Toll-like receptors and innate immune responses in systemic lupus erythematosus

A series of discoveries over the past several years has provided a new paradigm for understanding autoimmunity in systemic lupus erythematosus. The discoveries of pattern recognition receptors and of how these receptors can be recruited into autoimmune responses underpin this paradigm. The implications of these observations continue to unfold with ongoing investigation into the range and specificity of pattern recognition receptors, into how immune complexes containing nucleic acids trigger these receptors, into how endogenous macromolecular 'danger signals' stimulate innate immune responses, and into the effect of pattern recognition receptor activation on various cell types in initiating and perpetuating autoimmunity. The development of clinical trials using therapeutic agents that target components of the innate immune system suggests that these advances may soon culminate in new medications for treating patients with systemic lupus erythematosus

Toll-like Receptor 9–Dependent and –Independent Dendritic Cell Activation by Chromatin–Immunoglobulin G Complexes

Dendritic cell (DC) activation by nucleic acid–containing immunoglobulin (Ig)G complexes has been implicated in systemic lupus erythematosus (SLE) pathogenesis. However, the mechanisms responsible for activation and subsequent disease induction are not completely understood. Here we show that murine DCs are much more effectively activated by immune complexes that contain IgG bound to chromatin than by immune complexes that contain foreign protein. Activation by these chromatin immune complexes occurs by two distinct pathways. One pathway involves dual engagement of the Fc receptor FcγRIII and Toll-like receptor (TLR)9, whereas the other is TLR9 independent. Furthermore, there is a characteristic cytokine profile elicited by the chromatin immune complexes that distinguishes this response from that of conventional TLR ligands, notably the induction of BAFF and the lack of induction of interleukin 12. The data establish a critical role for self-antigen in DC activation and explain how the innate immune system might drive the adaptive immune response in SLE

Anatomy of autoantibody production: Dominant localization of antibody-producing cells to T cell zones in fas-deficient mice

AbstractThe goal of this study was to examine the in vivo site of autoantibody production in normal and autoimmune-prone mice. B cells were identified in tissue sections with IgM- and IgG2a-specific riboprobes that readily distinguished resting cells from antibody-forming cells (AFC). In normal mice, the few identifiable IgG2a-secreting cells were found in the red pulp. By contrast, in Ipr mice exceedingly high numbers of IgG2a and autoantibody-producing cells were found deep within the T cell-rich periarteriolar lymphoid sheaths (PALS). This unusual anatomic location of autoantibody-secreting B cells is unique to Fas dysregulated strains, since IgG2a-producing cells in MRL/+ and (SWR × NZB)F1 mice were found predominantly in the red pulp or outer PALS, similar to normal mice. Furthermore, analysis of spleens from Ipr and non-Ipr anti-DNA immunoglobulin transgenic mice revealed dramatic accumulation of Tg+ cells in the inner PALS only in Ipr mice. These data suggest that in the absence of Fas, autoreactive B cells accumulate in T cell-rich zones, and this anatomic feature may contribute to autoantibody production

Selective pathogenicity of murine rheumatoid factors of the cryoprecipitable IgG3 subclass

To analyze the involvement of rheumatoid factors (RF) in the generation of cryoglobulins and the development of related tissue injuries, we have established a panel of anti-IgG2a RF mAbs derived from MRL/MpJ-lpr/lpr (MRL-lpr), C3H/HeJ-lpr/lpr, and 129/Sv mice. After injection of hybridoma cells to normal mice, all four IgG3 RF mAbs induced cryoglobullnemia, and various degrees of glomerulonephritis and skin leukocytoclastic vasculitis. In contrast, none of the RF mAbs of the other isotypes generated cryoglobulins or tissue lesions. Since the same observation was obtained with another panel of five clonally related anti-IgG2a RF mAbs of MRL-lpr origin with almost Identical heavy and light chain variable (V) regions but five different Isotypes, it seems unlikely that the absence of pathogenicity of non-IgG3 RF mAbs was due to differences in fine specificity or V framework regions. In addition, the analysis of serum RF In MRL-lpr mice has demonstrated that a majority of 4 month old MRL-lpr mice produced substantial amounts of IgG3 RF with cryoglobulin activity. Because the cryoglobulin activity is associated with the murine IgG3 heavy chain constant region, RF of this subclass may play a significant role in the development of autoimmune-related tissue injuries, especially In MRL-lpr mic

Human nasal wash RNA-Seq reveals distinct cell-specific innate immune responses in influenza versus SARS-CoV-2

BACKGROUND Influenza A virus (IAV) and SARS-CoV-2 are pandemic viruses causing millions of deaths, yet their clinical manifestations are distinctly different. METHODS With the hypothesis that upper airway immune and epithelial cell responses are also distinct, we performed single-cell RNA sequencing (scRNA-Seq) on nasal wash cells freshly collected from adults with either acute COVID-19 or influenza or from healthy controls. We focused on major cell types and subtypes in a subset of donor samples. Results Nasal wash cells were enriched for macrophages and neutrophils for both individuals with influenza and those with COVID-19 compared with healthy controls. Hillock-like epithelial cells, M2-like macrophages, and age-dependent B cells were enriched in COVID-19 samples. A global decrease in IFN-associated transcripts in neutrophils, macrophages, and epithelial cells was apparent in COVID-19 samples compared with influenza samples. The innate immune response to SARS-CoV-2 appears to be maintained in macrophages, despite evidence for limited epithelial cell immune sensing. Cell-to-cell interaction analyses revealed a decrease in epithelial cell interactions in COVID-19 and highlighted differences in macrophage-macrophage interactions for COVID-19 and influenza. Conclusions Our study demonstrates that scRNA-Seq can define host and viral transcriptional activity at the site of infection and reveal distinct local epithelial and immune cell responses for COVID-19 and influenza that may contribute to their divergent disease courses. Funding Massachusetts Consortium on Pathogen Readiness, the Mathers Foundation, and the Department of Defense (W81XWH2110029) COVID-19 Expansion for AIRe Program

DNA-like class R inhibitory oligonucleotides (INH-ODNs) preferentially block autoantigen-induced B-cell and dendritic cell activation in vitro and autoantibody production in lupus-prone MRL-Faslpr/lpr mice in vivo

INTRODUCTION. B cells have many different roles in systemic lupus erythematosus (SLE), ranging from autoantigen recognition and processing to effector functions (for example, autoantibody and cytokine secretion). Recent studies have shown that intracellular nucleic acid-sensing receptors, Toll-like receptor (TLR) 7 and TLR9, play an important role in the pathogenesis of SLE. Dual engagement of rheumatoid factor-specific AM14 B cells through the B-cell receptor (BCR) and TLR7/9 results in marked proliferation of autoimmune B cells. Thus, strategies to preferentially block innate activation through TLRs in autoimmune B cells may be preferred over non-selective B-cell depletion. METHODS. We have developed a new generation of DNA-like compounds named class R inhibitory oligonucleotides (INH-ODNs). We tested their effectiveness in autoimmune B cells and interferon-alpha-producing dendritic cells in vitro and in lupus-prone MRL-Faslpr/lpr mice in vivo. RESULTS. Class R INH-ODNs have 10- to 30-fold higher inhibitory potency when autoreactive B cells are synergistically activated through the BCR and associated TLR7 or 9 than when stimulation occurs via non-BCR-engaged TLR7/9. Inhibition of TLR9 requires the presence of both CCT and GGG triplets in an INH-ODN, whereas the inhibition of the TLR7 pathway appears to be sequence-independent but dependent on the phosphorothioate backbone. This difference was also observed in the MRL-Faslpr/lpr mice in vivo, where the prototypic class R INH-ODN was more effective in curtailing abnormal autoantibody secretion and prolonging survival. CONCLUSIONS. The increased potency of class R INH-ODNs for autoreactive B cells and dendritic cells may be beneficial for lupus patients by providing pathway-specific inhibition yet allowing them to generate protective immune response when needed.National Institutes of Health (AI047374, AI064736); Alliance for Lupus Researc

cGAS-STING Pathway Does Not Promote Autoimmunity in Murine Models of SLE

Detection of DNA is an important determinant of host-defense but also a driver of autoinflammatory and autoimmune diseases. Failure to degrade self-DNA in DNAseII or III(TREX1)-deficient mice results in activation of the cGAS-STING pathway. Deficiency of cGAS or STING in these models ameliorates disease manifestations. However, the contribution of the cGAS-STING pathway, relative to endosomal TLRs, in systemic lupus erythematosus (SLE) is controversial. In fact, STING deficiency failed to rescue, and actually exacerbated, disease manifestations in Fas-deficient SLE-prone mice. We have now extended these observations to a chronic model of SLE induced by the i.p. injection of TMPD (pristane). We found that both cGAS- and STING-deficiency not only failed to rescue mice from TMPD-induced SLE, but resulted in increased autoantibody production and higher proteinuria levels compared to cGAS STING sufficient mice. Further, we generated cGAS(KO)Fas(lpr) mice on a pure MRL/Fas(lpr) background using Crispr/Cas9 and found slightly exacerbated, and not attenuated, disease. We hypothesized that the cGAS-STING pathway constrains TLR activation, and thereby limits autoimmune manifestations in these two models. Consistent with this premise, mice lacking cGAS and Unc93B1 or STING and Unc93B1 developed minimal systemic autoimmunity as compared to cGAS or STING single knock out animals. Nevertheless, TMPD-driven lupus in B6 mice was abrogated upon AAV-delivery of DNAse I, implicating a DNA trigger. Overall, this study demonstrated that the cGAS-STING pathway does not promote systemic autoimmunity in murine models of SLE. These data have important implications for cGAS-STING-directed therapies being developed for the treatment of systemic autoimmunity

Suppression of systemic autoimmunity by the innate immune adaptor STING

Cytosolic DNA-sensing pathways that signal via Stimulator of interferon genes (STING) mediate immunity to pathogens and also promote autoimmune pathology in DNaseII- and DNaseIII-deficient mice. In contrast, we report here that STING potently suppresses inflammation in a model of systemic lupus erythematosus (SLE). Lymphoid hypertrophy, autoantibody production, serum cytokine levels, and other indicators of immune activation were markedly increased in STING-deficient autoimmune-prone mice compared with STING-sufficient littermates. As a result, STING-deficient autoimmune-prone mice had significantly shorter lifespans than controls. Importantly, Toll-like receptor (TLR)-dependent systemic inflammation during 2,6,10,14-tetramethylpentadecane (TMPD)-mediated peritonitis was similarly aggravated in STING-deficient mice. Mechanistically, STING-deficient macrophages failed to express negative regulators of immune activation and thus were hyperresponsive to TLR ligands, producing abnormally high levels of proinflammatory cytokines. This hyperreactivity corresponds to dramatically elevated numbers of inflammatory macrophages and granulocytes in vivo. Collectively these findings reveal an unexpected negative regulatory role for STING, having important implications for STING-directed therapies