HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Related articles in The JI Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Guo, Z. Articles by Lederer, J. A. Search for Related Content PubMed PubMed Citation Articles by Guo, Z. Articles by Lederer, J. A.

Burn Injury Promotes Antigen-Driven Th2-Type Responses In Vivo ¹

Department of Surgery, Brigham and Women’s Hospital/Harvard Medical School, Boston, MA 02115

	Abstract

Top Abstract Introduction Materials and Methods Results Discussion References

 
Severe injury induces detrimental changes in immune function, often leaving the host highly susceptible to developing life-threatening opportunistic infections. Advances in our understanding of how injury influences host immune responses suggest that injury causes a phenotypic imbalance in the regulation of Th1- and Th2-type immune responses. We report in this study, using a TCR transgenic CD4⁺ T cell adoptive transfer approach, that injury skews T cell responses toward increased Th2-type reactivity in vivo without substantially limiting Ag-driven CD4⁺ T cell expansion. The increased Th2-type response did not occur unless injured mice were immunized with specific Ag, suggesting that the phenotypic switch is Ag dependent. These findings establish that severe injury induces fundamental changes in the induction of Ag-specific CD4⁺ Th cell responses favoring the development of Th2-type immune reactivity in vivo.

	Introduction

Top Abstract Introduction Materials and Methods Results Discussion References

 
The mammalian immune system responds to injury by rapidly producing proinflammatory cytokines and other mediators of acute inflammation. After this initial inflammatory response, a compensatory anti-inflammatory response ensues. Although this response scenario may have evolved as a means to protect the injured host from the harmful effects of injury-induced inflammation, many of the mediators of this type of counterinflammatory response also have strong immune suppressive activity (1, 2, 3, 4). Consequently, clinical observations along with numerous studies in animal models suggest that injury often leads to a transient state of immune suppression that predisposes the injured host to infections caused by opportunistic pathogens (5, 6, 7, 8). This effect of injury on host immunity remains a significant clinical problem today.

Research done using animal models for injury has shown that interfering with the counterinflammatory response to injury by promoting proinflammatory Th1 reactivity can prevent the development of injury-induced immune suppression. In particular, it has been shown that treating mice with Th1-inducing agents such as IL-12, anti-IL-6, or anti-IL-10 Abs can restore Th1-type responses in injured mice and improve their ability to survive polymicrobial sepsis (9, 10, 11, 12, 13, 14). As a group, these in vivo studies strongly suggest that injury causes suppressed Th1-type responses. In agreement with this hypothesis, we have shown that burn injury suppresses Th1-mediated Ab isotype switching in mice that were immunized with a T cell-dependent Ag at the time of injury (15). These studies also demonstrated that injury prevented the Ag-stimulated expansion of IFN--producing T cells, providing more evidence that injury inhibited Th1 responses in vivo. Moreover, treatments that enhance resistance to bacterial or viral infections after injury also restored Th1-type responses (15, 16). The findings from these studies and from clinical observations have reinforced the idea that suppressed Th1-type responses occurring after severe injury might be involved in the development and maintenance of injury-induced immune suppression.

To study the mechanisms involved in the injury-induced perturbations in T cell-mediated responses in vivo, we have chosen to track the differentiation and expansion of Ag-stimulated CD4⁺ Th cells using a technique that was first described by Kearney et al. (17). This involves the adoptive transfer of CD4⁺ T cells from DO-11.10 TCR transgenic mice into wild-type mice to measure sequentially the Ag responsiveness of a defined population of T cells in a nontransgenic environment. The DO-11.10 TCR recognizes the 323–339 aa peptide sequence of OVA (OVA_323–339) in a MHC class II-restricted fashion (18). When immunized with OVA_323–339 peptide, the transferred T cells undergo a primary T cell response that can be tracked using a mAb specific for the DO-11.10 TCR (clone KJ1-26) (17). In addition, peripheral lymphoid tissues can be harvested from recipient mice and stimulated ex vivo with OVA_323–339 peptide to phenotype the transferred T cells by Ag-induced cytokine production profiles. These features of this experimental approach allowed us to accurately study influences that injury may have on the expansion and differentiation of Ag-stimulated Th cells. In this report, we demonstrate that injury did not significantly limit the Ag-driven expansion of CD4⁺ T cells in peripheral lymphoid compartments of immunized mice. Instead, injury caused an Ag-dependent increase in Th2-type T cell responses. Therefore, it appears that severe injury modulates adaptive immunity by promoting a phenotypic switch toward increased Th2-type immune reactivity in vivo.

	Materials and Methods

Top Abstract Introduction Materials and Methods Results Discussion References

 
Animals

Five-week-old male BALB/c mice were purchased from The Jackson Laboratory (Bar Harbor, MA). The DO-11.10 TCR transgenic mice were originally obtained from Dr. K. Murphy (Washington University, St. Louis, MO) and maintained by standard breeding regimes (19). Mice were screened for the DO-11.10 transgene by PCR of tail DNA samples using transgene-specific primers: forward-CAGGAGGGATCCAGTGCCAGC, reverse-TGGCTCTACAGTGAGTTTGGT. Cell-surface DO-11.10 TCR expression was confirmed in each adoptive transfer study by two-color FACS analysis using PE-labeled anti-CD4 and FITC-labeled KJ1-26 mAb specific for the DO-11.10 TCR (Caltag Laboratories, Burlingame, CA) (20). Purchased mice were acclimated for at least 1 wk before use for experiments at 6–7 wk of age. Age-matched male DO-11.10 mice were used as donors for adoptive transfer studies. All mice were maintained in our accredited virus Ab-free animal facility in accordance with guidelines of the National Institutes of Health and the Harvard Medical School Standing Committee on Animal Research.

Reagents

Chicken OVA peptide, aa 323–339 sequence ISQAVHAAHAEINEAGR, was purchased from Research Genetics (Huntsville, AL). CFA, saponin, brefeldin A, and chemical reagents for ELISA were purchased from Sigma-Aldrich (St. Louis, MO). mAbs and reagents for FACS staining were obtained from the following sources: FITC-conjugated KJ1-26 mAb was from Caltag Laboratories; PE-conjugated mAb specific for mouse CD4, biotinylated mAb specific for IL-2, IFN-, IL-4, IL-10, and streptavidin-CyChrome were purchased from BD PharMingen (San Diego, CA). ELISA Abs specific for mouse cytokines were purchased from BD PharMingen, Caltag Laboratories, or R&D Systems (Minneapolis, MN). Culture medium referred to as complete-5 was prepared by supplementing RPMI 1640 with 5% heat-inactivated FCS, 1 mM glutamine, 1 mM sodium pyruvate, 100 µM nonessential amino acids, 10 mM HEPES, penicillin/streptomycin/fungizone, and 2.5 x 10^-5 M 2-ME, all purchased from Life Technologies (Grand Island, NY).

Mouse injury model

Mice were injured using a well-established thermal injury protocol that has been approved by the National Institutes of Health and Harvard Medical School Standing Committee on Animal Research. Before receiving sham or thermal injury, mice were randomized and anesthetized by i.p. injection with 70 mg/kg of pentobarbital obtained from Abbott Laboratories (Chicago, IL). Once fully anesthetized, the dorsum was shaved, and mice were placed in a plastic mold that exposed 25% of their total body surface area (TBSA). ³ The mice were then subjected to scald thermal injury of the exposed skin by immersion in 90°C water for 9 s. This treatment has been shown to cause full-thickness thermal injury (and thus an anesthetic injury) with a low mortality (<5%) (21). In our hands, the injured mice show a slight reduction in weight gain, but will survive long term unless given an infectious challenge such as cecal ligation and puncture (21). Sham mice were treated in the same fashion, except that they were exposed to room-temperature (24°C) water rather than 90°C water. Both sham- and thermal-injured mice were resuscitated by i.p. injection with 1 ml of sterile, normal saline solution.

Adoptive transfer

Inguinal, axillary, and brachial lymph nodes and spleens were harvested from 6- to 8-wk-old male DO-11.10 mice. Single-cell suspensions were prepared by mincing the tissue through a stainless steel mesh. Cells were washed three times in C-5, and then counted. The percentage of CD4⁺, DO-11.10 TCR transgenic T cells in the preparation was measured by two-color FACS analysis by staining 5 x 10⁵ cells with PE-labeled anti-CD4 mAb and FITC-labeled anti-DO-11.10 TCR (KJ1-26) mAb. A total of 2.5 x 10⁶ DO-11.10 transgenic T cells were suspended in 0.2 ml of PBS, and then transferred by intracardiac injection into anesthetized recipient age- and sex-matched BALB/c mice. While confirming our ability to use this method, we found that transferring cells into mice by cardiac injection rather than by the tail vein or retro-orbital injection routes resulted in much less mouse to mouse adoptive transfer variability. Although there was a slight increase in mortality (5–10%), all surviving mice had similar levels of DO-11.10 T cells in lymph nodes and spleens at 1 day after transfer (our unpublished observations). Recipient BALB/c mice were immunized at the time of sham or thermal injury with 10 µg of OVA_323–339 emulsified 1:1 in CFA (Sigma-Aldrich) by s.c. injection at a single site on the anterior abdomen.

Cell preparation and ex vivo stimulation

Mice were sacrificed by CO₂ asphyxiation. Spleens and lymph nodes were removed and then minced in C-5 medium to prepare single cell suspensions. RBC were lysed using tris-buffered ammonium chloride solution, and the spleen cells were then washed twice in C-5. Cells were counted using a hemacytometer and plated in 96-well U bottom plates (Corning Costar, Cambridge, MA) at a density of 5 x 10⁵ cells in the absence or presence of 1 µg/ml OVA_323–339 peptide. This dose of OVA peptide was used because it provided a maximal stimulatory response based upon the results of prior titration studies. After a 48-h incubation at 37°C in 7% CO₂, supernatants were harvested for subsequent ELISA analysis.

Cytokine ELISA

Cytokine ELISAs were used to detect IL-2, IFN-, IL-4, and IL-10 levels in culture supernatants. They were performed using the sandwich technique as described previously. Serial dilutions of cytokines standards were purchased from the Genzyme (Cambridge, MA), and the unknown samples were added to individual wells in triplicate. Upon completion of the ELISA protocol, an ELISA plate reader (Molecular Devices, Sunnyvale, CA) and its accompanying computer software program, SOFTmax PRO version 1.1, was used to analyze the results.

Intracellular cytokine detection

At 1 or 7 days after injury and immunization, cells were prepared from inguinal, axillary, and brachial lymph nodes and stimulated ex vivo with or without 1 µg/ml OVA_323–339 peptide in U-bottom 96-well tissue-culture plates at a concentration of 5 x 10⁵ cells per well. For the last 4 h of a 48-h incubation period, brefeldin A (Sigma-Aldrich) was added to cultures at a final concentration of 10 µg/ml to block cytokine release. The staining procedure was then initiated by washing the cells once with 100 µl of buffer containing 1% BSA and 0.1% sodium azide in PBS (pH 7.4) (PBA) by centrifugation of the 96-well plate. FC block (BD PharMingen) was added (2 µg/ml in 25 µl) for 30 min incubation at 4°C to reduce nonspecific Ab binding. After washing by centrifugation with PBA, the cells were surface stained at 4°C for 20 min with PE-labeled anti-CD4 mAb (BD PharMingen) and FITC-labeled KJ1-26 Ab (Caltag Laboratories). Cells were washed once by centrifugation and then fixed for exactly 20 min with 100 µl of 2% paraformaldehyde in PBS (pH 7.4) at 4°C. Following fixation, cells were washed once by centrifugation with PBA, then permeabilized in 100 µl of buffer containing 0.1% saponin, 1% BSA, and 0.1% sodium azide in PBS (pH 7.4) (permeabilization buffer). Cytokine staining was done by first pretreating the fixed and permeabilized cells with 25 µl of 1 µg/ml solution of normal rat IgG (Caltag Laboratories). After 30 min, PE-labeled anti-cytokine Abs specific for IL-2, IFN-, IL-4, or IL-10 (BD PharMingen) were added (25 µl of a 1 µg/ml solution) for an additional 30 min. Cells were washed twice with permeabilization buffer by centrifugation, then resuspended in 100 µl of PBS (pH 7.4). A FACSCaliber flow cytometer (BD Biosciences, San Jose, CA) was used to detect fluorescent staining cells, and the accompanying CellQuest computer software was used for FACS analysis.

	Results

Top Abstract Introduction Materials and Methods Results Discussion References

 
Injury effects on the expansion of Ag-driven CD4⁺ T cells in vivo

Because it is known that injury can alter T cell-mediated responses in vivo, we wanted first to determine whether burn injury might affect the normal expansion of Ag-activated CD4⁺ T cells. To address this question, we used an adoptive transfer approach that allows direct visualization of transgenic CD4⁺ T cells with known Ag specificity in vivo. As originally described by Kearney et al., transferring OVA_323–339 peptide-specific DO-11.10 TCR-transgenic T cells into syngeneic BALB/c mice followed by s.c. immunization with OVA_323–339 peptide causes significant in vivo expansion of the transferred DO-11.10 T cells. The unique anti-DO-11.10 TCR Ab, KJ1-26, then makes it possible to accurately detect adoptively transferred DO-11.10 T cells in the lymph nodes and spleen of OVA_323–339-immunized mice.

Although other research groups have used this approach, we performed several control studies to verify our ability to use the DO-11.10 transgenic T cell adoptive transfer technique. First, we determined whether we were able to specifically detect the expansion of adoptively transferred DO-11.10 CD4⁺ T cells in OVA_323–339 peptide-immunized recipient BALB/c mice. As illustrated in Fig. 1, significant expansion of DO-11.10 T cells was observed when BALB/c mice received DO-11.10 T cells by adoptive transfer and were immunized with OVA_323–339 peptide, whereas no KJ1-26⁺-staining CD4 T cells were detected in BALB/c mice that were given BALB/c T cells. We next tested whether BALB/c mice could directly respond to immunization with OVA_323–339 peptide emulsified 1:1 in CFA. This control experiment was performed to convince us that measured changes in OVA_323–339 peptide-specific CD4⁺ T cell reactivity were primarily dependent on the adoptively transferred TCR transgenic DO-11.10 T cells. Surprisingly, we observed a slight OVA_323–339 peptide-specific T cell response by immunized BALB/c mice as judged by OVA_323–339 peptide-stimulated IL-2 production by spleen and lymph node cells harvested 7 days after immunization (Table I). Burn injury did not alter this background IL-2 reactivity, and most importantly we did not detect any OVA_323–339-stimulated IFN-, IL-4, or IL-10 production (Table I). Furthermore, the IL-2 production levels observed in these control studies were >100-fold less than those observed when mice received DO-11.10 T cells by adoptive transfer (Fig. 3, A and B). The extremely low level IL-2 response, the lack of IFN-, IL-4, and IL-10 production, and the observation that burn injury had no effect on the IL-2 response convinced us that this background OVA_323–339 peptide response would not interfere with the OVA_323–339 peptide response measured in the adoptive transfer studies.

View larger version (20K): [in this window] [in a new window]   FIGURE 1. The expansion and detection of OVA323–339-specific DO-11.10 T cells in recipient BALB/c mice immunized with OVA323–339 peptide. Four BALB/c mice received BALB/c or DO-11.10 TCR transgenic T cells by intracardiac injection and were immunized s.c. with 10 µg of OVA323–339 peptide emulsified 1:1 in CFA. At 7 days after adoptive transfer and immunization, lymph node cells were prepared and stained with PE-anti-CD4 and FITC-KJ1-26 Abs. Stained cells were detected and analyzed by FACS. The number in the upper right quadrant indicates the percent CD4+ T cells expressing the DO-11.10 TCR in this representative FACS plot.

View larger version (23K): [in this window] [in a new window]   FIGURE 3. The influence of injury on in vivo Ag-stimulated CD4+ T cell expansion. Groups of BALB/c mice receiving DO-11.10 transgenic T cells by adoptive transfer (Materials and Methods) underwent sham or burn injury, and were immunized s.c. with 10 µg of OVA323–339 peptide emulsified 1:1 in CFA. At 1, 3, or 7 days after injury and immunization, lymph node and spleen cells were prepared and stained for two-color FACS analysis with PE-labeled anti-CD4 and FITC-labeled KJ1-26 Abs. A, Representative FACS plots illustrating CD4+ DO-11.10 TCR transgenic T cells present in the lymph node and spleen of sham- or burn-injured mice at 7 days following injury and immunization. The numbers in the upper right quadrant indicate the percentage of KJ1-26-positive staining cells present in the FACS-gated CD4+ cell population. B, Plot representing the levels DO-11.10 CD4+ T cells detected in the lymph nodes and spleens of OVA323–339-immunized DO-11.10 T cell recipient BALB/c mice at 1, 3, and 7 days after sham or burn injury. The results are plotted as the mean ± SEM of three experiments using four mice per group. C, Plot showing the levels of DO-11.10 CD4+ T cells detected in the lymph nodes (LN) and spleens of nonimmunized recipient BALB/c mice at 7 days after adoptive transfer and sham or burn injury. The results represent the mean ± SEM of two experiments using three individual mice per experiment.

View larger version (32K): [in this window] [in a new window]   FIGURE 2. The influence of injury on Th1- and Th2-type cytokine production profiles by adoptively transferred DO-11.10 TCR transgenic T cells. Groups of BALB/c mice that received DO-11.10 TCR transgenic T cells by adoptive transfer (Materials and Methods) underwent sham or burn injury and were immunized s.c. with 10 µg of OVA323–339 peptide emulsified 1:1 in CFA. At 1, 3, or 7 days after injury and immunization, lymph node (A) and spleen (B) cells were harvested, then cultured in the absence or presence of 1 µg/ml OVA323–339 peptide. Supernatants were harvested 48 h later and tested for IL-2, IFN-, IL-4, or IL-10 levels by ELISA. The results are plotted as the mean ± SEM of three experiments using four mice per group. The asterisk (*) indicates significant differences between sham- and burn-injured groups, p < 0.05, using a paired t test.

We next determined whether injury influences the phenotype of Ag-stimulated CD4⁺ T cells in vivo. To accomplish this, lymph node and spleen cells harvested from sham- or burn-injured mice were cultured with OVA_323–339 peptide, and supernatants were tested for IL-2, IFN-, IL-4, or IL-10 levels by ELISA. At 1 day after injury and immunization, we measured an injury-induced increase in IL-2 and IFN- levels produced by OVA_323–339-stimulated lymph node cells. At this same time point, burn injury did not affect Th2-type cytokine production by OVA_323–339 peptide-stimulated lymph node cells, and injury did not affect OVA_323–339-stimulated Th1-type cytokine production by spleen cells, but IL-4 production was significantly reduced (Fig. 3A).

By 3 days after injury and immunization, we detected significant reductions in IL-2 and IFN- production by OVA_323–339 peptide-stimulated lymph node cells (Fig. 3A) occurring in the absence of significant changes in IL-4 or IL-10 production. This findingsuggests that Th1-type reactivity is suppressed as early as 3 days after injury without a concomitant change in Th2-type reactivity. In contrast, OVA_323–339-stimulated spleen cell IL-2 and IFN- was not affected by burn injury at 3 days after injury, but IL-4 production was increased (Fig. 3B). This finding indicates that there is a significant tissue-compartmentalized effect of injury on the development of T cell responses.

The burn-induced reduction in OVA_323–339-stimulated IL-2, and IFN- production by lymph node cells became even more apparent by 7 days after injury (Fig. 3A). Furthermore, by 7 days after injury we detected a significant increase in the production of the Th2-type cytokines, IL-4 and IL-10, by OVA_323–339-stimulated lymph node and spleen cells harvested from burn- vs sham-injured mice (Fig. 3, A and B). Taken together, these observed changes in cytokine production profiles demonstrate that injury causes early enhanced Th1 reactivity in the lymph nodes draining the injury site and a later phenotypic switch defined by suppressed Th1 and increased Th2-type cytokine production.

Ag dependence of injury-induced enhanced Th2-type responses

The initiation of Ag-specific Th2-type responses usually requires TCR stimulation. To test whether Ag exposure is required for the in vivo injury-induced increase in Th2-type reactivity, we performed similar adoptive transfer studies in mice that were not immunized with OVA_323–339 peptide. At 1 day following injury, we observed significant increases in IL-2 and IFN- production by OVA_323–339 peptide-stimulated lymph node and spleen cells, without increased production of the Th2-type cytokines, IL-4 and IL-10 (Fig. 4A). By 7 days after injury, we observed decreased OVA_323–339-induced IL-2 and IFN- production by spleen cells along with decreased IFN- production by lymph node cells from burn-injured mice (Fig. 4B). However, we did not detect any OVA_323–339-induced IL-4 or IL-10 production by lymph node or spleen cells prepared from these unimmunized sham- or burn-injured mice that received DO-11.10 T cells by adoptive transfer. This finding suggests that the injury biasing effect on the development of Th2-type response is Ag dependent. Moreover, these findings support the concept that injury can enhance Th1-type reactivity at 1 day after injury.

View larger version (23K): [in this window] [in a new window]   FIGURE 4. Injury effects on DO-11.10 T cell cytokine production in unimmunized recipient mice. BALB/c mice received DO-11.10 TCR transgenic T cells by adoptive transfer (Materials and Methods), then underwent sham or burn injury. At 1 or 7 days after injury, lymph node and spleen cells were prepared, then stimulated in vitro with 1 µg/ml OVA323–339 peptide. Supernatants were harvested 48 h later and tested for IL-2, IFN-, IL-4, or IL-10 levels by ELISA. A, OVA323–339 peptide stimulated cytokine production at 1 day after sham or burn injury. The results are representative of three experiments using three mice per group. B, OVA323–339 peptide induced cytokine production at 7 days after sham or burn injury. The results are plotted as the mean ± SEM cytokine levels from two experiments using three mice per group.

Because we had demonstrated that injury induces a late enhancement in the production of Th2-type cytokines in an Ag-dependent fashion, we wanted to determine whether the Th2 cytokines were being made directly by T cells and not by the secondary stimulation of other cells present in culture. Thus, we attempted to use intracellular cytokine staining to determine the cellular source of OVA_323–339 peptide-induced cytokines. However, detection of intracellular cytokines in transferred OVA_323–339 peptide stimulated DO-11.10 T cells proved to be difficult because of low cell numbers. Therefore, we used intact DO-11.10 TCR transgenic mice for these studies, because the percentage of T cells that can be stimulated and tested for cytokine production is at least 10- to 20-fold higher. A prior study examining OVA_323–339 peptide responses in immunized DO-11.10 TCR transgenic mice demonstrated that DO-11.10 mice die when immunized with the same dose of OVA_323–339 used in our adoptive transfer studies. Thus, sham- and burn-injured DO-11.10 mice were immunized with a 10-fold lower concentration of OVA_323–339 peptide for these studies. Preliminary studies designed to optimize the intracellular cytokine staining technique demonstrated that we were able to detect intracellular IL-2, IFN-, IL-4, and IL-10 by two-color FACS in 48-h OVA_323–339 stimulated DO-11.10 T cells using PE-labeled anti-cytokine Abs and FITC-conjugated KJ1-26 Ab (Fig. 5A).

View larger version (24K): [in this window] [in a new window]   FIGURE 5. Assessment of injury effects on OVA323–339 peptide stimulated intracytoplasmic cytokine expression profiles. Groups of male DO-11.10 TCR transgenic mice underwent sham or burn injury and were immunized s.c. with 1 µg of OVA323–339 peptide in CFA. After 7 days, lymph node cells were prepared and cultured in the absence or presence of 1 µg/ml of OVA323–339 peptide for 48 h. Cells were stained for the indicated intracytoplasmic cytokines as described (Materials and Methods). A, A representative FACS plot of OVA323–339 peptide stimulated IL-2, IFN-, IL-4, and IL-10 expression in gated, KJ1-26+ staining cells harvested from sham or burn DO-11.10 mice. The quadrants for all FACS analyses were set relative to Ab isotype-stained controls. No intracytoplasmic cytokine staining was detected in unstimulated cultures. B, Intracytoplasmic cytokine staining results obtained at 1 day after injury and immunization of DO-11.10 mice. The plots illustrate the mean ± SEM percent cytokine staining levels from two experiments using three mice per group. The asterisk (*) indicates a significant difference between sham- and burn-injured IL-2 groups, p < 0.05 by a paired t test. C, Intracytoplasmic cytokine staining results obtained at 7 days after injury and immunization of DO-11.10 TCR transgenic mice. The plots illustrate the mean ± SEM percent cytokine staining levels from four experiments using three mice per group. The asterisk (*) indicates significant differences between sham- and burn-injured groups, p < 0.05 by a paired t test.

	Discussion

Top Abstract Introduction Materials and Methods Results Discussion References

 
Although injury is often thought of as an inflammatory host response, and thus driven by cells and mediators of the innate immune system, there is considerable experimental and clinical evidence indicating that injury can profoundly suppress host adaptive immune function (7, 22). Coincident with changes in adaptive immune function, the injured host can become highly susceptible to microbial infections (5, 23). For this reason, we initiated this study to provide a better understanding of how injury influences the development of adaptive immune responses in vivo, and to improve our limited understanding of how injury influences Ag-specific CD4⁺ T cell responses.

One unique feature of this study is that we used an animal model to provide a realistic view of how the mammalian immune system copes with severe injury. We use this injury model, because it demonstrates a low injury-induced mortality rate (<5%), yet causes many of the immunological changes associated with after-injury immune dysfunction. In fact, we and other research groups have demonstrated that burn-injured mice will survive long term, unless they are given a microbial or viral challenge at a time after injury when they are immune suppressed (21, 24, 25). The burn injury site is well demarcated, covering 25% of the TBSA. In our hands, using a >25% TBSA injury increases mortality significantly. An additional advantage of using this experimental model is that burn injury is full-thickness, and thus destroys the nerves at the injury site, making it more humane than other injury or infection animal models. Most important, early work using this mouse model showed that it mimicked many of the observations associated with immune dysfunction in severely injured patients or in patients that underwent major surgery. These observations included suppressed T cell proliferation in response to polyclonal stimulation, reduced IL-2 and IFN- production, and increased production of the Th2-type cytokines IL-4 and IL-10 by stimulated T cells, altered Ab responses, and reduced skin delayed-type hypersensitivity responses (5, 21, 26, 27, 28, 29, 30, 31, 32). Thus, we believe the mouse thermal injury model represents a valid and controlled approach to studying the host response to injury.

Although suppressed Th1-type responses have been shown to occur following burn injury in mice, it was not clear whether injury caused a phenotypic switch in the Th cell response or simply blocked T cell activation and expansion. To address this fundamental issue, we used an adoptive transfer approach that allowed us to directly compare the activation of naive CD4⁺ T cells with defined Ag specificity in immunized sham- vs burn-injured mice. After refining the DO-11.10 T cell adoptive transfer approach, we wanted to use it to determine whether injury significantly altered the expansion of CD4⁺ T cells in vivo. Comparing DO-11.10 T cell levels in lymph node and spleen cell suspensions harvested from sham- and burn-injured mice at early time points (1 and 3 days) after immunization showed no significant injury effect on the expansion or deletion of DO-11.10 T cells. This indicates that burn injury neither influenced the efficiency of the adoptive transfer procedure nor markedly altered the early OVA_323–339 peptide-induced CD4⁺ T cell proliferation in vivo. Although this finding may seem mundane, it suggests that injury does not act as an adjuvant to boost T cell proliferation following immunization, and it does not enhance Ag-induced T cell death. In contrast, by 7 days after injury and immunization, we observed a slight but not statistically significant reduction in the percentage of DO-11.10 CD4⁺ T cells detected in the lymph nodes of burn-injured mice. Interestingly, we observed a similar reduction in the level of transferred DO-11.10 CD4⁺ T cells in the lymph nodes of nonimmunized burn mice suggesting that injury might alter T cell maintenance or survival in the peripheral lymph nodes of mice by both Ag-dependent (activation-induced cell death) and independent (programmed cell death) mechanisms. Collectively, these findings provide the first direct evidence to suggest that the suppressive effect that injury has on adaptive immune reactivity cannot be simply explained by a substantial reduction in Ag-induced T cell expansion or deletion.

A major objective of this study was to define how injury influences the development of Th cell subset responses. This is an important question, because regulation of Th1- and Th2-type responses against infectious pathogens can markedly affect host survival (33). In general, a strong Th1-type response can provide protection from infections caused by most sepsis-causing pathogens by boosting innate immune cell microbicidal function and by promoting Ab responses associated with strong complement-fixing activity. We decided to use the TCR transgenic T cell adoptive transfer approach, because we believed it offered the best available method for studying Th cell subset differentiation in vivo (34, 35). In this series of experiments, BALB/c mice were given DO-11.10 TCR transgenic T cells, immunized s.c. with OVA_323–339 peptide, and exposed to sham or burn injury conditions. At 1, 3, or 7 days after injury, lymph node or spleen cell preparations were then tested for ex vivo OVA_323–339-stimulated IL-2, IFN-, IL-4, and IL-10 production. The early time points were included to help us determine when injury may elicit a phenotypic switch in Ag-induced Th1- and Th2-type cytokine production. Our findings on 1 day after injury demonstrated that injury did not cause an early increase in Th2-type responses, because the levels of IL-4 and IL-10 production were similar between sham- and burn-injured mice. Instead, we observed enhanced IL-2 and IFN- production by OVA_323–339-stimulated lymph node cells, suggesting that injury caused an early increase in Th1-type reactivity. This enhanced Th1-type reactivity at 1 day following injury was even more dramatic when mice were not immunized (Fig. 4). This observation is consistent with previous studies demonstrating that injury can augment Th1-type cytokine responses at 3 h or 1 day after injury as judged by bacterial superantigen reactivity (36). These findings also suggest that injury acts as an adjuvant to enhance early Th1-type responses in vivo. Furthermore, the adjuvant activity that injury may have on T cell responses supports the concept that the immune system mounts an inflammatory response to injured cells and tissues, thus supporting a role for "danger" in modulating adaptive immune responses (37, 38).

In contrast, by 3 days after injury we observed a significant reduction in IL-2 and IFN- being produced by OVA_323–339-stimulated lymph node cells, even though the level of DO-11.10 T cell expansion was similar between sham- and burn-injured mice at this time point after injury (Fig. 2B). Interestingly, this reduction in Th1-type cytokine production observed in the lymph nodes at 3 days after injury was not associated with an increase in OVA_323–339 peptide-induced IL-4 and IL-10 production. Instead, we observed lower levels of IL-4 and IL-10 produced by OVA_323–339 peptide-stimulated lymph node cells from both sham- and burn-injured mice. Thus, it appears that suppressed Th1-type reactivity may precede a phenotypic switch toward a Th2-type response. By 7 days after injury, Th1-type cytokine production by burn lymph node cells remained suppressed, while IL-4 and IL-10 production by OVA_323–339 peptide-stimulated lymph node and spleen cells was increased. Taken collectively, these findings suggest that injury causes a time-dependent skewing in Ag-induced CD4⁺ T cell responses toward increased Th2-type responses and suppressed Th1-type reactivity.

The observation that burn injury causes a phenotypic shift in Ag-specific CD4⁺ T cell differentiation toward increased Th2-type and decreased Th1-type responses indicates that injury provides a signal that skews T cell activation and differentiation. Similar type phenotypic switches have been observed in other studies using the TCR transgenic T cell adoptive transfer approach. In particular, BALB/c mice that received DO-11.10 TCR transgenic cells by adoptive transfer and were treated with tolerizing doses of OVA or OVA_323–339 peptide Ag by the oral or i.v. routes showed similar reductions in Th1-type responses or increased Th2-type cytokine production (39, 40). This suggests that there may be some commonality between injury effects on T cell-dependent immune responses and the induction of T cell tolerance. Changes in T cell costimulation or the induction of regulatory-type T cells, shown to be important in the induction or maintenance of T cell tolerance in these other experimental models, might contribute to the injury-induced change in Th1- and Th2-type responses described in this study (41, 42). Because it is not yet known how injury affects T cell costimulatory responses or the activation of CD4⁺CD25⁺ regulatory T cell populations, future studies will need to address how injury may alter these important immune regulatory mechanisms. Additional pathways associated with the injury response that may also skew CD4⁺ T cell differentiation include mediators such as IL-1, IL-6, or IL-10. These cytokines are known to be overproduced following injury and have been shown to bias Th2-type differentiation; and in the case of IL-6 and IL-10, blocking their activity in vivo following burn injury restores Ag-specific delayed type hypersensitivity responses and Th1 Ab isotype formation, respectively (14, 16, 43, 44, 45, 46).

Our findings also differ significantly from others that addressed the influence of inflammation on the development of Ag-specific or superantigen-induced CD4⁺ T cell responses (47). In those studies, inflammation induced by injecting mice with Escherichia coli-derived LPS caused augmented and prolonged CD4⁺ T cell expansion and boosted Th1-type responses. Moreover, in experiments using the same adoptive transfer approach selected for our studies, it was shown that LPS and bacteria that expressed OVA protein could enhance Th1-type reactivity of immunized mice and prevent the development of T cell tolerance in mice injected i.v. with high doses of OVA_323–339 peptide (35, 47, 48). One major difference between our study and these prior reports is that the host inflammatory response was induced by substantial damage to self tissues, whereas the inflammatory response elicited by the injection of LPS resulted from a nonself stimulus. This suggests that the host response to the tissue destruction that occurs after injury may send a dominant signal to the immune system to promote Th2-type differentiation of Ag-stimulated T cells, whereas signals delivered by inflammatory adjuvants such as LPS, CFA, or invading pathogens send a signal that promotes Th1-type reactivity. Therefore, the host adaptive immune response to noninfectious tissue injury appears to be fundamentally distinct from inflammatory responses triggered by pathogen-associated Ags that act via innate immune receptors such as Toll-like receptors. An increased understanding of these differing responses will certainly aid in the development of treatments aimed at controlling injury-induced changes in adaptive immune function, with the goal of improving the survival of critically injured patients.

In summary, the findings from this study contribute to our understanding of how injury affects the development of a specific adaptive immure response in vivo. We demonstrate, using a well-established adoptive transfer approach, that injury does not prevent Ag-dependent proliferation and expansion of naive CD4⁺ T cells. However, as early as 3 days after injury, we demonstrate reduced Ag-specific Th1-type cytokine production followed by a shift to a Th2-type cytokine production profile by 7 days after injury. In aggregate, these results provide a clearer understanding of how injury influences the adaptive immune response and support the theory that severe injury induces fundamental changes in the induction of Ag-specific CD4⁺ Th cell responses, favoring the development of Th2-type immune reactivity in vivo.

	Acknowledgments

 
We acknowledge the technical advice and support from Dr. Marc Jenkins (University of Minnesota, MN), Dr. Abul Abbas (University of California, San Francisco, CA), and Dr. Tom Kupper (Brigham and Women’s Hospital and Harvard Medical School, Boston, MA), without whom this work would not have been possible. We would also like to acknowledge the technical assistance of Dr. Malcolm Kell (University College, Cork, Ireland), Adam Delisle, and Chris Soberg.

	Footnotes

 
¹ This work was supported by grants from the National Institutes of Health (GM57664 and GM35633), the Brook Fund, and the Julian and Eunice Cohen Fund for Surgical Research.

² Address correspondence and reprint requests to Dr. James A. Lederer, Department of Surgery (Immunology), Brigham and Women’s Hospital, 75 Francis Street, Boston, MA 02115. E-mail address: jlederer{at}rics.bwh.harvard.edu

³ Abbreviation used in this paper: TBSA, total body surface area.

Received for publication April 21, 2003. Accepted for publication August 13, 2003.

	References

Top Abstract Introduction Materials and Methods Results Discussion References

 

1. Moore, K. W., R. de Waal Malefyt, R. L. Coffman, A. O’Garra. 2001. Interleukin-10 and the interleukin-10 receptor. Annu. Rev. Immunol. 19:683.[Medline]
2. Rivas, J. M., S. E. Ullrich. 1994. The role of IL-4, IL-10, and TNF- in the immune suppression induced by ultraviolet radiation. J. Leukocyte Biol. 56:769.[Abstract]
3. Oberholzer, A., C. Oberholzer, L. L. Moldawer. 2002. Interleukin-10: a complex role in the pathogenesis of sepsis syndromes and its potential as an anti-inflammatory drug. Crit. Care Med. 30:S58.[Medline]
4. Wahl, S. M.. 1999. TGF- in the evolution and resolution of inflammatory and immune processes: introduction. Microbes Infect. 1:1247.[Medline]
5. Angele, M. K., E. Faist. 2002. Clinical review: immunodepression in the surgical patient and increased susceptibility to infection. Crit. Care 6:298.[Medline]
6. Mannick, J. A., M. L. Rodrick, J. A. Lederer. 2001. The immunologic response to injury. J. Am. Coll. Surg. 193:237.[Medline]
7. Lederer, J. A., M. L. Rodrick, J. A. Mannick. 1999. The effects of injury on the adaptive immune response. Shock 11:153.[Medline]
8. Bone, R. C.. 1996. Immunologic dissonance: a continuing evolution in our understanding of the systemic inflammatory response syndrome (SIRS) and the multiple organ dysfunction syndrome (MODS). Ann. Intern. Med. 125:680.[Abstract/Free Full Text]
9. Goebel, A., E. Kavanagh, A. Lyons, I. B. Saporoschetz, C. Soberg, J. A. Lederer, J. A. Mannick, M. L. Rodrick. 2000. Injury induces deficient interleukin-12 production, but interleukin-12 therapy after injury restores resistance to infection. Ann. Surg. 231:253.[Medline]
10. Kobayashi, H., M. Kobayashi, T. Utsunomiya, D. N. Herndon, R. B. Pollard, F. Suzuki. 1999. Therapeutic protective effects of IL-12 combined with soluble IL-4 receptor against established infections of herpes simplex virus type 1 in thermally injured mice. J. Immunol. 162:7148.[Abstract/Free Full Text]
11. O’Suilleabhain, C., S. T. O’Sullivan, J. L. Kelly, J. Lederer, J. A. Mannick, M. L. Rodrick. 1996. Interleukin-12 treatment restores normal resistance to bacterial challenge after burn injury. Surgery 120:290.[Medline]
12. Lyons, A., A. Goebel, J. A. Mannick, J. A. Lederer. 1999. Protective effects of early interleukin 10 antagonism on injury-induced immune dysfunction. Arch. Surg. 134:1317.[Abstract/Free Full Text]
13. Gennari, R., J. W. Alexander, T. Pyles, S. Hartmann, C. K. Ogle. 1994. Effects of antimurine interleukin-6 on bacterial translocation during gut-derived sepsis. Arch. Surg. 129:1191.[Abstract]
14. Fontanilla, C. V., D. E. Faunce, M. S. Gregory, K. A. Messingham, E. A. Durbin, L. A. Duffner, E. J. Kovacs. 2000. Anti-interleukin-6 antibody treatment restores cell-mediated immune function in mice with acute ethanol exposure before burn trauma. Alcohol. Clin. Exp. Res. 24:1392.[Medline]
15. Kelly, J. L., C. B. O’Suilleabhain, C. C. Soberg, J. A. Mannick, J. A. Lederer. 1999. Severe injury triggers antigen-specific T-helper cell dysfunction. Shock 12:39.[Medline]
16. Kelly, J. L., A. Lyons, C. C. Soberg, J. A. Mannick, J. A. Lederer. 1997. Anti-interleukin-10 antibody restores burn-induced defects in T-cell function. Surgery 122:146.[Medline]
17. Kearney, E. R., K. A. Pape, D. Y. Loh, M. K. Jenkins. 1994. Visualization of peptide-specific T cell immunity and peripheral tolerance induction in vivo. Immunity 1:327.[Medline]
18. Robertson, J. M., P. E. Jensen, B. D. Evavold. 2000. DO11.10 and OT-II T cells recognize a C-terminal ovalbumin 323–339 epitope. J. Immunol. 164:4706.[Abstract/Free Full Text]
19. Hsieh, C. S., S. E. Macatonia, A. O’Garra, K. M. Murphy. 1993. Pathogen-induced Th1 phenotype development in CD4⁺ -TCR transgenic T cells is macrophage dependent. Int. Immunol. 5:371.[Abstract/Free Full Text]
20. Haskins, K., R. Kubo, J. White, M. Pigeon, J. Kappler, P. Marrack. 1983. The major histocompatibility complex-restricted antigen receptor on T cells. I. Isolation with a monoclonal antibody. J. Exp. Med. 157:1149.[Abstract/Free Full Text]
21. Moss, N. M., D. B. Gough, A. L. Jordan, J. T. Grbic, J. J. Wood, M. L. Rodrick, J. A. Mannick. 1988. Temporal correlation of impaired immune response after thermal injury with susceptibility to infection in a murine model. Surgery 104:882.[Medline]
22. Faist, E., C. Schinkel, S. Zimmer. 1996. Update on the mechanisms of immune suppression of injury and immune modulation. World J. Surg. 20:454.[Medline]
23. Baker, C. C., C. L. Miller, D. D. Trunkey. 1979. Predicting fatal sepsis in burn patients. J. Trauma 19:641.[Medline]
24. Zapata-Sirvent, R. L., J. F. Hansbrough, E. M. Bender, E. J. Bartle, M. A. Mansour, W. H. Carter. 1986. Postburn immunosuppression in an animal model. IV. Improved resistance to septic challenge with immunomodulating drugs. Surgery 99:53.[Medline]
25. Kobayashi, M., K. Mori, H. Kobayashi, R. B. Pollard, F. Suzuki. 1998. The regulation of burn-associated infections with herpes simplex virus type 1 or Candida albicans by a non-toxic aconitine-hydrolysate, benzoylmesaconine. Part 1. Antiviral and anti-fungal activities in thermally injured mice. Immunol. Cell Biol. 76:202.[Medline]
26. O’Sullivan, S. T., J. A. Lederer, A. F. Horgan, D. H. Chin, J. A. Mannick, M. L. Rodrick. 1995. Major injury leads to predominance of the T helper-2 lymphocyte phenotype and diminished interleukin-12 production associated with decreased resistance to infection. Ann. Surg. 222:482.[Medline]
27. Lyons, A., J. L. Kelly, M. L. Rodrick, J. A. Mannick, J. A. Lederer. 1997. Major injury induces increased production of interleukin-10 by cells of the immune system with a negative impact on resistance to infection. Ann. Surg. 226:450.[Medline]
28. Messingham, K. A., C. V. Fontanilla, A. Colantoni, L. A. Duffner, E. J. Kovacs. 2000. Cellular immunity after ethanol exposure and burn injury: dose and time dependence. Alcohol 22:35.[Medline]
29. Teodorczyk-Injeyan, J. A., B. G. Sparkes, G. B. Mills, W. J. Peters, R. E. Falk. 1986. Impairment of T cell activation in burn patients: a possible mechanism of thermal injury-induced immunosuppression. Clin. Exp. Immunol. 65:570.[Medline]
30. De, A. K., K. M. Kodys, J. Pellegrini, B. Yeh, R. K. Furse, P. Bankey, C. L. Miller-Graziano. 2000. Induction of global anergy rather than inhibitory Th2 lymphokines mediates posttrauma T cell immunodepression. Clin. Immunol. 96:52.[Medline]
31. Wolfe, J. H., I. Saporoschetz, A. E. Young, N. E. O’Connor, J. A. Mannick. 1981. Suppressive serum, suppressor lymphocytes, and death from burns. Ann. Surg. 193:513.[Medline]
32. Wood, J. J., J. B. O’Mahony, M. L. Rodrick, R. Eaton, R. H. Demling, J. A. Mannick. 1986. Abnormalities of antibody production after thermal injury: an association with reduced interleukin 2 production. Arch. Surg. 121:108.[Abstract]
33. Abbas, A. K., K. M. Murphy, A. Sher. 1996. Functional diversity of helper T lymphocytes. Nature 383:787.[Medline]
34. Kearney, E. R., T. L. Walunas, R. W. Karr, P. A. Morton, D. Y. Loh, J. A. Bluestone, M. K. Jenkins. 1995. Antigen-dependent clonal expansion of a trace population of antigen-specific CD4⁺ T cells in vivo is dependent on CD28 costimulation and inhibited by CTLA-4. J. Immunol. 155:1032.[Abstract]
35. Reinhardt, R. L., A. Khoruts, R. Merica, T. Zell, M. K. Jenkins. 2001. Visualizing the generation of memory CD4 T cells in the whole body. Nature 410:101.[Medline]
36. Kell, M. R., E. G. Kavanaugh, A. Goebel, C. C. Soberg, J. A. Lederer. 1999. Injury primes the immune system for an enhanced and lethal T-cell response against bacterial superantigen. Shock 12:139.[Medline]
37. Matzinger, P.. 1994. Tolerance, danger, and the extended family. Annu. Rev. Immunol. 12:991.[Medline]
38. Matzinger, P.. 2002. The danger model: a renewed sense of self. Science 296:301.[Abstract/Free Full Text]
39. Pape, K. A., R. Merica, A. Mondino, A. Khoruts, M. K. Jenkins. 1998. Direct evidence that functionally impaired CD4⁺ T cells persist in vivo following induction of peripheral tolerance. J. Immunol. 160:4719.[Abstract/Free Full Text]
40. Chen, Y., J. Inobe, H. L. Weiner. 1997. Inductive events in oral tolerance in the TCR transgenic adoptive transfer model. Cell Immunol. 178:62.[Medline]
41. Cottrez, F., S. D. Hurst, R. L. Coffman, H. Groux. 2000. T regulatory cells 1 inhibit a Th2-specific response in vivo. J. Immunol. 165:4848.[Abstract/Free Full Text]
42. Rouleau, M., F. Cottrez, M. Bigler, S. Antonenko, J. M. Carballido, A. Zlotnik, M. G. Roncarolo, H. Groux. 1999. IL-10 transgenic mice present a defect in T cell development reminiscent of SCID patients. J. Immunol. 163:1420.[Abstract/Free Full Text]
43. Schmitz, N., M. Kurrer, M. Kopf. 2003. The IL-1 receptor 1 is critical for Th2 cell type airway immune responses in a mild but not in a more severe asthma model. Eur. J. Immunol. 33:991.[Medline]
44. Rincon, M., J. Anguita, T. Nakamura, E. Fikrig, R. A. Flavell. 1997. Interleukin (IL)-6 directs the differentiation of IL-4-producing CD4⁺ T cells. J. Exp. Med. 185:461.[Abstract/Free Full Text]
45. Groux, H., A. O’Garra, M. Bigler, M. Rouleau, S. Antonenko, J. E. de Vries, M. G. Roncarolo. 1997. A CD4⁺ T-cell subset inhibits antigen-specific T-cell responses and prevents colitis. Nature 389:737.[Medline]
46. Dodge, I. L., M. W. Carr, M. Cernadas, M. B. Brenner. 2003. IL-6 production by pulmonary dendritic cells impedes Th1 immune responses. J. Immunol. 170:4457.[Abstract/Free Full Text]
47. Pape, K. A., A. Khoruts, A. Mondino, M. K. Jenkins. 1997. Inflammatory cytokines enhance the in vivo clonal expansion and differentiation of antigen-activated CD4⁺ T cells. J. Immunol. 159:591.[Abstract]
48. Chen, Z. M., M. K. Jenkins. 1998. Revealing the in vivo behavior of CD4⁺ T cells specific for an antigen expressed in Escherichia coli. J. Immunol. 160:3462.[Abstract/Free Full Text]

Related articles in The JI:

IN THIS ISSUE

The JI 2003 171: 3911-3912. [Full Text]  

This article has been cited by other articles:

				D. D. Shao, R. Suresh, V. Vakil, R. H. Gomer, and D. Pilling Pivotal Advance: Th-1 cytokines inhibit, and Th-2 cytokines promote fibrocyte differentiation J. Leukoc. Biol., June 1, 2008; 83(6): 1323 - 1333. [Abstract] [Full Text] [PDF]

				J. Bohannon, W. Cui, R. Cox, R. Przkora, E. Sherwood, and T. Toliver-Kinsky Prophylactic Treatment with Fms-Like Tyrosine Kinase-3 Ligand after Burn Injury Enhances Global Immune Responses to Infection J. Immunol., March 1, 2008; 180(5): 3038 - 3048. [Abstract] [Full Text] [PDF]

				J. Unsinger, J. M. Herndon, C. G. Davis, J. T. Muenzer, R. S. Hotchkiss, and T. A. Ferguson The Role of TCR Engagement and Activation-Induced Cell Death in Sepsis-Induced T Cell Apoptosis J. Immunol., December 1, 2006; 177(11): 7968 - 7973. [Abstract] [Full Text] [PDF]

				R. Maile, C. M. Barnes, A. I. Nielsen, A. A. Meyer, J. A. Frelinger, and B. A. Cairns Lymphopenia-Induced Homeostatic Proliferation of CD8+ T Cells Is a Mechanism for Effective Allogeneic Skin Graft Rejection following Burn Injury. J. Immunol., June 1, 2006; 176(11): 6717 - 6726. [Abstract] [Full Text] [PDF]

				J. W. Smith, R. L. Gamelli, S. B. Jones, and R. Shankar Immunologic responses to critical injury and sepsis. J Intensive Care Med, May 1, 2006; 21(3): 160 - 172. [Abstract] [PDF]

				N. N. Choileain, M. MacConmara, Y. Zang, T. J. Murphy, J. A. Mannick, and J. A. Lederer Enhanced Regulatory T Cell Activity Is an Element of the Host Response to Injury J. Immunol., January 1, 2006; 176(1): 225 - 236. [Abstract] [Full Text] [PDF]

				J. Patenaude, M. D'Elia, C. Hamelin, D. Garrel, and J. Bernier Burn injury induces a change in T cell homeostasis affecting preferentially CD4+ T cells J. Leukoc. Biol., February 1, 2005; 77(2): 141 - 150. [Abstract] [Full Text] [PDF]

				T. E. Toliver-Kinsky, W. Cui, E. D. Murphey, C. Lin, and E. R. Sherwood Enhancement of Dendritic Cell Production by Fms-Like Tyrosine Kinase-3 Ligand Increases the Resistance of Mice to a Burn Wound Infection J. Immunol., January 1, 2005; 174(1): 404 - 410. [Abstract] [Full Text] [PDF]

				Y. Zang, S. M. Dolan, N. N. Choileain, S. J. Kriynovich, T. J. Murphy, P. Sayles, J. A. Mannick, and J. A. Lederer Burn Injury Initiates a Shift in Superantigen-Induced T Cell Responses and Host Survival J. Immunol., April 15, 2004; 172(8): 4883 - 4892. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Related articles in The JI