SUMMARY On the lupus-prone MRL-lpr/lpr (MRL-lpr) background AM14 rheumatoid factor (RF) B cells are activated, differentiate into plasmablasts, and undergo somatic hypermutation outside of follicles. Using multiple strategies to impair T cells, we found that such AM14 B cell activation did not require T cells, but could be modulated by them. In vitro, the signaling adaptor MyD88 is required for IgG anti-chromatin to stimulate AM14 B cell proliferation when T cells are absent. However the roles of Toll-like receptors (TLRs) in AM14 B cell activation in vivo have not been investigated. We found that activation, expansion and differentiation of AM14 B cells depended on MyD88; however, mice lacking either TLR7 or TLR9 displayed partial defects, indicating complex roles for these receptors. T-independent activation of certain autoreactive B cells, which instead can gain stimuli via endogenous TLR ligands, may be the initial step in the generation of canonical autoantibodies.

Antibodies to myelin components are routinely detected in multiple sclerosis patients. However, their presence in some control subjects has made it difficult to determine their contribution to disease pathogenesis. Immunization of C57BL͞6 mice with either rat or human myelin oligodendrocyte glycoprotein (MOG) leads to experimental autoimmune encephalomyelitis (EAE) and comparable titers of anti-MOG antibodies as detected by ELISA. However, only immunization with human (but not rat) MOG results in a B cell-dependent EAE. In this study, we demonstrate that these pathogenic and nonpathogenic anti-MOG antibodies have a consistent array of differences in their recognition of antigenic determinants and biological effects. Specifically, substituting proline at position 42 with serine in human MOG (as in rat MOG) eliminates the B cell requirement for EAE. All MOG proteins analyzed induced high titers of anti-MOG (tested by ELISA), but only antisera from mice immunized with unmodified human MOG were encephalitogenic in primed B cell-deficient mice. Nonpathogenic IgGs bound recombinant mouse MOG and deglycosylated MOG in myelin (tested by Western blot), but only pathogenic IgGs bound glycosylated MOG. Only purified IgG to human MOG bound to live rodent oligodendrocytes in culture and, after cross-linking, induced repartitioning of MOG into lipid rafts, followed by dramatic changes in cell morphology. The data provide a strong link between in vivo and in vitro observations regarding demyelinating disease, further indicate a biochemical mechanism for anti-MOGinduced demyelination, and suggest in vitro tools for determining autoimmune antibody pathogenicity in multiple sclerosis patients. multiple sclerosis ͉ experimental autoimmune encephalomyelitis ͉ lipid rafts ͉ B cell-deficient mice ͉ encephalitogenicity M ultiple sclerosis (MS) is a chronic demyelinating disease of the central nervous system (CNS) in which both T cells and antibodies against myelin antigens are routinely detected (1, 2). B cell responses in MS pathogenesis are implicated by the presence of Ig deposits and myelin debris in demyelinating lesions (3-8), and the observation that plasma exchange dramatically reduces clinical disease in a subset of patients (9). Of particular interest to the present study, antibodies to myelin oligodendrocyte glycoprotein (MOG) are detected in the sera and plaques of MS patients (10), and thus are possible predictors of disease progression (11). However, because some control subjects can also harbor anti-myelin antibodies (1, 2, 12), their contribution to MS pathogenesis has been controversial and difficult to identify in individual patients. Further complicating the issue, MS may be several diseases of differing etiologies (5), whereby anti-myelin antibodies may be pathogenic in some forms of MS but merely a reflection of tissue damage in others. Thus, an understanding of whether anti-myelin antibodies are in fact pathogenic, and if so, by what mechanisms they operate, could provide important information for novel diagnosti...

Protective high-affinity antibody responses depend on competitive selection of B cells carrying somatically mutated B-cell receptors by follicular helper T (TFH) cells in germinal centres. The rapid T-B-cell interactions that occur during this process are reminiscent of neural synaptic transmission pathways. Here we show that a proportion of human TFH cells contained dense-core granules marked by chromogranin B, which are normally found in neuronal presynaptic terminals storing catecholamines such as dopamine. TFH cells produce high amounts of dopamine and released it upon cognate interaction with B cells. Dopamine causes rapid translocation of intracellular ICOSL (inducible T-cell co-stimulator ligand, also known as ICOSLG) to the B-cell surface, which enhances accumulation of CD40L and chromogranin B granules at the human TFH cell synapse and increases the synapse area. Mathematical modelling suggests that faster dopamine-induced T-B-cell interactions increase total germinal centre output and accelerate it by days. Delivery of neurotransmitters across the T-B-cell synapse may be advantageous in the face of infection.

Extrafollicular (EF) B-cell responses are increasingly being recognized as an alternative pathway of B-cell activation, particularly in autoimmunity. Critical cellular interactions required for the EF Bcell response are unclear. A key question in autoimmunity, in which Toll-like receptor (TLR) signals are costimulatory and could be sufficient for B-cell activation, is whether T cells are required for the response. This is pivotal, because autoreactive B cells are considered antigen-presenting cells for autoreactive T cells, but where such interactions occur has not been identified. Here, using AM14 site-directed transgenic rheumatoid factor (RF) mice, we report that B cells can be activated, differentiate, and isotypeswitch independent of antigen-specific T-cell help, αβ T cells, CD40L signaling, and IL-21 signaling to B cells. However, T cells do dramatically enhance the response, and this occurs via CD40L and IL-21 signals. Surprisingly, the response is completely inducible T-cell costimulator ligand independent. These results establish that, although not required, T cells substantially amplify EF autoantibody production and thereby implicate T-independent autoreactive B cells as a potential vector for breaking T-cell tolerance. We suggest that these findings explain why autoreactivity first focuses on self-components for which B cells carry TLR ligands, because these will uniquely be able to activate B cells independently of T cells, with subsequent T-B interactions activating autoreactive T cells, resulting in chronic autoimmunity.systemic lupus | autoantibodies