Roquin-2 shares functions with its paralog Roquin-1 in the repression of mRNAs controlling T follicular helper cells and systemic inflammation.

Accumulation of T follicular helper (Tfh) cells and proinflammatory cytokines drive autoantibody-mediated diseases. The RNA-binding protein Roquin-1 (Rc3h1) represses the inducible costimulator ICOS and interferon-γ (IFN-γ) in T cells to prevent Tfh cell accumulation. Unlike Rc3h1(san) mice with a mutation in the ROQ domain of Roquin-1, mice lacking the protein, paradoxically do not display increased Tfh cells. Here we have analyzed mice with mutations that eliminate the RING domain from Roquin-1 or its paralog, Roquin-2 (Rc3h2). RING or ROQ mutations both disrupted Icos mRNA regulation by Roquin-1, but, unlike the ROQ mutant that still occupied mRNA-regulating stress granules, RING-deficient Roquin-1 failed to localize to stress granules and allowed Roquin-2 to compensate in the repression of ICOS and Tfh cells. These paralogs also targeted tumor necrosis factor (TNF) in nonlymphoid cells, ameliorating autoantibody-induced arthritis. The Roquin family emerges as a posttranscriptional brake in the adaptive and innate phases of antibody responses.

LIGHT (TNFSF14/CD258) is a decisive factor for recovery from experimental autoimmune encephalomyelitis.

The TNF superfamily ligand LIGHT (lymphotoxin-like, exhibits inducible expression and competes with HSV glycoprotein D for herpesvirus entry mediator [HVEM], a receptor expressed by T lymphocytes) has been shown to play a role in T cell costimulation and be involved in apoptosis of mononuclear cells. As both T cells and monocytes are key components in the development and progression of experimental autoimmune encephalomyelitis (EAE), we studied the role of LIGHT in EAE. Following immunization with myelin oligodendrocyte glycoprotein peptide (35-55), LIGHT-deficient mice developed severe EAE that resulted in an atypically high mortality rate. Histological examinations revealed intensive activation of microglia/macrophages in the CNS and higher numbers of apoptotic cells within the CNS parenchyma of LIGHT-deficient mice. However, myelin oligodendrocyte glycoprotein peptide-specific CD4(+) T cells from LIGHT-deficient mice showed reduced IFN-γ and IL-17 production and migration. Serum levels of reactive nitrogen intermediates and CNS transcripts of several proinflammatory cytokines and chemokines were also substantially decreased in the absence of LIGHT. EAE adoptive transfer experiments and bone marrow chimeras indicated that expression of LIGHT on donor cells is not required for disease induction. However, its expression on CNS host cells is a decisive factor to limit disease progression and tissue damage. Together, these data show that LIGHT expression is crucially involved in controlling activated macrophages/microglia during autoimmune CNS inflammation.

Neuronal nitric oxide synthase plays a key role in CNS demyelination.

Nitric oxide (NO) is a small, short-lived molecule released from a variety of cells that is implicated in a multitude of biological processes. In pathological conditions, overproduction of NO may lead to the generation of highly reactive species, such as peroxynitrite and stable nitrosothiols, that may cause irreversible cell damage. Accordingly, several studies have suggested that NO may be involved in the pathogenesis of various neuroinflammatory/degenerative diseases. Increased concentrations of NO in the CNS in such cases are usually attributed to an increase in the inducible isoform of NO synthase (iNOS) usually produced by inflammatory cells. However, recent reports have suggested that the constitutive isoforms of NOS, neuronal (nNOS) and endothelial (eNOS), can also play a role. Here we examined the role that the constitutive isoforms of NOS might play in the cuprizone-induced model of demyelination/remyelination. Our results demonstrate that demyelination was greatly prevented in mice lacking nNOS. Protection was associated with a dramatic increase in mature oligodendrocyte survival and a decrease in apoptosis. Moreover, nNOS-/- mice did not respond to cuprizone with the extensive recruitment of microglia/macrophages and astrocytes, which is a typical feature in wild-type mice. Although demyelinating less, nNOS-/- mice exhibited a delay in remyelination. In eNOS-/- mice, demyelination progressed to the same extent as in wild type, but they showed a slight delay in spontaneous remyelination. In conclusion, this study highlights the importance of considering the source of NO when assessing its role in neuroinflammation/degeneration and emphasizes the differing pathological effects driven by the different NOS isoforms.

Music, immunity and cancer.

The effects of music on the immune system and cancer development were evaluated in rodents subjected to sound stress. Animals were exposed daily to broad band noise around midnight and/or music for 5 hours on the following morning. Thymus and spleen cellularity, peripheral T lymphocyte population, the proliferative response of spleen cells to mitogen concanavalin A and natural killer cell activity were calculated in BALB/c mice. Sprague Dawley rats were injected i.v. with Walker 256 carcinosarcoma cells; 8 days later the rats were sacrificed and the number of metastatic nodules on the surface of the lungs was calculated macroscopically. Music reduced the suppressive effects of stress on immune parameters in mice and decreased the enhancing effects of stress on the development of lung metastases provoked by carcinosarcoma cells. Music enhanced the immune parameters and the anti-tumor response in unstressed rodents. Our data at present demonstrates that music can effectively reverse adverse effects of stress on the number and capacities of lymphocytes that are required for an optimal immunological response against cancer in rodents.

Demyelination caused by the copper chelator cuprizone halts T cell mediated autoimmune neuroinflammation.

Myelin reactive T cells are central in the development of the autoimmune response leading to CNS destruction in Multiple Sclerosis and Experimental Autoimmune Encephalomyelitis (EAE). Investigations on the mechanisms underlying the activation and expansion of myelin reactive T have stressed the importance of non-autoimmune conditions impinging the autoimmune repertoire potentially involved in the disease. Here, we show that CNS injury caused by the toxic cuprizone results in the generation of immunoreactivity towards several myelin components. Paradoxically, exposure to CNS injury does not increase the susceptibility to develop EAE, but render mice protected to the pathogenic autoimmune response against myelin antigens.

Deleterious role of IFNgamma in a toxic model of central nervous system demyelination.

Interferon-gamma (IFNgamma) is a pleiotropic cytokine that plays an important role in many inflammatory processes, including autoimmune diseases such as multiple sclerosis (MS). Demyelination is a hallmark of MS and a prominent pathological feature of several other inflammatory diseases of the central nervous system, including experimental autoimmune encephalomyelitis, an animal model of MS. Accordingly, in this study we followed the effect of IFNgamma in the demyelination and remyelination process by using an experimental autoimmune encephalomyelitis model of demyelination/remyelination after exposure of mice to the neurotoxic agent cuprizone. We show that demyelination in response to cuprizone is delayed in mice lacking the binding chain of IFNgamma receptor. In addition, IFNgammaR(-/-) mice exhibited an accelerated remyelination process after cuprizone was removed from the diet. Our results also indicate that the levels of IFNgamma were able to modulate the microglia/macrophage recruitment to the demyelinating areas. Moreover, the accelerated regenerative response showed by the IFNgammaR(-/-) mice was associated with a more efficient recruitment of oligodendrocyte precursor cells in the demyelinated areas. In conclusion, this study suggests that IFNgamma regulates the development and resolution of the demyelinating syndrome and may be associated with toxic effects on both mature oligodendrocytes and oligodendrocyte precursor cells.

A BAFF antagonist suppresses experimental autoimmune encephalomyelitis by targeting cell-mediated and humoral immune responses.

BAFF [B cell-activating factor of the tumour necrosis factor (TNF) family] and APRIL (a proliferation-inducing ligand) are two TNF family members with shared receptors. While, physiological roles for APRIL are not fully understood, BAFF is critical for B cell homeostasis and also acts as a co-stimulator of T cells. Using a B and T cell-mediated mouse model of multiple sclerosis (MS), myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE), we observed that a BAFF/APRIL antagonist (soluble BCMA-Fc) inhibited central nervous system inflammation and demyelination such that it suppressed the onset and progression of clinical symptoms of EAE. In addition to dramatically reducing the titre of MOG-specific auto-antibodies, this treatment also induced a switch in the subtype of the T(h) cell population characterized by marked alterations in cytokine production following re-stimulation with MOG in vitro. Indeed, hBCMA-Fc therapy led to significant increases in the level of transforming growth factor beta, while the levels of T(h)1 cytokines were markedly diminished. These results not only identify BAFF as a critical factor in maintaining humoral immunity in EAE but also support its role in T lymphocyte responses. Our findings demonstrate that hBCMA-Fc acts on both effector arms of the immune response in EAE, a characteristic that may be of significant therapeutic value in the treatment of MS.

Single-step purification and refolding of recombinant mouse and human myelin oligodendrocyte glycoprotein and induction of EAE in mice.

The extracellular domain of human and rat MOG (ED-MOG) induces experimental autoimmune encephalomyelitis (EAE) when injected into susceptible animals. EAE is a T cell-mediated disease of the central nervous system commonly used as an animal model for human multiple sclerosis. Here, we describe a straightforward procedure for the purification and refolding of mouse and human ED-MOG overexpressed in Escherichia coli as inclusion bodies. Following solubilization and purification using Ni-NTA resin chromatography under denaturing conditions, a column-based refolding proceeded in renaturation buffer supplemented with a glutathione redox buffer system. Using this approach up to 33 mg of highly pure soluble proteins was obtained per liter of expression culture. The ability of purified proteins to induce EAE was evaluated in three strains of mice. We believe that the strategy described here would facilitate researchers to carry out encephalitogenic as well as structure-function studies of this autoantigen. Additionally, we show for the first time that mouse ED-MOG induces severe disease in mice.

Tolerance induction by molecular mimicry: prevention and suppression of experimental autoimmune encephalomyelitis with the milk protein butyrophilin.

Multiple sclerosis (MS) is a common inflammatory disease of the central nervous system. Although the etiology of MS remains unknown, studies in experimental autoimmune encephalomyelitis (EAE) have suggested that foreign molecules, which show molecular mimicry with myelin antigens, may play an important role as causative agents of the human disease. In this study, we investigate the molecular mimicry between the extracellular Ig-like domain of the cow's milk protein butyrophilin (BTN) and the extracellular domain of myelin oligodendrocyte glycoprotein (MOG), a candidate autoantigen in MS. Interestingly, we found that as a result of a non-pathogenic cross-reactivity that is localized to a subdominant region of MOG, treatment of C57BL/6 mice with BTN either before or after immunization with MOG was shown to prevent and also suppress the clinical manifestations of EAE. BTN treatment resulted in a significant reduction in both proliferation and production of Th1-related cytokines (IFN-gamma, IL-2, IL-12 and granulocyte macrophage colony stimulating factor) in response to MOG. This specific inhibition was consistently associated with an up-regulation in IL-10 secretion. Furthermore, adoptive transfer of BTN-specific T cells prior to active immunization with MOG resulted in a transitory reduction of the clinical symptoms. Our results suggest that the clinical improvement associated with BTN treatment involved the combination of both anergy and regulatory cells secreting high levels of IL-10. In conclusion, we show that despite the traditional link between molecular mimicry and pathogenic immune response, environmental agents that share homology with autoantigens may also represent a source of cells with a protective phenotype.

The magnitude and encephalogenic potential of autoimmune response to MOG is enhanced in MOG deficient mice.

Myelin oligodendrocyte glycoprotein (MOG) is a minor component of central nervous system myelin presumably implicated in the pathogenesis of Multiple Sclerosis (MS). Immunization with MOG leads to the development of Experimental Autoimmune Encephalomyelitis (EAE), the experimental model of MS. It has been suggested that its encephalitogenic potential may be due to the lack of MOG self-immune tolerance. To clarify this, we have generated a MOG deficient mouse (MOG(-/-)) strain. Surprisingly, MOG(35-55)specific proliferation and Th1-type cytokine production were markedly enhanced in MOG(-/-)mice compared to wild type control. Furthermore, adoptive transfer of MOG(35-55)specific T cells, isolated from MOG deficient mice, into wild-type recipients resulted in the development of a more severe disease, indicating a high capacity of MOG(-/-)T cells to initiate effector responses. Interestingly, T cell reactivity to overlapping MOG peptides in MOG(-/-)mice did not reveal new potential immunodominant epitopes in H-2(b)mice. Taken together, our data suggests that MOG self-tolerance modulates the encephalitogenic potential of autoreactive MOG T cells in the periphery.