TLR9 deficiency breaks tolerance to RNA-associated antigens and upregulates TLR7 protein in Sle1 mice (original) (raw)
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Arthritis & Rheumatology, 2018
Objective. Toll-like receptors (TLRs) 7 and 9 are important innate signaling molecules with opposing roles in the development and progression of systemic lupus erythematosus (SLE). While multiple studies support the notion of a dependency on TLR-7 for disease development, genetic ablation of TLR-9 results in severe disease with glomerulonephritis (GN) by a largely unknown mechanism. This study was undertaken to examine the suppressive role of TLR-9 in the development of severe lupus in a mouse model. Methods. We crossed Sle1 lupus-prone mice with TLR-9-deficient mice to generate Sle1TLR-9 À/À mice. Mice ages 4.5-6.5 months were evaluated for severe autoimmunity by assessing splenomegaly, GN, immune cell populations, autoantibody and total Ig profiles, kidney dendritic cell (DC) function, and TLR-7 protein expression. Mice ages 8-10 weeks were used for functional B cell studies, Ig profiling, and determination of TLR-7 expression. Results. Sle1TLR-9 À/À mice developed severe disease similar to TLR-9-deficient MRL and Nba2 models. Sle1TLR-9 À/À mouse B cells produced more class-switched antibodies, and the autoantibody repertoire was skewed toward RNA-containing antigens. GN in these mice was associated with DC infiltration, and purified Sle1TLR-9 À/À mouse renal DCs were more efficient at TLR-7-dependent antigen presentation and expressed higher levels of TLR-7 protein. Importantly, this increase in TLR-7 expression occurred prior to disease development, indicating a role in the initiation stages of tissue destruction. Conclusion. The increase in TLR-7-reactive immune complexes, and the concomitant enhanced expression of their receptor, promotes inflammation and disease in Sle1TLR9 À/À mice.
B cell-intrinsic TLR9 expression is protective in murine lupus
Journal of Clinical Investigation
Toll-like receptor 9 (TLR9) is a regulator of disease pathogenesis in systemic lupus erythematosus (SLE). Why TLR9 represses disease while TLR7 and MyD88 have the opposite effect remains undefined. To begin to address this question, we created two alleles to manipulate TLR9 expression, allowing for either selective deletion or overexpression. We used these to test cell type-specific effects of Tlr9 expression on the regulation of SLE pathogenesis. Notably, Tlr9 deficiency in B cells was sufficient to exacerbate nephritis while extinguishing anti-nucleosome antibodies, whereas Tlr9 deficiency in dendritic cells (DCs), plasmacytoid DCs, and neutrophils had no discernable effect on disease. Thus, B cell-specific Tlr9 deficiency unlinked disease from autoantibody production. Critically, B cell-specific Tlr9 overexpression resulted in ameliorated nephritis, opposite of the effect of deleting Tlr9. Our findings highlight the non-redundant role of B cell-expressed TLR9 in regulating lupus and suggests therapeutic potential in modulating and perhaps even enhancing TLR9 signals in B cells.
Toll-like receptor 9 suppresses lupus disease in Fas-sufficient MRL Mice
PLOS ONE, 2017
Genetic deficiency in TLR9 accelerates pathogenesis in the spontaneous polygenic MRL. Fas lpr murine model of systemic lupus erythematosus, despite the absence of anti-nucleosome autoantibodies. However, it could be argued that this result was dependent on Fasdeficiency rather than lupus-promoting genes in the MRL genetic background. Here we report the effects of TLR9 deficiency on autoimmune disease independent of the lpr mutation in Fas by characterizing Tlr9-/and Tlr9 +/+ mice on the Fas-intact MRL/+ genetic background. By 30 weeks of age, Tlr9-deficient MRL/+ had more severe renal disease, increased T cell activation, and higher titers of anti-Sm and anti-RNA autoantibodies than Tlr9-intact animals, as had been the case in the MRL.Fas lpr model. In addition, Tlr9-deficient MRL/+ mice had increased numbers of germinal center phenotype B cells and an increase in splenic neutrophils and conventional dendritic cell populations. Thus, the disease accelerating effects of Tlr9 deficiency are separable from those mediated by the Fas mutation in the lupus-prone MRL genetic background. Nonetheless, disease acceleration in Tlr9-deficient MRL/+ mice was phenotypically distinct from that in Fas-deficient counterparts, which has important implications.
Toll-like receptor 9-independent aggravation of glomerulonephritis in a novel model of SLE
International Immunology, 2006
The generation of anti-DNA auto-antibodies is characteristic for the human autoimmune condition systemic lupus erythematosus (SLE) and its animal models. However, the contribution of the toll-like receptor (TLR) system of innate immunity receptors and, in particular, TLR9 to this B cell-mediated autoimmune process remains controversial. Here we report that in a novel murine model of SLE, based on hyper-reactive B cell activation mediated by mutant phospholipase Cg2, the genetic deficiency of TLR9 does not protect from spontaneous anti-DNA auto-antibody formation and glomerulonephritis. On the contrary, disease induction is aggravated and additional nucleolar antibody specificity develops in autoimmune TLR9-deficient mice. In vitro studies demonstrate that, in autoimmune-prone mice, dual signaling via the B cell receptor and non-CpG DNA results in synergistic B cell activation in a TLR9-independent manner. These results suggest that engagement of a TLR9-independent DNA activation pathway may promote autoimmunity, while TLR9 signaling can ameliorate SLE-like immune pathology in vivo.
Journal of the American Society of Nephrology, 2007
Small nuclear RNA and associated lupus autoantigens activate B cells and dendritic cells via Toll-like receptor-7 (TLR-7); therefore, TLR-7 may represent a potential therapeutic target in lupus. MRL lpr mice were administered an injection of either saline or synthetic oligodeoxynucleotides with immunoregulatory sequences (IRS) that specifically block signaling via TLR-7 (IRS 661) or via TLR-7 and TLR-9 (IRS 954, which uses a active sequence from IRS 661 along with a TLR-9 inhibitory sequence) from weeks 11 to 24 of age. IRS 661 and IRS 954 both significantly reduced the weight of spleen and lymph nodes as well as serum levels of TNF as compared with saline-treated MRL lpr mice. Only IRS 661 but not IRS 954 significantly reduced serum levels of IL-12p40, anti-dsDNA IgG 2a , IgG 2b , and anti-Smith IgG. Both IRS localized to the kidney after intraperitoneal injection and significantly improved the activity index and chronicity index for lupus nephritis in MRL lpr mice. This was associated with significant reduction of renal glomerular and interstitial macrophage infiltrates and the number of interstitial T cells. Autoimmune lung injury was also attenuated with IRS 661 and IRS 954. These data demonstrate that TLR-7 antagonism, initiated after the onset of autoimmunity, can prevent autoimmune kidney and lung injury in MRL lpr mice. Concomitant blockade of TLR-9 with IRS 954 neutralized the effect of TLR-7 blockade on dsDNA IgG 2a , dsDNA IgG 2b , and Smith antigen autoantibodies but had neither additive nor opposing effects on autoimmune lung and kidney injury. Hence, TLR-7 is proposed as a novel and potential therapeutic target in systemic lupus erythematosus.
Nature Immunology
In lupus, TLR7 and TLR9 mediate loss of tolerance to RNA and DNA, respectively. Yet, TLR7 promotes while TLR9 protects from disease, implying differences in signaling. To dissect this "TLR paradox", we generated two TLR9 point mutants-lacking either ligand (TLR9 K51E) or MyD88 (TLR9 P915H) binding-in lupus-prone MRL/lpr mice. Ameliorated disease of Tlr9 K51E mice compared to Tlr9 −/− controls revealed a TLR9 "scaffold" protective function that is ligand-and MyD88-independent. Unexpectedly, Tlr9 P915H mice were more protected than both Tlr9 K51E and Tlr9 WT mice, suggesting that TLR9 also possesses ligand-dependent, but MyD88-independent, regulatory signaling and MyD88-mediated proinflammatory signaling. Triple mixed bone marrow chimeras showed that TLR9-MyD88 independent regulatory roles were B cell-intrinsic and restrained differentiation into pathogenic age-associated B cells (ABC) and plasmablasts (PB). These studies reveal MyD88-independent regulatory roles of TLR9, shedding light on the biology of endosomal TLRs.
Immunity, 2006
Antibodies (Abs) to RNA-and DNA-containing autoantigens are characteristic of systemic lupus erythematosus (SLE). We showed previously that Toll-like receptor (TLR) 9, recognizing DNA, is required for the spontaneous generation of DNA autoantibodies, but not for the development of lupus nephritis in susceptible mice. We report that lupus-prone mice deficient in TLR7, a receptor for ssRNA, failed to generate Abs to RNA-containing antigens (Ags) such as Smith (Sm) Ag. TLR9 and TLR7 also had dramatic effects on clinical disease in lupus-prone mice. In the absence of TLR9, autoimmune disease was exacerbated, lymphocytes and plasmacytoid DCs were more activated, and serum IgG and IFN-a were increased. In contrast, TLR7-deficient mice had ameliorated disease, decreased lymphocyte activation, and decreased serum IgG. These findings reveal opposing inflammatory and regulatory roles for TLR7 and TLR9, despite similar tissue expression and signaling pathways. These results have important implications for TLR-directed therapy of autoimmune disease.