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*Bacterial Infections
*Burns
The Journal of Immunology, 2002, 169: 4437-4442.
Copyright © 2002 by The American Association of Immunologists

IFN-{gamma} Production from Liver Mononuclear Cells of Mice in Burn Injury As Well As in Postburn Bacterial Infection Models and the Therapeutic Effect of IL-18

Katsunori Ami1,*,§, Manabu Kinoshita1,*, Akira Yamauchi{dagger}, Tetsuro Nishikage§, Yoshiko Habu{ddagger}, Nariyoshi Shinomiya{ddagger}, Takehisa Iwai§, Hoshio Hiraide* and Shuhji Seki2,{ddagger}

* Division of Basic Traumatology, National Defense Medical College Research Institute, and Departments of {dagger} Surgery 1 and {ddagger} Microbiology, National Defense Medical College, Namiki, Tokorozawa, Japan; and § First Department of Surgery, Tokyo Medical and Dental University, Tokyo, Japan


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Hosts after severe burn injury are known to have a defect in the Th1 immune response and are susceptible to bacterial infections. We herein show that liver NK cells are potent IFN-{gamma} producers early after burn injury. However, when mice were injected with LPS 24 h after burn injury, IFN-{gamma} production from liver mononuclear cells (MNC; which we previously showed to be NK cells) was suppressed, and the serum IFN-{gamma} concentration did not increase, while serum IL-10 conversely increased compared with control mice. Interestingly, a single injection of IL-18 simultaneously with LPS greatly restored the serum IFN-{gamma} concentration in mice with burn injury and also increased IFN-{gamma} production from liver MNC. Nevertheless, a single IL-18 injection into mice simultaneously with LPS was no longer effective in the restoration of serum IFN-{gamma} and IFN-{gamma} production from the liver MNC at 7 days after burn injury, when mice were considered to be the most immunocompromised. However, IL-18 injections into mice on alternate days beginning 1 day after burn injury strongly up-regulated LPS-induced serum IFN-{gamma} levels and IFN-{gamma} production from liver and spleen MNC of mice 7 days after burn injury and down-regulated serum IL-10. Furthermore, similar IL-18 therapy up-regulated serum IFN-{gamma} levels in mice with experimental bacterial peritonitis 7 days after burn injury and greatly decreased mouse mortality. Thus, IL-18 therapy restores the Th1 response and may decrease the susceptibility to bacterial infection in mice with burn injury.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
It is well known that hosts with severe burn injury are exceedingly susceptible to bacterial infections, including sepsis. Not only bacterial infection in situ of burn injury but also bacterial translocation from the gut sometimes cause sepsis in the hosts with burn injury (1, 2, 3). Various reports to date have indicated that the functional impairment of Th1 lymphocytes is responsible for the susceptibility to bacterial infections of burned hosts (4). NK cell activity is thus compromised (5), and the production of IFN-{gamma}, which is a representative Th1 cytokine and is essential for the host defense against microbial infections, is decreased (6, 7). In contrast, a Th2 cytokine, IL-10 production, was augmented (8, 9). In line with these findings, recent reports have indicated that IL-12 and IFN-{gamma} prevent bacterial infections associated with burn injury (10, 11), and anti-IL-10 Ab also attenuates infections (8, 9), whereas another report indicated that IFN-{gamma} is not effective in the prevention of burn-related bacterial infections (12). However, why and how the Th1 immune response is compromised after burn injury and cytokine behavior in burn injury itself are largely unknown.

We have recently shown liver mononuclear cells (MNC),3 especially NK cells and/or NK1.1+ T (NKT) cells, to produce large amounts of IFN-{gamma} by various stimuli, including IL-12 (13), LPS (14), {alpha}-galactosylceramide (15), and bacterial peritonitis made by cecal ligation and puncture (CLP) (16). In addition, Kupffer cells (resident liver macrophages) produce IL-12 and IL-18 (14, 17), and hepatocytes produce acute phase proteins and LPS binding protein in bacterial infections (14, 18, 19). We therefore proposed the liver to be an important organ for host defense (reviewed in Ref. 14). Furthermore, hepatocytes have been reported to be the main producers of acute phase proteins in burn injury (20, 21).

On the other hand, IL-18 is a recently cloned cytokine that has various biological effects and is mainly produced by macrophages and Kupffer cells (17, 22, 23). IL-18 was originally described as a Th1 cytokine and induces potent IFN-{gamma} production from NK cells in the presence of IL-12 (22, 23). In addition, however, IL-18 plays a functional role in Th2 responses, because IL-18 alone has been recently reported to induce IgE production as a result of IL-4 production from T cells (17). Therefore, although the function of IL-18 is not as simple (17), it is now generally accepted that IL-18 enhances the IL-12-driven Th1 immune responses in bacterial and viral infections and therefore is essential for host defense (17).

In light of these findings, we herein focus on burn injury of mice and IFN-{gamma} production from liver MNC either in burn injury itself or in postburn bacterial infection models. We show that liver NK cells produce IFN-{gamma} in mice with burn injury, while the impaired Th1 response to bacterial infection after burn injury is related to the diminished IFN-{gamma} production from liver NK cells. In addition, we show the effect of IL-18 treatment on the impaired Th1 response in the bacterial infection. We hope that our findings will provide new insights into the immune function of the liver and burn injury.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Mice and mouse burn model

Eight-week-old male C57BL/6 mice were obtained from SLC (Hamamatsu, Japan). The mice were anesthetized by i.p. pentobarbital (1 mg/mouse) and were shaved over the dorsum. The full-thickness burn injury (20% of total body surface area) was made by a heated brass blade, and 1 ml PBS was i.p. injected. Control mice were also anesthetized. This study was approved by the ethical committee of National Defense Medical College.

CLP procedure

CLP was performed essentially as previously described (24). Briefly, after i.p. pentobarbital anesthetization of the mice with or without prior burn injury, the anterior abdominal walls of the mice were shaved, and a small incision was made to expose the cecum, which was ligated at its base with 3.0 silk. The cecum was punctured once with a 23-gauge needle, and a small volume of feces was placed on the exterior. The cecum was then returned to the peritoneal cavity, and the abdomen was closed.

Reagents

LPS (Escherichia coli 0111:B4) was purchased from Sigma-Aldrich (St. Louis, MO). Mouse IL-18 was purchased from MBL (Nagoya, Japan).

Isolation of MNC

Under deep ether anesthesia, the mice were euthanized by exsanguination from the subclavian artery and vein, and then the liver was removed. Liver MNC were obtained as previously described (13, 25). The liver was passed through a stainless steel mesh and then suspended in RPMI 1640 medium. After one washing, the cells were resuspended in osmolarity- and pH-adjusted 33% Percoll solution containing 100 U/ml heparin and were centrifuged at 500 x g for 15 min at room temperature. The pellet was resuspended in a RBC lysis solution, then washed twice in 10% FBS/RPMI 1640. The degree of contamination by Kupffer cells or hepatocytes was minimal. Splenocytes were passed through a 200-gauge stainless steel mesh, and were treated with RBC lysis solution, and washed twice in 10% FBS/RPMI 1640. Peripheral blood MNC were obtained from heparinized blood by Ficoll-Hypaque density gradient centrifugation. To obtain lung MNC, the lung was minced, suspended in medium containing 0.05% collagenase (Wako, Tokyo, Japan) and 0.01% trypsin inhibitor (Sigma), and then shaken for 20 min in a 37°C water bath. Thereafter, the lung specimens were passed through a stainless steel mesh, and MNC were obtained with Percoll solution as described above.

Cell cultures

At 4 h after burn injury and/or LPS injection, 5 x 105 liver MNC, 5 x 105 splenocytes, 5 x 105 PBMC, or 5 x 105 lung MNC in 200 µl 10% FBS/RPMI 1640 medium were cultured in 96-well flat-bottom plates in 5% CO2 at 37°C for 24 h, and then the culture supernatants were stocked at -80°C and subjected to ELISA.

In vivo NK cell depletion

Anti-NK1.1 Ab (PK136; 200 µg/mouse) or anti-asialo-GM1 Ab (AGM1; 50 µg/mouse) was i.v. injected into mice 4 days before burn injury. PK136 hybridoma was grown in our laboratory, and anti-AGM1 Ab was purchased from Wako. Anti-NK1.1 Ab depletes both NK cells and NK1.1+T cells, and anti-AGM1 Ab depletes NK cells alone for ~1 wk, as we previously reported (15, 26).

ELISA of sera and culture supernatants

IFN-{gamma} and IL-10 levels in sera or culture supernatants were measured by cytokine-specific ELISA kits (Endogen, Woburn, MA). Serum IL-18 levels were assayed using an ELISA kit (MBL). The sera were usually 20-fold diluted by the assay buffer included in the ELISA kit and used to measure the IFN-{gamma}, IL-10, and IL-18 levels.

Statistical analysis

Differences between the groups were analyzed by Mann-Whitney U test, ANOVA with Fisher’s protected least significant difference test, or Scheffé’s F test using the StatView program (Abacus Concepts, Berkeley, CA) on an Apple computer (Cupertino, CA). The mouse survival rate was determined in accordance with the product-limited (Kaplan-Meier) estimate, and survival curves were compared with one another using the log-rank (Mantel-Cox) test. Differences were considered significant at p < 0.05.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Liver MNC produce a large amount of IFN-{gamma} after burn injury

When liver MNC from mice that had received burn injury 12 h previously were cultured in vitro for 24 h, they produced a large amount of IFN-{gamma} in vitro without any additional stimulation (Fig. 1GoA). However, PBMC and splenocytes produced little IFN-{gamma} (Fig. 1GoA). Lung MNC also did not produce a significant amount IFN-{gamma} (data not shown). A time-course analysis after burn injury showed that liver MNC produced the largest amount of IFN-{gamma} 12 h after burn injury, while depletion of NK cells or both NK cells and NKT cells in mice with anti-AGM1 Ab or anti-NK1.1 Ab in vivo significantly reduced the IFN-{gamma} level (Fig. 1GoB), suggesting that NK1.1+ cells, especially NK cells, produce IFN-{gamma}. The depletion of NK cells by either anti-AGM1 Ab or anti-NK1.1 Ab also greatly reduced serum IFN-{gamma} levels, except at 3 h after burn injury (Fig. 1GoC). Although the source of the high levels of IFN-{gamma} at 3 h after burn injury is unclear, macrophages may produce IFN-{gamma} (27) in the early phase after burn injury.



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  		FIGURE 1. A, IFN-{gamma} production from liver MNC after burn injury. Twelve hours after burn injury of mice, liver MNC, splenocytes, and PBMC were obtained and cultured for 24 h, and culture supernatants were subjected to ELISA to determine IFN-{gamma} levels. All data are the mean ± SE from the indicated number of mice. B, Time-course analysis of in vitro IFN-{gamma} production from liver MNC after burn injury and the effect of NK cell depletion on IFN-{gamma} production by liver MNC. The mice were injected i.v. with anti-AGM1 Ab (50 µg/200 µl PBS), anti-NK1.1 Ab (200 µg/200 µl PBS), or PBS as a control 4 days before burn injury, and liver MNC were obtained at the indicated time points after burn injury and then cultured for 24 h. The culture supernatants were subjected to ELISA to determine IFN-{gamma} levels. The data are the mean ± SE from the indicated number of mice. *, p < 0.05; **, p < 0.01 (vs the control (PBS) group). C, Serum IFN-{gamma} concentrations after burn injury and the effect of the NK cell depletion. The mice were injected i.v. with anti-AGM1 Ab (50 µg), anti-NK1.1 Ab (200 µg), or PBS 4 days before burn injury, and sera in the blood samples obtained from retroorbital plexus at the indicated time points after burn injury were subjected to ELISA. The data are the mean ± SE from five mice. *, p < 0.05 (vs control (PBS) group).

 
LPS-stimulated serum IFN-{gamma} and IFN-{gamma} production from liver MNC decreased, while serum IL-10 increased in mice after burn injury

At 24 h after burn injury the mice were i.v. injected with LPS, and serum IFN-{gamma} and IL-10 levels were followed for 24 h. The serum IFN-{gamma} concentration in control unburned mice markedly increased, peaked at 6 h after LPS injection, and then reverted to basal levels at 24 h after LPS injection (Fig. 2GoA). However, IFN-{gamma} did not increase in the burned mice after LPS injection (Fig. 2GoA). In contrast, the serum IL-10 concentration increased more obviously in the burned mice than in the control unburned mice (Fig. 2GoB) after LPS injection, especially 3 and 6 h after LPS injection. In addition, when liver MNC obtained from burned mice or control mice 6 h after LPS injection were cultured for 24 h in vitro, IFN-{gamma} production from liver MNC from burned mice was severely suppressed compared with that in unburned control mice (Fig. 2GoC). We previously reported that NK cells are the main IFN-{gamma} producers in the liver MNC of mice injected with LPS (14).



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  		FIGURE 2. Serum IFN-{gamma} (A) and serum IL-10 (B) concentrations in mice with or without burn injury injected with LPS. Twenty-four hours after burn injury, mice with burn injury and control unburned mice were i.v. injected with LPS (50 µg/mouse), and serum IFN-{gamma} and IL-10 concentrations were determined in samples of blood obtained from the retro-orbital plexus of mice at the indicated times after LPS injection. The data are the mean ± SE from eight mice in each group. *, p < 0.05; **, p < 0.01 (vs the corresponding values). C, IFN-{gamma} production from liver MNC with burn injury. LPS was injected into the control mice and burned mice 24 h after burn injury. Six hours after LPS injection liver MNC were obtained from each mouse group and cultured for 24 h, and culture supernatants were subjected to ELISA. The data are the mean ± SE from the indicated number of mice. **, p < 0.01 (vs the control (PBS) group).

 
Recovery of the IFN-{gamma} response to LPS by IL-18 treatment in mice early after burn injury, but failure of IL-18 to recover IFN-{gamma} response in mice 7 days after burn injury

At 24 h after burn injury the mice were i.v. injected with LPS or LPS plus IL-18 (0.2 µg/mouse i.p.), and serum IFN-{gamma} levels 6 h after LPS injection were compared with those in the control mice. The serum IFN-{gamma} concentration in the control unburned mice markedly increased after LPS injection (Fig. 3GoA). In contrast, IFN-{gamma} did not increase in burned mice after LPS injection (Fig. 3GoA). However, serum IFN-{gamma} concentrations 6 h after LPS injection remarkably increased by simultaneous IL-18 injection with LPS (Fig. 3GoA). We confirmed that injection of mice with 0.2 µg IL-18 sufficiently elevated serum IL-18 levels (~30 ng/ml) at 2 h after the injection, and thereafter IL-18 levels gradually decreased and returned to basal levels by 48 h after the injection (data not shown).



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  		FIGURE 3. The effect of a single IL-18 injection on serum IFN-{gamma} (A and B) and IFN-{gamma} production from liver MNC (C) of mice with burn injury injected with LPS or LPS plus IL-18. A, Twenty-four hours after burn injury, mice with burn injury and control unburned mice were injected with LPS (50 µg i.v.) or LPS plus IL-18 (200 µl in PBS i.p.). Six hours thereafter, sera were obtained, and IFN-{gamma} concentrations were measured. B, At the indicated times after burn injury, LPS or LPS plus IL-18 was injected into mice. Sera were obtained 6 h after injection, and serum IFN-{gamma} concentrations were measured. C, At the indicated times after burn injury, LPS or LPS plus IL-18 were injected into mice, and liver MNC were obtained 6 h after injection and cultured for 24 h. Thereafter, IFN-{gamma} levels in culture supernatants were measured.

 
Next, a time-course analysis of the effect of a single injection of IL-18 simultaneously with LPS on the indicated days after burn injury on serum IFN-{gamma} concentrations was conducted. The results showed that the effect of IL-18 on LPS-induced elevation of serum IFN-{gamma} was almost abolished in mice 7 days after burn injury (Fig. 3GoB). Consistent with these results, IL-18 did not significantly restore IFN-{gamma} production in liver MNC from mice 7 days after burn injury (Fig. 3GoC). The time-course analysis of serum IL-18 in burned mice after LPS injection (Fig. 4Go) revealed that serum IL-18 levels were not lower in burned mice compared with control mice, thus indicating that IL-18 production from macrophages in burned mice did not differ from that in control mice. Serum IL-18 levels 12 h after LPS injection at the earlier time point (1 day after burn injury), when the single injection of IL-18 (with LPS) enhanced IFN-{gamma} production, also did not significantly differ from those in control mice (not shown). We observed that the lymphocyte subsets (i.e., NK cells, NKT cells, and T cells) in the liver did not significantly change after burn injury (not shown). These findings suggest that the impaired IFN-{gamma} production in response to LPS in mice at 7 days after burn injury is not related to the proportional change in liver MNC subsets or the IL-18 production capacity in mice.



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  		FIGURE 4. Serum IL-18 concentrations at the indicated time points in mice after LPS injection at 7 days after burn injury and in unburned control mice. The data are the mean ± SE from five mice in each group.

 
Multiple injections of IL-18 restored LPS-induced serum IFN-{gamma} in mice 7 days after burn injury, restored IFN-{gamma} production from liver and spleen MNC, and inhibited the LPS-induced serum IL-10 elevation

If mice with burn injury were injected with IL-18 on days 1, 3, and 5 and were injected with LPS simultaneously with IL-18 on day 7 after burn injury, the serum IFN-{gamma} concentration greatly increased (Fig. 5GoA). The decreased IFN-{gamma} production from liver and spleen MNC of burned mice injected with LPS was also enhanced by multiple IL-18 injections (Fig. 5Go). In addition, elevated serum IL-10 levels at 6 h after LPS injection in burned mice reverted to levels comparable to those in control mice injected with LPS (Fig. 6Go). These findings suggest that the impaired IFN-{gamma} production in mice 7 days after burn injury may be related to the unresponsiveness of liver and spleen MNC to the ordinary level of endogenous IL-18 induced by LPS.



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  		FIGURE 5. The effect of multiple injections of IL-18 (mIL-18) on LPS-induced serum IFN-{gamma} concentrations (A) and IFN-{gamma} production from liver MNC (B) and spleen MNC (C) of mice at 7 days after burn injury. The mice with burn injury were i.p. injected with PBS or IL-18 (0.2 µg) on days 1, 3, 5, and 7 after burn injury, and LPS was i.v. injected on day 7 simultaneously with PBS or IL-18 injection on day 7. The sera were obtained from blood samples 6 h after LPS injection and were subjected to ELISA. The data are the mean ± SE from five mice in each group. Liver and spleen MNC were also obtained and cultured for 24 h. The culture supernatants were subjected to ELISA. The data are the mean ± SE from seven mice in each group.

 


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  		FIGURE 6. The effect of multiple injections of IL-18 (mIL-18) on LPS-induced serum IL-10 concentrations. Mice with burn injury were i.p. injected with PBS or IL-18 (0.2 µg) on days 1, 3, 5, and 7 after burn injury, and LPS was i.v. injected on day 7 simultaneously with PBS or IL-18 injection on day 7. The sera were obtained from blood samples 6 h after LPS injection and were subjected to ELISA. The data are the mean ± SE from five mice in each group.

 
Decrease in the mortality of mice with bacterial peritonitis following burn injury and up-regulation of IFN-{gamma}

To examine the therapeutic effect of IL-18 on bacterial infections, bacterial peritonitis was induced by CLP in mice 7 days after burn injury. Although most control unburned mice survived after CLP (80%), only 40% of the burned mice survived after CLP (Fig. 7GoA). However, when mice with burn injury were injected with IL-18 (0.2 µg/mouse) on days 1, 3, 5, 7, and 8, it greatly improved the mouse survival rate (70%) after CLP (Fig. 7GoA). Consistent with these findings, the therapy of multiple IL-18 (0.2 µg each) injections led to an elevation of serum IFN-{gamma} in mice with burn injury after CLP (Fig. 7GoB) and increased IFN-{gamma} production from liver MNC (data not shown), in which NK cells are the main IFN-{gamma} producers (16).



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  		FIGURE 7. A, The effect of the treatment with multiple IL-18 injections on the survival rate of mice subjected to CLP following burn injury. Mice with burn injury were i.p. injected with IL-18 (0.2 µg) or PBS on days 1, 3, 5, and 7, and CLP was made on day 7 after burn injury and 2 h after the IL-18 injection on day 7. IL-18 or PBS was also s.c. injected into burned mice on day 8. CLP was also made in unburned control mice. The data are from 10 mice in each group. B, The effect of treatment with multiple IL-18 injections on serum IFN-{gamma} concentrations in burned mice after CLP. Mice with burn injury were i.p. injected with IL-18 (0.2 µg) or PBS on days 1, 3, 5, and 7. Then CLP was made in PBS-injected, burned mice, in IL-18-injected, burned mice (2 h after IL-18 injection on day 7) 7 days after burn injury, and in unburned control mice. Serum IFN-{gamma} concentrations were determined by ELISA from blood samples obtained from the retro-orbital plexus before (pre) CLP and at the indicated time points after CLP. The data are the mean ± SE from six mice in each group.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
In the present study we found that liver MNC, especially NK cells, are the major IFN-{gamma} producers in burn injury itself. However, liver MNC in burned mice subsequently became unresponsive (anergic) to LPS or bacterial infection and did not effectively produce IFN-{gamma}. The impaired IFN-{gamma} production of liver MNC was most evident at 7 days after burn injury and recovered by 14 days after burn injury. We have also shown that IL-18 therapy restored LPS-induced IFN-{gamma} production while it inhibited IL-10 production in burned mice and was effective against bacterial peritonitis after burn injury, thereby remarkably decreasing mouse mortality.

In the hosts with severe burn injury, not only bacterial infection in situ but also systemic bacterial infection (sepsis) frequently occur and can sometimes be fatal. One of the major causes of sepsis is believed to be bacterial translocation from the gut (1, 2, 3). Mesenteric lymph nodes and liver indeed contain bacteria after burn injury in mice (1). On the other hand, 70% of bacteria that enter the bloodstream are reported to accumulate in liver and are trapped and removed from the blood by Kupffer cells and hepatocytes, suggesting the important role of the liver in host defense as a reticuloendothelial organ (14, 28). Consistent with these findings, we recently found that liver NK cells produce a large amount of IFN-{gamma} in mice with septic bacterial peritonitis or mice injected with bacterial superantigens, while splenocytes, peripheral blood MNC, or lung MNC produce only low amounts of IFN-{gamma} (16, 25). The production of IFN-{gamma} from liver MNC is largely dependent on IL-12 and IL-18 produced by Kupffer cells (14, 17), and IFN-{gamma} further activates the phagocytosis and cytokine production of Kupffer cells in a positive feedback loop. Therefore, the impaired IFN-{gamma} production in liver NK cells in response to bacterial stimulation indicates the impaired immune function, including phagocytosis, in the liver and may thus induce the susceptibility to systemic bacterial infection in burned mice.

Despite these findings, it was surprising and somewhat unexpected that only liver MNC among those tested produced IFN-{gamma} after burn injury itself. However, it should be noted that hepatocytes are the main producers of acute phase proteins, including C-reactive protein, in burn injury (21, 29), similar to LPS injection or bacterial infections (21). In addition, the acute phase proteins, {alpha}1-acid glycoprotein and {alpha}1-antitripsin mRNA, were detected in the liver of rats, but not in the kidney or spleen at the early phase (within 48 h) of either burn injury or after LPS injection (21). Furthermore, {alpha}1-acid glycoprotein and {alpha}1-antitripsin expression or production is reportedly correlated to the rat’s resistance to burn injury, and {alpha}1-acid glycoprotein treatment accelerates healing of the burn wound and decreases the incidence of complications (21). Since monocytes express receptors for acute phase proteins (29, 30) and are indeed activated by acute phase proteins (29), the activation of Kupffer cells and subsequently of liver lymphocytes, especially NK cells, and their IFN-{gamma} production may also take place early after burn injury. We believe that IFN-{gamma} production in liver NK cells in burn injury may thus have a beneficial effect on the host.

IL-18 therapy up- and down-regulated serum IFN-{gamma} and IL-10 concentrations, respectively, in mice with burn injury in response to LPS regardless of the fact that the IL-18 response to LPS in mice after burn injury was seemingly not impaired. However, it can be speculated that the usual IL-18 concentration after LPS stimulation is not enough to induce IFN-{gamma} production from anergic liver and spleen MNC, and a larger amount of IL-18 is needed to restore IFN-{gamma} production. It should also be noted that the degree of up-regulation of the IFN-{gamma}-producing capacity in splenocytes of burned mice was larger than that in liver MNC. Therefore, the role of splenocytes in IL-18 treatment of burned mice should also be important for host defense.

IL-12 as well as IL-18 are potent stimulators of IFN-{gamma} production from NK cells and NKT cells in bacterial infections (13, 23). It was recently reported that IL-12 therapy was effective in mice with burn injury against bacterial peritonitis induced by CLP and in reducing mouse mortality (4, 11, 31), although to determine the appropriate dose that induces a beneficial effect in burned mice is difficult (11). Whereas the effectiveness of IFN-{gamma} treatment against bacterial infection in mice with burn injury has been controversial (10, 12), it may be related to its short {beta} half-life (20 min) (32). IL-18 therapy elevated serum IFN-{gamma} levels for at least several hours and inhibited IL-10 production and may thus be effective for the treatment of bacterial infections in hosts with burn injury. A detailed study of the therapeutic effect of IL-18 on systemic E. coli infection (including E. coli clearance from the liver) and serum cytokine concentrations in burned mice and cytokine production from liver and spleen MNC will be reported (M. Kinoshita et al., manuscript in preparation).

Collectively, liver MNC produce IFN-{gamma} early after burn injury and are subsequently unresponsive to bacterial stimulation, and the impaired IFN-{gamma} production from liver MNC and spleen MNC in response to ordinary levels of IL-18 in mice with burn injury may be related to the susceptibility to bacterial infections. IL-18 therapy restores IFN-{gamma} production from liver and spleen MNC and may prevent bacterial infection while reducing infection-related mortality in mice with burn injury.


   Footnotes
 
1 K.A. and M.K. are equal contributors to this manuscript. Back

2 Address correspondence and reprint requests to Dr. Shuhji Seki, Department of Microbiology, National Defense Medical College, Namiki 3-2, Tokorozawa 359-8513, Japan. E-mail address: btraums{at}res.ndmc.ac.jp Back

3 Abbreviations used in this paper: MNC, mononuclear cells; AGM1, asialo-GM1; CLP, cecal ligation and puncture; NKT, NK1.1+ T. Back

Received for publication April 22, 2002. Accepted for publication August 12, 2002.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Maejima, K., E. Deitch, R. Berg. 1984. Promotion by burn stress of the translocation of bacteria from the gastrointestinal tracts of mice. Arch. Surg. 119:166.[Abstract/Free Full Text]
  2. Deitch, E. A., J. Winterton, R. Berg. 1986. Thermal injury promotes bacterial translocation from the gastrointestinal tract in mice with impaired T-cell-mediated immunity. Arch. Surg. 121:97.[Abstract/Free Full Text]
  3. Eaves-Pyles, T., J. W. Alexander. 1998. Rapid and prolonged impairment of gut barrier function after thermal injury in mice. Shock 9:95.[Medline]
  4. 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]
  5. Bender, B. S., R. A. Winchurch, J. N. Thupari, J. J. Proust, W. H. Adler, A. M. Munster. 1988. Depressed natural killer cell function in thermally injured adults: successful in vivo and in vitro immunomodulation and the role of endotoxin. Clin. Exp. Immunol. 71:120.[Medline]
  6. Suzuki, F., R. B. Pollard. 1982. Alterations of interferon production in a mouse model of thermal injury. J. Immunol. 129:1806.[Abstract]
  7. Suzuki, F., R. B. Pollard. 1982. Mechanism for the suppression of {gamma}-interferon responsiveness in mice after thermal injury. J. Immunol. 129:1811.[Abstract]
  8. 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]
  9. 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]
  10. Gennari, R., J. W. Alexander, T. Eaves-Pyles. 1994. IFN-{gamma} decreases translocation and improves survival following transfusion and thermal injury. J. Surg. Res. 56:530.[Medline]
  11. 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]
  12. Wasserman, D., J. D. Ioannovich, R. D. Hinzmann, G. Deichsel, G. G. Steinmann. 1998. Interferon-{gamma} in the prevention of severe burn-related infections: a European phase III multicenter trial: The Severe Burns Study Group. Crit. Care. Med. 26:434.[Medline]
  13. Ogasawara, K., K. Takeda, W. Hashimoto, M. Satoh, R. Okuyama, N. Yanai, M. Obinata, K. Kumagai, H. Takada, H. Hiraide, et al 1998. Involvement of NK1+ T cells and their IFN-{gamma} production in the generalized Shwartzman reaction. J. Immunol. 160:3522.[Abstract/Free Full Text]
  14. Seki, S., Y. Habu, T. Kawamura, K. Takeda, H. Dobashi, T. Ohkawa, H. Hiraide. 2000. The liver as a crucial organ in the first line of host defense: the roles of Kupffer cells, natural killer (NK) cells and NK1.1 Ag+ T cells in T helper 1 immune responses. Immunol. Rev. 174:35.[Medline]
  15. Nakagawa, R., I. Nagafune, Y. Tazunoki, H. Ehara, H. Tomura, R. Iijima, K. Motoki, M. Kamishohara, S. Seki. 2001. Mechanisms of the antimetastatic effect in the liver and of the hepatocyte injury induced by {alpha}-galactosylceramide in mice. J. Immunol. 166:6578.[Abstract/Free Full Text]
  16. Seki, S., S. Osada, S. Ono, S. Aosasa, Y. Habu, T. Nishikage, H. Mochizuki, H. Hiraide. 1998. Role of liver NK cells and peritoneal macrophages in {gamma} interferon and interleukin-10 production in experimental bacterial peritonitis in mice. Infect. Immun. 66:5286.[Abstract/Free Full Text]
  17. Nakanishi, K., T. Yoshimoto, H. Tsutsui, H. Okamura. 2001. Interleukin-18 regulates both Th1 and Th2 responses. Annu. Rev. Immunol. 19:423.[Medline]
  18. Grube, B. J., C. G. Cochane, R. D. Ye, C. E. Green, M. E. McPhail, R. J. Ulevitch, P. S. Tobias. 1994. Lipopolysaccharide binding protein expression in primary human hepatocytes and HepG2 hepatoma cells. J. Biol. Chem. 269:8477.[Abstract/Free Full Text]
  19. Kawamoto, S., Y. Matsumoto, K. Mizuno, K. Okubo, K. Matsubara. 1996. Expression profiles of active genes in human and mouse livers. Gene 174:151.[Medline]
  20. Sevaljevic, L., M. Glibetic, G. Poznanovic, J. Savic, M. Petrovic. 1991. Effect of lethal scald on the mechanisms of acute-phase protein synthesis in rat liver. Circ. Shock. 33:98.[Medline]
  21. Xia, Z. F., M. I. Coolbaugh, F. He, D. N. Herndon, J. Papaconstantinou. 1992. The effects of burn injury on the acute phase response. J. Trauma 32:245.[Medline]
  22. Okamura, H., H. Tsutsi, T. Komatsu, M. Yutsudo, A. Hakura, T. Tanimoto, K. Torigoe, T. Okura, Y. Nukada, K. Hattori, et al 1995. Cloning of a new cytokine that induces IFN-{gamma} production by T cells. Nature 378:88.[Medline]
  23. Okamura, H., H. Tsutsui, S. Kashiwamura, T. Yoshimoto, K. Nakanishi. 1998. Interleukin-18: a novel cytokine that augments both innate and acquired immunity. Adv. Immunol. 70:281.[Medline]
  24. Baker, C. C., I. H. Chaudry, H. O. Gaines, A. E. Baue. 1983. Evaluation of factors affecting mortality rate after sepsis in a murine cecal ligation and puncture model. Surgery 94:331.[Medline]
  25. Dobashi, H., S. Seki, Y. Habu, T. Ohkawa, S. Takeshita, H. Hiraide, I. Sekine. 1999. Activation of mouse liver natural killer cells and NK1.1+ T cells by bacterial superantigen-primed Kupffer cells. Hepatology 30:430.[Medline]
  26. Tsukahara, A., S. Seki, T. Iiai, T. Moroda, H. Watanabe, S. Suzuki, T. Tada, H. Hiraide, K. Hatakeyama, T. Abo. 1997. Mouse liver T cells: their change with aging and in comparison with peripheral T cells. Hepatology 26:301.[Medline]
  27. Fultz, M. J., S. A. Barber, C. W. Dieffenbach, S. N. Vogel. 1993. Induction of IFN-{gamma} in macrophages by lipopolysaccharide. Int. Immunol. 5:1383.[Abstract/Free Full Text]
  28. Gregory, S. H., L. K. Barczynski, E. J. Wing. 1992. Effector function of hepatocytes and Kupffer cells in the resolution of systemic bacterial infections. J. Leukocyte Biol. 51:421.[Abstract]
  29. Schultz, D. R., P. I. Arnold. 1990. Properties of four acute phase proteins: C-reactive protein, serum amyloid A protein, {alpha}1-acid glycoprotein, and fibrinogen. Semin. Arthritis. Rheum. 20:129.[Medline]
  30. Bharadwaj, D., M. P. Stein, M. Volzer, C. Mold, T. W. Du Clos. 1999. The major receptor for C-reactive protein on leukocytes is Fc{gamma} receptor II. J. Exp. Med. 190:585.[Abstract/Free Full Text]
  31. 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]
  32. Rutenfranz, I., H. Kirchner. 1988. Pharmacokinetics of recombinant murine interferon-{gamma} in mice. J. Interferon Res. 8:573.[Medline]



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