentation is inadequate, so only a small number of cells
become activated and express the proper adhesion molecules for
extravasation into the infected tissues. To test whether phagocytic
cells from the lungs of neonatal mice are capable of transporting
particles to the draining lymph nodes, we inoculated mice with
fluorescent 1-µm beads. Neonatal and adult BALB/c mice were given
i.n. inoculations of 1.2 x 107 beads/g of
body weight. Two days later, TBLN were isolated from the mice, and the
fluorescent bead-associated cells were enumerated using a fluorescent
microscope. The percentage of TBLN cells that were bead-associated is
shown in Fig. 3. There was a significant
difference in the proportion of bead-positive cells in the TBLN of
adult mice compared with neonatal mice. For adult mice, an average of
>2% of cells contained beads, whereas four of six neonatal mice had
no detectable bead-positive cells in the TBLN (Fig. 3). In contrast,
adult and neonatal mice had 36.8 ± 6.1 and 40.0 ± 7.8%
bead-positive cells in lung lavages, respectively, suggesting that the
cells in the alveolar spaces were able to phagocytose particles with
the same efficiency. Similar results were found when P.
carinii organisms were labeled with the fluorescent dye
carboxyfluorescein succinimidyl ester before inoculation into pups
(data not shown). These data suggest that due to either low numbers of
APCs or inefficient migration to draining lymph nodes, inadequate APC
activity may explain the delayed clearance of P. carinii in
neonatal mice.
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, there was a significantly
higher percentage of CD11c+ cells in the lungs of
adult mice compared with pups on day 7 postinfection. Similarly, the
percentage of nonlymphoid cells that expressed Ia was significantly
higher in the lungs and TBLN of adult mice compared with neonatal mice
(Fig. 4). These data suggest that migration of dendritic cells and Ag
presentation in the TBLN of neonatal mice may be much reduced compared
with that in adults.
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It has been reported that P. carinii infection
stimulates increased expression of cytokine and chemokine mRNA in the
lungs of adult reconstituted SCID mice (18, 19). Increases
in lung TNF-, IL-1, IL-6, and IFN- mRNA as well as increased lung
lymphotactin, RANTES, MCP-1, MIP-1, and MIP-1 mRNA have been
demonstrated. This increased cytokine and chemokine production precedes
clearance of the organisms and corresponds to lung inflammation
(18, 19).
It is possible that the lack of an inflammatory response in the lungs
of P. carinii-infected neonates is due to the lack of
cytokine and/or chemokine signals for migration to the site of
infection. Chemokine mRNA expression over time in SCID mice infected
with P. carinii as neonates or adults and reconstituted with
splenocytes from immunocompetent adult BALB/c donors was detected by
RPA. Consistent with the delayed inflammatory response in mice infected
as neonates, there was delayed expression of cytokine and chemokine
mRNA in P. carinii-infected SCID pups reconstituted with
adult lymphocytes compared with reconstituted adults (Fig. 5). On day 13 postreconstitution, low,
but detectable, amounts of mRNA for RANTES, MIP-1, MIP-1, and
MCP-1 was found in the pup lungs (Fig. 5). In contrast, lung
lymphotactin, RANTES, MIP-1, MIP-1, and MCP-1 mRNA had peaked in
the lungs of adult mice by day 13 or 24 postreconstitution, whereas
chemokine mRNA expression did not peak until day 37 in the lungs of
reconstituted pups (Figs. 5 and 6). This
delayed expression of chemokine mRNA in mice infected as neonates was
consistent with the delayed inflammatory response and resolution of the
infection.
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is of particular interest
because it has been shown that this proinflammatory cytokine is
critical for resolution of PCP in murine models of infection
(20, 21, 22). As shown in Figs. 5
and 7A, TNF- mRNA expression in
adult mice had already peaked by day 13 postreconstitution. TNF-
mRNA expression decreased almost 2-fold between days 13 and 24
postreconstitution in adult SCID mice and remained level thereafter. In
contrast, TNF- levels in neonatal mice were low on day 13 and
increased significantly through day 37 postreconstitution (Fig. 7A). The delayed changes in TNF- mRNA expression observed
in pups corresponded to the delayed inflammation in the lungs of
P. carinii-infected pups compared with adults as well as the
delayed clearance of the infection. Similarly, delayed expression of
IL-6 mRNA was seen in reconstituted pups compared with adults (data not
shown). Lung IFN- mRNA expression had already peaked in
reconstituted adult mice by day 13 (Fig. 7B). However, mice
infected as neonates had increasing levels of IFN- mRNA expression
through day 48 postreconstitution (Fig. 7B). Expression of
IL-4 and IL-5 mRNA was so low so as to not be confidently quantitated
in both pups and adults after reconstitution (data not shown). The
kinetics of cytokine mRNA expression corresponded with those of
chemokine mRNA expression in the lungs of pups and adults. Furthermore,
the delayed cytokine and chemokine responses in the lungs of pups were
consistent with the delayed inflammatory response and resolution of
P. carinii, suggesting that altered cytokine and/or
chemokine responses may contribute to the defects in the neonatal lung
environment that result in delayed P. carinii clearance.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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There are several possible reasons for the inability of mice infected as neonates to resolve PCP with the same kinetics as adults. These include delayed migration of immature dendritic cells into the lungs, inability of immature dendritic cells to mature from phagocytic cells to mature APCs, inability of mature dendritic cells to migrate to the draining lymph nodes, and inability of T cells to respond to Ag presentation and migrate back to the lungs. Because neonatal mice were reconstituted with competent adult splenocytes, they should have been fully able to respond to appropriate stimuli for activation and migration to the site of inflammation. These data indicate that the lung phagocytic cells did not recognize the infectious agent, they were not sufficiently stimulated to maturity, and/or they did not migrate properly. Additionally, it is possible that lung chemoattractants or adhesion molecules were not sufficiently expressed in neonatal lungs for extravasation of mature T cells to the site of infection.
It has been reported in neonatal rats that there is delayed maturation
of respiratory tract dendritic cells characterized by decreased
expression of Ia, decreased Ag presentation activity in mixed
lymphocyte reactions, and decreased recruitment of dendritic cells into
the airway epithelium in response to heat-killed Moraxella
catarrhalis (16). Consistent with these previous
findings, our data indicated that there was a lower percentage of
dendritic cells in the lungs and TBLN of P. carinii-infected
neonatal mice compared with adult mice. In addition, there was
decreased expression of Ia on nonlymphoid cells of both lungs and TBLN
of neonatal mice compared with adults. Phagocytic cells from neonatal
lungs were significantly less efficient at migrating to draining lymph
nodes than were cells from adult lungs. Two days after i.n. challenge
with either fluorescent beads or fluorescent P. carinii
organisms, there were detectable numbers of fluorescent cells in the
TBLN of the adult mice, but not in the neonatal mice. The decreased
migration was not necessarily due to decreased phagocytosis, because
the percentages of lung lavage cells containing beads were similar in
neonatal and adult mice. It is possible that the signals necessary for
migration were either not present or not at a sufficient concentration
for attracting phagocytic cells to the TBLN. These signals could
include TNF-, up-regulation of chemokine receptors such as CCR7, or
expression of adhesion molecules. We are currently examining the
expression of chemokine receptors and adhesion molecules in the lungs
and immune cells of neonatal mice.
It is not possible to determine unequivocally from the data presented whether dendritic cells in the TBLN of neonatal mice were capable of effective Ag presentation. However, there were 200-fold fewer activated CD4+ T cells in the TBLN of neonates than in adults on day 13 postreconstitution, even though fully competent adult splenocytes had been adoptively transferred into both adults and pups. This may have been due to decreased expression of adhesion molecules in the lymph nodes or decreased chemokine production, such as lymphotactin. Both could be a result of the inability of Ag-loaded dendritic cells or macrophages to reach the draining lymph nodes. Additionally, the reduced numbers of CD11c+ dendritic cells in the lungs, the reduced expression of Ia in nonlymphoid cells of the lungs and TBLN, as well as literature reports from rat models (15, 16) suggest that Ag presentation is depressed in neonatal mice compared with that in adults. The depressed Ag presentation is likely to be at least partially responsible for the delayed host response to P. carinii in mice infected as neonates compared with that in adults.
Immature dendritic cells are dependent on inducible chemokines such as
RANTES, MIP-1, MIP-1, and MCP-1 to migrate from the circulation
to sites of inflammation (23). Once in the lungs, immature
dendritic cells mature from phagocytic cells to APCs. The signals
required for maturation include proinflammatory cytokines such as IL-1
and TNF-, viral or bacterial products, and CD40 ligand
(23). Mature dendritic cells down-regulate chemokine
receptors such as CCR1, -2, and -5 and up-regulate CCR7, causing them
to migrate to draining lymph nodes in response to chemokines such as
EBV-induced molecule 1 ligand chemokine (ELC)/MIP-3 (24, 25). It is possible that any or all of these steps in
mobilization of dendritic cells could be impaired in the lungs of
P. carinii-infected neonates. Specifically, we found that
lung mRNAs from the inducible chemokines RANTES, MIP, and MCP were all
at significantly lower levels in neonates compared with adults. This
may explain the reduced numbers of CD11c+
dendritic cells in the lungs of neonates compared with adults.
Additionally, expression of lymphotactin mRNA, a lymphocyte-specific
chemoattractant, was delayed in neonates compared with adults. This may
account in part for the delayed lymphocyte response to P.
carinii in the neonatal mice. It has been shown in adult mice
infected with Crytococcus neoformans that MCP-1 is critical
for attracting CD4+ cells to the lungs
(26). Because CD4+ cells are
required for host defense against P. carinii (6, 7), the delayed expression of lymphotactin and MCP-1 in neonatal
mice probably contributes significantly to delayed resolution of
PCP.
In contrast to the data presented here, it has been reported that
exposure to hyperoxia significantly increased RANTES, MIP-1, and
MCP-1 mRNA levels in the lungs of 4-day-old C57BL/6 mice
(27). These data suggest that given the proper stimulus,
the lungs of neonatal mice are capable of increasing chemokine mRNA
expression. Consistent with this idea, we have found that neonatal mice
infected with Streptococcus pneumoniae mount a neutrophil
response that is as intense at that of adult mice (28).
The delayed response to P. carinii infection may be due to
low levels of TNF- or IFN- production in neonates. Although
IFN- is not required for clearance of P. carinii
(29), it has been shown to be critical for the expression
of RANTES and MIP-1 in a model of murine lung infection with
Cryptococcus neoformans (30), suggesting that
IFN- may has critical functions in controlling inflammation in the
lungs.
TNF- has been shown to be a stimulus for maturation of dendritic
cells resulting in increased CCR7 expression and migration toward
EBV-induced molecule 1 ligand chemokine (ELC)/MIP-3
(24). In addition, both TNF- and IFN- increase
expression of MHC class II on APCs and stimulate expression of
costimulatory and adhesion molecules. The presence of TNF- has been
shown to be required early in the immune response to P.
carinii for resolution to take place (20).
Furthermore, double IFN- and TNF- receptor knockout mice are
susceptible to PCP (21), indicating that in concert
TNF- and IFN- have important roles in host defense to P.
carinii that probably include controlling the inflammatory
response.
The data suggest a model in which P. carinii infection in
neonatal mice fails to stimulate production of proinflammatory
cytokines such as TNF-, resulting in decreased chemokine production
in the lungs. As a consequence, immature dendritic cells either do not
migrate to the lungs promptly or are not stimulated to maturity once
there. Therefore, mature Ag-loaded dendritic cells do not migrate to
the draining lymph nodes, and so Ag presentation and activation of
CD4+ T cells are delayed. Additionally, delayed
chemokine production may further delay migration of
CD4+ cells to the lungs. The inability of
dendritic cells and T cells to traffic appropriately results in delayed
clearance of the organisms. Although these data do not rule out the
possibility that immaturity of the lymphoid compartment in neonatal
mice also contributes to delayed resolution of PCP, we have found that
adult SCID mice reconstituted with CD4+ cells
from 10-day-old BALB/c mice are capable of resolving PCP as efficiently
as mice reconstituted with equal numbers of CD4+
cells from adult donors (manuscript in preparation). These data clearly
indicate that the neonatal lung environment is inadequate for clearance
of P. carinii, even in the presence of mature T cells.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Beth A. Garvy, Department of Internal Medicine, University of Kentucky Medical Center, Room MN668, 800 Rose Street, Lexington, KY 40536.
3 Abbreviations used in this paper: PCP, Pneumocystis carinii pneumonia; i.n., intranasal; RPA, RNase protection assay; MIP, macrophage inflammatory protein; MCP, monocyte chemoattractant protein; TBLN, tracheobronchial lymph nodes.
Received for publication June 12, 2000. Accepted for publication September 1, 2000.
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cell-deficient mice: diagnosis and therapy. Lab. Anim. Sci. 50:49.
and interferon in host resistance against Pneumocystis carinii infection. Infect. Immun. 60:1279. and type 1 and 2 TNF receptor genes. J. Immunol. 161:360. in the host immune and inflammatory response to P. carinii infection. Infect. Immun. 65:373.[Abstract]
(IFN-)-dependent protection and synthesis of chemoattractants for mononuclear leucocytes caused by IL-12 in the lungs of mice infected with Cryptococcus neoformans. Clin. Exp. Immunol. 117:113.[Medline]This article has been cited by other articles:
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  C. Ptaschinski and R. Rochford Infection of neonates with murine gammaherpesvirus 68 results in enhanced viral persistence in lungs and absence of infectious mononucleosis syndrome J. Gen. Virol., May 1, 2008; 89(5): 1114 - 1121. [Abstract] [Full Text] [PDF]
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 M. Hollifield, E. B. Ghanem, W. J. S. de Villiers, and B. A. Garvy Scavenger Receptor A Dampens Induction of Inflammation in Response to the Fungal Pathogen Pneumocystis carinii Infect. Immun., August 1, 2007; 75(8): 3999 - 4005. [Abstract] [Full Text] [PDF]
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K. M. Empey, M. Hollifield, and B. A. Garvy Exogenous Heat-Killed Escherichia coli Improves Alveolar Macrophage Activity and Reduces Pneumocystis carinii Lung Burden in Infant Mice Infect. Immun., July 1, 2007; 75(7): 3382 - 3393. [Abstract] [Full Text] [PDF]
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 L. Huang, A. Morris, A. H. Limper, J. M. Beck, and on behalf of the ATS Pneumocystis Workshop Partici An Official ATS Workshop Summary: Recent Advances and Future Directions in Pneumocystis Pneumonia (PCP) Proceedings of the ATS, November 1, 2006; 3(8): 655 - 664. [Abstract] [Full Text] [PDF]
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 M. H. Qureshi, K. M. Empey, and B. A. Garvy Modulation of Proinflammatory Responses to Pneumocystis carinii f. sp. muris in Neonatal Mice by Granulocyte-Macrophage Colony-Stimulating Factor and IL-4: Role of APCs J. Immunol., January 1, 2005; 174(1): 441 - 448. [Abstract] [Full Text] [PDF]
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K. M. Empey, M. Hollifield, K. Schuer, F. Gigliotti, and B. A. Garvy Passive Immunization of Neonatal Mice against Pneumocystis carinii f. sp. muris Enhances Control of Infection without Stimulating Inflammation Infect. Immun., November 1, 2004; 72(11): 6211 - 6220. [Abstract] [Full Text] [PDF]
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 R. L. Chelvarajan, S. M. Collins, I. E. Doubinskaia, S. Goes, J. Van Willigen, D. Flanagan, W. J. S. de Villiers, J. S. Bryson, and S. Bondada Defective macrophage function in neonates and its impact on unresponsiveness of neonates to polysaccharide antigens J. Leukoc. Biol., June 1, 2004; 75(6): 982 - 994. [Abstract] [Full Text] [PDF]
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M. H. Qureshi, J. Cook-Mills, D. E. Doherty, and B. A. Garvy TNF-{alpha}-Dependent ICAM-1- and VCAM-1-Mediated Inflammatory Responses Are Delayed in Neonatal Mice Infected with Pneumocystis carinii J. Immunol., November 1, 2003; 171(9): 4700 - 4707. [Abstract] [Full Text] [PDF]
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M. H. Qureshi, A. G. Harmsen, and B. A. Garvy IL-10 Modulates Host Responses and Lung Damage Induced by Pneumocystis carinii Infection J. Immunol., January 15, 2003; 170(2): 1002 - 1009. [Abstract] [Full Text] [PDF]
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M. H. Qureshi and B. A. Garvy Neonatal T Cells in an Adult Lung Environment Are Competent to Resolve Pneumocystis carinii Pneumonia J. Immunol., May 1, 2001; 166(9): 5704 - 5711. [Abstract] [Full Text] [PDF]
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