concanavalin-a and gadolinium-chloride

Increasing evidence suggests that a close interaction of Kupffer cells with T cells plays a central role in concanavalin A-induced hepatic injury in mice, but the underlying mechanisms remain obscure. The present study aimed to determine the relative roles of Th1 and Th17 type responses in concanavalin A-induced hepatic injury in mice, and to investigate whether or not Kupffer cells contribute to hepatic injury via a Th1 or Th17 type response-dependent pathway.. Immune-mediated hepatic injury was induced in C57BL/6 mice by intravenous injection of concanavalin A. Kupffer cells were inactivated by pretreatment with gadolinium chloride 24 hours before the concanavalin A injection. The interferon-gamma (IFN-gamma) and interleukin-17 (IL-17) pathways were blocked by specific neutralizing antibodies. Hepatic injury was assessed using serum transferase activity and pathological analysis. Expression of inflammatory cytokines within the liver was detected by real-time polymerase chain reaction and immunohistochemistry.. Neutralization of IFN-gamma significantly attenuated concanavalin A-induced hepatic injury. However, neutralization of IL-17 failed to suppress the injury. Inactivation of Kupffer cells by gadolinium chloride pretreatment protected against concanavalin A-induced injury and significantly reduced hepatic cytokine levels including TNF-alpha, IL-6 and IFN-gamma but not IL-17.. Our findings suggest that Kupffer cells contribute to concanavalin A-induced hepatic injury via a Th1 type response-dependent pathway and production of inflammatory cytokines including TNF-alpha, IL-6 and IFN-gamma.

Topics: Animals; Chemical and Drug Induced Liver Injury; Concanavalin A; Female; Gadolinium; Interferon-gamma; Interleukin-17; Kupffer Cells; Mice; Mice, Inbred C57BL; Th1 Cells; Th17 Cells

Although concanavalin A (Con-A)-induced experimental hepatitis is thought to be induced by activated T cells, natural killer T (NKT) cells, and cytokines, precise mechanisms are still unknown. In the current study, we investigated the roles of Kupffer cells, NKT cells, FasL, tumor necrosis factor (TNF), and superoxide in Con-A hepatitis in C57BL/6 mice. Removal of Kupffer cells using gadolinium chloride (GdCl(3)) from the liver completely inhibited Con-A hepatitis, whereas increased serum TNF and IFN-gamma levels were not inhibited at all. Unexpectedly, anti-FasL antibody pretreatment did not inhibit Con-A hepatitis, whereas it inhibited hepatic injury induced by a synthetic ligand of NKT cells, alpha-galactosylceramide. Furthermore, GdCl(3) pretreatment changed neither the activation-induced down-regulation of NK1.1 antigens as well as T cell receptors of NKT cells nor the increased expression of the CD69 activation antigen of hepatic T cells. CD68(+) Kupffer cells greatly increased in proportion in the early phase after Con-A injection; this increase was abrogated by GdCl(3) pretreatment. Anti-TNF antibody (Ab) pretreatment did not inhibit the increase of Kupffer cells, but it effectively suppressed superoxide/reactive oxygen production from Kupffer cells and the resulting hepatic injury. Conversely, depletion of NKT cells in mice by NK1.1 Ab pretreatment did suppress both the increase of CD68(+) Kupffer cells and Con-A hepatitis. Consistently, the diminution of oxygen radicals produced by Kupffer cells by use of free radical scavengers greatly inhibited Con-A hepatitis without suppressing cytokine production. However, adoptive transfer experiments also indicate that a close interaction/cooperation of Kupffer cells with NKT cells is essential for Con-A hepatitis.. Superoxide produced by Kupffer cells may be the essential effector in Con-A hepatitis, and TNF and NKT cells support their activation and superoxide production.

Topics: Animals; Anti-Inflammatory Agents; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Cell Communication; Chemical and Drug Induced Liver Injury; Concanavalin A; Cytokines; Disease Models, Animal; Gadolinium; Interferon-gamma; Killer Cells, Natural; Kupffer Cells; Male; Mice; Mice, Inbred C57BL; Reactive Oxygen Species; Superoxides; T-Lymphocytes; Tumor Necrosis Factor-alpha

Cytochrome P450 (CYP) gene expression in the livers of mice with concanavalin A-induced hepatitis was examined. Treatment of mice with concanavalin A (10 mg/kg, i.v.) elevated plasma alanine aminotransaminase activity. In normal liver, CYP1A2, 3A and 2E1 mRNAs were expressed, and concanavalin A treatment differentially suppressed the expression of these CYP genes. Gadolinium chloride (40 mg/kg, i.p.) treatment, which inhibited the concanavalin A-induced elevation of plasma alanine aminotransferase activity without affecting concanavalin A-induced cytokine expression, counteracted the concanavalin A-induced suppression of CYP gene expression in the liver. Kupffer cell function or hepatic injury might contribute to the concanavalin A-induced suppression of CYP gene expression.

Topics: Animals; Chemical and Drug Induced Liver Injury; Concanavalin A; Cytochrome P-450 Enzyme System; Female; Gadolinium; Gene Expression Regulation, Enzymologic; Liver; Mice; Mice, Inbred BALB C

The effect of Kupffer cell depression on concanavalin A (Con A)-induced cytokine mRNA expression in the liver was studied. Gadolinium chloride (GdCl3) is a commonly used Kupffer cell inhibitor. GdCl3 (40 mg/kg, i.p.) was injected into each mouse, and 24 hr later, Con A (0.2 mg/mouse) was administered. Plasma was obtained at 24 hr after Con A treatment for alanine aminotransferase (ALT) measurement. GdCl3 treatment inhibited Con A-induced elevation of ALT. However, it did not inhibit Con A-induced interleukin-2 or tumor necrosis factor-alpha mRNA expression. The present results suggest that Kupffer cells are not responsible for Con A-induced cytokine expression in the liver.

Topics: Alanine Transaminase; Animals; Anti-Inflammatory Agents; Concanavalin A; Cytokines; Female; Gadolinium; Gene Expression; Gene Expression Regulation; Interleukin-2; Kupffer Cells; Liver; Mice; Mice, Inbred BALB C; RNA, Messenger; Tumor Necrosis Factor-alpha

CD4+ T lymphocytes have been identified as being responsible for organ damage in the murine model of experimental liver injury induced by intravenous injection of concanavalin A (Con A). Liver sinusoidal endothelial cells (SEC) and Kupffer's cells (KC) are among the first cells that come into contact with lymphocytes in the liver sinusoid. We aimed to investigate the respective role of these cell populations in the initial steps of T-cell-mediated liver injury in Con A-induced hepatitis. By electron microscopy, we could show that intravenously applied Con A bound predominantly to SEC but not to KC. KC depletion by gadolinium chloride treatment of mice did not result in protection from liver injury, indicating that KCs are not primarily involved in the generation of liver injury. We could show that a CD4+ T-cell line (LNC.2) displayed selective cytotoxicity toward SEC (>50%) but not KC (12%) or fibroblasts (5%) in the presence of Con A in vitro. Microscopic observation revealed that the SEC monolayer was rapidly destroyed by LnC2 in the presence of Con A. Specificity of the Con A-induced cytotoxicity was shown by the ability of a competitive ligand, methyl-alpha-D-mannopyranoside, to reduce T-cell-mediated cytotoxicity to SEC by more than 50%. Tumor necrosis factor alpha (TNF-alpha) was produced by LnC2 in high amounts after Con A stimulation (>6 ng/mL), but antiserum to TNF-alpha did not reduce LnC2-mediated cytotoxicity toward SEC. In conclusion, we could show for the first time that liver SECs have accessory function and are selectively destroyed by CD4+ T lymphocytes in the presence of Con A. We speculate that SEC damage is an early event in T-cell-mediated liver injury recruiting T lymphocytes from the sinusoidal circulation. Loss of the SEC barrier function then exposes underlying hepatocytes to further attack by activated T lymphocytes. These results offer a model of initiating events in T-cell-mediated liver diseases, such as viral or autoimmune hepatitis, and suggest an important role for sinusoidal endothelial cells.

Topics: Animals; CD4-Positive T-Lymphocytes; Concanavalin A; Endothelium; Female; Fibroblasts; Gadolinium; Hepatic Encephalopathy; Injections, Intravenous; Kupffer Cells; Mice; Mice, Inbred BALB C; Organ Specificity; T-Lymphocytes, Cytotoxic; Tumor Necrosis Factor-alpha