Toll-like receptors are important in the activation of innate immunity, and CD40 is a molecule critical for many T and B cell responses. Whereas agonists for either pathway have been used as vaccine adjuvants, we show that a combination of Toll-like receptor (TLR)7 and CD40 agonists synergize to stimulate CD8+ T cell responses 10–20-fold greater than the use of either agonist alone. Antigen-specific CD8+ T cells elicited from combination CD40/TLR7 treatment demonstrated both lytic activities and interferon (IFN)γ production and an enhanced secondary response to antigenic challenge. Agonists for TLRs 2/6, 3, 4, and 9 also synergized with CD40 stimulation, demonstrating that synergy with the CD40 pathway is a property of TLR-derived stimuli in general. The CD8+ T cell expansion induced by CD40/TLR7 triggering was independent of CD4+ T cells, IFNγ, and IL-12 but dependent on B7-mediated costimulation and surprisingly on type I IFN. These studies provide the rational basis for the use of TLR and CD40 agonists together as essential adjuvants to optimize vaccines designed to elicit protective or therapeutic immunity.

SlltninlaryK46J B lymphomas express a T cell costimulatory activity that is not inhibited by CTLA-4Ig, anti-B7-1, anti-B7-2, anti-intercellular adhesion molecule 1 or antibodies to heat stable antigen. In this paper we report that this costimulatory activity is mediated at least in part by 4-1BB ligand, a member of the tumor necrosis factor (TNF) gene family that binds to 4-1BB, a T cell activation antigen with homology to the TNF/nerve growth factor receptor family. A fusion protein between 4-1BB and alkaline phosphatase (4-1BB-AP) blocks T cell activation by K46J lymphomas in both an antigen-specific system and with polyclonally (anti-CD3) activated T cells. 4-1BB-AP also blocks antigen presentation by normal spleen cells. When the antigen-presenting cells express B7 molecules as well as 4-1BB ligand, we find that B7 molecules and 4-1BB-AP both contribute to T cell activation. These data suggest that 4-1BB ligand plays an important role in costimulation of IL-2 production and proliferation by T cells. The B lymphoma M12 expresses low levels of 4-1BB-L but can be induced to express higher levels by treatment of the B ceils with cAMP, which also induces B7-1 and B7-2 in these cells. Thus cAMP appears to coordinately induce several costimulatory molecules on B cells.

Neuromyelitis optica (NMO) is a severe, debilitating human disease that predominantly features immunopathology in the optic nerves and the spinal cord. An IgG1 autoantibody (NMO-IgG) that binds aquaporin 4 (AQP4) has been identified in the sera of a significant number of NMO patients, as well as in patients with two related neurologic conditions, bilateral optic neuritis (ON), and longitudinal extensive transverse myelitis (LETM), that are generally considered to lie within the NMO spectrum of diseases. NMO-IgG is not the only autoantibody found in NMO patient sera, but the correlation of pathology in central nervous system (CNS) with tissues that normally express high levels of AQP4 suggests NMO-IgG might be pathogenic. If this is the case, it is important to identify and understand the mechanism(s) whereby an immune response is induced against AQP4. This review focuses on open questions about the "events" that need to be understood to determine if AQP4 and NMO-IgG are involved in the pathogenesis of NMO. These questions include: 1) How might AQP4-specific T and B cells be primed by either CNS AQP4 or peripheral pools of AQP4? 2) Do the different AQP4-expressing tissues and perhaps the membrane structural organization of AQP4 influence NMO-IgG binding efficacy and thus pathogenesis? 3) Does prior infection, genetic predisposition, or underlying immune dysregulation contribute to a confluence of events which lead to NMO in select individuals? A small animal model of NMO is essential to demonstrate whether AQP4 is indeed the incipient autoantigen capable of inducing NMO-IgG formation and NMO. If the NMO model is consistent with the human disease, it can be used to examine how changes in AQP4 expression and blood-brain barrier (BBB) integrity, both of which can be regulated by CNS inflammation, contribute to inductive events for anti-AQP4-specific immune response. In this review, we identify reagents and experimental questions that need to be developed and addressed to enhance our understanding of the pathogenesis of NMO. Finally, dysregulation of tolerance associated with autoimmune disease appears to have a role in NMO. Animal models would allow manipulation of hormone levels, B cell growth factors, and other elements known to increase the penetrance of autoimmune disease. Thus an AQP4 animal model would provide a means to manipulate events which are now associated with NMO and thus demonstrate what set of events or multiplicity of events can push the anti-AQP4 response to be pathogenic.

Synthetic antigens that mimic the terminal hexasaccharide epitope of the O-specific polysaccharide of Vibrio cholerae O1, serotype Ogawa, were conjugated to bovine serum albumin (BSA). Conjugates with carbohydrate-to-carrier molar ratios of 15.5:1, 9.2:1, and 4.6:1 were tested for immunogenicity and efficacy in mice. The role of preimmunity to BSA and the use of adjuvant in the generation of the serologic response to the O-specific polysaccharide and protection against virulent V. cholerae was examined. Preimmunity to BSA did not affect the anti-Ogawa titers but seemed to enhance the protective capacity of antiserum. All 3 conjugates were immunogenic, but adjuvant was effective at inducing higher and earlier antibody responses. In tertiary serum samples, a correlation was found between vibriocidal activity and protection. The protective capacity of antiserum was evident in serum from mice immunized with all conjugates, but it was highest in the groups that received the conjugate with the lowest level of substitution. Further studies are required to increase understanding of the reason for differential protection.

Inflammatory mediators facilitate the maturation of dendritic cells (DC), enabling them to induce the activation, proliferation and differentiation of cognate T cells. The role of CD40 on DC and CD154 on T cells has been studied by the co‐adoptive transfer of antigen‐pulsed DC and TCR‐transgenic (Tg) T cells in vivo. It is shown that in the absence of CD40‐CD154 interactions, initial Tg T cell expansion occurs in vivo, but over time, T cell expansion cannot be sustained. The basis for the demise of the T cell population is likely due to the disappearance of the antigen‐pulsed DC in the draining lymph nodes when CD154‐CD40 interactions are interrupted. These findings show that both T cell and DC persistence in vivo is dependent on CD40‐CD154 interactions. In addition to the physical persistence of the DC, CD40 triggering of DC also greatly increases the period for which they can productively present antigen to Tg T cells. Hence DC persistence and antigen‐presenting cell capacity are both dependent on CD40 signaling. While TNF‐α can mature DC as measured by a variety of criteria, the unique capacity of CD40 signaling to sustain T cell responses and induce DC maturation is underscored by the inability of TNF‐α to rescue the immune deficiency of CD40–/– DC. Hence, the profound impact of CD154 deficiency on cell‐mediated immunity may be due to its ability to limit the duration of antigen presentation in vivo and cause the premature demise of antigen‐specific T cells.