RESUMO
Antibodies to double-stranded DNA (dsDNA) are prevalent in systemic lupus erythematosus (SLE), particularly in patients with lupus nephritis, yet the nature and regulation of antigenic cell-free DNA (cfDNA) are poorly understood. Null mutations in the secreted DNase DNASE1L3 cause human monogenic SLE with anti-dsDNA autoreactivity. We report that >50% of sporadic SLE patients with nephritis manifested reduced DNASE1L3 activity in circulation, which was associated with neutralizing autoantibodies to DNASE1L3. These patients had normal total plasma cfDNA levels but showed accumulation of cfDNA in circulating microparticles. Microparticle-associated cfDNA contained a higher fraction of longer polynucleosomal cfDNA fragments, which bound autoantibodies with higher affinity than mononucleosomal fragments. Autoantibodies to DNASE1L3-sensitive antigens on microparticles were prevalent in SLE nephritis patients and correlated with the accumulation of cfDNA in microparticles and with disease severity. DNASE1L3-sensitive antigens included DNA-associated proteins such as HMGB1. Our results reveal autoantibody-mediated impairment of DNASE1L3 activity as a common nongenetic mechanism facilitating anti-dsDNA autoreactivity in patients with severe sporadic SLE.
Assuntos
Anticorpos Antinucleares/imunologia , Autoanticorpos/imunologia , DNA/imunologia , Endodesoxirribonucleases/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Nefrite Lúpica/imunologia , Adulto , Animais , Anticorpos Antinucleares/sangue , Autoanticorpos/sangue , Micropartículas Derivadas de Células/imunologia , Micropartículas Derivadas de Células/metabolismo , Ácidos Nucleicos Livres/sangue , Ácidos Nucleicos Livres/imunologia , Criança , Endodesoxirribonucleases/sangue , Endodesoxirribonucleases/genética , Endodesoxirribonucleases/metabolismo , Feminino , Células HEK293 , Proteína HMGB1/imunologia , Proteína HMGB1/metabolismo , Humanos , Lúpus Eritematoso Sistêmico/metabolismo , Nefrite Lúpica/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Índice de Gravidade de DoençaRESUMO
Antibodies to DNA and chromatin drive autoimmunity in systemic lupus erythematosus (SLE). Null mutations and hypomorphic variants of the secreted deoxyribonuclease DNASE1L3 are linked to familial and sporadic SLE, respectively. We report that DNASE1L3-deficient mice rapidly develop autoantibodies to DNA and chromatin, followed by an SLE-like disease. Circulating DNASE1L3 is produced by dendritic cells and macrophages, and its levels inversely correlate with anti-DNA antibody response. DNASE1L3 is uniquely capable of digesting chromatin in microparticles released from apoptotic cells. Accordingly, DNASE1L3-deficient mice and human patients have elevated DNA levels in plasma, particularly in circulating microparticles. Murine and human autoantibody clones and serum antibodies from human SLE patients bind to DNASE1L3-sensitive chromatin on the surface of microparticles. Thus, extracellular microparticle-associated chromatin is a potential self-antigen normally digested by circulating DNASE1L3. The loss of this tolerance mechanism can contribute to SLE, and its restoration may represent a therapeutic opportunity in the disease.
Assuntos
Autoanticorpos/imunologia , Micropartículas Derivadas de Células/química , Cromatina/imunologia , DNA/imunologia , Endodesoxirribonucleases/genética , Lúpus Eritematoso Sistêmico/imunologia , Animais , Micropartículas Derivadas de Células/metabolismo , Modelos Animais de Doenças , Endodesoxirribonucleases/deficiência , Endodesoxirribonucleases/metabolismo , Humanos , Células Jurkat , Lúpus Eritematoso Sistêmico/enzimologia , Lúpus Eritematoso Sistêmico/genética , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Eicosanoids are inflammatory mediators that play a key but incompletely understood role in linking the innate and adaptive immune systems. Here, we show that cytotoxic effector T cells (CTLs) are capable of both producing and responding to cysteinyl leukotrienes (CystLTs), allowing for the killing of target cells in a T cell receptor-independent manner. This process is dependent on the natural killer receptor NKG2D and exposure to IL-15, a cytokine induced in distressed tissues. IL-15 and NKG2D signaling drives the up-regulation of key enzymes implicated in the synthesis of CystLTs, as well as the expression of CystLT receptors, suggesting a positive feedback loop. Finally, although the CystLT pathway has been previously linked to various allergic disorders, we provide unexpected evidence for its involvement in the pathogenesis of celiac disease (CD), a T helper 1 cell-mediated enteropathy induced by gluten. These findings provide new insights into the cytolytic signaling pathway of NKG2D and the pathogenesis of organ-specific immune disorders. Furthermore, they suggest that the blockade of CystLT receptors may represent a potent therapeutic target for CD or potentially other autoimmune disorders in which NKG2D has been implicated.
Assuntos
Doença Celíaca/imunologia , Cisteína/metabolismo , Interleucina-15/imunologia , Leucotrienos/metabolismo , Subfamília K de Receptores Semelhantes a Lectina de Células NK/imunologia , Linfócitos T Citotóxicos/imunologia , Adulto , Araquidonato 5-Lipoxigenase/genética , Araquidonato 5-Lipoxigenase/metabolismo , Estudos de Casos e Controles , Doença Celíaca/fisiopatologia , Células Cultivadas , Cisteína/imunologia , Feminino , Humanos , Interleucina-15/metabolismo , Leucotrienos/imunologia , Masculino , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Receptores de Leucotrienos/genética , Receptores de Leucotrienos/metabolismo , Células Th1/imunologia , Células Th1/metabolismo , Regulação para CimaRESUMO
The NKG2 family of NK receptors includes activating and inhibitory members. With the exception of the homodimer-forming NKG2D, NKG2 receptors recognize the nonclassical MHC class I molecule HLA-E, and they can be subdivided into two groups: those that associate with and signal through DAP12 to activate cells, and those that contain an ITIM motif to promote inhibition. The function of NKG2 family member NKG2E is unclear in humans, and its surface expression has never been conclusively established, largely because there is no Ab that binds specifically to NKG2E. Seeking to determine a role for this molecule, we chose to investigate its expression and ability to form complexes with intracellular signaling molecules. We found that NKG2E was capable of associating with CD94 and DAP12 but that the complex was retained intracellularly at the endoplasmic reticulum instead of being expressed on cell surfaces, and that this localization was dependent on a sequence of hydrophobic amino acids in the extracellular domain of NKG2E. Because this particular sequence has emerged and been conserved selectively among higher order primates evolutionarily, this observation raises the intriguing possibility that NKG2E may function as an intracellular protein.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Citoplasma/metabolismo , Proteínas de Membrana/metabolismo , Subfamília C de Receptores Semelhantes a Lectina de Células NK/metabolismo , Subfamília D de Receptores Semelhantes a Lectina de Células NK/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Sequência de Aminoácidos , Aminoácidos/química , Aminoácidos/genética , Aminoácidos/metabolismo , Animais , Sítios de Ligação/genética , Western Blotting , Linhagem Celular , Retículo Endoplasmático/metabolismo , Células HEK293 , Humanos , Interações Hidrofóbicas e Hidrofílicas , Proteínas de Membrana/genética , Microscopia Confocal , Dados de Sequência Molecular , Mutação , Subfamília C de Receptores Semelhantes a Lectina de Células NK/classificação , Subfamília C de Receptores Semelhantes a Lectina de Células NK/genética , Subfamília D de Receptores Semelhantes a Lectina de Células NK/genética , Filogenia , Ligação Proteica , Homologia de Sequência de AminoácidosRESUMO
NK cells are large granular lymphocytes that form a critical component of the innate immune system, whose functions include the killing of cells expressing stress-induced molecules. It is increasingly accepted that despite being considered prototypical effector cells, NK cells require signals to reach their full cytotoxic potential. We previously showed that IL-15 is capable of arming CD8 effector T cells to kill independently of their TCR via NKG2D in a cPLA2-dependent process. As NK cells also express NKG2D, we wanted to investigate whether this pathway functioned in an analogous manner and if resting NK cells could be primed to the effector phase by IL-15. Furthermore, to establish relevance to human disease we studied a possible role for this pathway in the pathogenesis of psoriatic arthritis, since there are aspects of this disease that suggest a potential effector role for the innate immune system. We found that PsA patients had upregulated IL-15 and MIC in their affected synovial tissues, and that this unique inflammatory environment enabled NK cell activation and killing via NKG2D and cPLA2. Moreover, we were able to reproduce the phenotype of joint NK cells from blood NK cells by incubating them with IL-15. Altogether, these findings suggest a destructive role for NK cells when activated by environmental stress signals during the pathogenesis of PsA and demonstrate that IL-15 is capable of priming resting NK cells in tissues to the effector phase.