| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
* Oral Infection and Immunity Branch, National Institute of Dental and Craniofacial Research, and
Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892;
Department of Pathology, George Washington University, Washington, DC 20037, and
Department of Microbiology, Colorado State University, Fort Collins, CO 80523
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
, and
MIP-1
with donor-dependent changes in monocyte chemotactic
protein-1. Pyrrolidine dithiocarbamate, an antioxidant, inhibited the
activation of NF-
B and significantly diminished the MAC-induced
chemotaxis, concurrently lowering the levels of monocyte chemotactic
protein-1 and MIP-1. These data demonstrate that MAC induces
macrophage production of multiple chemotactic factors via NF-B to
promote monocyte migration to sites of MAC infection. In vivo,
opportunistic infection may act as a recruitment mechanism in which
newly arrived monocytes serve as naive hosts for both MAC and HIV-1,
thus perpetuating both infections. |
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
An increase in viral burden is often seen in sera from patients with
M. avium coinfection, which correlates with increased levels
of circulating TNF- (21). The hosts’ natural defense
of producing inflammatory cytokines such as TNF- to rid the body of
infection instead becomes the machinery for increased viral and
bacterial infection. In LN from patients coinfected with MAC,
macrophages emerge as hosts, in addition to CD4+
T cells for HIV-1 (22). Augmentation through the NF-B
pathway in both the production of inflammatory cytokines and the
expression of CCR5 (the macrophage-tropic coreceptor for HIV-1) by
MAC-infected or M. avium Ag (MAg)-induced macrophages
provides a mechanism for increased HIV-1 replication observed in this
population (23, 24, 25). Another potential device used for the
amplification of viral burden during coinfection is the pronounced
increase in numbers of macrophages present within these tissues that
may magnify the population of virus-producing cells. Moreover,
localized release of recruitment factors may alter the trafficking of
cells bearing virus in addition to naive populations, further
exacerbating viral replication. Understanding the underlying mechanisms
whereby OI and/or inflammatory lesions (26) promote the
recruitment of naive and/or infected host cells may provide strategies
for disengaging this cyclic infectious process.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Lymphoid tissue biopsies (LN) were obtained with consent from
patients with AIDS-defining OI, HIV-1-seropositive subjects without
evidence of OI, and HIV-1-seronegative donors. Tissues were fixed in
10% neutral buffered Formalin, paraffin embedded, and sectioned. In
situ hybridization using 35S-labeled sense and
antisense probes for HIV-1 RNA was used to assess the
HIV-1+ cells in the LN, as described (22, 25). For identification of macrophages, deparaffinized sections
were blocked with 1.5% normal serum and subsequently incubated with
Abs against CD68+ (DAKO, Carpinteria, CA).
Sections were washed, incubated with secondary Abs, followed by
peroxidase staining (ABC elite; Vector Laboratories, Burlingame, CA).
Enzymatic visualization was achieved using 3-amino-9-ethylcarbazole
(Vector Laboratories) and counterstaining with methyl green. The
expression of macrophage-inflammatory protein (MIP)-1, monocyte
chemotactic protein (MCP)-1, RANTES, and MIP-1 (mAb; R&D Systems,
Minneapolis, MN) in these tissues was detected with enhanced
immunohistochemical staining using tyramide signal amplification (NEN
Life Science Products, Boston, MA), followed by colorimetric detection
of streptavidin peroxidase with Vector Red (Vector Laboratories), and
counterstained with methyl green. Similarly, detection of HIV-1 was
assessed using an Ab against p24 (DAKO) and Ag retrieval methods with
tyramide signal amplification enhancement, and then visualized with
diaminobenzidine and hematoxylin. Quantification of positive cells in
LN sections was performed using MetaMorph Imaging Software (Universal
Imaging, West Chester, PA). For each LN, two independent images
captured at x20 were used to determine the average number of positive
cells. Cells within two 900 x 350-µm regions per image were
counted for a total of four fields for each section. The final cell
numbers were defined per 75.9 x 105
µm2 (one rectangle).
M. avium complex
MAC, a virulent smooth transparent morphotype strain 2-151, was grown, as previously described (25, 27), and viable organisms were added to adherent monocyte-derived macrophages (MDM) at a ratio of 5:1 or 2.5:1. To acquire MAg, 108 CFU/ml MAC underwent freeze-thaw lysis. After three cycles, the lysate was disrupted by three 10-s pulses with a sonicator probe (Heat Systems, Farmingdale, NY). The protein concentration was determined by bicinchoninic acid assay (28). M. avium lipoarabinomannan (LAM) was generated as previously described (29). In this study, LAM isolated from M. avium, strain 2-151, smooth transparent morphotype was used.
Monocytes and MDM cultures
Human peripheral blood cells, obtained by leukapheresis of healthy volunteers (Department of Transfusion Medicine, National Institutes of Health), were density sedimented, and the monocytes were purified from the mononuclear cell population by elutriation (30, 31). Freshly elutriated monocytes were resuspended in DMEM containing 2 mM L-glutamine and 50 µg/ml gentamicin (BioWhittaker, Walkersville, MD) and used as nonadherent monocytes in the chemotaxis assays or plated into 24-well plates (Corning, Costar, Corning, NY) at a concentration of 1 x 106 cells/well or into 100 x 20-mm petri dishes (Falcon; BD Biosciences, Franklin Lakes, NJ) at 50 x 106/dish. The plated monocytes were allowed to adhere for 3 h at 37°C with 5% CO2 before FCS (Life Technologies, Grand Island, NY) was added for a final concentration of 10%. The monocytes were differentiated into MDM by culturing for 7 days at 37°C with 5% CO2.
Treatment of MDM
After seven days, MDM culture supernatants were removed and one
of the following added: fresh medium, MAC at a ratio of 5:1 or 2.5:1,
1–25 µg/ml MAg, 0.1–10 µg/ml LAM, or 1 x
103 tissue culture infectious
dose50 BaL (HIV-1; Advanced Biotechnologies,
Columbia, MD). MDM cultures were incubated for 1 h at 37°C.
After the incubation, wells were washed three times with PBS, and 1 ml
fresh DMEM containing gentamicin, L-glutamine, and 10% FCS
was added (complete medium). In some experiments, MDM were treated for
2 h with cycloheximide (10 µg/ml; Sigma-Aldrich, St. Louis, MO)
and washed in PBS before MAg exposure. Pyrrolidine dithiocarbamate
(PDTC; Sigma-Aldrich), an antioxidant that has been shown to inhibit
NF-B pathways, was added from 0.6 to 60 µM to the cultures 30 min
before MAg introduction (32, 33). Mitogen-activated
protein kinase (MAPK) p38 inhibitor SB203508 (Calbiochem, San Diego,
CA) was added at 5 µM to MDM cultures 30 min before MAg. In
additional experiments, MDM were first treated with MAg for 1 h
and washed, and fresh medium was added so that only newly released
chemoattractants would be present at the time of coculture.
Neutralizing Abs against the following were then added to the MDM
cultures in the lower chambers for 30 min before performing the
chemotaxis assay, thus allowing time for Abs to bind to any released
chemoattractant before the addition of naive monocytes to the upper
chambers: RANTES (1 µg/ml; BioSource International, Camarillo, CA),
MCP-1, MIP-1, MIP-1 (all 1 µg/ml; R&D Systems), IL-8 (10
µg/ml; R&D Systems), and mouse IgG1 isotype control (1 or 10 µg/ml;
Vector Laboratories). Abs against MCP-1, MIP-1, and MIP-1 were
added in combination in some experiments. All experimental conditions
were done in duplicate. Viability of cells after treatment was assessed
using trypan blue dye.
Chemotaxis assay
Sterile 3-µm polycarbonate transwell inserts (Corning, Costar) were placed into the 24-well plates that contained treated 7-day adherent MDM. Monocytes (1 x 106) that had been fluorescently labeled using Red Fluorescent Cell Linker kit (Sigma-Aldrich) were then added into the top of the transwells and cocultured for 1 h at 37°C. The transwells were removed, and migrated cells were viewed and quantified using an inverted fluorescence microscope (Olympus, Melville, NY). Following photomicrography, the supernatants were collected for further analysis. For some experiments, the monocytes were pretreated for 1 h with 10 ng/ml pertussis toxin (Biomol Research Laboratories, Plymouth Meeting, PA) or MAC, or 30 min with PDTC (60 µM) or SB203508 (5 µM) before fluorescent labeling. All experimental conditions were performed in duplicate.
To determine chemotaxis due to the stimulants themselves, 0.4 ml medium containing MAC (5:1), and MAg (25 µg/ml) was added to the bottom of the transwell system. MCP-1 (NCI Biological Resources Branch, Frederick, MD) or FMLP (Sigma-Aldrich; 2 nM) was used as the positive control, and medium alone as a negative control. Monocytes were resuspended at 3 x 106 in chemotaxis buffer (1x HBSS with calcium and magnesium and 0.5% BSA) and placed in the upper transwell before the 90-min incubation at 37°C. The transwells were removed, and 40 µl 10x chemotaxis fixative (PBS containing 100 mM EDTA plus 10% formaldehyde) was added to the lower chamber; the cells were collected and counted using a Corixa (Seattle, WA) counter.
EMSA
MDM cultured for 7 days in 100 x 20-mm petri
dishes (50 x 106/plate) were treated with
PDTC, MAg, and PDTC with MAg, SB203508, SB with MAg, or medium, as
described earlier. Nuclear extracts were prepared and the EMSA run
using a radiolabeled NF-B consensus oligonucleotide probe (Promega,
Madison, WI), as described previously (25). The binding
reactions were run on nondenaturing 6% polyacrylamide gels (Novex, San
Diego, CA) in 0.25x Tris borate EDTA buffer, dried, and analyzed with
a PhosphorImager using IMAGEQUANT software (Molecular Dynamics,
Sunnyvale, CA).
RNase protection assay (RPA)
Total cellular RNA was extracted using the RNeasy minikit (Qiagen, Chatsworth, CA) from 6 x 106 cells/well of control or treated MDM. Three micrograms of total RNA were used with the hCK-5 template of the BD PharMingen Riboquant MultiProbe RPA system (San Diego, CA) and developed using phosphor imaging. Band densities were normalized to GAPDH housekeeping gene using IMAGEQUANT software (Molecular Dynamics).
Cytokine/chemokine ELISA
The supernatants from the 7-day MDM cultures after 1-h MAC
treatment were analyzed for IL-8, MIP-1, MCP-1, RANTES, TGF-,
MCP-3 (BioSource International), and MIP-1 (R&D Systems) production
by ELISA. Additionally, supernatants were analyzed for MIP-1, MCP-1,
and MIP-1 after 15, 30, 60, or 120 min of MAC, LAM, or MAg
exposure.
Statistical analysis
Nonparametric statistical analysis was determined by the two-tailed, Mann-Whitney t test at 95% confidence intervals.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
By immunohistochemical analysis, dramatically increased numbers of
CD68+ macrophages were evident within LN from
patients coinfected with MAC and HIV-1 as compared with those HIV-1
infected with no OI or with neither HIV-1 nor OI (Fig. 1
, A–D). Consistent with this
finding was the increased presence of HIV-1-infected macrophages in the
MAC tissues and around the high endothelial venules within these
coinfected LN as compared with the LN from the
HIV-1+ donor with no OI (Fig. 1, E–H)
(22, 25), suggesting an active process of adhesion and
recruitment. Because at the time of OI infection there is diminished T
cell function, we focused on the role of macrophages in enlisting new
hosts to carry out viral replication. To ascertain whether M.
avium induces migration of circulating monocytes into the LN,
chemotaxis studies were performed (Fig. 2A). In the coculture system,
macrophages that were treated with MAC for 1 h before the
chemotaxis assay released a soluble factor within the subsequent 60 min
of the assay that induced nonadherent monocytes to migrate an average
of 4-fold over control MDM (Fig. 2B). The presence of
migrating monocytes in the control wells in this dynamic system is
consistent with the fact that macrophages may constitutively express
low levels of some chemoattractants (Fig. 2A) (34, 35). The MAC-induced response did not significantly increase in
the subsequent 4 h from that observed after MDM were stimulated
for 2 h; however, there was a significant difference with MAC
infection after 8 h (data not shown), presumably due to continued
MAC infection and replication (T. Greenwell-Wild, unpublished
observations), and/or release of de novo synthesized recruitment
factors. Because of the rapid response to MAC, we focused on this
initial generation of chemotactic activity.
|
|
B), suggesting that bacterial components could initiate the
rapid MDM response. Based on the dose-response curves, 1 µg/ml lysate
was sufficient to induce the secretion of chemotactic factors (Fig. 2C), but higher doses (25 µg/ml) provided consistent
stimulation in all donors. Purified LAM, a cell wall lipoglycan of MAC
with known immunomodulatory effects (reviewed in Ref. 29),
was assessed for its ability to induce the MDM production of
chemoattractants. Monocytic migration into the LAM-treated MDM wells
was on average 2-fold greater than control MDM (Fig. 2B),
with induction of activity at 0.1 µg/ml and optimal at 10 µg/ml
(Fig. 2C). Exposure of macrophages to a macrophage-tropic
strain of HIV-1, BaL, did not induce chemotaxis above control levels
within this 60-min time interval (Fig. 2B). The question of
whether MAC or MAg was directly chemotactic or induced a chemotactic
factor(s) was addressed by making dilutions of intact bacteria or
bacterial Ags in chemotaxis buffer and adding these dilutions in the
absence of MDM into the bottom transwell, and monocyte migration was
measured. Neither induced chemotaxis independently (data not shown),
indicating that MAC initiates signaling in MDM to release a
chemoattractant factor(s). Because MAg was as effective as viable
M. avium in inducing the MDM to produce chemotactic
activity, subsequent experiments used MAg for defining the recruitment
factors.
To further define the mechanism whereby M. avium triggers
the release of chemoattractants, cycloheximide, an inhibitor of protein
translation, was added before MAg in the MDM cultures. This inhibitor
decreased the chemotactic activity, implying that the M.
avium-stimulated chemoattractant(s) released from MDM is at least
in part synthesized de novo (Fig. 3A). Because MAg activates
NF-B pathways (25), an antioxidant that has been shown
to inhibit this pathway, PDTC (32), was incubated with the
MDM before MAg activation. Although PDTC did not completely eliminate
the response, it caused a significant dose-dependent reduction (Fig. 3A, 60 µM shown) in the MAg-mediated production of
chemotactic molecules, which corresponded with the reduced MAg
activation of NF-B (Fig. 3B, 60 µm shown). The
possibility that activation of p38 MAPK signal transduction induced
chemotactic factors that may be independent of NF-B binding was also
assessed (36, 37). Addition of SB203508, an inhibitor of
p38 MAPK, to the cultures did not significantly decrease MAg-induced
chemotaxis nor influence NF-B (Fig. 3, A and
B). These data provide evidence that in macrophages,
activation of the NF-B pathway and subsequent cellular events are
involved in the regulation of rapid MAg-induced MDM chemoattractant
production.
|
To define the molecules involved in recruitment, the migrating
monocytes were pretreated with pertussis toxin, an inhibitor of G
protein-mediated pathways. The pertussis toxin-treated monocytes failed
to migrate in response to the MAg-induced chemoattractant, suggesting
that the receptor mediating this response is pertussis toxin sensitive
(Fig. 3C), and thus, pointing to a chemokine receptor
(35). Surprisingly, treatment of monocytes with PDTC
blocked both the MAg-induced, as well as the background chemotaxis,
indicating that the NF-B pathway may be involved in monocyte
movement in this system. The p38 MAPK inhibitor also reduced monocytic
chemotaxis, albeit to a lesser extent, thus implicating several
pathways in the monocytic response to the soluble chemoattractant(s)
generated by MAg activation of macrophages.
The production of several members of the chemokine family, including
MCP-1, RANTES, and MIP-1, is known to occur rapidly, and their
production is inhibited by cycloheximide (reviewed in Refs. 34, 35 , and 38). Additionally, the receptors for these
chemokines are sensitive to pertussis toxin (34, 35, 38),
which inhibited the monocytic response to the MDM-derived factor.
Because the data suggested a role for G protein-mediated signaling,
MAg-stimulated MDM mRNA was analyzed by RPA using probes for known
chemokines. RNA analysis indicated that MAg up-regulated mRNA for MIP-1
, MIP-1, and IL-8 (Fig. 4A) from 5- to 76-fold
compared with the control during the first 3 h. Similar results
were also obtained with LAM treatment of MDM (Fig. 4A,
lower panel). Increases in RANTES and MCP-1 mRNA were donor
dependent (Fig. 4), being highly up-regulated in some donors, but not
induced in others.
|
, MIP-1, and IL-8 proteins were measured in the MDM treated
with MAg supernatants (Fig. 4B). The increase for MIP-1
averaged 18.3 ± 2.3-fold in eight independent experiments,
MIP-1 from 11.9 ± 4.5-fold (n = 10), and IL-8
from 7.1 ± 3.77-fold (n = 8) over controls.
Similar to the RPA data, MCP-1 levels varied with donor, dependent in
part on the constitutive levels of expression. Neither RANTES protein
nor mRNA levels were typically elevated and did not significantly
increase above the control in this 2-h assay. MCP-3 and TGF- were
not detected in control supernatants or after MAg stimulation. To
determine the kinetics of the rapid expression of MCP-1, MIP-1, or
MIP-1 after MAC, LAM, or MAg treatment, supernatants were analyzed
after 15-, 30-, 60-, and 120-min intervals. MCP-1 levels were present
in the control supernatants at 15 min, and these levels did not change
significantly after treatment at these early time points (data not
shown). Detectable levels of MIP-1 and MIP-1 (Fig. 4C)
were observed only after 60 min of exposure with MAC, LAM, or MAg.
These results suggest that MCP-1 may be preformed and ready to ignite
early migratory events. However, MIP-1 and MIP-1 appear to be
made rapidly in response to MAC or its components. Taken together,
results from the neutralizing Ab and the kinetic studies suggest that
these multiple chemokines may work interdependently to recruit
monocytes.
NF-B-dependent chemokine production
Because several chemokines and cytokines, including MCP-1, are
regulated through NF-B (34, 35), and inhibition of the
NF-B pathway decreased MAg-stimulated MDM recruitment of monocytes,
we evaluated the supernatants from the PDTC-treated MDM for chemokine
protein expression (Fig. 5). The decrease
seen in the levels of MIP-1 and MCP-1, but not MIP-1 after PDTC
treatment corresponded to the levels of functional chemotactic activity
(Fig. 3). Moreover, when neutralizing Abs against MIP-1, MIP-1,
IL-8, MCP-1, RANTES, and an irrelevant isotype control were
preincubated with the MAg-treated MDM before the chemotaxis assay was
performed, anti-MIP-1 and anti-MCP-1, but not anti-RANTES,
anti-IL-8, anti-MIP-1, or the isotype control, significantly
inhibited the MAg-induced chemotaxis (Fig. 5 and data not shown),
consistent with the role of multiple factors induced by MAg. The
combination of Abs against MCP-1 and MIP-1 on average reduced the
monocyte chemotaxis to background levels (Fig. 5). This would suggest
that at least, in the in vitro system, MCP-1 and MIP-1 are the prime
chemokines involved in monocyte recruitment.
|
Tissue sections of LN from MAC-coinfected, HIV-1-infected, and
HIV-1-negative individuals were examined by immunohistochemical
analysis and quantified for the expression of MCP-1, MIP-1, and
MIP-1 (Fig. 6). In MAC-coinfected LN,
MCP-1 and MIP-1 expression was significantly increased as compared
with the other LN sections (Fig. 6). Expression of MIP-1 in the
MAC-coinfected tissues appeared to occur in multiple cell types,
including multinucleated giant cells. Interestingly, MIP-1
expression was higher in the HIV-1-infected than in the MAC LN, whereas
the expression of MIP-1 and MCP-1 was not.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
B. Functional data suggest that both MCP-1 and MIP-1 are
essential in the rapid recruitment of monocytes. MIP-1 has not
previously been shown to be a predominant mediator of
mycobacterial-induced recruitment. In nonimmunocompromised individuals,
this fast response may enable a rapid clearance of invading bacteria in
sites of inflammation by recruiting reinforcement phagocytes into the
area to help control the infection, whereas in immunosuppressed or
HIV-1-infected individuals, it may foster a permissive milieu for viral
infection.
Because both HIV-1 and MAC infection appear to be associated with the
recruitment of mononuclear cells into LN and other tissues, it was
important to delineate the role of MAC in this process. Although
MAC infection or LAM treatment of macrophages induces production of
multiple cytokines (21, 25, 29, 39, 40, 41, 42, 43, 44), investigations
into chemokines and recruitment are limited (45).
Knowledge concerning cell migration and chemokine production has been
primarily derived from work with the more virulent Mycobacterium
tuberculosis (46, 47, 48, 49, 50). However, due to the innate
differences in the infection processes and virulence of
Mycobacterium spp. as well as differences in receptor use
(51), cytokine and chemokine induction may not be
comparable (52, 53). Previous reports indicate that
M. tuberculosis can induce neutrophil, eosinophil, monocyte,
and lymphocyte migration (46, 48, 49, 52), as well as
chemokine up-regulation in infected monocytes/macrophages (47, 49, 50). Mycobacterial infections have been shown to activate
NF-B via Toll-like receptors, resulting in the enhanced expression
of TNF-, CCR5, and IL-8 (25, 54, 55, 56, 57). In this
investigation, we report that MAC or its cellular constituents quickly
activate NF-B and the induction of several chemokines. We observed
significant increases in the production of multiple chemokines,
including MIP-1, MIP-1, IL-8, and MCP-1. At least two chemokines
investigated, MIP-1 and MCP-1, appear to coordinately promote the
recruitment of blood monocytes both in vitro and in vivo, consistent
with the striking accumulation of mononuclear phagocytes in the
MAC-infected tissues.
These data are compatible with signaling via Toll-like receptor 2, a
recently identified mycobacterial receptor (51, 58), which
can lead to NF-B signaling (56). We demonstrate that
the activation of NF-B pathway by MAC is required for the induction
of recruitment factors, including MCP-1 and MIP-1, and plays a role
in the responding monocytes. The p38 MAPK, however, appears not
essential for induction of chemotactic factors, but may be involved in
the signaling pathways induced in the responding monocytes. It is
conceivable that the activation or cross talk of multiple signaling
pathways, including those involving p38 MAPK, occurs due to the
complexity of chemokines and other mediators in initiating this
cellular migration.
Previous reports suggest that HIV-1 induces both T cells and monocytes
to migrate in response to viral infection or Ags, including Nef, gp120,
and Tat (59, 60, 61, 62, 63, 64). In our system, HIV-1 binding and
early entry events do not trigger MDM to produce detectable chemotactic
factors within 60 min. However, from in vitro kinetic assays of the
HIV-1-induced chemotactic response, we found that as HIV-1 begins to
replicate (as evidenced by p24 levels), chemotaxis to HIV-1-infected
MDM occurs (64, 65). Kinetic analyses also revealed that
MAC replication and reinfection of macrophages sustained the production
of elevated chemotactic factors, including an increase in MCP-1
production (H. Hale-Donze, unpublished data), consistent with in vivo
observations (Fig. 6). These findings would indicate that dual
infection, in vitro and in vivo, maintains the production of a
chemoattractant gradient over the course of infection.
That neutralizing individual chemokines significantly affected the MAC-induced chemotaxis, even in the presence of other chemokines that can themselves elicit monocytic migration, is consistent with studies in the MCP-1-deficient mouse. Even in the presence of other inducible chemokines, monocyte recruitment to sites of infection was impaired in these mice (66). Despite the modest detection of MAC-induced MCP-1 protein expression above the constitutive expression, Abs against this chemokine markedly reduced the MAC-elicited chemotaxis. Potentially, complex formation or structural alterations of MCP-1 may render it undetectable by ELISA. Posttranslational modifications in MCP-1, which occur naturally, can influence its chemotactic potency (67), because glycosylated MCP-1 was shown to be less chemotactic than nonglycosylated forms, whereas NH2-terminally truncated forms lost bioactivity (68). Less severe modifications to the NH2-terminal domains cannot only change the chemopotency, but also alter the effector cell population (69). Taken together, posttranslational regulation of MCP-1 may be as critical as increased production for inflammatory responses.
Our findings implicate an essential role for multiple factors in a
systematic pathway to set up chemotactic gradients for the recruitment
of monocytes into sites of inflammation. Some of these factors may act
directly as chemokines; others may act indirectly to cause the release
of additional chemokines, recruitment of other cell types, or, in the
case of mycobacterial infections, the induction of granulomas. For
instance, induction of IL-8 and MIP-1 in Mycobacterium
infections has been postulated to control the recruitment of T cells
and neutrophils involved in granuloma formation (49, 70, 71, 72). MIP-1 and IL-8 levels were also reportedly elevated
in adherent blood monocyte cultures after infection with M.
tuberculosis; however, neutralizing Abs against these chemokines
did not significantly reduce the migration of monocytes
(49). In addition, elevated MIP-1 levels were not
detected in bronchoalveolar lavage fluid from patients with active
pulmonary tuberculosis (49), nor did we find increased
expression in our MAC LN. These findings indicate that MIP-1 may be
less critical in recruitment of monocytes and/or function in other
processes of Mycobacterium infection. In this regard, IL-8
along with MCP-1 were recently found to induce firm adhesion of
monocytes to vascular epithelium, indicating that IL-8 may indeed be
important for monocytic extravasation from the circulation into sites
of inflammation (73).
The coculture chemotaxis system has allowed the initial dissection of
the complex mechanism involved in recruitment of monocytes by infected
macrophages. Multiple factors, including MIP-1, MIP-1, MCP-1,
RANTES, IL-8, and TNF-, as well as other yet undefined components,
are likely to interact in blood monocyte recruitment. These results
have clearly demonstrated the need to further define the interactions
of multiple factors that initiate monocyte chemotaxis to understand
potential interventions that would prevent the spread of viral or
bacterial infections.
M. avium infection has been an OI that plagued patients with HIV-1 before the advent of HAART (74). Clinical data suggest that T cell function together with phagocytic cells are needed to control MAC and other OI infections (8, 9, 10, 11, 12, 15). Unfortunately, HAART, which often partially restores T cell function in infected individuals, is not taken or accessible to all HIV-1 patients throughout the world; therefore, the threat from this pathogen still persists for these and other immunocompromised individuals (75). In the later stages of HIV-1 infection, when T cell function is diminished, the composition of immunoregulatory cells is dramatically changed and OIs including MAC become a serious health risk (74). Macrophages are the primary target for infection and replication of M. avium and become an additional reservoir for HIV-1 (22, 23, 25). Understanding the impact of the bacteria on this cell population is fundamental to developing strategies of intervention.
|
Acknowledgments |
|---|
|
Footnotes |
|---|
2 Current address: Department of Tuberculosis Immunology, Statens Serum Institute, Copenhagen, Denmark 2300S.
3 Address correspondence and reprint requests to Dr. Sharon M. Wahl, Oral Infection and Immunity, National Institute of Dental and Craniofacial Research, 30 Convent Drive, MSC 4352, Building 30/332, Bethesda, MD 20892-4352. E-mail address: smwahl{at}dir.nidcr.nih.gov
4 Abbreviations used in this paper: MAC, Mycobacterium avium complex; HAART, highly active antiretroviral therapy; LAM, M. avium lipoarabinomannan; LN, lymph node; MAg, M. avium Ag; MAPK, mitogen-activated protein kinase; MCP, monocyte chemotactic protein; MDM, monocyte-derived macrophage; MIP, macrophage-inflammatory protein; OI, opportunistic infection; PDTC, pyrrolidine dithiocarbamate; RPA, RNase protection assay.
Received for publication November 20, 2000. Accepted for publication July 24, 2002.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
B activation in intact cells. J. Exp. Med. 175:1181.B/Rel in induction of nitric oxide synthase. J. Biol. Chem. 269:4705.. A. R. Mire-Sluis, and R. Thorpe, eds. Cytokines 467. Academic Press, New York.
B- dependent gene expression: the role of TATA-binding protein (TBP). J. Biol. Chem. 274:30858. and IL-6. J. Immunol. 147:3942.[Abstract]
1 by purified protein derivative of Mycobacterium tuberculosis. Infect. Immun. 63:224.[Abstract]
and IL6 from human monocytes infected with Mycobacterium kansasii: a comparison to Mycobacterium avium. Infection 22:326.[Medline]
B via induction of inflammatory cytokines. Cell. Immunol. 191:117.[Medline]
B in different cell types through reactive oxygen intermediates. J. Immunol. 161:4834.B-dependent network. J. Immunol. 163:3936. in Schistosoma mansoni egg-induced granulomatous inflammation. J. Exp. Med. 177:1551. (MIP-1) and MIP-1 chemokines attract distinct populations of lymphocytes. J. Exp. Med. 177:1821.This article has been cited by other articles:
|
|
 |
|
  J. Fischer, J. West, N. Agochukwu, C. Suire, and H. Hale-Donze Induction of Host Chemotactic Response by Encephalitozoon spp. Infect. Immun., April 1, 2007; 75(4): 1619 - 1625. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. M. Wahl, T. Greenwell-Wild, and N. Vazquez HIV accomplices and adversaries in macrophage infection J. Leukoc. Biol., November 1, 2006; 80(5): 973 - 983. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Vazquez, T. Greenwell-Wild, S. Rekka, J. M. Orenstein, and S. M. Wahl Mycobacterium avium-induced SOCS contributes to resistance to IFN-{gamma}-mediated mycobactericidal activity in human macrophages J. Leukoc. Biol., November 1, 2006; 80(5): 1136 - 1144. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M.C. Herzberg, A. Weinberg, and S.M. Wahl (C3) The Oral Epithelial Cell and First Encounters with HIV-1 Advances in Dental Research, April 1, 2006; 19(1): 158 - 166. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. Ma, T. Greenwell-Wild, K. Lei, W. Jin, J. Swisher, N. Hardegen, C. T. Wild, and S. M. Wahl Secretory Leukocyte Protease Inhibitor Binds to Annexin II, a Cofactor for Macrophage HIV-1 Infection J. Exp. Med., November 15, 2004; 200(10): 1337 - 1346. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. M. Wahl, T. Greenwell-Wild, G. Peng, G. Ma, J. M. Orenstein, and N. Vazquez Viral and host cofactors facilitate HIV-1 replication in macrophages J. Leukoc. Biol., November 1, 2003; 74(5): 726 - 735. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. Greenwell-Wild, N. Vazquez, D. Sim, M. Schito, D. Chatterjee, J. M. Orenstein, and S. M. Wahl Mycobacterium avium Infection and Modulation of Human Macrophage Gene Expression J. Immunol., December 1, 2002; 169(11): 6286 - 6297. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
0.001.
