The role of toll-like receptors in systemic autoimmune disease
Introduction
Systemic autoimmune diseases such as SLE are frequently associated with the production of high titers of autoantibodies specific for nuclear and/or cytoplasmic cellular components. Such antibodies subsequently form immune complexes (ICs), deposit in the blood vessel walls, joints, and glomeruli, and thereby contribute to the vasculitis, arthritis, and glomerulonephritis characteristic of systemic disease. Remarkably, in both animal models and human disease, the autoantibody repertoire consistently targets a relatively limited set of DNA and RNA associated autoantigens [1], [2]. A better understanding of the mechanisms by which these particular molecules activate autoreactive B cells is critical to the design of more effective therapeutics.
A number of recent studies have linked the antigenicity of particular autoantigens to the process of apoptotic cell death and subsequent conformational changes of many of the prominent autoantigens. Apoptosis can be induced by environmental insults or by cytotoxic effector cells; in either case, many potential autoantigens are cleaved by caspases or granzymes, or undergo post-translational modifications such as oxidation, citrullination, or phosphorylation [3], [4]. It has been proposed that such alterations may lead to the creation of neo-epitopes that are then recognized as foreign by the adaptive immune system. The process of epitope spreading can then effectively break tolerance to the actual self-determinants. However, it has recently been shown that autoantibodies specific for granzyme B target proteins are just as prevalent in granzyme B-deficient autoimmune mice as in granzyme B-sufficient autoimmune prone mice [5]. It follows that autoantigen cleavage may be a fingerprint of apoptosis but not necessarily a trigger for autoantibody production.
Alternatively, some aspect of the apoptotic process may directly prime the innate immune system to initiate an adaptive response, perhaps by converting autoantigens into “autoadjuvants”, or by promoting the redistribution or aberrant accumulation of pre-existing autoadjuvants. In this context, normally benign self-determinants may appear foreign. The potential adjuvanticity of mammalian nucleic acids was originally revealed by the pivotal studies of Ronnblom et al. who first demonstrated the potent effects of DNA- and RNA-containing ICs on IFNα-producing dendritic cells [6], [7]. Our laboratory has extended this analysis by taking advantage of a B cell receptor transgenic line designated AM14 [8].
AM14 B cells recognize IgG2a with relatively low affinity and can be activated in vitro only by IgG2a presented as a multivalent IC. Importantly, the type of antigen contained in the IC plays a critical role in the cell activation process. IgG2a mAbs specific for histones, nucleosomes, or DNA bind to DNAse sensitive material released from dead or dying cells that is present in the supernatant of primary B cell cultures [9], [10]. These “chromatin” ICs are potent ligands for AM14 B cells under conditions in which IgG2a bound to protein antigens, or protein ICs, have little effect. The stimulatory capacity of the chromatin ICs can be explained by their ability to effectively engage both the AM14 B cell receptor and one or more chloroquine/bafilomycin sensitive receptors. Ongoing studies from our laboratory have focused on the identification or the relevant receptor(s) and the nature of the mammalian DNA ligand.
Section snippets
The role of TLR9 in the activation of AM14 B cells
The first indication that TLRs might be involved in B cell autoantibody production came from the observation that MyD88-deficient AM14 B cells completely failed to proliferate in response to anti-nucleosome/histone monoclonal antibodies such as PL2-3. In addition, the PL2-3 response of wild-type AM14 B cells could be very effectively blocked by the phosphorothioate oligodeoxynucleotide (s-ODN) 2088 [10]. Even though 2088 incorporates a CpG dinucleotide, the upstream CCT and 5′GGGG render this
TLRs and mammalian DNA
TLR9 was first identified as a pattern recognition receptor capable of distinguishing bacterial DNA from mammalian DNA. These initial studies were carried out with non-transgenic B cells via a mechanism that did not depend on the B cell receptor for DNA uptake [13]. However, even in experimental systems involving the BCR-mediated delivery of bacterial and mammalian genomic DNA to the relevant endosomal/lysosomal compartments, we have found a dramatic difference in the stimulatory capacity of
TLR activation as a result of DNA modification
There is also evidence that certain modifications in mammalian DNA structure may enhance TLR reactivity. Loxoribine, and other guanosine analogs, have been found to activate TLR7 [23]. Loxoribine is structurally related to 8-hydroxy-2′-deoxyguanosine (8OHdG), and its presence correlates with ROS-mediated DNA damage. Intriguingly, 8OHdG has been detected in the circulating ICs of SLE patient sera [24]. ROS-modified DNA has been shown to be more immunogenic than unmodified DNA [25], [26] and to
RNA-associated autoantigens and TLR activity
Just as TLR9 plays a critical role in the recognition of bacterial and viral DNA, TLR3 and TLR7 have been shown to respond to double stranded and single-stranded RNA, respectively [35], [36], [37]. At the moment, the sequence specificity of the RNA-reactive receptors is somewhat unresolved. Particularly strong RNA stimulatory motifs have not yet been identified although it is known that TLR7-expressing cells appear to preferentially recognize GU-rich stretches of RNA. Although it might be
Summary and speculation
Extrapolating from the AM14 chromatin IC data and what is currently known about the cell biology of TLR7 and TLR9 expression, it is reasonable to propose that the AM14 receptor (specific for IgG2a) can bind IgG2a specific for either a chromatin-associated protein (such as histone) (Fig. 2, left), or an RNA-associated protein, (such as the Sm antigen) (Fig. 2, right). Uptake by the AM14 BCR then directs the complex to a vesicular compartment where the nucleic acid in the complex can engage
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