Caigan Du

EXPRESSION OF IL-12 IN CNS AND LYMPHOID ORGANS OF MICE WITH EXPERIMENTAL ALLERGIC ENCEPHALITIS

EAE is a Th1 cell-mediated inflammatory autoimmune demyelinating disease of the central nervous system. IL-12 is a 70 kd heterodimeric cytokine, capable of regulating a wide range of immune functions. In view of its crucial role in the development of Th1 immune responses, we studied the expression of IL-12 p40 in the CNS and lymphoid organs of mice with EAE. RT-PCR analysis showed an increase in the expression of IL-12 p40 in brain and spinal cord during the acute paralytic phase of EAE and that decreased upon clinical recovery. The expression of p40 mRNA was also increased in spleen, lymph node and liver along with an elevated levels of circulating serum IL-12 during the height of disease. In vivo administration of rIL-12 increased the proliferative response and IFN-gamma production of MBP sensitized T cells and that was decreased following treatment with anti-IL-12 antibody. The expression of IL-12 in the target and lymphoid organs of animals with EAE, the induction of a Th1 type immune response following immunization with neuronal antigens and the inhibition of clinical disease upon treatment with anti-IL-12 antibody, suggest the crucial role of IL-12 in the pathogenesis of EAE.

Administration of Dehydroepiandrosterone Suppresses Experimental Allergic Encephalomyelitis in SJL/J Mice

Experimental allergic encephalomyelitis (EAE) is a Th1-mediated inflammatory demyelinating disease in the CNS, an animal model of multiple sclerosis. We have examined the effect of dehydroepiandrosterone (DHEA) on the development of EAE in mice. The addition of DHEA to cultures of myelin basic protein-primed splenocytes resulted in a significant decrease in T cell proliferation and secretion of (pro)inflammatory cytokines (IFN-γ, IL-12 p40, and TNF-α) and NO in response to myelin basic protein. These effects were associated with a decrease in activation and translocation of NF-κB. In vivo administration of DHEA significantly reduced the severity and incidence of acute EAE, along with a decrease in demyelination/inflammation and expressions of (pro)inflammatory cytokines in the CNS. These studies suggest that DHEA has potent anti-inflammatory properties, which at least are in part mediated by its inhibition of NF-κB activation.

Mechanism of inhibition of LPS-induced IL-12p40 production by IL-10 and TGF-beta in ANA-1 cells.

IL‐12, a macrophage‐derived proinflammatory cytokine, consists of two polypeptide subunits (p40 and p35) encoded by two separate genes. The p35 subunit is constitutively expressed, whereas the p40 subunit is induced after activation. The bioactive interleukin‐12 (IL‐12; p70) influences the development of Th1 responses and is a potent activator of natural killer (NK) and T cells. In contrast to IL‐12, transforming growth factor β (TGF‐β) and IL‐10 inhibit production of proinflammatory cytokines, including IL‐12, and attenuate Th1‐mediated immune response. We have examined the molecular mechanisms by which TGF‐β and IL‐10 inhibit production of the IL‐12p40 subunit in LPS‐stimulated murine macrophage cell line. We show that both IL‐10 and TGF‐β suppress IL‐12p40 production by inhibiting the transcription of IL‐12p40 gene. At equal concentrations, IL‐10 was more potent than TGF‐β in inhibiting IL‐12p40 gene transcription. TGF‐β also reduces the stability of IL‐12p40 mRNA, accounting thereby to an additional mechanism of inhibition of IL‐12 production. J. Leukoc. Biol. 64: 92–97; 1998.

Chlamydia pneumoniae Infection of the Central Nervous System Worsens Experimental Allergic Encephalitis

Experimental allergic encephalitis (EAE) is considered by many to be a model for human multiple sclerosis. Intraperitoneal inoculation of mice with Chlamydia pneumoniae, after immunization with neural antigens, increased the severity of EAE. Accentuation of EAE required live infectious C. pneumoniae, and the severity of the disease was attenuated with antiinfective therapy. After immunization with neural antigens, systemic infection with C. pneumoniae led to the dissemination of the organism into the central nervous system (CNS) in mice with accentuated EAE. Inoculation with Chlamydia trachomatis did not worsen EAE and infectious organisms were not seen in the CNS. These observations suggest that dissemination of C. pneumoniae results in localized infection in CNS tissues in animals with EAE. We propose that infection of the CNS by C. pneumoniae can amplify the autoreactive pool of lymphocytes and regulate the expression of an autoimmune disease.

Stimulation of Th2 response by high doses of dehydroepiandrosterone in KLH-primed splenocytes.

Although dehydroepiandrosterone (DHEA) has long been considered as a precursor for steroid hormones, it has also been shown to have regulatory effects in immune homeostasis. We have examined the effect of high DHEA doses on T cell proliferation, differentiation, and cytokine secretion patterns following stimulation with mitogens and soluble antigens. DHEA profoundly inhibited T cell receptor-mediated T cell proliferation in the upstream of IL-2R signaling. Addition of DHEA to KLH-primed splenocytes stimulated Th2 response, indicated by an increase of IL-4 or a decrease of IFN-γ production in the cultures. Further studies showed that DHEA enhanced IL-4, but inhibited IL-12-mediated T cell proliferation and IL-12 production in antigen-presenting cells (APCs). Our data demonstrated that supraphysiologic levels of DHEA favored Th2 immune responses in vitro by inhibition of IL-12 production from APCs and/or stimulation of Th2 proliferation during the interactions of T cells with APCs.

Increased Severity of Experimental Allergic Encephalomyelitis in lyn−/− Mice in the Absence of Elevated Proinflammatory Cytokine Response in the Central Nervous System

lyn, a member of the src kinase family, is an important signaling molecule in B cells. lyn−/− mice display hyperactive B-1 cells and IgM hyperglobulinemia. The role of lyn on T cell function and development of Th1-mediated inflammatory disease is not known. Therefore, we examined the effect of disruption of the lyn gene on the development of experimental allergic encephalomyelitis (EAE), a well-established Th1-mediated autoimmune disease. Following immunization with myelin oligodendrocyte protein (MOG) p35-55, lyn−/− mice had higher clinical and pathological severity scores of EAE when compared with wild type (WT). The increase in the severity of EAE in lyn−/− mice was not associated with a commensurate increase in the production of proinflammatory cytokines in the CNS. lyn−/− mice with EAE showed elevation in serum anti-IgM MOG Ab levels over that seen in WT mice, along with a modest increase in the mRNA levels of complement C5 and its receptor, C5aR, in the spinal cord. Transfer of serum from MOG-immunized lyn−/− mice worsened EAE in WT mice, suggesting a pathogenic role for anti-MOG IgM Abs in EAE. These observations underscore the potential role of lyn in regulation of Th1-mediated disease and the role of autoantibodies and complement in the development of EAE.

Inhibition of CD40 signaling pathway by tyrphostin A1 reduces secretion of IL-12 in macrophage, Th1 cell development and experimental allergic encephalomyelitis in SJL/J mice.

Activation of antigen presenting cells through the interaction of CD40 with its ligand is a critical co-stimulatory signal for IL-12 production and Th1 differentiation. Tyrphostins are organic molecules that inhibit the phosphorylation of protein tyrosine kinases. We show that tyrphostin A1 inhibits CD40L-stimulated IL-12 production in macrophage cultures and antigen-induced generation of Th1 cells. Our data also show that tyrphostin A1 blocks CD40L-induced translocation of NF-kappaB to the nucleus, and reduces the activation of IL-12 p40 gene. In vivo therapy with A1 leads to decrease in generation of myelin basic protein (MBP) specific encephalitogenic T cells. In addition, treatment of SJL/J mice with A1 results in attenuation of experimental allergic encephalomyelitis (EAE).

Amelioration of CR-EAE with lisofylline: effects on mRNA levels of IL-12 and IFN-γ in the CNS

Abstract CR-EAE is a Th1-mediated inflammatory autoimmune demyelinating disease of the CNS and serves as a model of human multiple sclerosis. Our previous studies have shown the protective effect of orally administered lisofylline in the prevention of active and passively induced acute EAE. In our present studies we have examined the efficacy and mechanism of action of lisofylline on CR-EAE. Lisofylline decreased the number and severity of paralytic attacks in mice with relapsing EAE. The reduction of clinical disease correlated with decreased levels of mRNA levels of IFN-γ but not of mRNA levels of IL-12. These studies suggest that lisofylline may be an effective therapeutic for established Th1 mediated autoimmune disease and that it acts by blocking IL-12R signaling and not IL-12 production in vivo.