Inhibition of concanavalin A-induced hepatic injury of mice by bacterial lipopolysaccharide via the induction of IL-6 and the subsequent reduction of IL-4: the cytokine milieu of concanavalin A hepatitis

doi:10.1016/S0168-8278(99)80159-7

Journal of Hepatology

Volume 31, Issue 1, July 1999, Pages 18-26

https://doi.org/10.1016/S0168-8278(99)80159-7 Get rights and content

Abstract

Background/Aims: Liver natural killer 1.1 antigen (NK1)⁺ T cells and IL-4 play a crucial role in concanavalin-A (Con-A)-induced hepatic injury in mice, and a T helper (Th) 2 immune response was thus suggested to be involved. This study was designed to examine the effect of bacterial lipopolysaccharide (LPS), a strong inducer of a Th 1 immune response, on Con-A hepatic injury and also to clarify further the cytokine milieu of Con-A hepatitis.

Methods: LPS were injected into mice before Con-A injection to evaluate the effect on hepatic injury. The effect of the pretreatment with various T1 and Th2 cytokines or anti-cytokine antibodies on Con-A hepatitis was also examined.

Results: LPS in quantities ≧500 ng/mouse, when injected 24 h before Con-A injection, abrogated the Con-A-induced elevation of transaminases, hepatocyte destruction and serum IL-4 elevation. This LPS inhibitory effect was blocked when the mice were injected with either anti-IL-6 antibody before LPS injection or IL-4 before Con-A injection. IL-6, but neither IL-10 nor IL-12 pretreatment suppressed Con-A-induced IL-4 production and hepatitis. NK1⁺ T cells produced IL-4 while both NK1⁺ T cells and NK1⁻ T cells produced IFN-γ. Not only anti-IL-4 antibody but also the anti-IFN-γ antibody pretreatment inhibited Con-A hepatitis. However, although the anti-IL4 antibody suppressed IL-4 alone, the anti-IFN-γ Ab unexpectedly inhibited both IFN-γ and IL-4 elevation, while IL-4 injection evoked a moderate Con-A hepatitis even in the anti-IFN-γ antibody-treated mice. Furthermore, the IL-4 mutant mice did not develop Con-A hepatitis.

Conclusion: LPS inhibited Con-A hepatitis by inducing IL-6 and thereby inhibited IL-4 synthesis from NK1⁺ T cells. Although both IL-4 and IFN-γ were required for the full induction of Con-A hepatic injury, exogenous IL-4 evoked a moderate Con-A hepatitis, even in the absence of IFN-γ.

Section snippets

Mice

Six-eight-week-old male C57BL/6 (B6) mice (H-2^b) were purchased from SLC Japan Inc. (Hamamatsu, Japan). IL-4 gene-disrupted (IL-4 −/−) mice of B6 background (14) were kindly provided by Dr. F. Sendo at Yamagata University School of Medicine, Yamagata, Japan. All mice were fed under specific pathogen-free conditions.

Reagents

Con-A was purchased from Vector Laboratories, Inc. (Burlingame, CA, USA). Escherichia coli-derived LPS was purchased from Sigma (St. Louis, MO, USA). Recombinant murine IL-4, IL-6,

LPS inhibited Con-A hepatic injury in a dosedependent manner

When the mice were pretreated with 5 μg LPS (i.p.) 24 h before i.v. Con-A injection (0.5 mg/mouse), Con-A hepatitis was thus greatly suppressed (Fig. 1a). When various doses of LPS were injected, more than 500 ng/mouse of LPS inhibited transaminase elevation (Fig. 1b). A histological examination confirmed that 5 μg LPS inhibited hepatocyte destruction (Fig. 2). The protective effect of LPS diminished when injected within 10 h before the Con-A injection (data not shown).

LPS inhibited the IL-2 receptor α (IL-2Rα) expression of hepatic T cells

We recently reported

Discussion

In the present study, we have demonstrated that LPS inhibits Con-A hepatitis by inducing IL-6 production and subsequently inhibiting IL-4 production. LPS also inhibited the IL-2R expression of liver T cells. IL-6, but not IL-10 nor IL-12 pretreatment inhibited Con-A hepatitis. Although pretreatment of the mice with either anti-IL-4 Ab or anti-IFN-γ Ab suppressed Con-A-induced hepatic injury, anti-IFN-γ Ab inhibited both IFN-γ and IL-4 production, while anti-IL-4 Ab inhibited only IL-4

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TNFα, IL-6, and IL-10 are known to play important roles in the modulation of injury at larger doses of Con A. For example, TNFα, a cytokine released primarily by activated macrophages and T cells (Gantner et al., 1996), is a major effector of liver injury induced by large, hepatotoxic doses of Con A; neutralization of TNFα completely protected against hepatotoxicity (Gantner et al., 1995; Mizuhara et al., 1994). IL-6 plays contrasting roles in the development of injury after Con A. IL-6 was protective if administered or induced prior to Con A treatment (Nishikage et al., 1999), whereas recombinant IL-6 exacerbated injury when administered after Con A (Mizuhara et al., 1994). The anti-inflammatory cytokine IL-10 was induced early after Con A administration.
Inflammation plays a major role in immune-mediated liver injury, and exposure to environmental pollutants such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) has been reported to alter the inflammatory response as well as affect immune cell activity. In this study, we tested the hypothesis that TCDD pretreatment exacerbates hepatotoxicity in a murine model of immune-mediated liver injury induced by concanavalin A (Con A) administration. Mice were pretreated with 30 μg/kg TCDD or vehicle control on day zero and then given either Con A or saline intravenously on day four. Mice treated with TCDD did not develop liver injury; however, TCDD pretreatment increased liver injury resulting from moderate doses of Con A (4–10 mg/kg). TCDD-pretreated mice had altered plasma concentrations of inflammatory cytokines, including interferon gamma (IFNγ), and TCDD/Con A-induced hepatotoxicity was attenuated in IFNγ knockout mice. At various times after treatment, intrahepatic immune cells were isolated, and expression of cell activation markers as well as cytolytic proteins was determined. TCDD pretreatment increased the proportion of activated natural killer T (NKT) cells and the percent of cells expressing Fas ligand (FasL) after Con A administration. In addition FasL knockout mice and mice treated with CD18 antiserum were both protected from TCDD/Con A-induced hepatotoxicity, suggesting a requirement for direct cell–cell interaction between effector immune cells and parenchymal cell targets in the development of liver injury from TCDD/Con A treatment. In summary, exposure to TCDD increased NKT cell activation and exacerbated immune-mediated liver injury induced by Con A through a mechanism involving IFNγ and FasL expression.
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However, we recently observed that platelets are protective in Fas-mediated hepatitis, when the hepatitis is local or not severe [21]. A very interesting point to note is that although LPS induces HPA, as described above, LPS-pretreatment has been shown to protect mice against the hepatitis induced by either ConA or GalN + LPS [21–25]. As yet, the precise mechanisms underlying this protection remain unclear.
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Inhibition of concanavalin A-induced hepatic injury of mice by bacterial lipopolysaccharide via the induction of IL-6 and the subsequent reduction of IL-4: the cytokine milieu of concanavalin A hepatitis

Abstract

Section snippets

Mice

Reagents

LPS inhibited Con-A hepatic injury in a dosedependent manner

LPS inhibited the IL-2 receptor α (IL-2Rα) expression of hepatic T cells

Discussion

Hepatology

J Immunol Methods

J Immunol Methods

Cell

Blood

Gastroenterology

Hepatology

Relationship between intermediate TCR cells and NK1.1+ T cells in various immune organs. NK1+ T cells are present within a population of intermediate TCR cells

J Immunol

Mouse liver T cells: their change with aging and in comparison with peripheral T cells

Hepatology

NK1.1+CD4+CD8− thymocytes with specific lymphokine secretion

Eur J Immunol

Murine thymic CD4+ T cell subset (Thy0) that secretes diverse cytokines and overexpress the Vβ8 T cell receptor gene family

J Exp Med

Cytotoxic NK1 Ag+ αβ T cells with intermediate TCR induced in the liver of mice by IL-12

J Immunol

Liver NK1.1+ CD4+ cells activated by IL-12 as a major effect or in inhibition of experimental tumor metastasis

J Immunol

IL-12 induces cytotoxic NK1+ αβ T cells in the lung of euthymic and athymic mice

Immunology

Involvement of NK1.1+ T cells and their IFN-γ production in the generalized Shwartzman reaction

J Immunol

A T cell-dependent experimental liver injury in mice inducible by Concanavalin A.

J Clin Invest

Relationship between intermediate TCR cells and NK1.1⁺ T cells in various immune organs. NK1⁺ T cells are present within a population of intermediate TCR cells

NK1.1⁺CD4⁺CD8⁻ thymocytes with specific lymphokine secretion

Murine thymic CD4⁺ T cell subset (Thy0) that secretes diverse cytokines and overexpress the Vβ8 T cell receptor gene family

Cytotoxic NK1 Ag⁺ αβ T cells with intermediate TCR induced in the liver of mice by IL-12

Liver NK1.1⁺ CD4⁺ cells activated by IL-12 as a major effect or in inhibition of experimental tumor metastasis

IL-12 induces cytotoxic NK1⁺ αβ T cells in the lung of euthymic and athymic mice

Involvement of NK1.1⁺ T cells and their IFN-γ production in the generalized Shwartzman reaction