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Infection and Immunity, January 2007, p. 306-313, Vol. 75, No. 1
0019-9567/07/$08.00+0     doi:10.1128/IAI.01220-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Staphylococcal Enterotoxin B In Vivo Modulates both Gamma Interferon Receptor Expression and Ligand-Induced Activation of Signal Transducer and Activator of Transcription 1 in T Cells{triangledown}

R. Plaza, J. L. Rodriguez-Sanchez, and C. Juarez*

Department of Immunology, Hospital de la Santa Creu i Sant Pau, Universitat Autònoma de Barcelona, Avda. Sant Antoni Maria Claret 167, Barcelona 08025, Spain

Received 2 August 2006/ Returned for modification 15 September 2006/ Accepted 19 October 2006


   ABSTRACT
Top
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Superantigens (SAg) are bacterial exotoxins that provoke extreme responses in the immune system; for example, the acute hyperactivation of SAg-reactive T cells that leads to toxic shock syndrome is followed within days by strong immunosuppression. The gamma interferon (IFN-{gamma}) response is deeply affected in both extremes. The implication of IFN-{gamma} in the pathophysiology of lethal shock induced in mice after a secondary challenge with the SAg staphylococcal enterotoxin B (SEB) prompted us to study the regulation of IFN-{gamma} secretion and the intracellular response. We demonstrate in this study that a rechallenge with SEB becomes lethal only when given inside a critical time window after SEB priming and is associated with an increase of IFN-{gamma} serum release 72 h after priming. However, at this time, a selective blockade of IFN-{gamma}/STAT1 signaling develops in spleen cells, correlating with a lack of expression of the IFN-{gamma} receptor beta subunit and STAT1 in the T-cell population. Selective blockade of the STAT1 signaling pathway—while simultaneously maintaining STAT3 signaling and expression—may be a protective mechanism that shortens IFN-{gamma} production during the Th1 effector response. This blockade may also have consequences on switching towards a suppressor phenotype with chronic exposure to the superantigen.


   INTRODUCTION
Top
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Superantigens (SAgs) are highly immunostimulatory exotoxins produced by several microorganisms, mainly the gram-positive bacteria Staphylococcus aureus and Streptococcus pyogenes (34, 43). The pathophysiology associated with SAgs is linked to their ability to bypass conventional antigen peptide presentation by binding to major histocompatibility complex class II molecules on antigen-presenting cells without being internally processed and then interacting with particular external Vß domains in the T-cell receptors of reactive T lymphocytes, thus stimulating T-cell proliferation without regard for the antigen specificity of these cells (25, 32). The massive T-cell activation induced triggers the systemic release of proinflammatory cytokines, which has been associated with the development of toxic shock syndrome. Common toxic shock syndrome symptoms are fever, rash, hypotension, and multiple organ system dysfunction, which can eventually lead to death (4, 34).

Administration of the SAg staphylococcal enterotoxin B (SEB) to BALB/c mice has been a useful model for studying some in vivo effects of superantigens. While the excessive gamma interferon (IFN-{gamma}) release induced by SEB has been associated with the development of a life-threatening systemic inflammation (17) and the prevention of a protective humoral response (5), the following inhibition of IFN-{gamma} production (15, 39) might favor bacterial dissemination and susceptibility to reinfections. We therefore believe that the study of the physiological consequences of SEB on the modulation of the IFN-{gamma} response in vivo will be rewarding in unraveling some aspects of SAg-induced lethality and immunosuppression. An accurate regulation of the IFN-{gamma} response from the host is required for the orchestration of effective Th1 and macrophage-rich inflammatory reactions against microbial infections (8). IFN-{gamma} produced by T and natural killer (NK) cells interacts with a specific cell surface receptor (IFN-{gamma}R) which is expressed on all nucleated cells at modest levels (8). IFN-{gamma}R is a heterodimeric receptor composed of two subunits, namely, the {alpha} chain, which exhibits high-affinity ligand binding properties, and the ß chain (accessory factor), which is required primarily for signaling (8). Binding to the receptor activates signaling through the JAK/STAT pathway, which leads to STAT1{alpha} phosphorylation, dimerization, and translocation to the nucleus, where it binds to the gamma activation site (GAS) elements in the promoters of IFN-{gamma}-inducible genes and modulates transcription (8). IFN-{gamma} signaling can be inhibited by other cytokines, such as interleukin-6 (IL-6) or IL-10, through STAT3-mediated activation (11, 12). IL-6 and IL-10 have been found to be able to inhibit Th1 differentiation and terminate inflammatory responses (12, 44). IL-10 can even further promote the differentiation and function of T regulatory (Treg) cells to suppress inflammatory responses or induce tolerance (15, 37, 39).

The hyperresponse and the immunosuppression that follows upon SEB exposure have been studied extensively. However, these studies focused on the cellular level and on cytokine secretion but not on the modulation of the intracellular response. To gain a better understanding of the mechanism by which SAgs condition the immune responsiveness state of the host, we examined the modulation of IFN-{gamma} production and its intracellular signaling in the in vivo response to SEB. Here we show that mouse acquisition of susceptibility to a lethal shock upon rechallenge with SEB correlates with the development of the T-cell clonal expansion phase and is associated with excessive IFN-{gamma} production. In contrast, the IFN-{gamma}/STAT1 signaling pathway, but not the STAT3 pathway, is selectively down-regulated in splenic T cells 72 h after the first exposure to SEB. Differential modulation of the STAT1/STAT3 signaling pathways in Th1 cells may be involved in T-cell survival and in acquisition of a suppressor phenotype upon further stimulation with the superantigen.


   MATERIALS AND METHODS
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Animals and in vivo treatments. BALB/c (H-2d) mice were purchased from Harlan Interfauna Ibérica (Barcelona, Spain). They were bred and housed at the animal facilities of our hospital. Female mice of 8 to 16 weeks of age were utilized in all experiments. Handling of mice and experimental procedures were conducted in accordance with institutional guidelines for animal care and use. Mice were injected intraperitoneally (i.p.) with 100 µg of SEB or 10 µg of lipopolysaccharide (LPS) in 100 µl of sterile physiological saline.

Reagents and antibodies. SEB and LPS from Escherichia coli serotype O217:B8 (L3129) were purchased from Sigma (St. Louis, MO). Cyclosporine A (CsA) was purchased from Sandoz (Basel, Switzerland). The hybridoma R4-6A2 producing an anti-IFN-{gamma} monoclonal antibody (MAb) (rat immunoglobulin G1 [IgG1]) was obtained from the American Type Culture Collection (Rockville, MD). Protein G affinity columns to purify anti-IFN-{gamma} MAb from culture supernatants were purchased from Pharmacia (Freiburg, Germany). Murine recombinant IFN-{gamma} was purchased from Boehringer Mannheim (Mannheim, Germany).

Preparation of splenocytes. Spleens were removed and prepared in single-cell suspensions. Erythrocytes were osmotically lysed by a brief ammonium chloride treatment. The cells were washed three times with Tris-buffered saline and resuspended in culture medium. Cell viability was determined by trypan blue exclusion. For some experiments, T-cell-enriched spleen cells were separated according to their expression of CD90 (Thy1.2). Briefly, cells were incubated with magnetic beads coated with anti-CD90 MAb and loaded onto MACS separation columns (Miltenyi Biotec, Bergisch Gladbach, Germany). The CD90– spleen cells passed through the column (T-cell-depleted fraction). The retained cells were eluted as CD90+ T cells (T-cell-enriched fraction).

In vitro assays. Spleen cells were isolated as described above, plated in 24-well plates at a density of 5 x 106 cells/well in supplemented RPMI 1640 (final volume, 200 µl), and incubated with the indicated stimuli for 30 min at 37°C in a humidified 5% CO2 atmosphere. Cells were then harvested, and whole-cell protein extracts were prepared for activated STAT1 complex analysis by electrophoretic mobility shift assays (EMSAs).

Cell extract preparation and EMSA. After culture with appropriate stimuli, whole-cell extracts from splenocytes were prepared according to the method of Hibi et al. (24) as reported previously (3), with some modification. Briefly, cells were seeded, washed twice with cold Tris-buffered saline, and lysed in 40 µl of cold high-salt buffer (20 mM HEPES, pH 7.9, 400 mM NaCl, 1 mM EDTA, pH 8, 1 mM EGTA, pH 7.9, 1 mM dithiothreitol) that contained 0.2% Nonidet P-40, to which 0.5 mM phenylmethylsulfonyl fluoride and 1x protease inhibitor mixture (Boehringer, Mannheim, Germany) had been freshly added. After 1 h of incubation on ice, the extracts were spun for 5 min in an Eppendorf centrifuge at 4°C to pellet cellular debris. The supernatants were removed to a new tube, aliquoted, and frozen at –80°C until needed.

In another set of experiments, mice were killed, the spleens were removed, and nuclear extracts from spleen cells (10 x 107 cells) were prepared as described by Gimeno et al. (20), but using the protease inhibitors described above.

For some experiments, livers were also harvested, and nuclear extracts were prepared by a modification of Shapiro et al.'s method (19, 49). The livers were weighed, and 500 mg was excised in 3 ml of cold hypotonic buffer (10 mM HEPES, pH 7.9, 10 mM KCl, 0.1 mM EGTA, pH 7.9, 5 mM MgCl2, 1 mM dithiothreitol, and the protease inhibitors described above) to be disrupted with 10 strokes of a tight-fitting Teflon plunger, using a Potter homogenizer from Braun (Melsungen AG, Germany). Cell lysates were centrifuged at 500 x g for 5 min at 4°C, and supernatants were discarded. The pellet was resuspended in 3 ml of the same hypotonic buffer containing 0.2% Nonidet P-40 and carefully laid over 4 ml of a sucrose solution (hypotonic buffer with 0.2% Nonidet P-40 and 0.1 M sucrose) in a 15-mm centrifuge tube. After centrifugation at 1,000 x g for 20 min at 4°C in a swing-out rotor, the supernatant was discarded, and the nuclear pellet was resuspended in 500 µl of the cold high-salt buffer used for whole-cell extracts (see above) containing protein inhibitors. The nuclear suspension was incubated at 4°C for 1 h with gentle mixing by rotation. After incubation, nuclear debris was removed by centrifugation at 10,000 x g for 5 min at 4°C, and the supernatant was collected, aliquoted, and stored frozen at –80°C.

EMSAs were performed using cell extracts prepared in this manner, exactly as described by Gimeno et al. (20), using the Fc{gamma}RI and high-affinity sis-inducible element (hSIE) GAS (IFN-{gamma}-activated sequence)-like oligonucleotide probes. The Fc{gamma}RI oligonucleotide probe (5'-GTATTTCCCAGAAAAGGAAC-3') is derived from a GAS-like sequence located in the promoter of the Fc{gamma}RI gene (20), and the hSIE oligonucleotide probe (5'-GATCGTGCATTTCCCGTAAATCTTGTCTACAATTC-3') is derived from the c-fos promoter (47). Supershift assays were performed using anti-STAT1 (Transduction Systems, Lexington, KY) or anti-STAT3 (Santa Cruz Biotechnology, Santa Cruz, CA) antibodies as reported previously (20, 55).

RNA extraction and RT-PCR. RNA extraction and reverse transcription-PCR (RT-PCR) were carried out as described previously (42). The following primers were used: IFN-{gamma} sense, 5'-AACGCTACACACTGCATCTTGG-3'; IFN-{gamma} antisense, 5'-GACTTCAAAGAGTCTGAGG-3' (237 bp); IL-6 sense, 5'-TTCCATCCAGTTGCCTTCTTGG-3'; IL-6 antisense, 5'-CTTCATGTACTCCAGGTAG-3' (360 bp); IFN-{gamma}R ß-chain sense, 5'-GAACAAATCGAAGAGTATCT-3'; and IFN-{gamma}R ß-chain antisense, 5'-AATACTTGTAGCATCCAGAA-3' (281 bp). Primer sequences for STAT1, STAT3, and ß-actin were described previously (42).

Determination of IFN-{gamma} by enzyme-linked immunosorbent assay. Serum IFN-{gamma} levels were measured using a commercial kit from Endogene Inc. (Woburn, MA) according to the manufacturer's instructions.


   RESULTS
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Involvement of IFN-{gamma} in time window of susceptibility to lethal shock for mice primed with SEB. Although mice are resistant to even high doses of SEB, they become sensitized to a lethal shock when rechallenged after 48 h with the superantigen (17). This susceptibility to a rechallenge with SEB coincides with the development of the clonal expansion phase, as observed by several authors, in the activated Vß8+ T-cell subpopulation 48 h after SEB priming (28, 36, 45). These cells expand until day 3 after SEB treatment (36, 45) and decay thereafter, between days 4 and 7, to values below pretreatment values (28, 36, 45). To test the temporal window in which mortality could be achieved, mice were rechallenged with SEB 24 h, 72 h, and 120 h after being primed with SEB. A lethal shock developed only when primed mice where rechallenged after 72 h, not after 24 h or 120 h (Table 1). These results suggest that sensitization of mice to a lethal shock is indeed associated with the amplification phase of an effector T-cell response induced in SEB-primed mice. Furthermore, we also observed that mice pretreated with SEB for 72 h had highly increased IFN-{gamma} serum levels upon rechallenge with the exotoxin (Fig. 1), and blockade of IFN-{gamma} production with CsA (500 µg/mouse i.p. twice, 20 and 4 h prior to rechallenge) or neutralization with anti-IFN-{gamma} antibodies (1 mg/mouse i.p., 4 h prior to rechallenge) rescued mice from death (zero of three mice in each group died after rechallenge, in contrast to four of four mice who received no blockade of IFN-{gamma}). These data confirmed and extended previous observations where SEB-treated mice displayed an increased IFN-{gamma} response upon secondary exposure after a 48-h interval (17, 38) and neutralizing antibodies against IFN-{gamma} were able to prevent the lethal syndrome (17). Collectively, together with previous studies, these results determine a time window of susceptibility of mice to a lethal shock upon rechallenge with SEB of between 48 and 72 h after the primary SEB exposure. The findings also demonstrate that IFN-{gamma} is critically involved in the lethality that follows after a second injection with SEB given after a 72-h interval.


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  		TABLE 1. Lethality after SEB rechallenge in vivo

 

Figure 1
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  		FIG. 1. Mice are primed for IFN-{gamma} production after SEB rechallenge. BALB/c mice were left untreated or were primed with SEB (100 µg/mouse i.p.) and, 24 h or 72 h later, were challenged with the same dose of SEB. Blood samples were taken 90 min later, and serum levels of IFN-{gamma} were determined by enzyme-linked immunosorbent assay. Data presented are representative of three independent experiments and are expressed as means ± standard deviations for at least three individual mice in each experimental group. The levels of IFN-{gamma} were obtained by subtracting the value for IFN-{gamma} control mice rechallenged with phosphate-buffered saline 24 h or 72 h after being primed with SEB.

 
SEB priming induces inhibition of STAT1 activation in the spleen but not in the liver upon secondary SEB or LPS exposure. In spite of the high secretion of IFN-{gamma} observed after a second injection with SEB, signaling through STAT1 is markedly impaired in the spleen, as observed by lowered DNA-binding activity and mRNA expression (42). To ascertain if this inactivation is specific or not for the tissue analyzed, we further determined STAT1 activation in the liver. Livers were obtained at different times after i.p. injection of SEB, and their nuclear extracts were tested for the presence of STAT1 and STAT3 complexes by EMSA. The oligonucleotide used was either the hSIE probe, to detect active complexes containing STAT1 and STAT3, or the Fc{gamma}RI probe, for better detection of the STAT1 homodimers. Supershift assays with antibodies to STAT1 and STAT3 determined the STAT composition of the different bands detected with the oligonucleotides, and their results have been omitted for simplicity (20; data not shown).

As expected, spleen cells from SEB-treated mice displayed a diminished STAT1 response upon rechallenge (Fig. 2A). In contrast, no inhibitory effect was found in liver extracts. The DNA-binding activity of STAT1 complexes in the liver was similarly induced in the primary and secondary responses to SEB (Fig. 2A). Note also that SEB priming selectively down-regulated the STAT1 response in the spleen, while STAT3 was again activated in response to SEB in both the liver and the spleen (Fig. 2A).


Figure 2
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  		FIG. 2. STAT1 element binding activity is inhibited in the spleen but not in the liver in SEB-primed animals upon secondary in vivo challenge with SEB or LPS. (A) Mice which were untreated or SEB primed for 72 h were injected with SEB (100 µg/mouse i.p.). After 90 min, liver and splenic nuclear extracts were analyzed by EMSA, using the Fc{gamma}RI or hSIE probe. The migration of STAT1 and STAT3 is indicated at the left. (B) Mice, with or without SEB pretreatment for 72 h, were treated with LPS (10 µg/mouse i.p.) for 90 min. Liver and splenic nuclear extracts were assayed by EMSA, using the Fc{gamma}RI or hSIE oligonucleotide. The mobilities of STAT1 and STAT3 are depicted by arrows. Data are representative of at least three experiments.

 
To further evaluate if the inhibition of the STAT1 response was specific for the stimulus given, STAT1 activation was analyzed after LPS challenge of SEB-primed mice. LPS is a bacterial compound that also induces the serum release of IFN-{gamma}, mainly by activating the NK cell population (14). In the primary response to LPS, mice displayed DNA-binding activity of STAT1 and STAT3 complexes within 90 min of treatment (Fig. 2B, lane 2). However, the response of STAT1 was induced to a lesser extent in the challenge with LPS than in that with SEB, which is consistent with the lower production of IFN-{gamma} induced by the endotoxin (data not shown). In agreement with the results observed upon SEB restimulation, STAT1 DNA-binding activity was inhibited in spleen but not in liver extracts (Fig. 2B). Moreover, the secondary response to LPS in SEB-primed mice also led to inhibition of the DNA-binding activity of STAT1 but not of STAT3 (Fig. 2B). The reactivation of STAT3 complexes correlates with the increased IL-6 and IL-10 serum production observed upon SEB rechallenge (42) (these cytokines are known to utilize the STAT3 signaling pathway in their intracellular responses [2]). When we further investigated cytokine expression within the tissue, we found that IFN-{gamma} and IL-6 mRNAs were induced in spleens from mice rechallenged with SEB at similar or even higher levels than those in the primary exposure to SEB. This occurred whether the secondary stimulus was SEB or LPS (Fig. 3A and B), in accordance with the serum levels of the cytokines (42).


Figure 3
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  		FIG. 3. SEB priming does not affect the IFN-{gamma} and IL-6 mRNA expression induced by SEB or LPS in spleen cells. (A) Mice were treated with a single injection of SEB (100 µg/mouse i.p.) for the indicated times or were rechallenged for 90 min with a second injection of SEB given 72 h after the first priming dose (lanes 8 to 10). Total mRNA was isolated from spleen cells, and IFN-{gamma} and IL-6 expression was determined by RT-PCR. (B) Mice were treated with a single injection of LPS (10 µg/mouse i.p.) for the indicated times or were rechallenged for 6 h with an injection of LPS given 72 h after SEB priming (lanes 7 to 9). Total mRNA was collected from spleen cells and amplified by RT-PCRs for IFN-{gamma} and IL-6.

 
Taken together, these results describe a specific down-regulation of the STAT1 response in spleens of SEB-primed animals. This down-regulation is non-Vß-specific, as observed in the secondary challenge with either SEB or LPS, and does not correspond with a IFN-{gamma} induction decrease.

Spleen cells from SEB-primed mice are unresponsive to IFN-{gamma} via the STAT1 signaling pathway. We have previously shown that sera from mice treated with SEB for 90 min can induce STAT1 DNA-binding activity in control spleen cells; this activity is blocked when the sera are previously incubated with neutralizing anti-IFN-{gamma} antibodies, demonstrating a direct relationship between IFN-{gamma} production and the activation of the STAT1 response (20). However, sera from mice challenged with SEB for 24 h are unable to activate STAT1 in control spleen cells because IFN-{gamma} has lost its activity (20). To evaluate the possible impairment of IFN-{gamma}'s capacity to induce a STAT1 response in the sera of mice rechallenged with SEB 72 h after SEB priming, control spleen cells were incubated with the sera, and STAT1 DNA-binding activity was analyzed by EMSA. The activation of STAT1 complexes was more strongly induced with the sera from SEB-rechallenged mice than with the sera of mice treated with a single injection of SEB for 90 min (Fig. 4). The increased STAT1 response observed was consistent with the greater serum production of IFN-{gamma} induced in the secondary response to SEB and suggested that the IFN-{gamma} released was indeed functional. To further analyze splenocyte responsiveness to IFN-{gamma} 72 h after SEB priming, spleen cells from SEB-primed mice and control mice were incubated ex vivo with IFN-{gamma} for 30 min, and STAT1 DNA-binding activity was determined by EMSA (Fig. 5). Whereas STAT1 activation was induced in control spleen cells after IFN-{gamma} incubation, it was almost completely abrogated in spleen cells from SEB-primed mice (Fig. 5). These results further confirm the STAT1 inhibition observed in vivo and show a state of IFN-{gamma} unresponsiveness in spleen cells from mice primed with SEB for 72 h.


Figure 4
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  		FIG. 4. Activation of STAT1 by serum. Spleen cells from control mice were incubated in vitro with sera obtained from untreated mice (lanes 1 and 2), from mice treated i.p. with a single injection of 100 µg of SEB for 90 min (lane 3) or 72 h (lane 4), or from mice rechallenged with SEB for 90 min with a second injection given 72 h after the first (lane 5). As a control for STAT1 activation, spleen cells were also incubated with IFN-{gamma} (10 U/ml) (lane 2). After 30 min of incubation, whole-cell extracts were obtained, and STAT1 activation was detected by EMSA using the Fc{gamma}RI probe. The migration of STAT1 is indicated on the left. All experiments were repeated three times with similar results.

 

Figure 5
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  		FIG. 5. In vivo priming with SEB for 72 h desensitizes IFN-{gamma}/STAT1 signaling in spleen cells. Splenocytes from naive mice (lanes 1 to 3) or mice treated i.p. with 100 µg of SEB for 72 h (lanes 4 to 6) were incubated in vitro with IFN-{gamma} (10 U/ml). After 30 min of incubation, whole-cell extracts were obtained, and STAT1 activation was detected by EMSA using the Fc{gamma}RI probe. The migration of STAT1 is indicated on the left. Data are representative of at least three experiments.

 
Inhibition of IFN-{gamma} receptor ß chain and STAT1 mRNA expression in splenic T cells upon in vivo administration of SEB. The in vivo administration of SEB induces the acute release of the cytokines IL-2, IFN-{gamma}, and IL-12 (16, 38), which might lead to the development of a Th1 effector response at around 48 to 72 h, when clonal expansion takes place (28, 36, 45). Th1 populations are characterized as being unresponsive to IFN-{gamma} by down-regulating the mRNA expression of the IFN-{gamma} receptor ß chain, which is required for signaling and for full receptor functionality (23, 48, 53). To investigate whether the inhibition of the IFN-{gamma}/STAT1 signaling pathway observed in spleen cells from SEB-rechallenged mice was due to the absence of any of the signaling cascade components, we analyzed the effects of SEB on the expression of the IFN-{gamma}R ß chain and STAT1 72 h after priming. As shown in Fig. 6A, SEB down-regulated the mRNA expression of the IFN-{gamma}R ß chain in spleen cells 72 h after priming. Furthermore, the expression of the IFN-{gamma}R ß subunit in splenic cells was specifically diminished in the T-cell population (Fig. 6B). Note that splenic T cells down-regulated not only the ß chain but also the expression of Stat1 mRNA, whereas the expression of Stat3 was not affected by priming with SEB (Fig. 6B). The selective down-regulation of the IFN-{gamma}R ß chain and of STAT1 expression observed in T cells 72 h after priming with SEB is consistent with the induction of a Th1 response.


Figure 6
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  		FIG. 6. IFN-{gamma}R ß chain and STAT1 expression is inhibited upon secondary in vivo challenge with SEB in splenic T cells 72 h after the priming injection. (A) Mice which were untreated or primed with SEB for 72 h were challenged with SEB for 90 min or 6 h. Total mRNA was extracted from spleen cells and amplified by RT-PCR for the IFN-{gamma}R ß chain. (B) Mice, with or without SEB priming for 72 h, were treated with SEB for 90 min (100 µg/mouse i.p.). T cells from spleens were purified by MACS (Miltenyi Biotec), and total mRNA was extracted from the T-cell-enriched fraction (lanes 1 to 3) and the T-cell-depleted fraction (lanes 4 to 6). mRNAs were amplified by RT-PCRs for the IFN-{gamma}R ß chain, STAT1, and STAT3. All experiments were repeated three times with similar results.

 

   DISCUSSION
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
Mice treated with SEB experience several acute but transient pathological manifestations, such as postration, pyloerection, diarrhea, hypoglycemia, hypothermia, and weight loss (18, 31, 33). Despite the severity of SEB-induced pathogenesis, it is rarely lethal, and mice resist even high doses of the superantigen. The resistance of mice to a lethal shock and the requirement of a much greater dose of SAgs than that required in humans in order to raise serum levels of proinflammatory cytokines to levels associated with pathogenesis may be the consequence of a weaker binding strength with the cellular receptors for these toxins, as previously reported (10, 31, 54). However, mice become sensitized to a lethal shock when rechallenged after 48 h with the superantigen (17). We elicited a lethal shock as described by Florquin et al. (17) by two consecutive i.p. injections of 100 µg of SEB to obtain clues about the mechanisms involved in the lethal syndrome that follows after SEB rechallenge in SEB-pretreated mice. The administration of lower doses of SEB diminished mortality scores (data not shown).

In agreement with previous observations, our results corroborate the implication of the massive production of IFN-{gamma} in the lethal shock induced in mice after a second challenge with SEB and extend the time window of susceptibility to a rechallenge with the superantigen from 48 (17) to 72 h after the priming treatment. The acquired susceptibility to lethal shock observed correlates in time with the development of the T-cell clonal expansion phase described for mice at around 48 to 72 h after primary exposure to SEB (28, 36, 45). Consistent with these observations, mice were able to resist the second injection of SEB when it was given 24 h after the first, shortly before clonal expansion has been described to begin (28, 36, 45), although the production of IFN-{gamma} was also highly induced. At this time, the secretion of tumor necrosis factor alpha (TNF-{alpha}) is inhibited (35; data not shown). The lack of synergistic effects in the absence of TNF-{alpha} and the presence of anti-inflammatory mediators, such as IL-10 and glucocorticoids, may protect mice from the pathogenic effects of the high IFN-{gamma} production induced upon SEB rechallenge 24 h after the priming injection. Glucocorticoids and IL-10, which are known to protect from SEB-mediated lethal shock (6, 16, 21), are no longer detectable in sera of mice 48 to 72 h after SEB pretreatment (35, 42). Furthermore, the inhibition of TNF-{alpha} production might be temporary, and high cocirculating levels of IFN-{gamma} and TNF-{alpha} could be induced during the T-cell clonal expansion phase upon SEB rechallenge. This could account for the mouse acquisition of susceptibility to lethal shock. Sundstedt et al. (50) found that secondary stimulation with staphylococcal enterotoxin A (SEA) triggers a primed TNF-{alpha} serum release in mice pretreated with SEA for 4 days, and Florquin et al. (17) observed that TNF-{alpha} production was again induced upon SEB rechallenge 48 h after priming with the SAg. The proinflammatory effects of the massive production of IFN-{gamma} together with the synergistic effects of a TNF-{alpha} response might ultimately lead to a lethal syndrome, as they do in other models of inflammation (13, 16, 40). However, we and others have shown that IFN-{gamma} neutralization is sufficient to rescue mice from the lethal shock induced by secondary SEB exposure (17), further supporting the critical implication of IFN-{gamma} in the pathogenesis induced by the SAg. Finally, as predicted, the restimulation of mice with SEB was not lethal when SEB was given 120 h after the priming injection. This was consistent with previous studies showing that the SEB-reactive Vß8+ T-cell population decays to less than basal levels between days 4 and 7, when clonal deletion takes place (27, 30).

We have previously shown that although SEB priming for 72 h induces high serum levels of IFN-{gamma} upon a second exposure to the SAg, the induction of STAT1 DNA-binding activity and mRNA expression in spleen cells is markedly impaired (42). In the present work, we found that the activation of STAT1 was diminished in the spleen but not in the liver, indicating a selective mechanism of IFN-{gamma} unresponsiveness. Moreover, spleen cells from SEB-rechallenged mice did not induce activation of STAT1 complexes when they were stimulated ex vivo with IFN-{gamma}; this was consistent with the in vivo results. The lack of IFN-{gamma} responsiveness correlated with a decrease of the IFN-{gamma}R ß chain mRNA expression on the splenic T-cell population. Previous work demonstrated that T cells at all stages of stimulation and expansion express similar amounts of the IFN-{gamma}R {alpha} chain protein or mRNA, and their ability to respond to IFN-{gamma} is dependent solely on the expression of the IFN-{gamma}R ß chain (48). Abrogation of ß chain expression in T cells represents a hallmark of Th1 populations because Th0/Th2 subsets constitutively express transcripts for the IFN-{gamma}R ß chain (23). Although initial IFN-{gamma}/STAT1 signaling would induce the development of a Th1 response (1), it should later be down-modulated in order to fully display the effector functions (52, 53). Therefore, the T-cell loss of ß chain expression observed 72 h after in vivo SEB priming would be consistent with the development of a Th1 response, as would be expected from the IL-2, IFN-{gamma}, and IL-12 serum release induced during the primary response to SEB (28, 38, 42). It has been postulated that blockade of the IFN-{gamma}/STAT1 signaling pathway would also allow clonal expansion of the Th1 population by preventing the apoptotic effects of STAT1 (48). However, forced expression of the ß chain in transgenic mice does not inhibit clonal expansion of the T cells, despite STAT1 activation (52, 53). IFN-{gamma}-induced apoptosis seems to be strictly dependent on the extent of STAT1 activation (7), and Th1 cells might down-regulate not only the IFN-{gamma}R ß chain but also Stat1 expression, as shown by our results. Blockade of Stat1 expression in the Th1 population would lead to STAT1 activation that could be too low to trigger apoptotic signals. Therefore, not only down-regulation of IFN-{gamma}R ß chain expression but also blockade of STAT1 expression might allow Th1 clonal expansion.

Another consequence of down-regulating the IFN-{gamma}/STAT1 signaling pathway may be the modulation of IFN-{gamma} production during the clonal expansion phase. Indeed, the secondary response to SEB in mice primed for 48 h is characterized by an increased acute production of IFN-{gamma}, which reaches basal levels already after 6 h (38), in contrast with the sustained IFN-{gamma} production observed after a single injection with the superantigen (28, 38, 42). It has been shown that IFN-{gamma} positively induces its own expression (8, 9), and the subsequent secretion of IL-12 from macrophages further amplifies the IFN-{gamma} response to SEB (28, 38). This secretion of IL-12 is transiently inhibited in mice 48 h after receiving the exotoxin (38). Therefore, the IFN-{gamma}R ß chain down-regulation, together with the inhibition of IL-12 production, might limit the IFN-{gamma} response in time in a secondary challenge with SEB during the Th1 effector response.

The rechallenge with SEB 48 h to 72 h after the priming injection not only triggers the secretion of high levels of IFN-{gamma} in mouse serum but also up-regulates levels of IL-10 (17, 39, 42). After a third injection of SEB and after subsequent injections given every other day, IFN-{gamma} is no longer produced (39), whereas IL-10 dominates the response (39, 50) due to the development of a regulatory T-cell population (15, 39, 41) able to suppress SEB-specific primary T-cell responses (15, 39). Induction of the Treg phenotype requires IL-10 (22, 29, 46) and seems to involve activation of STAT3 (26, 56). The different modulation of the IFN-{gamma} and IL-10 signaling pathways might be crucial for the development of active suppression. Although IFN-{gamma} and IL-10 production is primed upon secondary SEB exposure during the T-cell clonal expansion phase, the signaling pathways of these two cytokines were differentially regulated 72 h after SEB priming. Blockade of IFN-{gamma}/STAT1 signaling and persistent signaling through STAT3 may lead to the final establishment of a suppressor T-cell population with a dominant IL-10 response upon repetitive SAg exposure. Furthermore, when we analyzed the mRNA expression of STAT proteins 72 h after SEB priming, we observed that STAT1 was inhibited, whereas the expression of STAT3 was maintained in splenic T cells upon SEB rechallenge. Recent studies have demonstrated that activation through STAT3 can promote T-lymphocyte proliferation and survival after IFN-{alpha}/ß production, but only under conditions where STAT1 expression is abrogated (51). Activation of the STAT3 signaling pathway by IL-10 may therefore have consequences on the survival of effector Th1 cells and differentiation towards a suppressor phenotype upon SEB restimulation, and this may only take place when the STAT1 response is inhibited.

In summary, mice primed with SEB become sensitized to a second challenge during the T-cell clonal expansion phase, and the increased production of IFN-{gamma} from T cells is associated with lethality. Simultaneously, SEB priming for 72 h induces a selective IFN-{gamma}/STAT1 unresponsiveness in spleen cells which correlates with the down-regulation of the IFN-{gamma}R ß chain and Stat1 expression in the T-cell population. Whereas the STAT1 pathway is inhibited, the activation of STAT3 signaling and expression persists upon rechallenge with SEB. The ability of SAgs to differentially modulate the STAT1 and STAT3 signaling pathways in T cells may have profound effects on the initiation and subsequent maintenance of immunosuppression upon chronic exposure to the exotoxin.


   ACKNOWLEDGMENTS
 
This work was supported by grants from the Comision Interministerial de Ciencia y Tecnologia (Plan Nacional I+D, SAF 97/0081) and Comissionat per a Universitats i Recerca (II Pla de Recerca de Catalunya, no. 1997SGR 00352).

We thank Caroline Newey for her help with the preparation of the manuscript. We are also grateful to members of the Immunology Laboratory for their constant support.


   FOOTNOTES
 
* Corresponding author. Mailing address: Department of Immunology, Hospital de la Santa Creu i Sant Pau, Avda. Sant Antoni Maria Claret 167, 08025 Barcelona, Spain. Phone: 3493 2919017. Fax: 3493 2919066. E-mail:cjuarez{at}hsp.santpau.es. Back

{triangledown} Published ahead of print on 30 October 2006. Back

Editor: D. L. Burns


   REFERENCES
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
1. Afkarian, M., J. R. Sedy, J. Yang, N. G. Jacobson, N. Cereb, S. Y. Yang, T. L. Murphy, and K. M. Murphy. 2002. T-bet is a STAT1-induced regulator of IL-12R expression in naive CD4+ T cells. Nat. Immunol. 3:549-557. [Epub ahead of print.][CrossRef][Medline]
2. Akira, S. 2000. Roles of STAT3 defined by tissue-specific gene targeting. Oncogene 19:2607-2611.[CrossRef][Medline]
3. Anderson, P., A. Sundstedt, L. Li, E. J. O'Neill, S. Li, D. C. Wraith, and P. Wang. 2003. Differential activation of signal transducer and activator of transcription (STAT) 3 and STAT5 and induction of suppressors of cytokine signalling in T(h) 1 and T(h) 2 cells. Int. Immunol. 15:1309-1317.[Abstract/Free Full Text]
4. Andrews, M. M., E. M. Parent, M. Barry, and J. Parsonnet. 2001. Recurrent nonmenstrual toxic shock syndrome: clinical manifestations, diagnosis, and treatment. Clin. Infect. Dis. 32:1470-1479.[CrossRef][Medline]
5. Arad, G., D. Hillman, R. Levy, and R. Kaempfer. 2004. Broad-spectrum immunity against superantigens is elicited in mice protected from lethal shock by a superantigen antagonist peptide. Immunol. Lett. 91:141-145.[CrossRef][Medline]
6. Bean, A. G., R. A. Freiberg, S. Andrade, S. Menon, and A. Zlotnik. 1993. Interleukin 10 protects mice against staphylococcal enterotoxin B-induced lethal shock. Infect. Immun. 61:4937-4939.[Abstract/Free Full Text]
7. Bernabei, P., E. M. Coccia, L. Rigamonti, M. Bosticardo, G. Forni, S. Pestka, C. D. Krause, A. Battistini, and F. Novelli. 2001. Interferon-gamma receptor 2 expression as the deciding factor in human T, B, and myeloid cell proliferation or death. J. Leukoc. Biol. 70:950-960.[Abstract/Free Full Text]
8. Boehm, U., T. Klamp, M. Groot, and J. C. Howard. 1997. Cellular responses to interferon-gamma. Annu. Rev. Immunol. 15:749-795.[CrossRef][Medline]
9. Cockfield, S. M., V. Ramassar, J. Noujaim, P. H. van der Meide, and P. F. Halloran. 1993. Regulation of IFN-gamma expression in vivo. FN-gamma up-regulates expression of its mRNA in normal and lipopolysaccharide-stimulated mice. J. Immunol. 150:717-725.[Abstract]
10. Cole, B. C., A. D. Sawitzke, E. A. Ahmed, C. L. Atkin, and C. S. David. 1997. Allelic polymorphisms at the H-2A and HLA-DQ loci influence the response of murine lymphocytes to the Mycoplasma arthritidis superantigen MAM. Infect. Immun. 65:4190-4198.[Abstract]
11. Diehl, S., J. Anguita, A. Hoffmeyer, T. Zapton, J. N. Ihle, E. Fikrig, and M. Rincon. 2000. Inhibition of Th1 differentiation by IL-6 is mediated by SOCS1. Immunity 13:805-815.[CrossRef][Medline]
12. Ding, Y., D. Chen, A. Tarcsafalvi, R. Su, L. Qin, and J. S. Bromberg. 2003. Suppressor of cytokine signaling 1 inhibits IL-10-mediated immune responses. J. Immunol. 170:1383-1391.[Abstract/Free Full Text]
13. Dinges, M. M., and P. M. Schlievert. 2001. Role of T cells and gamma interferon during induction of hypersensitivity to lipopolysaccharide by toxic shock syndrome toxin 1 in mice. Infect. Immun. 69:1256-1264.[Abstract/Free Full Text]
14. Emoto, M., M. Miyamoto, I. Yoshizawa, Y. Emoto, U. E. Schaible, E. Kita, and S. H. Kaufmann. 2002. Critical role of NK cells rather than V alpha 14(+)NKT cells in lipopolysaccharide-induced lethal shock in mice. J. Immunol. 169:1426-1432.[Abstract/Free Full Text]
15. Feunou, P., L. Poulin, C. Habran, A. Le Moine, M. Goldman, and M. Y. Braun. 2003. CD4+CD25+ and CD4+CD25– T cells act respectively as inducer and effector T suppressor cells in superantigen-induced tolerance. J. Immunol. 171:3475-3484.[Abstract/Free Full Text]
16. Florquin, S., Z. Amraoui, D. Abramowicz, and M. Goldman. 1994. Systemic release and protective role of IL-10 in staphylococcal enterotoxin B-induced shock in mice. J. Immunol. 153:2618-2623.[Abstract]
17. Florquin, S., Z. Amraoui, and M. Goldman. 1995. T cells made deficient in interleukin-2 production by exposure to staphylococcal enterotoxin B in vivo are primed for interferon-gamma and interleukin-10 secretion. Eur. J. Immunol. 25:1148-1153.[Medline]
18. Florquin, S., and M. Goldman. 1996. Immunoregulatory mechanisms of T-cell-dependent shock induced by a bacterial superantigen in mice. Infect. Immun. 64:3443-3445.[Medline]
19. Gao, J. J., M. B. Filla, M. J. Fultz, S. N. Vogel, S. W. Russell, and W. J. Murphy. 1998. Autocrine/paracrine IFN-alphabeta mediates the lipopolysaccharide-induced activation of transcription factor Stat1alpha in mouse macrophages: pivotal role of Stat1alpha in induction of the inducible nitric oxide synthase gene. J. Immunol. 161:4803-4810.[Abstract/Free Full Text]
20. Gimeno, R., J. Codony-Servat, M. Plana, J. L. Rodriguez-Sanchez, and C. Juarez. 1996. Stat1 implication in the immune response to superantigens in vivo. J. Immunol. 156:1378-1386.[Abstract]
21. Gonzalo, J. A., A. Gonzalez-Garcia, C. Martinez, and G. Kroemer. 1993. Glucocorticoid-mediated control of the activation and clonal deletion of peripheral T cells in vivo. J. Exp. Med. 177:1239-1246.[Abstract/Free Full Text]
22. Groux, H., A. O'Garra, M. Bigler, M. Rouleau, S. Antonenko, J. E. de Vries, and M. G. Roncarolo. 1997. A CD4+ T-cell subset inhibits antigen-specific T-cell responses and prevents colitis. Nature 389:737-742.[CrossRef][Medline]
23. Groux, H., T. Sornasse, F. Cottrez, J. E. de Vries, R. L. Coffman, M. G. Roncarolo, and H. Yssel. 1997. Induction of human T helper cell type 1 differentiation results in loss of IFN-gamma receptor beta-chain expression. J. Immunol. 158:5627-5631.[Abstract]
24. Hibi, M., A. Lin, T. Smeal, A. Minden, and M. Karin. 1993. Identification of an oncoprotein- and UV-responsive protein kinase that binds and potentiates the c-Jun activation domain. Genes Dev. 7:2135-2148.[Abstract/Free Full Text]
25. Kappler, J., B. Kotzin, L. Herron, E. W. Gelfand, R. D. Bigler, A. Boylston, S. Carrel, D. N. Posnett, Y. Choi, and P. Marrack. 1989. V beta-specific stimulation of human T cells by staphylococcal toxins. Science 244:811-813.[Abstract/Free Full Text]
26. Kasprzycka, M., M. Marzec, X. Liu, Q. Zhang, and M. A. Wasik. 2006. Nucleophosmin/anaplastic lymphoma kinase (NPM/ALK) oncoprotein induces the T regulatory cell phenotype by activating STAT3. Proc. Natl. Acad. Sci. USA 103:9964-9969.[Abstract/Free Full Text]
27. Kawabe, Y., and A. Ochi. 1991. Programmed cell death and extrathymic reduction of Vbeta8+ CD4+ T cells in mice tolerant to Staphylococcus aureus enterotoxin B. Nature 349:245-248.[CrossRef][Medline]
28. Lauw, F. N., S. Florquin, P. Speelman, S. J. van Deventer, and T. van der Poll. 2001. Role of endogenous interleukin-12 in immune response to staphylococcal enterotoxin B in mice. Infect. Immun. 69:5949-5952.[Abstract/Free Full Text]
29. Levings, M. K., R. Sangregorio, C. Sartirana, A. L. Moschin, M. Battaglia, P. C. Orban, and M. G. Roncarolo. 2002. Human CD25+CD4+ T suppressor cell clones produce transforming growth factor beta, but not interleukin 10, and are distinct from type 1 T regulatory cells. J. Exp. Med. 196:1335-1346.[Abstract/Free Full Text]
30. MacDonald, H. R., S. Baschieri, and R. K. Lees. 1991. Clonal expansion precedes anergy and death of V beta 8+ peripheral T cells responding to staphylococcal enterotoxin B in vivo. Eur. J. Immunol. 21:1963-1966.[Medline]
31. Marrack, P., M. Blackman, E. Kushnir, and J. Kappler. 1990. The toxicity of staphylococcal enterotoxin B in mice is mediated by T cells. J. Exp. Med. 171:455-464.[Abstract/Free Full Text]
32. Marrack, P., and J. Kappler. 1990. The staphylococcal enterotoxins and their relatives. Science 248:1066.[Free Full Text]
33. Matthys, P., T. Mitera, H. Heremans, J. Van Damme, and A. Billiau. 1995. Anti-gamma interferon and anti-interleukin-6 antibodies affect staphylococcal enterotoxin B-induced weight loss, hypoglycemia, and cytokine release in D-galactosamine-sensitized and unsensitized mice. Infect. Immun. 63:1158-1164.[Abstract]
34. McCormick, J. K., J. M. Yarwood, and P. M. Schlievert. 2001. Toxic shock syndrome and bacterial superantigens: an update. Annu. Rev. Microbiol. 55:77-104.[CrossRef][Medline]
35. Miethke, T., C. Wahl, K. Heeg, and H. Wagner. 1993. Acquired resistance to superantigen-induced T cell shock. V beta selective T cell unresponsiveness unfolds directly from a transient state of hyperreactivity. J. Immunol. 150:3776-3784.[Abstract]
36. Miethke, T., C. Wahl, K. Heeg, and H. Wagner. 1995. Superantigens: the paradox of T-cell activation versus inactivation. Int. Arch. Allergy Immunol. 106:3-7.[Medline]
37. Moore, K. W., R. de Waal Malefyt, R. L. Coffman, and A. O'Garra. 2001. Interleukin-10 and the interleukin-10 receptor. Annu. Rev. Immunol. 19:683-765.[CrossRef][Medline]
38. Muraille, E., B. Pajak, J. Urbain, M. Moser, and O. Leo. 1999. Role and regulation of IL-12 in the in vivo response to staphylococcal enterotoxin B. Int. Immunol. 11:1403-1410.[Abstract/Free Full Text]
39. Noel, C., S. Florquin, M. Goldman, and M. Y. Braun. 2001. Chronic exposure to superantigen induces regulatory CD4(+) T cells with IL-10-mediated suppressive activity. Int. Immunol. 13:431-439.[Abstract/Free Full Text]
40. Ozmen, L., M. Pericin, J. Hakimi, R. A. Chizzonite, M. Wysocka, G. Trinchieri, M. Gately, and G. Garotta. 1994. Interleukin 12, interferon gamma, and tumor necrosis factor alpha are the key cytokines of the generalized Shwartzman reaction. J. Exp. Med. 180:907-915.[Abstract/Free Full Text]
41. Papiernik, M., M. do Carmo Leite-de-Moraes, C. Pontoux, A. M. Joret, B. Rocha, C. Penit, and M. Dy. 1997. T cell deletion induced by chronic infection with mouse mammary tumor virus spares a CD25-positive, IL-10-producing T cell population with infectious capacity. J. Immunol. 158:4642-4653.[Abstract]
42. Plaza, R., S. Vidal, J. L. Rodriguez-Sanchez, and C. Juarez. 2004. Implication of STAT1 and STAT3 transcription factors in the response to superantigens. Cytokine 25:1-10.[CrossRef][Medline]
43. Proft, T., and J. D. Fraser. 2003. Bacterial superantigens. Clin. Exp. Immunol. 133:299-306.[CrossRef][Medline]
44. Rennick, D. M., M. M. Fort, and N. J. Davidson. 1997. Studies with IL-10–/– mice: an overview. J. Leukoc. Biol. 61:389-396.[Abstract]
45. Renno, T., A. Attinger, S. Locatelli, T. Bakker, S. Vacheron, and H. R. MacDonald. 1999. Cutting edge: apoptosis of superantigen-activated T cells occurs preferentially after a discrete number of cell divisions in vivo. J. Immunol. 162:6312-6315.[Abstract/Free Full Text]
46. Roncarolo, M. G., R. Bacchetta, C. Bordignon, S. Narula, and M. K. Levings. 2001. Type 1 T regulatory cells. Immunol. Rev. 182:68-79.[CrossRef][Medline]
47. Sadowski, H. B., K. Shuai, J. E. Darnell, Jr., and M. Z. Gilman. 1993. A common nuclear signal transduction pathway activated by growth factor and cytokine receptors. Science 261:1739-1744.[Abstract/Free Full Text]
48. Sakatsume, M., and D. S. Finbloom. 1996. Modulation of the expression of the IFN-gamma receptor beta-chain controls responsiveness to IFN-gamma in human peripheral blood T cells. J. Immunol. 156:4160-4166.[Abstract]
49. Shapiro, D. J., P. A. Sharp, W. W. Wahli, and M. J. Keller. 1988. A high-efficiency HeLa cell nuclear transcription extract. DNA 7:47-55.[Medline]
50. Sundstedt, A., I. Hoiden, A. Rosendahl, T. Kalland, N. van Rooijen, and M. Dohlsten. 1997. Immunoregulatory role of IL-10 during superantigen-induced hyporesponsiveness in vivo. J. Immunol. 158:180-186.[Abstract]
51. Tanabe, Y., T. Nishibori, L. Su, R. M. Arduini, D. P. Baker, and M. David. 2005. Cutting edge: role of STAT1, STAT3, and STAT5 in IFN-alpha beta responses in T lymphocytes. J. Immunol. 174:609-613.[Abstract/Free Full Text]
52. Tau, G. Z., S. N. Cowan, J. Weisburg, N. S. Braunstein, and P. B. Rothman. 2001. Regulation of IFN-gamma signaling is essential for the cytotoxic activity of CD8(+) T cells. J. Immunol. 167:5574-5582.[Abstract/Free Full Text]
53. Tau, G. Z., T. von der Weid, B. Lu, S. Cowan, M. Kvatyuk, A. Pernis, G. Cattoretti, N. S. Braunstein, R. L. Coffman, and P. B. Rothman. 2000. Interferon gamma signaling alters the function of T helper type 1 cells. J. Exp. Med. 192:977-986.[Abstract/Free Full Text]
54. Yeung, R. S., J. M. Penninger, T. Kundig, W. Khoo, P. S. Ohashi, G. Kroemer, and T. W. Mak. 1996. Human CD4 and human major histocompatibility complex class II (DQ6) transgenic mice: supersensitivity to superantigen-induced septic shock. Eur. J. Immunol. 26:1074-1082.[Medline]
55. Zhong, Z., Z. Wen, and J. E. Darnell, Jr. 1994. Stat3: a STAT family member activated by tyrosine phosphorylation in response to epidermal growth factor and interleukin-6. Science 264:95-98.[Abstract/Free Full Text]
56. Zorn, E., E. A. Nelson, M. Mohseni, F. Porcheray, H. Kim, D. Litsa, R. Bellucci, E. Raderschall, C. Canning, R. J. Soiffer, D. A. Frank, and J. Ritz. 2006. IL-2 regulates FOXP3 expression in human CD4+CD25+ regulatory T cells through a STAT dependent mechanism and induces the expansion of these cells in vivo. Blood 27:27.

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