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B and STAT Signaling Pathways in Human Macrophages1
Department of Virology, National Public Health Institute, Helsinki, Finland
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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B and STAT
DNA-binding activity in human primary macrophages as analyzed by EMSA.
NF-B activation was rapid and was not inhibited by a protein
synthesis inhibitor cycloheximide, suggesting that these bacteria could
directly activate NF-B. STAT1, STAT3, and IFN regulatory factor-1
DNA binding was induced by both bacteria with delayed kinetics compared
with NF-B. In addition, streptococci induced the formation of
IFN-
-specific transcription factor complex and IFN-stimulated gene
factor-3 (ISGF3). STAT1 and STAT3 activation and ISGF3 complex
formation were inhibited by cycloheximide or by neutralization with
IFN-/ß-specific Abs. Streptococci were more potent than
lactobacilli in inducing STAT1, ISGF3, and IFN regulatory factor-1 DNA
binding. Accordingly, only streptococci induced IFN- production. The
activation of the IFN- signaling pathway by streptococci could play
a role in the pathogenesis of these bacteria. These results indicate
that extracellular Gram-positive bacteria activate transcription
factors involved in cytokine signaling by two mechanisms: directly,
leading to NF-B activation, and indirectly via cytokines, leading to
STAT activation. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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and IL-6, which are among the first produced
during the innate immune response toward bacteria. TNF- and IL-6
have pleiotropic effects such as induction of the acute phase response
and activation of macrophages (2, 3). An innate immune
response to viruses is characterized by rapid production of IFN- in
macrophages. In addition to its direct antiviral functions, IFN- has
a range of immunoregulatory functions, including NK cell activation and
enhancement of Th1-type immunity (4, 5, 6, 7). Although the role
of IFN- in viral and intracellular bacterial infections is well
established, in extracellular bacterial infections it remains poorly
characterized.
Cytokines initiate signaling cascades through their receptors, leading
to activation of transcription factors and target gene expression.
NF-B and STATs are both latent cytoplasmic transcription factors
activated by Ag or cytokine stimulation. They regulate transcription of
genes encoding proteins involved in immune, acute phase, and
inflammatory responses. In humans, activated NF-B dimers consist
mainly of the Rel family proteins p50 and p65 subunits. NF-B binds
to responsive B sites in the promoters and enhancers of target
genes, including TNF- and IL-6. TNF- is
known to initiate an autoregulatory feedback loop, where the activation
of NF-B is followed by the production of TNF- and further
activation of NF-B (8, 9). STAT activation occurs via
tyrosine phosphorylation; binding of cytokines to their receptors
results in autophosphorylation of receptor-associated JAK kinases that
phosphorylate and activate STATs. Activated STATs form homo- or
heterodimers that translocate into the nucleus and bind specific target
elements in the promoters of cytokine-inducible genes
(10, 11, 12). STAT1 and STAT3 can be activated by several
cytokines, including IFN- and IL-6, and bind to the IFN-
activation site (GAS)3
element. IFN- is the only known activator of STAT2 that, together
with STAT1 and p48, form the IFN-stimulated gene factor-3 (ISGF3) (3)
complex that binds to the IFN response element (ISRE) (13, 14).
Lactobacilli are nonpathogenic Gram-positive inhabitants of human
normal microflora (15); some of them have been postulated
to have health beneficial effects, such as stimulation of the immune
system (16). Strain Lactobacillus rhamnosus GG
has been extensively studied regarding safety and clinical effects
(17). Streptococcus pyogenes (group A
streptococci) is a major Gram-positive human pathogen causing a wide
range of infections. Since the 1980s, highly invasive strains
associated with shock and organ failure have emerged (18).
To analyze the mechanisms by which Gram-positive bacteria induce immune
responses we have compared nonpathogenic and pathogenic strains. We
have previously shown that L. rhamnosus GG and S.
pyogenes induce the production of several cytokines in human PBMC
(19). In this work we have compared and analyzed the role
of live L. rhamnosus GG and S. pyogenes in
activation of NF-B and STATs in human primary macrophages.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Lactobacillus rhamnosus GG (American Type Culture Collection 53103) was obtained from Valio R&D (Helsinki, Finland), and Streptococcus pyogenes serotype T1 (IH32030), isolated from a child with bacteremia, was obtained from the collection of the National Public Health Institute (Helsinki, Finland). Bacteria were stored in skimmed milk at -70°C and passaged three times as previously described (20) before their use in stimulation experiments. Lactobacilli were grown in MRS medium (Difco, Detroit, MI) and streptococci in TY medium supplemented with 0.2% glucose (21). For stimulation experiments bacteria were grown to logarithmic growth phase, and the number of bacterial cells was determined by counting in a Petroff Hausser counting chamber.
Cell culture
Freshly collected leukocyte-rich buffy coats from healthy blood donors were supplied by the Finnish Red Cross Blood Transfusion Service (Helsinki, Finland). PBMC were isolated by a density gradient centrifugation over Ficoll-Paque gradient (Pharmacia, Uppsala, Sweden). After washing, the cells were resuspended in RPMI 1640 medium (Sigma, St. Louis, MO) supplemented with 0.6 µg/ml penicillin, 60 µg/ml streptomycin, 2 mM L-glutamine, and 20 mM HEPES. For monocyte differentiation, PBMC were allowed to adhere to plastic six-well plates (Falcon, Becton Dickinson, Franklin Lakes, NJ) for 1 h at 37°C in RPMI 1640 medium supplemented with antibiotics, glutamine, and HEPES without FCS (10 x 106 cells/well). After incubation nonadherent cells were removed, and the wells were washed twice with PBS (pH 7.4). Adherent cells were then grown for 7 days in macrophage/serum-free medium (Life Technologies, Grand Island, NY) supplemented with antibiotics and recombinant GM-CSF at 10 ng/ml (Leucomax, Schering-Plough, Innishannon, Ireland). More than 90% of the cultured cells were macrophages as determined by their morphology and CD14 expression (data not shown).
Stimulation experiments
To minimize interindividual variation all experiments were
performed with cells obtained from four to six blood donors.
Stimulation experiments were conducted in RPMI 1640 medium either with
or without 10% heat-inactivated FCS (Integro, Zaandam, The
Netherlands). Macrophages were stimulated with live bacteria at a 1:1
ratio. Cycloheximide (CHX; Sigma) was used to inhibit protein synthesis
at concentration of 10 µg/ml. It was added to cell culture medium
0.5 h after the beginning of stimulations. Neutralizing sheep
anti-IFN-/ß (22) and goat anti-IL-6 (R & D
Systems, Abingdon, U.K.) Abs were used at concentrations of 2400/165
neutralizing IU/ml and 1 µg/ml, respectively. Cell culture
supernatants and cells were collected at different times after
stimulation and pooled. Cells were used for preparing nuclear extracts,
isolation of total cellular RNA, or SDS-PAGE sample preparation.
Supernatants stored at -20°C were used for cytokine
determinations.
EMSA
Nuclear extracts were prepared as previously described
(23). Nuclear protein/DNA-binding reactions were performed
in a volume of 20 µl containing 5 µg of nuclear extract protein, 10
mM HEPES-KOH (pH 7.9), 100 mM NaCl, 1 mM EDTA, 1 mM DTT, 10% glycerol,
and 2 µg poly(dI-dC) as a nonspecific competitor. NF-B
(5'-AGTTGAGGGGACTTTCCCAGG-3'), IRF1-GAS
(5'-AGCTTCAGCCTGATTTCCCCGAAATGACGGCA-3'), SIE
(5'-GATCTAGGGATTTCCGGGAAATGAAGCT-3'), ISRE15
(5'-AGCTTGATCGGGAAAGGGAAACCGAAACTGAAGCCA-3'), and PRDI
(5'-GATCAAGTGAAAGTGAAAGTGA-3') oligonucleotides were synthesized with
an IBI oligonucleotide synthesizer (Foster City, CA) and purified on
PAGE in the presence of 8 mol/l urea. The probes for NF-B, IRF1-GAS,
SIE, and ISRE15 were end labeled with
[-32P]dATP (3000 Ci/mol; Amersham,
Aylesbury, U.K.) by T4 polynucleotide kinase and the probe for PRDI
with the Klenow fill-in method. The binding reaction was performed at
room temperature for 30 min. For supershift assays, nuclear protein
extracts were incubated with Abs against NF-B p50 (sc-1190 x),
NF-B p65 (sc-372 x), STAT1 p91 (sc-345 x), STAT5 recognizing both
STAT5A and STAT5B (sc-835 x; all from Santa Cruz Biotechnology, Santa
Cruz, CA), and STAT3 (71-0900, Zymed, San Francisco, CA) for 1 h
on ice before addition of the radiolabeled probe. Nondenaturing low
ionic strength PAGE gels (6%) in 0.25 x Tris-borate-EDTA buffer
were used. Gels were dried, and bands were visualized by
autoradiography.
Western blotting
SDS-PAGE was conducted by using the Laemmli buffer system (24) on 10% polyacrylamide gels. Proteins separated on gels were transferred to Immobilon-P (polyvinylidene difluoride) membranes (Millipore, Bedford, MA) with an Isophor electrotransfer device (Hoefer Scientific Instruments, San Francisco, CA) at 200 mA for 2 h. Binding of the primary and secondary Abs was performed in PBS (pH 7.4) containing 5% nonfat milk for 1 h at room temperature. Primary Abs used in immunoblotting were guinea pig anti-human IRF-1 (1/2000 dilution) (25) and guinea pig anti-human MxA (1/2000 dilution) (26). Peroxidase-conjugated rabbit anti-guinea pig Igs (Dako, Copenhagen, Denmark) were used as secondary Abs (1/2000 dilution). The bands were visualized on Amersham HyperMax film using the ECL chemiluminescence system according to the manufacturer’s (Amersham) instructions. Protein concentrations in the samples were determined with the Bio-Rad protein assay kit (Hercules, CA).
RNA isolation and analysis
For isolation of total cellular RNA, stimulated cells were
collected, washed once with cold PBS (pH 7.4), and lysed in guanidium
isothiocyanate (stored at -70°C) (27) followed by
centrifugation through a CsCl cushion (28). RNA was
quantitated photometrically, and samples containing equal amounts (20
µg) of total cellular RNA were size-fractionated on 1%
formaldehyde-agarose gels, transferred to Hybond-N nylon membranes
(Amersham), and hybridized with human IRF-1 (29) and MxA
(30) cDNA probes. To control for equal RNA loading,
ethidium bromide staining or hybridization with ß-actin cDNA probe
was used. The probes were labeled with
[-32P]dCTP (3000 Ci/mmol; Amersham) using a
random primed DNA labeling kit (Roche, Indianapolis, IN).
Hybridizations were performed in a solution containing 50% formamide,
5x Denhardt’s solution, 5x SSPE, and 0.5% SDS at 42°C. Membranes
were washed twice with 1x SSC/0.1% SDS at 42°C for 30 min and once
at 65°C for 30 min. The membranes were exposed to Kodak X-OMAT AR
films (Eastman Kodak, Rochester, NY) at -70°C with intensifying
screens.
Cytokine-specific ELISAs and biological assay for IFN-/ß
TNF- and IL-6 levels in cell culture supernatants were
determined by ELISA methods as described previously (19)
with sensitivities of 20 pg/ml. The IFN-/ß assay was performed as
previously described (31) with a detection limit of 3
IU/ml. Briefly, cell culture supernatants were harvested and dialyzed
against acidic glycine buffer (pH 2) followed by two dialyses in PBS.
IFN- titers in samples were assayed by VSV plaque reduction in Hep2
cells.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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B DNA binding in human macrophages by
Gram-positive bacteria
To study lactobacilli- and streptococci-induced activation of
transcription factors, differentiated macrophages were stimulated with
live bacteria. Nuclear extracts from stimulated macrophages were
prepared and analyzed by EMSA using NF-B oligonucleotide. Both
bacteria activated NF-B DNA binding rapidly at 1 h after
stimulation, and this activation increased with time for up to 24
h (Fig. 1
A). As shown by the
supershift experiment, the NF-B DNA binding complex containing p50
and p65 was activated at 1 h after bacterial stimulation and was
most prominent at the 24 h point (Fig. 1B).
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Both NF-B and STATs are involved in cytokine signaling during
immune responses. We therefore studied the possible activation of STATs
induced by Gram-positive bacteria. STAT activation was analyzed using
nuclear extracts prepared from live bacteria-stimulated macrophages by
EMSA with IRF1-GAS, SIE, and ISRE15 oligonucleotides. STAT binding to
IRF1-GAS in response to streptococci was detected at 3 h after
stimulation, and the intensity of this complex increased up to 24
h. Lactobacilli-induced STAT binding to IRF1-GAS was weakly detectable
at 9 and 24 h after stimulation (Fig. 2A). Supershift experiments
with anti-STAT1, anti-STAT3 and anti-STAT5 Abs showed that
both bacteria activated STAT1 binding (Fig. 2B). As analyzed
using the SIE element, streptococci apparently activated the lowest of
the three STAT DNA binding complexes at 3 h after stimulation.
Both lactobacilli and streptococci activated the STAT DNA binding
complex at 6 h after stimulation, with the intensity of the
complexes increasing at least up to 24 h (Fig. 3A). The SIE binding complexes
activated by bacteria containing STAT1 and STAT3 homodimers (the lowest
and the uppermost of the three bands, respectively) as well as STAT1/3
heterodimers (the middle band). Also, STAT5 binding to the SIE element
induced by lactobacilli was seen at 24 h after stimulation (Fig. 3B). ISGF3 complex binding to the ISRE15 element was
detectable at 6 and 9 h after stimulation with streptococci, and
the complex started to disappear thereafter (Fig. 4).
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B
and STATs by Gram-positive bacteria
To analyze the requirement of ongoing protein synthesis for
NF-B and STAT activation, macrophages were stimulated with live
bacteria for 3 and 6 h in the presence or the absence of CHX.
NF-B activation by lactobacilli or streptococci was not inhibited by
CHX at either time point (Fig. 5A). Streptococci-induced
STAT1 DNA binding to IRF1-GAS or SIE elements was inhibited by CHX
(Fig. 5, B and C). Lactobacilli activated weak
STAT1 and STAT3 binding that was also inhibited by CHX (Fig. 5C). Streptococci-induced ISGF3 complex formation was
similarly blocked by CHX (Fig. 5D).
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in human
macrophages stimulated with Gram-positive bacteria
As CHX experiments showed, ongoing protein synthesis was required
for bacteria-induced STAT activation. This suggested that activation of
STATs is likely to be cytokine mediated. The cytokines involved in
STAT1 and STAT3 activation include macrophage-derived IFN- and IL-6,
while TNF- is the most prominent activator of NF-B. To quantitate
TNF- and IL-6 production we used ELISA, whereas IFN-ß levels
were measured by a biological assay. Both lactobacilli and streptococci
induced the production of TNF- at 3 h and IL-6 at 6 h
after stimulation. Bacteria-induced TNF- and IL-6 production
increased up to 24 h after stimulation. Detectable amounts of
IFN- were found only after 24 h of stimulation with
streptococci (Fig. 6).
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ß Abs inhibit Gram-positive bacteria-induced STAT
activation
To study the role of IFN- and IL-6 in bacteria-induced STAT
activation, we used neutralizing Abs. Macrophages were stimulated with
live bacteria for 8 h in the presence or the absence of
neutralizing Abs to IFN-ß or IL-6. STAT1 binding to IRF1-GAS by
both bacteria was reduced by anti-IFN-ß Abs, while
anti-IL-6 Abs had no detectable effect on STAT1 or STAT3 DNA
binding (Fig. 7A). Similarly,
bacteria-induced SIE DNA binding was reduced, while ISGF3 DNA binding
activity was completely blocked by anti-IFN-ß Abs (Fig. 7, B and C).
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To characterize in more detail bacteria-induced IFN- production
and IFN--mediated target gene activation, we studied
bacteria-induced IRF-1 and MxA mRNA and protein expression as well as
activation of IRF-1 DNA binding. Both lactobacilli and streptococci
induced IRF-1 mRNA at 2 h and MxA mRNA at 6 h after
stimulation (Fig. 8A).
Streptococci induced IRF-1 and MxA mRNAs more strongly than
lactobacilli. Expression of IRF-1 and MxA proteins by streptococci was
first detectable at 6 h after stimulation, continuing up to
24 h (Fig. 8B). Increasing the streptococcal dose to
10-fold slightly potentiated the expression of IRF-1, but decreased
that of MxA. Both lactobacilli and streptococci induced IRF-1 DNA
binding to PRDI oligonucleotide at 3 h after stimulation. However,
streptococci induced IRF-1 DNA binding more efficiently than
lactobacilli at all time points studied (Fig. 9).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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and IL-6 is
accomplished by NF-B. We have analyzed and compared the ability of
live nonpathogenic Lactobacillus rhamnosus GG and pathogenic
Streptococcus pyogenes to induce NF-B activation in human
primary macrophages. Lactobacilli and streptococci similarly induced
NF-B DNA binding with fast kinetics, and the intensity of the DNA
binding complex increased with time. As shown by the supershift
experiment, bacteria-induced NF-B consisted of at least of p50 and
p65 proteins, which form the most common active dimer in human cells
(8). Our data are in line with previous studies where
Streptococcus pneumoniae and its cell wall
(32), Listeria monocytogenes and cell wall
lipoteichoic acid (33), and Lactobacillus
crispatus (34) were shown to activate NF-B in
several cell lines. Lactobacilli and streptococci induced the
production of TNF- and IL-6 in macrophages with similar kinetics.
These cytokines were produced only after the initial activation of
NF-B DNA binding, and their production increased with time. The
increase in the intensity of NF-B DNA binding signal as a function
of time could thus result from the additive activating effect of
TNF- produced by macrophages stimulated with bacteria.
The rapid activation of NF-B DNA binding without measurable cytokine
production suggested that lactobacilli and streptococci were able to
directly activate NF-B in macrophages. NF-B DNA binding was
activated by both bacteria in the presence of protein synthesis
inhibitor, CHX, supporting this observation. Likewise, rapid NF-B
activation after binding of pathogenic bacteria to their target cells
has been observed with listeria, mycobacteria, Escherichia
coli, and salmonella (33, 35, 36). The possible
direct activation of NF-B as a result of binding of lactobacilli and
streptococci to macrophages raises the question of the nature of the
receptor binding these bacteria and mediating the signal. CD14 receptor
has been implicated in NF-B activation by Gram-positive bacteria and
their cell wall components (37). However, other receptors
have been suggested to mediate binding and signaling
(38, 39, 40). Human Toll-like receptor 2 (TLR2) and TLR4 were
first demonstrated to mediate signaling by LPS leading to NF-B
activation, and this response was shown to be enhanced by CD14
(41, 42, 43). TLR2 and TLR4 were then shown to have a role in
Gram-positive bacteria-mediated signaling, leading to NF-B
activation (44, 45, 46). Most recent data indicate that TLR2
is mainly involved in responses to Gram-positive bacteria, while TLR4
has a role in recognition and signaling in response to Gram-negative
bacteria (47, 48). It is thus likely that in our
experimental system with human primary macrophages TLRs in addition to
CD14 may play a role in mediating lactobacilli- and
streptococci-induced NF-B activation. TLR2 and TLR4 mRNAs are
expressed in human primary macrophages (data not shown), making our
hypothesis plausible. The marked similarity in the ability of
lactobacilli and streptococci regardless of their pathogenicity to
induce NF-B activation could be explained by the similarity in the
components and structure of the cell wall. Pepetidoglycan that is
present in abundance in the cell walls of all Gram-positive bacteria
and has been shown to induce NF-B activation through CD14
(37) and TLR2 (44, 45, 48) receptors is a
potential component in the cell walls of lactobacilli and streptococci
mediating NF-B activation.
STATs are activated as a result of cytokine binding to their receptors
and, like NF-B, are involved in activating immune responses.
Streptococci-induced STAT1 DNA binding was more pronounced compared
with that induced by lactobacilli. In contrast, STAT3 was more strongly
activated by lactobacilli than by streptococci. Bacteria-induced
activation of STAT1 and STAT3 appeared to be dependent on protein
synthesis. This suggested that STAT activation by Gram-positive
bacteria was cytokine mediated. The kinetics of STAT activation and
IL-6 production induced by bacteria coincided, suggesting that IL-6 has
a role in bacteria-induced STAT activation. Interestingly,
neutralization of IL-6 did not have any effect on bacteria-induced
activation of these STATs. Although the role of STAT1 in intracellular
bacterial infection (listeria) in mice is well established
(49), to our knowledge the only reported work on
bacteria-induced STAT activation in human cells concerns Listeria
monocytogenes-induced IL-6-dependent activation of STAT3 in
hepatocytes (50). It could be that neutralization of IL-6
in our experimental setting was incomplete, and a small amount of IL-6
was sufficient to exert an activating effect. Neutralization of IFN-
clearly reduced STAT activation. IFN- thus appears to have a
significant role in both STAT1 and STAT3 activation, even though we
were unable to detect IFN- production in 8 h supernatants (data
not shown). However, we were able to measure IFN- in 24 h
supernatants of streptococci-stimulated macrophages.
IRF-1 is a transcriptional regulator participating in the up-regulation
of IFN-- and IFN-inducible genes by binding to their ISRE elements.
The expression of IRF-1 is induced, e.g., by viruses, IFNs, TNF-,
and IL-6 (51). While acting as a positive regulator of
innate immune responses, IRF-1 also mediates Th1-type immune responses,
thus being of importance in both viral and bacterial infections
(52, 53). Lactobacilli and streptococci induced rapid
expression of IRF-1 mRNA that coincided with TNF- and IL-6 mRNA
expression (data not shown). Based on our data it remains open whether
bacteria-induced TNF- and IL-6 have a role in IRF-1 induction.
Likewise, IFN- production at early time points after bacterial
stimulation could not be detected. It is thus possible that these
bacteria directly induce the expression of IRF-1, as has been shown to
be the case with LPS (54). Rapid induction of IRF-1
production could also suggest the involvement of NF-B in
bacteria-induced IRF-1 activation. Since it is known that IRF-1 gene
expression is regulated by both STAT1 and NF-B (55, 56), it is likely that IFN-, TNF-, and IL-6 produced at
later time points after bacterial stimulation may then contribute to
activation of IRF-1 gene expression. Both bacteria induced
IRF-1 DNA binding. However, streptococci activated IRF-1 DNA binding
more efficiently than lactobacilli, which correlated to IFN-
production. Bacteria-induced IRF-1 activation was inhibited by CHX
(data not shown), suggesting that the activation of IRF-1 DNA binding
is not induced directly by these bacteria but, instead, could involve
bacteria-induced cytokines, IFN-, TNF-, or IL-6.
Formation of ISGF3 complex is specifically induced by IFN-
(57). Streptococci induced the formation of ISGF3 complex,
which was inhibited by CHX and abolished with anti-IFN- Ab,
while lactobacilli-induced ISGF3 complex formation was variably
detectable. These results together with those regarding
bacteria-induced STAT1 and STAT3 activation, strongly suggest that
despite the lack of measurable IFN- production, bacteria-induced
IFN- is involved in STAT activation. In some experiments we have
been able to measure the production of 3–10 IU/ml IFN- induced by
lactobacilli; however, streptococci was always a better inducer. In
previous studies in mice, it has been shown that nonpathogenic
lactobacilli are able to induce the production of IFN- (58, 59). It is known that as little as 0.3 IU/ml of IFN- has
biological effects in human PBMC (60). Thus, it is likely
that a small amount of IFN- is produced and used up by the
macrophages at early time points after stimulation with both bacteria.
The observation that both bacteria induced the expression of MxA, that
is strictly under regulation of IFN- (61, 62), further
supports the idea that, in addition to streptococci, lactobacilli
induce the production of small, yet biologically significant, amounts
of IFN-.
Analysis of the activation of transcription factors involved in immune
responses sheds light on the molecular mechanisms of interaction
between bacteria and immune cells, especially those between
Gram-positive bacteria and human leukocytes, which have been poorly
characterized. This work elucidates factors involved in cytokine
signaling and regulation of immune responses by nonpathogenic and
pathogenic Gram-positive bacteria. We show that lactobacilli and
streptococci similarly induce the activation of transcription factor
NF-B in primary macrophages. We are also the first to report that
both nonpathogenic and pathogenic extracellular bacteria can induce the
activation of STATs. Our results suggest that NF-B activation is
direct, while STAT activation is mediated mainly by IFN-.
Nonpathogenic and pathogenic bacteria, or at least the studied strains,
differ with regard to STAT activation. Pathogenic streptococci potently
induce IFN- production, which is likely to contribute to the
activation of STATs and IRF-1. It can be speculated that the efficient
induction of IFN- production and transcription factor activation by
streptococci would lead to fast and effective immune responses and
could play a role in the pathogenesis. The factors in lactobacilli and
streptococci mediating different interactions with macrophages remain
uncharacterized; however, it could be that streptococci have surface
components, such as lipoteichoic acids or proteins, that more avidly
interact with macrophages. We have previously shown that S.
pyogenes is a better inducer of Th1 type cytokines IL-12 and
IFN- than L. rhamnosus GG (19). The ability
of streptococci to induce IFN- production better than lactobacilli
could further contribute to its ability to direct immune responses
toward the Th1 type.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Minja Miettinen, Department of Virology, National Public Health Institute, Mannerheimintie 166, 00300 Helsinki, Finland. E-mail address:
3 Abbreviations used in this paper: GAS, IFN- activation site; ISGF3, IFN-stimulated gene factor-3; ISRE, IFN response element; CHX, cycloheximide; IRF, IFN regulatory factor; TLR, Toll-like receptor; GAS, group A streptococci; SIE, c-sis-inducible element.
Received for publication July 19, 1999. Accepted for publication January 21, 2000.
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V. Veckman and I. Julkunen Streptococcus pyogenes activates human plasmacytoid and myeloid dendritic cells J. Leukoc. Biol., February 1, 2008; 83(2): 296 - 304. [Abstract] [Full Text] [PDF]
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 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]
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S. J. Salminen, M. Gueimonde, and E. Isolauri Probiotics That Modify Disease Risk J. Nutr., May 1, 2005; 135(5): 1294 - 1298. [Abstract] [Full Text] [PDF]
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 J. A. Pena, A. B. Rogers, Z. Ge, V. Ng, S. Y. Li, J. G. Fox, and J. Versalovic Probiotic Lactobacillus spp. Diminish Helicobacter hepaticus-Induced Inflammatory Bowel Disease in Interleukin-10-Deficient Mice Infect. Immun., February 1, 2005; 73(2): 912 - 920. [Abstract] [Full Text] [PDF]
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M. C. Noverr and G. B. Huffnagle Regulation of Candida albicans Morphogenesis by Fatty Acid Metabolites Infect. Immun., November 1, 2004; 72(11): 6206 - 6210. [Abstract] [Full Text] [PDF]
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D. Ma, P. Forsythe, and J. Bienenstock Live Lactobacillus reuteri Is Essential for the Inhibitory Effect on Tumor Necrosis Factor Alpha-Induced Interleukin-8 Expression Infect. Immun., September 1, 2004; 72(9): 5308 - 5314. [Abstract] [Full Text] [PDF]
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O. Goldmann, M. Rohde, G. S. Chhatwal, and E. Medina Role of Macrophages in Host Resistance to Group A Streptococci Infect. Immun., May 1, 2004; 72(5): 2956 - 2963. [Abstract] [Full Text] [PDF]
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V. Veckman, M. Miettinen, J. Pirhonen, J. Siren, S. Matikainen, and I. Julkunen Streptococcus pyogenes and Lactobacillus rhamnosus differentially induce maturation and production of Th1-type cytokines and chemokines in human monocyte-derived dendritic cells J. Leukoc. Biol., May 1, 2004; 75(5): 764 - 771. [Abstract] [Full Text] [PDF]
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 G. Reid, J. Jass, M. T. Sebulsky, and J. K. McCormick Potential Uses of Probiotics in Clinical Practice Clin. Microbiol. Rev., October 1, 2003; 16(4): 658 - 672. [Abstract] [Full Text] [PDF]
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V. Veckman, M. Miettinen, S. Matikainen, R. Lande, E. Giacomini, E. M. Coccia, and I. Julkunen Lactobacilli and streptococci induce inflammatory chemokine production in human macrophages that stimulates Th1 cell chemotaxis J. Leukoc. Biol., September 1, 2003; 74(3): 395 - 402. [Abstract] [Full Text] [PDF]
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 J. Rincon, N. T. Viera, M. J. Romero, and J. A. Mosquera Increased production of chemotactic cytokines and elevated proliferation and expression of intercellular adhesion molecule-1 in rat mesangial cells treated with erythrogenic toxin type B and its precursor isolated from nephritogenic streptococci Nephrol. Dial. Transplant., June 1, 2003; 18(6): 1072 - 1078. [Abstract] [Full Text] [PDF]
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 T. Matsuguchi, A. Takagi, T. Matsuzaki, M. Nagaoka, K. Ishikawa, T. Yokokura, and Y. Yoshikai Lipoteichoic Acids from Lactobacillus Strains Elicit Strong Tumor Necrosis Factor Alpha-Inducing Activities in Macrophages through Toll-Like Receptor 2 Clin. Vaccine Immunol., March 1, 2003; 10(2): 259 - 266. [Abstract] [Full Text] [PDF]
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 N Borruel, M Carol, F Casellas, M Antolin, F de Lara, E Espin, J Naval, F Guarner, and J R Malagelada Increased mucosal tumour necrosis factor {alpha} production in Crohn's disease can be downregulated ex vivo by probiotic bacteria Gut, November 1, 2002; 51(5): 659 - 664. [Abstract] [Full Text] [PDF]
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D. Haller, P. Serrant, D. Granato, E. J. Schiffrin, and S. Blum Activation of Human NK Cells by Staphylococci and Lactobacilli Requires Cell Contact-Dependent Costimulation by Autologous Monocytes Clin. Vaccine Immunol., May 1, 2002; 9(3): 649 - 657. [Abstract] [Full Text] [PDF]
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