| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
*
Pulmonary Center, Boston University School of Medicine, and
Infectious Disease Section, Boston Medical Center, Boston, MA 02118
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
B. The central role played by NF-B in
signal transduction pathways activated by cytokines, and in the
regulation of cytokine genes themselves, implicates mammalian Toll
proteins in cellular responses similar to those evoked by cytokines.
Third, the intracellular domains of Toll proteins share significant
sequence similarity with the type I IL-1 receptor, the known mammalian
Toll-like receptors, and the cytosolic adapter protein MyD88
(4, 5, 6).
Recent data have demonstrated that mammalian TLR proteins participate
in intracellular signaling initiated by Gram-negative bacterial LPS.
CD14 has been recognized for many years as the major receptor
responsible for the effects of LPS on macrophages, monocytes, and
neutrophils (reviewed in Ref. 7). Two groups independently
reported that TLR2 could function as a signaling receptor for LPS in
the presence of CD14 (8, 9). These investigators reported
that human HEK293 cells stably transfected with TLR2 could respond to
LPS in the presence of CD14 and LPS-binding protein, as judged by
activation of a reporter gene under the control of the
NF-B-dependent ELAM-1 promoter. Deletion mutants of TLR2 that lack a
region of the intracellular domain that shares sequence homology with
the IL-1 receptor failed to mediate LPS responsiveness in this
assay.
Subsequent to these findings, two other groups identified the gene responsible for the LPS-hyporesponsive phenotype of the C3H/HeJ mouse (10, 11). Macrophages from this mouse are hyporesponsive to LPS, even though they express normal amounts of CD14 on their surface. The gene locus responsible for this defect (Lpsd) mapped to the Tlr4 gene. In the C3H/HeJ mice, a single missense mutation within the Tlr4 coding sequence was identified (P712H). Supporting evidence for the hypothesis that this mutation is responsible for the LPS-hyporesponsive phenotype of the C3H/HeJ mouse comes from the finding that the C57BL/10ScCr LPS-nonresponsive mouse does not express TLR4. These results raised the possibility that, at least in mice, TLR2 is not sufficient to confer LPS responsiveness. Data from our own laboratories have shown that TLR4 is predominantly responsible for LPS signaling in murine and hamster cells, whereas TLR2 mediates cellular activation by a distinct CD14 ligand, the mycobacterial cell wall glycolipid lipoarabinomannan (LAM).4 This conclusion was further supported by our demonstration that LPS hyporesponsive macrophages from C3H/HeJ mice were not hyporesponsive to LAM.4 Most recently, Chow et al. (13) reported that human TLR4 could mediate LPS responsiveness in HEK293 cells. Together, these results suggest that TLR2 and TLR4 mediate CD14-dependent signals in both a ligand- and species-specific manner.
We have extended these earlier studies to test the hypothesis that Mycobacterium tuberculosis, bacteria that do not synthesize LPS, might also be recognized by TLR proteins. It was recently found that cellular activation by Gram-positive bacteria and Mycobacterium avium was mediated by TLR2, but not TLR4 (Ref. 14 and data not shown).5 Here we report that M. tuberculosis activates cells in a TLR-dependent manner, but unlike Gram-positive bacteria and M. avium, these organisms utilize both TLR2 and TLR4 proteins. Unlike Gram-positive bacteria, TLR-dependent cellular activation by M. tuberculosis does not appear to depend on the presence of CD14. Furthermore, the mycobacterial ligands responsible for this activation appear to be distinct from LAM.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
M. tuberculosis strains H37Rv (ATCC 25618), H37Ra (ATCC 25177), and Mycobacterium bovis BCG (ATCC 35734) were purchased from the American Type Culture Collection (ATCC, Manassas, VA). Bacterial cultures were grown in Middlebrook 7H9 medium supplemented with Tween 80 and ADC (Difco, Detroit, MI) at 37°C under biosafety level 3 conditions. Bacterial culture medium was prepared in LPS-free flasks using LPS-free water (Baxter-Travenol, Deerfield, IL). Bacterial cultures were grown to midlogarithmic phase (OD620 nm = 0.4), and CFU per ml were determined by growth on Middlebrook agar plates. The CHO-K1 fibroblast (CCL-61) and RAW264.7 murine macrophage cell lines (TIB-71) were purchased from the ATCC. RAW264.7 cells were maintained in DMEM (BioWhittaker, Walkersville, MD) supplemented with 10% heat-inactivated fetal bovine serum (HyClone Laboratories, Logan, UT), 10 mM HEPES, 2 mM L-glutamine, 100 U/ml penicillin, and 100 µg/ml streptomycin (BioWhittaker). Chinese hamster ovary (CHO) cells were maintained in Ham’s F-12 culture medium (BioWhittaker) supplemented with 10% heat-inactivated FBS (HyClone, Logan, UT), 10 mM HEPES, 2 mM L-glutamine, 100 U/ml penicillin, and 100 µg/ml streptomycin (BioWhittaker). LPS levels in all medium components were <10 pg/ml final concentration as indicated by BioWhittaker or measured by the Limulus amebocyte lysate kit (BioWhittaker). Cells were cultured at 37°C in the presence of 5% CO2 in a humidified incubator. CHO/CD14, 3E10 (CHO/CD14/ELAM-CD25), 3E10/TLR2, 3E10/TLR4, and CHO/TLR2 were previously described4 (14, 15) and cultured as described above. All of these stable cell lines used expressed a similar level of surface CD14, TLR2, and TLR4. As described below, cells that were stably transfected with the TLR2 and TLR4 expression plasmids express FLAG-tagged TLR proteins. After transfection and selection of stable lines, clones were selected that expressed similar levels of TLR proteins, based on mean fluorescence intensity determined with the same anti-FLAG Ab. Furthermore, the TLR2- and TLR4-expressing cell lines were generated from the same CHO/CD14 parental clonal cell line, and thus each line also expresses the same amount of CD14.
LPS (purified from Escherichia coli 055:B5) was purchased from Sigma (St. Louis, MO). Mycobacterial LAM, purified from rapidly growing avirulent mycobacteria (AraLAM), M. bovis BCG, and M. tuberculosis strains H37Rv and H37Ra were all provided by Dr. John Belisle (Colorado State University, Fort Collins, CO) under the provisions of National Institutes of Health Contract NO1 AI25147. Levels of contaminating LPS in the LAM preparations were determined using a quantitative Limulus lysate assay (BioWhittaker) and were <1 pg/ml final concentration in all experiments. A neutralizing anti-LAM IgG3 mAb that recognizes LAM from these different mycobacteria (CS-35) was also provided by Dr. Belisle and was previously described (16). Recombinant human IL-1ß was purchased from Genzyme (Cambridge, MA). FITC- and PE-conjugated anti-human CD25 mAbs were purchased from Becton Dickinson (Bedford, MA).
Electrophoretic mobility shift assays (EMSA)
Nuclear extracts were prepared essentially as described by Schreiber et al. (17). Approximately 1.0 x 107 CHO or RAW264.7 cells were washed and harvested by scraping in Ca2+- and Mg2+-free PBS (BioWhittaker). Cells were pelleted by centrifugation at 800 x g for 10 min at 4°C. Cell pellets were resuspended in 400 µl of a buffer containing 10 mM KCl, 10 mM Tris-HCl (pH 7.8), 5 mM MgCl2, 0.5 mM DTT, 0.3 M sucrose, 10 mM ß-glycerol phosphate, 0.1 mM EGTA, 1 mM PMSF, and 5 µg/ml each of aprotinin, leupeptin, chymostatin, and antipain and incubated on ice for 10 min. Subsequently, 25 µl of 10% Nonidet P-40 (Sigma) were added to each sample before vortexing. The nuclei were centrifuged for 1 min at 5000 x g to pellet the nuclei. Nuclear pellets were resuspended in a nuclear extraction buffer containing 320 mM KCl, 10 mM Tris-HCl (pH 7.8), 5 mM MgCl2, 0.5 mM DTT, 10 mM ß-glycerol phosphate, 0.1 mM EGTA, 25% glycerol, 1 mM PMSF, and 5 µg/ml each of aprotinin, leupeptin, antipain, and chymostatin. Samples were extracted on ice for 15 min followed by centrifugation at 16,000 x g for 10 min at 4°C. Protein concentration was determined using the Bio-Rad assay kit (Bio-Rad, Hercules, CA). All nuclear extracts were stored at -70°C, and multiple freeze-thawing cycles were avoided.
A double-stranded oligonucleotide containing a single copy of the IL-2
receptor 
-chain NF-B (GGGGAATTCC) was used as an EMSA probe. DNA
probes were labeled with [-32P]dNTPs
(DuPont-NEN, Boston, MA) using E. coli DNA polymerase Klenow
fragment (U.S. Biochemicals, Cleveland, OH) as recommended by the
manufacturer. Unincorporated nucleotides were removed with Sephadex
G-25 columns (5 Prime
3 Prime, Boulder, CO). Nuclear extracts
(typically 3 µg) were incubated with radiolabeled probe DNA (0.1 ng,
typically 10,000 cpm) in the presence of 2 µg poly(dI-dC) (Pharmacia,
Piscataway, NJ), 1.0 mM EDTA, 10 mM Tris-HCl (pH 7.9), 25 mM glycerol,
and 0.5 mM DTT in a final volume of 20 µl, as previously described
(18). Binding reactions were then incubated at room
temperature for 30 min. After incubation, a portion of each binding
reaction (typically 6 µl) was loaded onto 7% nondenaturing low ionic
strength polyacrylamide gel. The gels were then dried and visualized by
autoradiography.
Plasmids
The ELAM-luc reporter plasmid was generated by subcloning the promoter of pUMS(ELAM)-Tac (15) into the promoterless pGL3 luciferase reporter plasmid (Promega, Madison, WI). The human TLR2 and TLR4 cDNAs cloned into the pFlag-CMV-1 mammalian expression plasmid were gifts of Drs. Carsten Kirschning and Mike Rothe (Tularik, South San Francisco, CA) and were previously described (9). These gene products are expressed as fusion proteins containing an N-terminal FLAG epitope tag. A second TLR4 expression plasmid that expresses native TLR4 (termed hToll) was a gift of Dr. Charles Janeway (Yale University, New Haven, CT) and was previously described (2). Plasmids were prepared using Qiagen (Valencia, CA) plasmid DNA purification columns, DNA was eluted from the columns using LPS-free buffers, and contaminating LPS levels were found to be <10 pg/ml. Furthermore, all plasmid preparations were unable to activate the LPS-sensitive CHO/CD14 cells, demonstrating that the plasmids were not contaminated with LPS.
Transfection and reporter assays
Transient transfections were performed using SuperFect reagent (Qiagen, Valencia, CA) as per the manufacturer’s instructions. Briefly, cells were plated on six-well dishes 1–2 days before transfection, and transfections were performed when cells plated reached 80% confluence. Plasmid DNA was added to 100 µl of Opti-Mem reduced serum media (Life Technologies, Gaithersburg, MD). All transfections utilized a total of 4 µg of plasmid DNA consisting of 2 µg of reporter plasmid, 1 µg of each expression vector, and the balance made up with empty vector described above. SuperFect, 10 µl, was added to the DNA-medium mixture and incubated for 10 min at ambient temperature. Subsequently, 600 µl of serum-containing medium were added to the reaction mixture and added to the individual wells. Each reaction was prepared individually, and each condition was performed in triplicate. Reactions were incubated with the cells for 2–3 h, whereupon the reaction was removed from the cells and fresh medium containing serum was added. On the following day, individual wells were left untreated or were stimulated with either LPS or LAM as indicated in the figures. Cells were then incubated for an additional 5 h before harvesting. Luciferase assays were performed as described below. All transfection experiments were repeated at least three times with different plasmid preparations, and a single representative experiment is shown. Each single experiment represents triplicate independent transfections and data are expressed as average values ± S.D.
Luciferase activity was measured using the Luciferase Assay System (Promega), according to manufacturer’s instructions. Briefly, cells were washed and scraped on ice in cold PBS, pelleted by centrifugation, and resuspended in 100 µl of reporter lysis buffer. Samples were freeze-thawed once and centrifuged at 14,000 x g for 10 min at 4°C to remove cellular debris. Supernatants were recovered and assayed for total protein using the Bio-Rad protein assay according to manufacturer’s instructions. An equal amount of total protein from each lysate was assayed for luciferase activity as measured by light emissions in a scintillation counter.
In experiments with 3E10 cells, which contain a stably transfected CD25 reporter gene under the control of the ELAM-1 promoter, reporter gene expression was measured by flow cytometry as previously described4 (14, 15). Data were collected using FACScan software (Becton Dickinson, Mountain View, CA) and expressed as the ratio (fold activation) of the percent of CD25+ cells in unstimulated and stimulated cell populations. The 95% confidence limit for nonspecific fluorescence was established with the use of isotype control Abs. Each experiment was repeated at least three times in all cases.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Previous studies demonstrated that both bacterial cell wall
components (LPS, LAM, and peptidoglycan) and heat-killed Gram-positive
bacteria (Staphylococcus aureus and Streptococcus
pneumoniae) could activate cells in a TLR-dependent
manner4 (14, 19). Here we tested whether live
M. tuberculosis bacilli (Mtb) could also activate cells in a
TLR-dependent manner. To measure TLR-dependent activation, we used CHO
fibroblast cell lines that were stably transfected with expression
plasmids encoding human CD14 (CHO/CD14) and human TLR2 proteins
(CHO/CD14/TLR2). These lines were also stably transfected with a
reporter plasmid consisting of a human CD25 cDNA under the control of
an NF-B-dependent ELAM-1 promoter. As we previously published,
activation of these cells in a CD14-dependent manner leads to the rapid
activation of NF-B and the subsequent expression of CD25 on the cell
surface4 (14, 15). Furthermore, CD14-
and TLR-independent stimuli that activate NF-B (e.g., IL-1ß
protein) also activate CD25 expression. Here we incubated viable Mtb
with CHO/CD14 and CHO/CD14/TLR2 cells (10 Mtb/CHO cell) for 16 h
and subsequently measured CD25 expression by flow cytometry. As shown
in Fig. 1
, both the virulent H37Rv and
attenuated H37Ra strains of Mtb did not activate the CHO/CD14 cells,
and overexpression of TLR2 conferred Mtb responsiveness on these cells.
This TLR-dependent activation did not appear to vary with the virulence
of the Mtb strain used. Similar data were obtained with avirulent
M. bovis BCG bacilli (data not shown). CD25 expression was
also measured at 1, 4, 8, 16, and 24 h after stimulation by H37Ra
and H37Rv. At each time point analyzed, we found that both Mtb strains
activated the cells to a similar extent (data not shown). Lastly,
surface expression of either CD14 or TLR proteins did not alter Mtb
binding or uptake by the CHO cells compared with untransfected CHO
cells (data not shown).
|
Our earlier data showed that TLR-dependent activation of cells by
the mycobacterial cell wall glycolipid LAM was substantially enhanced
in the presence of LPS-binding protein4 (LBP (20)). To
determine whether LBP present in serum was required for TLR-dependent
activation of cells by Mtb, we repeated the preceding experiment in the
presence and absence of serum. As shown in Fig. 2, the presence of serum did not
significantly affect responsiveness of cells to Mtb, suggesting that
LBP is not required for TLR-dependent activation by Mtb. Because the
FBS used in these studies was heat inactivated, our data also
demonstrate that neither opsonization nor activated complement was
required for this TLR-dependent activation. This is not surprising
because CHO cells do not express either complement receptors or other
receptors known to mediate the binding of Mtb to macrophages (e.g.,
macrophage mannose receptor). Because LBP, at least in part, transfers
bacterial glycolipids to CD14, our data also suggested that CD14 was
not required for this TLR-mediated activation of cells by Mtb.
|
We recently showed that TLR-dependent activation of cells by
either LAM or Gram-positive bacteria required the presence of CD14 as
either a membrane-bound or soluble protein4
(14). To test the hypothesis that Mtb activate cells in a
TLR-dependent, but CD14-independent manner, we compared the capacity of
Mtb to activate CHO/TLR2 cells (i.e., CHO fibroblasts that express
TLR2, but not CD14) in the absence and presence of serum, which serves
as a source of soluble CD14. Nuclear extracts were prepared from the
CHO/TLR2 cells 1 h after Mtb stimulation, and NF-B levels were
subsequently measured by EMSA. As shown in Fig. 3A, similar levels of NF-B
were induced in a TLR-dependent manner in the presence and absence of
soluble CD14. NF-B levels in CHO/CD14 cells were not altered by
exposure to Mtb. Furthermore, we compared the capacity of Mtb to
activate NF-B in CHO/CD14/TLR2 vs CHO/TLR2 cells and found that the
presence of membrane-bound CD14 did not augment Mtb responsiveness
(Fig. 3B). These data demonstrate that TLR-dependent
activation by Mtb is not mediated by CD14. This contrasts with
Gram-positive bacteria where activation is dependent on both TLR2 and
CD14 (14, 19).
|
Recent studies revealed that both Gram-positive bacteria and
M. avium can activate cells via TLR2, but not TLR4 (Ref. 14
and data not shown).5 We sought to determine
whether Mtb also activated cells via the same selective use of TLR
proteins. Mtb were added to CHO/CD14/TLR cells. As shown in Fig. 4A, we found that Mtb could
activate the cells via both TLR2 and TLR4. Thus, Mtb differed from
Gram-positive bacteria and M. avium in their utilization of
TLR proteins during cellular activation. Virtually identical results
were obtained with either H37Ra or H37Rv Mtb, or when the experiment
was performed in the presence or absence of serum (data not shown).
Furthermore, activation via either TLR2 or TLR4 was not dependent on
the presence of membrane-bound CD14 (data not shown). We subsequently
examined whether Mtb could also activate macrophages via both TLR
proteins. To test this possibility, we transiently cotransfected
RAW264.7 murine macrophages with a luciferase reporter plasmid under
the control of the NF-B-dependent ELAM-1 promoter and the various
TLR expression plasmids. After transfection, the cells were stimulated
with Mtb (10 Mtb/RAW264.7 cell) and harvested 5 h later. Lysates
were prepared from the harvested cells, and luciferase activity was
measured as described in Materials and Methods. As shown in
Fig. 4B, Mtb activated the macrophages in a TLR-dependent
manner. Like the CHO cells, this activation could be mediated by both
TLR2 and TLR4.
|
Our previous studies revealed that LAM purified from rapidly
growing Mycobacteria could activate cells in a TLR-dependent
manner.4 This form of LAM contains highly branched
arabinofuranosyl side chains (termed AraLAM), whereas Mtb and M.
bovis BCG contain LAM that is terminally capped with mannose
residues (ManLAM). These chemical differences are believed to be
responsible for the different biological activities of these
glycolipids (reviewed in Ref. 21). We sought to determine
whether LAM on the surface of the mycobacteria was the ligand
responsible for TLR-dependent activation of the cells. First, we
compared the capacity of LAM isolated from rapidly growing
Mycobacteria (AraLAM), Mtb (ManLAM), and M. bovis
BCG (BCG LAM) to activate cells in a TLR-dependent manner. As shown in
Fig. 5, AraLAM was capable of activating
CHO/CD14 cells that expressed TLR2, but not TLR4. In contrast to
AraLAM, neither Mtb LAM nor BCG LAM were capable of activating any of
the cell lines tested. This finding is consistent with the known
relative inability of these ManLAMs to activate macrophages
(22, 23, 24).
|
To confirm that TLR-dependent cellular activation by Mtb was not
mediated by LAM, we determined whether an anti-LAM Ab could block
activation of cells by Mtb bacilli. Mtb were pretreated with the
neutralizing CS-35 anti-LAM mAb (107 H37Ra
bacilli treated with 500 µg/ml CS-35 for 15 min), and then added to
the CHO/CD14/TLR cells (10 Mtb/CHO cell, 12.5 µg/ml CS-35 final
concentration). As shown in Fig. 6A, the anti-LAM mAb was
incapable of blocking Mtb activation of cells via TLR2 or TLR4. The
ability of CS-35 to block LAM-induced cellular activation was confirmed
by the finding that this mAb could block TLR2-dependent activation by
AraLAM (Fig. 6B). Normal rabbit serum had no effect on
either Mtb- or AraLAM-induced cellular activation (data not shown).
Thus, using two distinct experimental approaches (Figs. 5
and 6), we
have demonstrated that TLR-dependent activation of cells by Mtb was not
mediated by the cell wall glycolipid LAM. Although we have shown that
CD14 does not appear to play a role in TLR-dependent cellular
activation by Mtb, it remains to be determined whether the
mycobacterial ligands activate TLR proteins directly or in association
with other cell surface receptors.
|
We sought to determine whether soluble factors released by
cultured Mtb could activate the cells in a TLR-dependent manner. H37Ra
Mtb were grown to mid-log phase as described in Materials and
Methods, bacilli were removed by centrifugation and filtration,
and the Mtb-free culture medium was used to stimulate the CHO/CD14/TLR
cells. As shown in Fig. 7A, we
found that Mtb-conditioned culture medium activated the cells via TLR2,
but not TLR4. Fresh culture medium did not activate the cells under any
conditions tested (data not shown). We subsequently examined whether
the soluble factors responsible for TLR-dependent activation were
proteins. The Mtb-conditioned culture medium was digested with
proteinase K (50 µg/ml, 55°C, 1 h) and boiled for 5 min to
inactivate the protease. This protein-free Mtb-conditioned culture
medium was then evaluated for its capacity to activate cells in a
TLR2-dependent manner. As shown in Fig. 7B, the protein-free
and heat-stable preparation was still capable of inducing
TLR2-dependent cellular activation. Thus, it appears that a
nonproteinaceous heat-stable factor is the TLR2 ligand. Based on the
chemical nature of other known TLR ligands (e.g LPS, LAM,
peptidoglycan), it is likely that this factor is a polysaccharide or a
glycolipid. It is also possible that this preparation contains several
distinct ligands for TLR2.
|
).
Because viable Mtb bacilli could induce both TLR2- and TLR4-dependent
signaling, but the soluble mycobacterial factor only activated cells
via TLR2, we sought to determine whether it was a cell-associated
factor that induced TLR4-dependent signaling. As shown in Fig. 7C, we found that heat-killed Mtb that were washed free of
culture medium activated cells via TLR2. In contrast to the viable Mtb,
the heat-killed bacilli could not activate cells via TLR4. These
differences imply that the ligand for TLR4-dependent activation is a
heat-labile cell-associated mycobacterial factor. Cellular activation
via TLR4 was not caused by LPS contamination because all materials used
to perform this experiment were essentially LPS free and cellular
activation by the bacilli was not CD14 dependent. Thus, soluble
mycobacterial factors selectively activate cells in a TLR2-dependent
manner, whereas heat-labile cell-associated mycobacterial factors
selectively activate cells in a TLR4-dependent manner. These findings
demonstrate that whole Mtb and soluble mycobacterial factors differ in
their utilization of TLR proteins leading to cellular activation.
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
M. tuberculosis, the etiological agent of tuberculosis, is a major worldwide public health threat. Mtb bacilli are uniquely adapted to survive and grow within macrophages (reviewed in Ref. 25). Paradoxically, macrophages are the primary effector cells of the innate immune response. On binding microbial pathogens, these cells are activated to release a variety of cytokines, nitric oxide, and reactive oxygen intermediates. Cytokines released by activated macrophages can augment both innate and cell-mediated immune responses. The precise mechanisms by which Mtb activate macrophages remain unclear. Our data revealed two unexpected features of these Mtb-induced responses. The first feature is that CD14 does not appear to be required for cellular activation by Mtb. This contrasts with both Gram-positive and Gram-negative bacteria that predominantly activate cells in a CD14-dependent manner (14, 24, 26). Although CD14 has been reported to function as an adhesion receptor for Mtb bacilli in some cell types (27), blocking Abs against the complement receptors CR3, CR4, the macrophage mannose receptor, and the class A scavenger receptor can together prevent almost all binding of Mtb to macrophages (28, 29, 30). It is possible that complement receptors, like CD14, can serve as coreceptors for TLR proteins. Support for this possibility comes from the finding that the CR3 ligand taxol, a plant-derived antitumor agent, cannot activate macrophages from the TLR4-deficient C3H/HeJ mouse whereas taxol can activate macrophages from the normal C3H/FeJ mouse (31). The second novel feature of our findings is that Mtb differs in the utilization of TLR proteins to initiate intracellular signaling compared with Gram-positive bacteria (14).5 Whether these differences in TLR signaling induced by Mtb and other bacteria result in different qualitative or quantitative antimicrobial responses remains to be determined.
Like Gram-positive bacteria, the cell wall of Mtb does not contain LPS. Nevertheless, both types of bacteria can activate cells in a TLR-dependent manner via chemically distinct bacterial products that serve as ligands to mediate this activation. Previous studies have shown that LAM, LPS, and peptidoglycan all activate TLR-dependent signaling in association with CD14 (14, 19).4 In contrast, we have shown that TLR proteins can be activated by CD14-independent ligands. We found both soluble and cell-associated mycobacterial factors can activate TLR-dependent signaling in a CD14-independent manner. A heat-labile cell-associated mycobacterial factor activated cells in a TLR4-dependent manner, whereas a soluble heat-stable mycobacterial factor activated cells in a TLR2-dependent manner. This soluble factor was also resistant to protease treatment, suggesting that it is a polysaccharide or a glycolipid. In addition, we found that a heat-stable cell-associated factor could activate cells in a TLR2-dependent manner (data not shown), although it remains to be determined whether this factor is identical with the soluble heat-stable factor. Studies are currently under way to identify these factors. Interestingly, mycobacterial envelopes contain peptidoglycan, a known CD14 ligand that can activate cells in a TLR2-dependent manner (14, 19). Our finding that Mtb activates cells in a CD14-independent manner suggests that peptidoglycan is not the heat-stable factor we found that activated cells via TLR2. The extensive waxy cell wall of Mtb may mask the underlying peptidoglycan, although this possibility remains to be tested.
Pathogenic mycobacteria, such as Mtb, are part of a family of slow-growing mycobacteria that contain mannose-capped LAM (i.e., ManLAM) in their cell walls. In contrast, rapidly growing mycobacteria are nonpathogenic in immunocompetent hosts and possess arabinofuranosyl-capped LAM (i.e., AraLAM). Part of the survival strategy of Mtb may depend on bacilli gaining entry into the host macrophage without evoking a strong antimicrobial response. Our data demonstrate that Mtb can induce TLR-dependent cellular activation, but the extent of this activation may differ qualitatively from TLR signaling induced by other types of bacteria. CD14 ligands present in the cell walls of Gram-positive and Gram-negative bacilli that activate TLR signaling may induce potent antimicrobial responses that are not induced by Mtb. Our studies have only assessed signaling via TLR2 and TLR4, but how Mtb and other types of bacteria might differentially utilize other TLR proteins remain to be determined.
The Drosophila Toll and 18-wheeler proteins are receptors that mediate antifungal and antibacterial immune responses, respectively (3, 32). These findings support the possibility that mammalian TLR proteins also participate in innate immunity. Mammalian cells possess at least 10 distinct TLR genes, and it is possible that these proteins can bind a wide variety of bacterial products (Patent Cooperation Treaty Publication number WO9850547A3). Given the dimeric nature of the Drosophila Toll receptor, it is also possible that mammalian TLR proteins might heterodimerize, thereby further extending the variety of ligands that might be capable of inducing TLR signaling. Furthermore, mammalian ligands for TLR proteins may also exist. In Drosophila, the natural ligand for Toll is the Spatzle protein (33). Spatzle, a primitive member of the NGF family of cysteine-knot proteins (34), is secreted as an inactive precursor protein that is cleaved into a biologically active form by the serine protease Easter (12). Similar paradigms are well known in mammalian innate immune responses, and natural mammalian ligands for TLR proteins may yet be discovered. Our data do not exclude the possibility that Mtb, and mycobacterial factors do not bind directly to TLR proteins, but instead stimulate the production of a mammalian Spatzle-like factor that binds directly to the TLR proteins.
|
Footnotes |
|---|
2 Address correspondence and reprint requests to Dr. Matthew J. Fenton, Pulmonary Center, Room R-220, Boston University School of Medicine, 80 E. Concord Street, Boston, MA 02118. E-mail address:
3 Abbreviations used in this paper: TLR, Toll-like receptor; LAM, lipoarabinomannan; Mtb, Mycobacterium tuberculosis; CHO, Chinese hamster ovary; LBP, LPS-binding protein; BCG, bacillus Calmette-Guérin; EMSA, electrophoretic mobility shift assays.
4 T. K. Means, E. Lien, A. Yoshimura, S. Wang, D. T. Golenbock, and M. J. Fenton. 1999. The CD14 ligands lipoarabinomannan and lipopolysaccharide differ in their requirement for Toll-like receptors. Submitted for publication.
5 E. Lien, T. J. Sellati, A. Yoshimura, J. D. Carroll, R. R. Ingalls, J. D. Radolf, and D. T. Golenbock. 1999. Toll-like receptor 2 mediates pattern recognition by Borrelia burgdorferi, Treponema pallidum, and Mycobacterium avium. Submitted for publication.
Received for publication April 23, 1999. Accepted for publication July 19, 1999.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
, signal transduction. J. Immunol. 161:3001.. J. Immunol. 150:1886.[Abstract]
This article has been cited by other articles:
|
|
 |
|
  A. Tanne, B. Ma, F. Boudou, L. Tailleux, H. Botella, E. Badell, F. Levillain, M. E. Taylor, K. Drickamer, J. Nigou, et al. A murine DC-SIGN homologue contributes to early host defense against Mycobacterium tuberculosis J. Exp. Med., September 28, 2009; 206(10): 2205 - 2220. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Bourgarit, G. Carcelain, A. Samri, C. Parizot, M. Lafaurie, S. Abgrall, V. Delcey, E. Vicaut, D. Sereni, B. Autran, et al. Tuberculosis-Associated Immune Restoration Syndrome in HIV-1-Infected Patients Involves Tuberculin-Specific CD4 Th1 Cells and KIR-Negative {gamma}{delta} T Cells J. Immunol., September 15, 2009; 183(6): 3915 - 3923. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. A. S. Andersen, I. Rosenkrands, A. W. Olsen, P. Nordly, D. Christensen, R. Lang, C. Kirschning, J. M. Gomes, V. Bhowruth, D. E. Minnikin, et al. Novel Generation Mycobacterial Adjuvant Based on Liposome-Encapsulated Monomycoloyl Glycerol from Mycobacterium bovis Bacillus Calmette-Guerin J. Immunol., August 15, 2009; 183(4): 2294 - 2302. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. M. P. Chan, B. K. W. Cheung, J. C. B. Li, L. L. Y. Chan, and A. S. Y. Lau A role for glycogen synthase kinase-3 in antagonizing mycobacterial immune evasion by negatively regulating IL-10 induction J. Leukoc. Biol., August 1, 2009; 86(2): 283 - 291. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. E. Almeida, A. R. Silva, C. M. Maya-Monteiro, D. Torocsik, H. D'Avila, B. Dezso, K. G. Magalhaes, H. C. Castro-Faria-Neto, L. Nagy, and P. T. Bozza Mycobacterium bovis Bacillus Calmette-Guerin Infection Induces TLR2-Dependent Peroxisome Proliferator-Activated Receptor {gamma} Expression and Activation: Functions in Inflammation, Lipid Metabolism, and Pathogenesis J. Immunol., July 15, 2009; 183(2): 1337 - 1345. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. Sweet, W. Zhang, H. Torres-Fewell, A. Serianni, W. Boggess, and J. Schorey Mycobacterium avium Glycopeptidolipids Require Specific Acetylation and Methylation Patterns for Signaling through Toll-like Receptor 2 J. Biol. Chem., November 28, 2008; 283(48): 33221 - 33231. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. S. Dharmadhikari and E. A. Nardell What Animal Models Teach Humans about Tuberculosis Am. J. Respir. Cell Mol. Biol., November 1, 2008; 39(5): 503 - 508. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. T. Spencer, G. Abate, A. Blazevic, and D. F. Hoft Only a Subset of Phosphoantigen-Responsive {gamma}9{delta}2 T Cells Mediate Protective Tuberculosis Immunity J. Immunol., October 1, 2008; 181(7): 4471 - 4484. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. A. B. Joosten, B. Heinhuis, S. Abdollahi-Roodsaz, G. Ferwerda, L. LeBourhis, D. J. Philpott, M.-A. Nahori, C. Popa, S. A. Morre, J. W. M. van der Meer, et al. Differential function of the NACHT-LRR (NLR) members Nod1 and Nod2 in arthritis PNAS, July 1, 2008; 105(26): 9017 - 9022. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Gerosa, B. Baldani-Guerra, L. A. Lyakh, G. Batoni, S. Esin, R. T. Winkler-Pickett, M. R. Consolaro, M. De Marchi, D. Giachino, A. Robbiano, et al. Differential regulation of interleukin 12 and interleukin 23 production in human dendritic cells J. Exp. Med., June 9, 2008; 205(6): 1447 - 1461. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Nigou, T. Vasselon, A. Ray, P. Constant, M. Gilleron, G. S. Besra, I. Sutcliffe, G. Tiraby, and G. Puzo Mannan Chain Length Controls Lipoglycans Signaling via and Binding to TLR2 J. Immunol., May 15, 2008; 180(10): 6696 - 6702. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Schaefer, N. Reiling, C. Fessler, J. Stephani, I. Taniuchi, F. Hatam, A. O. Yildirim, H. Fehrenbach, K. Walter, J. Ruland, et al. Decreased Pathology and Prolonged Survival of Human DC-SIGN Transgenic Mice during Mycobacterial Infection J. Immunol., May 15, 2008; 180(10): 6836 - 6845. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. Lampropoulou, K. Hoehlig, T. Roch, P. Neves, E. C. Gomez, C. H. Sweenie, Y. Hao, A. A. Freitas, U. Steinhoff, S. M. Anderton, et al. TLR-Activated B Cells Suppress T Cell-Mediated Autoimmunity J. Immunol., April 1, 2008; 180(7): 4763 - 4773. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. D. Chakravarty, G. Zhu, M. C. Tsai, V. P. Mohan, S. Marino, D. E. Kirschner, L. Huang, J. Flynn, and J. Chan Tumor Necrosis Factor Blockade in Chronic Murine Tuberculosis Enhances Granulomatous Inflammation and Disorganizes Granulomas in the Lungs Infect. Immun., March 1, 2008; 76(3): 916 - 926. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Ferwerda, M. B. B. McCall, S. Alonso, E. J. Giamarellos-Bourboulis, M. Mouktaroudi, N. Izagirre, D. Syafruddin, G. Kibiki, T. Cristea, A. Hijmans, et al. From the Cover: TLR4 polymorphisms, infectious diseases, and evolutionary pressure during migration of modern humans PNAS, October 16, 2007; 104(42): 16645 - 16650. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Ferwerda, B. J. Kullberg, D. J. de Jong, S. E. Girardin, D. M. L. Langenberg, R. van Crevel, T. H. M. Ottenhoff, J. W. M. Van der Meer, and M. G. Netea Mycobacterium paratuberculosis is recognized by Toll-like receptors and NOD2 J. Leukoc. Biol., October 1, 2007; 82(4): 1011 - 1018. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Young Goo, Y. S. Han, W. H. Kim, K.-H. Lee, and S.-J. Park Vibrio vulnificus IlpA-induced Cytokine Production Is Mediated by Toll-like Receptor 2 J. Biol. Chem., September 21, 2007; 282(38): 27647 - 27658. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Doz, S. Rose, J. Nigou, M. Gilleron, G. Puzo, F. Erard, B. Ryffel, and V. F. J. Quesniaux Acylation Determines the Toll-like receptor (TLR)-dependent Positive Versus TLR2-, Mannose Receptor-, and SIGNR1-independent Negative Regulation of Pro-inflammatory Cytokines by Mycobacterial Lipomannan J. Biol. Chem., September 7, 2007; 282(36): 26014 - 26025. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Z. Kincaid, A. J. Wolf, L. Desvignes, S. Mahapatra, D. C. Crick, P. J. Brennan, M. S. Pavelka Jr., and J. D. Ernst Codominance of TLR2-Dependent and TLR2-Independent Modulation of MHC Class II in Mycobacterium tuberculosis Infection In Vivo J. Immunol., September 1, 2007; 179(5): 3187 - 3195. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. M. Fremond, D. Togbe, E. Doz, S. Rose, V. Vasseur, I. Maillet, M. Jacobs, B. Ryffel, and V. F. J. Quesniaux IL-1 Receptor-Mediated Signal Is an Essential Component of MyD88-Dependent Innate Response to Mycobacterium tuberculosis Infection J. Immunol., July 15, 2007; 179(2): 1178 - 1189. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. A. Stanley, J. E. Johndrow, P. Manzanillo, and J. S. Cox The Type I IFN Response to Infection with Mycobacterium tuberculosis Requires ESX-1-Mediated Secretion and Contributes to Pathogenesis J. Immunol., March 1, 2007; 178(5): 3143 - 3152. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Winkler, D. Ghadimi, J. Schrezenmeir, and J.-P. Kraehenbuhl Molecular and Cellular Basis of Microflora-Host Interactions J. Nutr., March 1, 2007; 137(3): 756S - 772S. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. D. Chakravarty, J. Xu, B. Lu, C. Gerard, J. Flynn, and J. Chan The Chemokine Receptor CXCR3 Attenuates the Control of Chronic Mycobacterium tuberculosis Infection in BALB/c Mice J. Immunol., February 1, 2007; 178(3): 1723 - 1735. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. D. Pecora, A. J. Gehring, D. H. Canaday, W. H. Boom, and C. V. Harding Mycobacterium tuberculosis LprA Is a Lipoprotein Agonist of TLR2 That Regulates Innate Immunity and APC Function J. Immunol., July 1, 2006; 177(1): 422 - 429. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S.-B. Jung, C.-S. Yang, J.-S. Lee, A-R. Shin, S.-S. Jung, J. W. Son, C. V. Harding, H.-J. Kim, J.-K. Park, T.-H. Paik, et al. The Mycobacterial 38-Kilodalton Glycolipoprotein Antigen Activates the Mitogen-Activated Protein Kinase Pathway and Release of Proinflammatory Cytokines through Toll-Like Receptors 2 and 4 in Human Monocytes. Infect. Immun., May 1, 2006; 74(5): 2686 - 2696. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Demissie, L. Wassie, M. Abebe, A. Aseffa, G. Rook, A. Zumla, P. Andersen, T. M. Doherty, and the VACSEL Study Group The 6-kilodalton early secreted antigenic target-responsive, asymptomatic contacts of tuberculosis patients express elevated levels of interleukin-4 and reduced levels of gamma interferon. Infect. Immun., May 1, 2006; 74(5): 2817 - 2822. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I. Fricke, D. Mitchell, J. Mittelstadt, N. Lehan, H. Heine, T. Goldmann, A. Bohle, and S. Brandau Mycobacteria Induce IFN-{gamma} Production in Human Dendritic Cells via Triggering of TLR2 J. Immunol., May 1, 2006; 176(9): 5173 - 5182. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Fan, Y. Li, Y. Vodovotz, T. R. Billiar, and M. A. Wilson Hemorrhagic shock-activated neutrophils augment TLR4 signaling-induced TLR2 upregulation in alveolar macrophages: role in hemorrhage-primed lung inflammation Am J Physiol Lung Cell Mol Physiol, April 1, 2006; 290(4): L738 - L746. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J.-S. Chang, J. F. Huggett, K. Dheda, L. U. Kim, A. Zumla, and G. A. W. Rook Myobacterium tuberculosis Induces Selective Up-Regulation of TLRs in the Mononuclear Leukocytes of Patients with Active Pulmonary Tuberculosis. J. Immunol., March 1, 2006; 176(5): 3010 - 3018. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Banaiee, E. Z. Kincaid, U. Buchwald, W. R. Jacobs Jr., and J. D. Ernst Potent Inhibition of Macrophage Responses to IFN-{gamma} by Live Virulent Mycobacterium tuberculosis Is Independent of Mature Mycobacterial Lipoproteins but Dependent on TLR2. J. Immunol., March 1, 2006; 176(5): 3019 - 3027. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. D'Avila, R. C. N. Melo, G. G. Parreira, E. Werneck-Barroso, H. C. Castro-Faria-Neto, and P. T. Bozza Mycobacterium bovis Bacillus Calmette-Guerin Induces TLR2-Mediated Formation of Lipid Bodies: Intracellular Domains for Eicosanoid Synthesis In Vivo. J. Immunol., March 1, 2006; 176(5): 3087 - 3097. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Viriyakosol, M. A. Matthias, M. A. Swancutt, T. N. Kirkland, and J. M. Vinetz Toll-Like Receptor 4 Protects against Lethal Leptospira interrogans Serovar Icterohaemorrhagiae Infection and Contributes to In Vivo Control of Leptospiral Burden Infect. Immun., February 1, 2006; 74(2): 887 - 895. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. A. W. Verreck, T. de Boer, D. M. L. Langenberg, L. van der Zanden, and T. H. M. Ottenhoff Phenotypic and functional profiling of human proinflammatory type-1 and anti-inflammatory type-2 macrophages in response to microbial antigens and IFN-{gamma}- and CD40L-mediated costimulation J. Leukoc. Biol., February 1, 2006; 79(2): 285 - 293. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. K. W. Cheung, D. C. W. Lee, J. C. B. Li, Y.-L. Lau, and A. S. Y. Lau A Role for Double-Stranded RNA-Activated Protein Kinase PKR in Mycobacterium-Induced Cytokine Expression J. Immunol., December 1, 2005; 175(11): 7218 - 7225. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. O. Omueti, J. M. Beyer, C. M. Johnson, E. A. Lyle, and R. I. Tapping Domain Exchange between Human Toll-like Receptors 1 and 6 Reveals a Region Required for Lipopeptide Discrimination J. Biol. Chem., November 4, 2005; 280(44): 36616 - 36625. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. M. Doherty and P. Andersen Vaccines for Tuberculosis: Novel Concepts and Recent Progress Clin. Microbiol. Rev., October 1, 2005; 18(4): 687 - 702. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I. Olsen, P. Boysen, S. Kulberg, J. C. Hope, G. Jungersen, and A. K. Storset Bovine NK Cells Can Produce Gamma Interferon in Response to the Secreted Mycobacterial Proteins ESAT-6 and MPP14 but Not in Response to MPB70 Infect. Immun., September 1, 2005; 73(9): 5628 - 5635. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Shi, A. Blumenthal, C. M. Hickey, S. Gandotra, D. Levy, and S. Ehrt Expression of Many Immunologically Important Genes in Mycobacterium tuberculosis-Infected Macrophages Is Independent of Both TLR2 and TLR4 but Dependent on IFN-{alpha}{beta} Receptor and STAT1 J. Immunol., September 1, 2005; 175(5): 3318 - 3328. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Takamura, K. Matsuo, Y. Takebe, and Y. Yasutomi Ag85B of Mycobacteria Elicits Effective CTL Responses through Activation of Robust Th1 Immunity as a Novel Adjuvant in DNA Vaccine J. Immunol., August 15, 2005; 175(4): 2541 - 2547. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Roura-Mir, L. Wang, T.-Y. Cheng, I. Matsunaga, C. C. Dascher, S. L. Peng, M. J. Fenton, C. Kirschning, and D. B. Moody Mycobacterium tuberculosis Regulates CD1 Antigen Presentation Pathways through TLR-2 J. Immunol., August 1, 2005; 175(3): 1758 - 1766. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Bulut, K. S. Michelsen, L. Hayrapetian, Y. Naiki, R. Spallek, M. Singh, and M. Arditi Mycobacterium Tuberculosis Heat Shock Proteins Use Diverse Toll-like Receptor Pathways to Activate Pro-inflammatory Signals J. Biol. Chem., June 3, 2005; 280(22): 20961 - 20967. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. A. Efron, H. Tsujimoto, F. R. Bahjat, R. Ungaro, J. Debernardis, C. Tannahill, H. V. Baker, C. K. Edwards, and L. L. Moldawer Differential maturation of murine bone-marrow derived dendritic cells with lipopolysaccharide and tumor necrosis factor-{alpha} Innate Immunity, June 1, 2005; 11(3): 145 - 160. [Abstract] [PDF]
|
|
|
|
|
|
J. Uehori, K. Fukase, T. Akazawa, S. Uematsu, S. Akira, K. Funami, M. Shingai, M. Matsumoto, I. Azuma, K. Toyoshima, et al. Dendritic Cell Maturation Induced by Muramyl Dipeptide (MDP) Derivatives: Monoacylated MDP Confers TLR2/TLR4 Activation J. Immunol., June 1, 2005; 174(11): 7096 - 7103. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Rehli{section}, S. Sulzbacher, S. Pape, T. Ravasi, C. A. Wells, S. Heinz, L. Sollner, C. El Chartouni, S. W. Krause, E. Steingrimsson, et al. Transcription Factor Tfec Contributes to the IL-4-Inducible Expression of a Small Group of Genes in Mouse Macrophages Including the Granulocyte Colony-Stimulating Factor Receptor J. Immunol., June 1, 2005; 174(11): 7111 - 7122. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Maeda, T. Mukai, J. Spencer, and M. Makino Identification of an Immunomodulating Agent from Mycobacterium leprae Infect. Immun., May 1, 2005; 73(5): 2744 - 2750. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Wang, H. M. Curry, B. S. Zwilling, and W. P. Lafuse Mycobacteria Inhibition of IFN-{gamma} Induced HLA-DR Gene Expression by Up-Regulating Histone Deacetylation at the Promoter Region in Human THP-1 Monocytic Cells J. Immunol., May 1, 2005; 174(9): 5687 - 5694. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. E. Geisel, K. Sakamoto, D. G. Russell, and E. R. Rhoades In Vivo Activity of Released Cell Wall Lipids of Mycobacterium bovis Bacillus Calmette-Guerin Is Due Principally to Trehalose Mycolates J. Immunol., April 15, 2005; 174(8): 5007 - 5015. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Sendide, N. E. Reiner, J. S. I. Lee, S. Bourgoin, A. Talal, and Z. Hmama Cross-Talk between CD14 and Complement Receptor 3 Promotes Phagocytosis of Mycobacteria: Regulation by Phosphatidylinositol 3-Kinase and Cytohesin-1 J. Immunol., April 1, 2005; 174(7): 4210 - 4219. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Tobias and L. K. Curtiss Thematic review series: The Immune System and Atherogenesis. Paying the price for pathogen protection: toll receptors in atherogenesis J. Lipid Res., March 1, 2005; 46(3): 404 - 411. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. W. Wieland, S. Knapp, S. Florquin, A. F. de Vos, K. Takeda, S. Akira, D. T. Golenbock, A. Verbon, and T. van der Poll Non-Mannose-capped Lipoarabinomannan Induces Lung Inflammation via Toll-like Receptor 2 Am. J. Respir. Crit. Care Med., December 15, 2004; 170(12): 1367 - 1374. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Nicolle, C. Fremond, X. Pichon, A. Bouchot, I. Maillet, B. Ryffel, and V. J. F. Quesniaux Long-Term Control of Mycobacterium bovis BCG Infection in the Absence of Toll-Like Receptors (TLRs): Investigation of TLR2-, TLR6-, or TLR2-TLR4-Deficient Mice Infect. Immun., December 1, 2004; 72(12): 6994 - 7004. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Hyakushima, H. Mitsuzawa, C. Nishitani, H. Sano, K. Kuronuma, M. Konishi, T. Himi, K. Miyake, and Y. Kuroki Interaction of Soluble Form of Recombinant Extracellular TLR4 Domain with MD-2 Enables Lipopolysaccharide Binding and Attenuates TLR4-Mediated Signaling J. Immunol., December 1, 2004; 173(11): 6949 - 6954. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. Vora, A. Youdim, L. S. Thomas, M. Fukata, S. Y. Tesfay, K. Lukasek, K. S. Michelsen, A. Wada, T. Hirayama, M. Arditi, et al. {beta}-Defensin-2 Expression Is Regulated by TLR Signaling in Intestinal Epithelial Cells J. Immunol., November 1, 2004; 173(9): 5398 - 5405. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Agrawal, C. Bettegowda, I. Cheong, J.-F. Geschwind, C. G. Drake, E. L. Hipkiss, M. Tatsumi, L. H. Dang, L. A. Diaz Jr., M. Pomper, et al. Bacteriolytic therapy can generate a potent immune response against experimental tumors PNAS, October 19, 2004; 101(42): 15172 - 15177. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. FITNESS, S. FLOYD, D. K. WARNDORFF, L. SICHALI, L. MWAUNGULU, A. C. CRAMPIN, P. E. M. FINE, and A. V. S. HILL LARGE-SCALE CANDIDATE GENE STUDY OF LEPROSY SUSCEPTIBILITY IN THE KARONGA DISTRICT OF NORTHERN MALAWI Am J Trop Med Hyg, September 1, 2004; 71(3): 330 - 340. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. FITNESS, S. FLOYD, D. K. WARNDORFF, L. SICHALI, S. MALEMA, A. C. CRAMPIN, P. E. M. FINE, and A. V. S. HILL LARGE-SCALE CANDIDATE GENE STUDY OF TUBERCULOSIS SUSCEPTIBILITY IN THE KARONGA DISTRICT OF NORTHERN MALAWI Am J Trop Med Hyg, September 1, 2004; 71(3): 341 - 349. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Jang, S. Uematsu, S. Akira, and P. Salgame IL-6 and IL-10 Induction from Dendritic Cells in Response to Mycobacterium tuberculosis Is Predominantly Dependent on TLR2-Mediated Recognition J. Immunol., September 1, 2004; 173(5): 3392 - 3397. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. J. Gehring, K. M. Dobos, J. T. Belisle, C. V. Harding, and W. H. Boom Mycobacterium tuberculosis LprG (Rv1411c): A Novel TLR-2 Ligand That Inhibits Human Macrophage Class II MHC Antigen Processing J. Immunol., August 15, 2004; 173(4): 2660 - 2668. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. A. Darrah, M. C. G. Monaco, S. Jain, M. K. Hondalus, D. T. Golenbock, and D. M. Mosser Innate Immune Responses to Rhodococcus equi J. Immunol., August 1, 2004; 173(3): 1914 - 1924. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. S. Michelsen, M. H. Wong, P. K. Shah, W. Zhang, J. Yano, T. M. Doherty, S. Akira, T. B. Rajavashisth, and M. Arditi Lack of Toll-like receptor 4 or myeloid differentiation factor 88 reduces atherosclerosis and alters plaque phenotype in mice deficient in apolipoprotein E PNAS, July 20, 2004; 101(29): 10679 - 10684. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. M. Fortune, A. Solache, A. Jaeger, P. J. Hill, J. T. Belisle, B. R. Bloom, E. J. Rubin, and J. D. Ernst Mycobacterium tuberculosis Inhibits Macrophage Responses to IFN-{gamma} through Myeloid Differentiation Factor 88-Dependent and -Independent Mechanisms J. Immunol., May 15, 2004; 172(10): 6272 - 6280. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Cornelie, J. Hoebeke, A.-M. Schacht, B. Bertin, J. Vicogne, M. Capron, and G. Riveau Direct Evidence that Toll-like Receptor 9 (TLR9) Functionally Binds Plasmid DNA by Specific Cytosine-phosphate-guanine Motif Recognition J. Biol. Chem., April 9, 2004; 279(15): 15124 - 15129. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. N. Dao, L. Kremer, Y. Guerardel, A. Molano, W. R. Jacobs Jr., S. A. Porcelli, and V. Briken Mycobacterium tuberculosis Lipomannan Induces Apoptosis and Interleukin-12 Production in Macrophages Infect. Immun., April 1, 2004; 72(4): 2067 - 2074. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. A. Fulton, S. M. Reba, R. K. Pai, M. Pennini, M. Torres, C. V. Harding, and W. H. Boom Inhibition of Major Histocompatibility Complex II Expression and Antigen Processing in Murine Alveolar Macrophages by Mycobacterium bovis BCG and the 19-Kilodalton Mycobacterial Lipoprotein Infect. Immun., April 1, 2004; 72(4): 2101 - 2110. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. A. Scanga, A. Bafica, C. G. Feng, A. W. Cheever, S. Hieny, and A. Sher MyD88-Deficient Mice Display a Profound Loss in Resistance to Mycobacterium tuberculosis Associated with Partially Impaired Th1 Cytokine and Nitric Oxide Synthase 2 Expression Infect. Immun., April 1, 2004; 72(4): 2400 - 2404. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H.-J. Anders, B. Banas, and D. Schlondorff Signaling Danger: Toll-Like Receptors and their Potential Roles in Kidney Disease J. Am. Soc. Nephrol., April 1, 2004; 15(4): 854 - 867. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Z. Darieva, E. B. Lasunskaia, M. N. N. Campos, T. L. Kipnis, and W. D. da Silva Activation of phosphatidylinositol 3-kinase and c-Jun-N-terminal kinase cascades enhances NF-{kappa}B-dependent gene transcription in BCG-stimulated macrophages through promotion of p65/p300 binding J. Leukoc. Biol., April 1, 2004; 75(4): 689 - 697. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. J. Quesniaux, D. M. Nicolle, D. Torres, L. Kremer, Y. Guerardel, J. Nigou, G. Puzo, F. Erard, and B. Ryffel Toll-Like Receptor 2 (TLR2)-Dependent-Positive and TLR2-Independent-Negative Regulation of Proinflammatory Cytokines by Mycobacterial Lipomannans J. Immunol., April 1, 2004; 172(7): 4425 - 4434. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. A. W. Verreck, T. de Boer, D. M. L. Langenberg, M. A. Hoeve, M. Kramer, E. Vaisberg, R. Kastelein, A. Kolk, R. de Waal-Malefyt, and T. H. M. Ottenhoff Human IL-23-producing type 1 macrophages promote but IL-10-producing type 2 macrophages subvert immunity to (myco)bacteria PNAS, March 30, 2004; 101(13): 4560 - 4565. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. E. Weir, G. F. Black, H. M. Dockrell, S. Floyd, P. E. M. Fine, S. D. Chaguluka, S. Stenson, E. King, B. Nazareth, D. K. Warndorff, et al. Mycobacterial Purified Protein Derivatives Stimulate Innate Immunity: Malawians Show Enhanced Tumor Necrosis Factor Alpha, Interleukin-1{beta} (IL-1{beta}), and IL-10 Responses Compared to Those of Adolescents in the United Kingdom Infect. Immun., March 1, 2004; 72(3): 1807 - 1811. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Branger, J. C. Leemans, S. Florquin, S. Weijer, P. Speelman, and T. van der Poll Toll-like receptor 4 plays a protective role in pulmonary tuberculosis in mice Int. Immunol., March 1, 2004; 16(3): 509 - 516. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. J. Murphy, H. M. Paterson, J. A. Mannick, and J. A. Lederer Injury, sepsis, and the regulation of Toll-like receptor responses J. Leukoc. Biol., March 1, 2004; 75(3): 400 - 407. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B.-C. Chiu, C. M. Freeman, V. R. Stolberg, J. S. Hu, E. Komuniecki, and S. W. Chensue The Innate Pulmonary Granuloma: Characterization and Demonstration of Dendritic Cell Recruitment and Function Am. J. Pathol., March 1, 2004; 164(3): 1021 - 1030. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A.C. Ogus, B. Yoldas, T. Ozdemir, A. Uguz, S. Olcen, I. Keser, M. Coskun, A. Cilli, and O. Yegin The Arg753Gln polymorphism of the human Toll-like receptor 2 gene in tuberculosis disease Eur. Respir. J., February 1, 2004; 23(2): 219 - 223. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. A. Campos, G. M. S. Rosinha, I. C. Almeida, X. S. Salgueiro, B. W. Jarvis, G. A. Splitter, N. Qureshi, O. Bruna-Romero, R. T. Gazzinelli, and S. C. Oliveira Role of Toll-Like Receptor 4 in Induction of Cell-Mediated Immunity and Resistance to Brucella abortus Infection in Mice Infect. Immun., January 1, 2004; 72(1): 176 - 186. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. B. Drennan, D. Nicolle, V. J. F. Quesniaux, M. Jacobs, N. Allie, J. Mpagi, C. Fremond, H. Wagner, C. Kirschning, and B. Ryffel Toll-Like Receptor 2-Deficient Mice Succumb to Mycobacterium tuberculosis Infection Am. J. Pathol., January 1, 2004; 164(1): 49 - 57. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. R. Alvarez, B. S. Zwilling, and W. P. Lafuse Mycobacterium avium Inhibition of IFN-{gamma} Signaling in Mouse Macrophages: Toll-Like Receptor 2 Stimulation Increases Expression of Dominant-Negative STAT1{beta} by mRNA Stabilization J. Immunol., December 15, 2003; 171(12): 6766 - 6773. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A.-S. W. Moller, R. Ovstebo, A.-B. Westvik, G. B. Joo, K.-B. F. Haug, and P. Kierulf Effects of bacterial cell wall components (PAMPs) on the expression of monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein-1{alpha} (MIP-1{alpha}) and the chemokine receptor CCR2 by purified human blood monocytes Innate Immunity, December 1, 2003; 9(6): 349 - 360. [Abstract] [PDF]
|
|
|
|
|
|
C. L. Fuller, J. L. Flynn, and T. A. Reinhart In Situ Study of Abundant Expression of Proinflammatory Chemokines and Cytokines in Pulmonary Granulomas That Develop in Cynomolgus Macaques Experimentally Infected with Mycobacterium tuberculosis Infect. Immun., December 1, 2003; 71(12): 7023 - 7034. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. L. Fisette, S. Ram, J. M. Andersen, W. Guo, and R. R. Ingalls The Lip Lipoprotein from Neisseria gonorrhoeae Stimulates Cytokine Release and NF-{kappa}B Activation in Epithelial Cells in a Toll-like Receptor 2-dependent Manner J. Biol. Chem., November 21, 2003; 278(47): 46252 - 46260. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. G. Feng, C. A. Scanga, C. M. Collazo-Custodio, A. W. Cheever, S. Hieny, P. Caspar, and A. Sher Mice Lacking Myeloid Differentiation Factor 88 Display Profound Defects in Host Resistance and Immune Responses to Mycobacterium avium Infection Not Exhibited by Toll-Like Receptor 2 (TLR2)- and TLR4-Deficient Animals J. Immunol., November 1, 2003; 171(9): 4758 - 4764. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. J. C. Gibson, L. Eggeling, W. N. Maughan, K. Krumbach, S. S. Gurcha, J. Nigou, G. Puzo, H. Sahm, and G. S. Besra Disruption of Cg-Ppm1, a Polyprenyl Monophosphomannose Synthase, and the Generation of Lipoglycan-less Mutants in Corynebacterium glutamicum J. Biol. Chem., October 17, 2003; 278(42): 40842 - 40850. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Shi, C. Nathan, D. Schnappinger, J. Drenkow, M. Fuortes, E. Block, A. Ding, T. R. Gingeras, G. Schoolnik, S. Akira, et al. MyD88 Primes Macrophages for Full-Scale Activation by Interferon-{gamma} yet Mediates Few Responses to Mycobacterium tuberculosis J. Exp. Med., October 6, 2003; 198(7): 987 - 997. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. K. Sukumaran, H. Shimada, and N. V. Prasadarao Entry and Intracellular Replication of Escherichia coli K1 in Macrophages Require Expression of Outer Membrane Protein A Infect. Immun., October 1, 2003; 71(10): 5951 - 5961. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Kariyone, T. Tamura, H. Kano, Y. Iwakura, K. Takeda, S. Akira, and K. Takatsu Immunogenicity of Peptide-25 of Ag85B in Th1 development: role of IFN-{gamma} Int. Immunol., October 1, 2003; 15(10): 1183 - 1194. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Fujita, T. Into, M. Yasuda, T. Okusawa, S. Hamahira, Y. Kuroki, A. Eto, T. Nisizawa, M. Morita, and K.-i. Shibata Involvement of Leucine Residues at Positions 107, 112, and 115 in a Leucine-Rich Repeat Motif of Human Toll-Like Receptor 2 in the Recognition of Diacylated Lipoproteins and Lipopeptides and Staphylococcus aureus Peptidoglycans J. Immunol., October 1, 2003; 171(7): 3675 - 3683. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Guerardel, E. Maes, V. Briken, F. Chirat, Y. Leroy, C. Locht, G. Strecker, and L. Kremer Lipomannan and Lipoarabinomannan from a Clinical Isolate of Mycobacterium kansasii: NOVEL STRUCTURAL FEATURES AND APOPTOSIS-INDUCING PROPERTIES J. Biol. Chem., September 19, 2003; 278(38): 36637 - 36651. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. Sandor, E. Latz, F. Re, L. Mandell, G. Repik, D. T. Golenbock, T. Espevik, E. A. Kurt-Jones, and R. W. Finberg Importance of extra- and intracellular domains of TLR1 and TLR2 in NF{kappa}B signaling J. Cell Biol., September 15, 2003; 162(6): 1099 - 1110. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Gilleron, V. F. J. Quesniaux, and G. Puzo Acylation State of the Phosphatidylinositol Hexamannosides from Mycobacterium bovis Bacillus Calmette Guerin and Mycobacterium tuberculosis H37Rv and Its Implication in Toll-like Receptor Response J. Biol. Chem., August 8, 2003; 278(32): 29880 - 29889. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Uehori, M. Matsumoto, S. Tsuji, T. Akazawa, O. Takeuchi, S. Akira, T. Kawata, I. Azuma, K. Toyoshima, and T. Seya Simultaneous Blocking of Human Toll-Like Receptors 2 and 4 Suppresses Myeloid Dendritic Cell Activation Induced by Mycobacterium bovis Bacillus Calmette-Guerin Peptidoglycan Infect. Immun., August 1, 2003; 71(8): 4238 - 4249. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. J. Gehring, R. E. Rojas, D. H. Canaday, D. L. Lakey, C. V. Harding, and W. H. Boom The Mycobacterium tuberculosis 19-Kilodalton Lipoprotein Inhibits Gamma Interferon-Regulated HLA-DR and Fc{gamma}R1 on Human Macrophages through Toll-Like Receptor 2 Infect. Immun., August 1, 2003; 71(8): 4487 - 4497. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. I. Tapping and P. S. Tobias Mycobacterial lipoarabinomannan mediates physical interactions between TLR1 and TLR2 to induce signaling Innate Immunity, August 1, 2003; 9(4): 264 - 268. [Abstract] [PDF]
|
|
|
|
 |
|
 J. Indrigo, R. L. Hunter Jr, and J. K. Actor Cord factor trehalose 6,6'-dimycolate (TDM) mediates trafficking events during mycobacterial infection of murine macrophages Microbiology, August 1, 2003; 149(8): 2049 - 2059. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. A. Heldwein, M. D. Liang, T. K. Andresen, K. E. Thomas, A. M. Marty, N. Cuesta, S. N. Vogel, and M. J. Fenton TLR2 and TLR4 serve distinct roles in the host immune response against Mycobacterium bovis BCG J. Leukoc. Biol., August 1, 2003; 74(2): 277 - 286. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Bafica, C. A. Scanga, M. L. Schito, S. Hieny, and A. Sher Cutting Edge: In Vivo Induction of Integrated HIV-1 Expression by Mycobacteria Is Critically Dependent on Toll-Like Receptor 2 J. Immunol., August 1, 2003; 171(3): 1123 - 1127. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. A. R. Tobian, N. S. Potter, L. Ramachandra, R. K. Pai, M. Convery, W. H. Boom, and C. V. Harding Alternate Class I MHC Antigen Processing Is Inhibited by Toll-Like Receptor Signaling Pathogen-Associated Molecular Patterns: Mycobacterium tuberculosis 19-kDa Lipoprotein, CpG DNA, and Lipopolysaccharide J. Immunol., August 1, 2003; 171(3): 1413 - 1422. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. S. Van Amersfoort, T. J. C. Van Berkel, and J. Kuiper Receptors, Mediators, and Mechanisms Involved in Bacterial Sepsis and Septic Shock Clin. Microbiol. Rev., July 1, 2003; 16(3): 379 - 414. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
