Microparticles as a source of extracellular DNA (original) (raw)
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Microparticles as autoantigens in human and murine lupus
Arthritis Research & Therapy, 2012
Background: CD4 T cells help B cells produce antibodies following antigen challenge. This response classically occurs in germinal centers (GC) located in B-cell follicles of secondary lymphoid organs (SLO), a site of immunoglobulin isotype switching and affinity maturation. GC formation requires specialized CD4 T cells, T-follicular helper (Tfh) cells, which localize to follicles and provide B cells with survival and differentiation signals that are essential for B-cell maturation into memory and long-lived plasma cells. Pathogenic autoantibodies in human and murine lupus arise in a like manner. Although Tfh cells are critical for GC development, their genesis in humans, role in promotion of autoimmunity, and potential as therapeutic targets in SLE are incompletely understood. To address these issues, we dissected Tfh cell development and function, defining their transcriptional regulation, migration, and function in vivo in normal and lupus-prone mice and ex vivo in normal humans and patients with SLE. Methods: We used a combination of approaches-flow cytometry, confocal microscopy, microarrays, quantitative chromatin immunoprecipitation and DNA sequencing (ChIP-seq), retroviral overexpression, and T-cell-B-cell helper assays-to characterize Tfh cells in normal mice and in lupus-prone strains, and from the tonsils of normal humans and the blood of patients with SLE. Results: We found that the transcription factor Bcl6 (B-cell CLL/lymphoma 6) is necessary and sufficient for Tfh development and function, via genetic control of Tfh proteins that are essential for their migration to B-cell follicles and GC and subsequent B-cell maturation. We dissected steps in Tfh development within SLO, beginning with their genesis in the T-cell zone followed by emigration to sites of B-cell interaction outside the B-cell follicle, where we have shown that B cells serve to provide signals for continued Tfh expansion and follicular migration. We have now begun to tease apart the factors that mediate T-cell-B-cell collaboration in the follicle; these represent therapeutic targets in SLE. Finally, we have shown that patients with SLE have expansion of Tfh cells in the blood, a finding that highlights their potential role in the pathogenesis of SLE and as likely therapeutic targets. Conclusion: These studies help define the developmental pathways for Tfh cells, and the steps that enable these cells to function in the B-cell follicle to promote immunoglobulin and autoantibody production. They have also helped define markers of Tfh cells in normals and autoimmune individuals, and suggest that they are a promising therapeutic target in patients.
Scientific Reports, 2016
Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease characterized by circulating autoantibodies and the formation of immune complexes. In these responses, the selecting self-antigens likely derive from the remains of dead and dying cells, as well as from disturbances in clearance. During cell death/activation, microparticles (MPs) can be released to the circulation. Previous MP studies in SLE have been limited in size and differ regarding numbers and phenotypes. Therefore, to characterize MPs more completely, we investigated 280 SLE patients and 280 individually matched controls. MPs were measured with flow cytometry and phenotyped according to phosphatidylserine expression (PS+/PS−), cellular origin and inflammatory markers. MPs, regardless of phenotype, are 2–10 times more abundant in SLE blood compared to controls. PS− MPs predominated in SLE, but not in controls (66% vs. 42%). Selectively in SLE, PS− MPs were more numerous in females and smokers. MP numbers dec...
The role of microparticles in the generation of immune complexes in murine lupus
Clinical Immunology, 2013
Systemic lupus erythematosus is a systemic inflammatory disease characterized by antibodies to nuclear molecules in association with immune complex deposition. As shown previously, microparticles (MPs), which are small membrane-bound vesicles released from dying and activated cells, contain nucleic acids and can form immune complexes found in patient blood. To assess the role of MPs in murine lupus, we used flow cytometry to measure the presence of MPs with bound IgG in the blood of MRL-lpr/lpr and NZB/W mice. These studies showed much higher numbers of MPs with bound IgG in the blood of MRL lpr/lpr compared to NZB/W mice. Furthermore, these studies showed that antibodies from MRL-lpr/lpr mice bound better to MPs from apoptotic cells than those from NZB/W mice. Together, these studies indicate important differences in the serological features of the two strains as reflected by the capacity of antibodies to bind to MPs.
Scandinavian Journal of Immunology, 2013
Microparticles (MPs) are small membrane‐bound vesicles with potent biological activities that can promote the pathogenesis of rheumatoid arthritis and systemic lupus erythematosus (SLE). These particles contain diverse cellular components and are shed from cells during apoptosis or activation. MPs can drive inflammation and autoimmunity by multiple mechanisms reflecting their content of bioactive molecules and ability to engage multiple receptor systems simultaneously. In the rheumatoid joint, particles can stimulate synovitis via their display of cytokines, chemokines, complement and angiogenesis factors. In SLE, particles can serve as an important source of extracellular nuclear molecules to signal ‘danger’ and form pathogenic immune complexes. Future studies will define the pathways by which particles promote pathogenesis, strategies to block their activity and their utility as biomarkers to assess disease activity and the response to therapy.
Unique protein signature of circulating microparticles in systemic lupus erythematosus
Arthritis & Rheumatism, 2013
Objective. To characterize the unique qualities of proteins associated with circulating subcellular material in systemic lupus erythematosus (SLE) patients compared with healthy controls and patients with other chronic autoimmune diseases. Methods. Using differential centrifugation and high-sensitivity nano-liquid chromatography tandem mass spectrometry, we systematically profiled proteins of microparticles (MPs) from SLE patients (n ؍ 12), systemic sclerosis (SSc) patients (n ؍ 6), and rheumatoid arthritis (RA) patients (n ؍ 6), as well as healthy controls (n ؍ 12). Results. We identified 531 unique proteins and showed that the differences between healthy controls and patients with SLE with regard to the abundance of 248 proteins were highly statistically significant. Almost half of the proteins that were increased by >2-fold were complement proteins and Ig (increased by 100-4,000 times). MP Ig and complement loads also distinguished SLE from RA and SSc and correlated strongly with clinical SLE severity. Subsets of microtubule proteins, fibronectin, 14-3-3, and desmosomal proteins as well as ficolin 2 and galectin 3 binding protein were also highly increased. In SLE MPs, levels of cytoskeletal, mitochondrial, and organelle proteins, including lysosome-associated membrane protein 1 and transforming growth factor 1, were decreased. Conclusion. The data show that SLE patients have increased numbers of MPs that are heavily tagged for removal and fewer MPs with normal protein composition. SLE MPs are unique and specific proteins that represent novel leads for our understanding of SLE and for the development of new treatments of the disease. The autoimmune disease systemic lupus erythematosus (SLE) is characterized by DNA/RNA-Tolllike receptor-dependent activation of innate immunity, with increased expression of type I interferon (IFN) response genes, autoantibody production resulting from loss of tolerance, and chronic, severe, and fluctuating morbidity due to thrombophilia and immune complex (IC)-mediated inflammatory damage of multiple tissues and organs. In SLE, the normal noninflammatory handling of ICs and material released from dead or dying cells is defective and instead is associated with a sustained type I IFN response leading to chronic inflammation (1,2). Consistent with the involvement of clearance defects in SLE pathogenesis, lupus-like disease evolves in patients with complement C1q deficiency. Defects of C1q-, DNA-, and apoptotic cell-binding molecules, such as C-reactive protein, serum amyloid P component, natural IgM, milk fat globule epidermal growth factor 8, annexins, protein S, Gas-6,  2-glycoprotein I ( 2 GPI), or deficiencies in proteins involved in the phagocytosis of ingested apoptotic material, such as the Mer receptor tyrosine kinase and DNase I, also lead to lupus-like manifestations in humans or in mice (3-9). Recent studies using flow cytometry show alterations in the numbers and types of subcellular particles circulating in SLE patients (10-12). The characterization of circulating subcellular particles is challenging because of their Supported by the Danish Rheumatism Association (grants R99-A1937 and R97-A1028), the Lundbeck Foundation, and the Novo
Arthritis & rheumatology (Hoboken, N.J.), 2015
Objective - Circulating chromatin-containing apoptotic material and/or neutrophil extracellular traps (NETs) have been proposed as an important driving force for the anti-chromatin autoimmune response in patients with systemic lupus erythematosus (SLE). We aimed to determine the exact nature of microparticles that circulate in SLE patients, and assess their effects on the immune system. Methods - We analyzed microparticles isolated from plasma of patients with SLE, rheumatoid arthritis (RA) or systemic sclerosis (SSc), and healthy subjects, and determined their effects on blood-derived DCs and neutrophils. Results - In SLE patients, we identified microparticles highly positive for annexin V and apoptosis-modified chromatin, not present in healthy subjects or RA and SSc patients. These microparticles were mostly CD31(+) /CD45(-) (endothelial), partially CD45(+) /CD66b(+) (granulocyte), and negative for B and T cell markers. Microparticles isolated from plasma of SLE patients increase...
Distinct proteome pathology of circulating microparticles in systemic lupus erythematosus
Clinical Proteomics, 2017
Background: The pathogenesis of systemic lupus erythematosus (SLE) is poorly understood but has been linked to defective clearance of subcellular particulate material from the circulation. This study investigates the origin, formation, and specificity of circulating microparticles (MPs) in patients with SLE based on comprehensive MP proteome profiling using patients with systemic sclerosis (SSc) and healthy donors (HC) as controls. Methods: We purified MPs from platelet-poor plasma using differential centrifugation of samples from SLE (n = 45), SSc (n = 38), and two sets of HC (n = 35, n = 25). MP proteins were identified and quantitated after trypsin digestion by liquid chromatography-tandem mass spectrometry. The abundance of specific proteins was compared between the groups using univariate statistics and false discovery rate correction for multiple comparisons. Specific proteins and protein ratios were explored for diagnostic and disease activity information using receiver-operating characteristic curves and by analysis of correlations of protein abundance with disease activity scores. Results: We identify and quantitate more than 1000 MP proteins and show that a subpopulation of SLE-MPs (which we propose to call luposomes) are highly specific for SLE, i.e. not found in MP preparations from HC or patients with another autoimmune, systemic disease, SSc. In SLE-MPs platelet proteins and mitochondrial proteins are significantly diminished, cytoskeletal proteins deranged, and glycolytic enzymes and apoptotic proteins significantly increased. Conclusions: Normal MPs are efficiently removed in SLE, but aberrant MPs, derived from non-lymphoid leukocytes, are less efficiently removed and abundantly produced leading to an altered MP proteome in SLE. The data suggest that an abnormal generation of MPs may partake in the pathology of SLE and that new diagnostic, monitoring, and treatment strategies targeting these processes may be advantageous.
Nucleosomes in the pathogenesis of systemic lupus erythematosus
Rheumatic Disease Clinics of North America, 2004
Antibodies that are directed against components of the cell nucleus almost always accompany systemic lupus erythematosus (SLE). Formation of antibodies against double-stranded DNA is a hallmark of SLE. For many years, the development of antibodies against cell components that normally are sequestered into the nucleus remained puzzling to researchers who were working in this field of rheumatology. A decade ago, it became clear that these intracellular antigens could become exposed at the cell surface during physiologic cell death (apoptosis), and, under particular conditions, become immunogenic. The primary antigen in SLE has been believed to be naked DNA. DNA is a poor immunogen, however, and recent evidence suggests that nucleosomes, that comprise DNA and histones, constitute the primary inciting antigen in SLE. We review up-to-date data on the mechanisms by which nucleosomes and other lupus autoantigens may become immunogenic in lupus and the role of antinucleosome antibodies in the development of glomerulonephritis, a common and severe complication that occurs in approximately 60% of patients who have SLE. The apoptotic cell: a source of lupus antigens Human and murine SLE are characterized by the appearance in the blood of an array of autoantibodies that are directed against nuclear components. Frequently-targeted antigens include nucleosomes (the elementary unit of chroma
Apoptosis, 2010
Systemic lupus erythematosus (SLE) is a complex disease resulting from inflammatory responses of the immune system against several autoantigens. Inflammation is conditioned by the continuous presence of autoantibodies and leaked autoantigens, e.g. from not properly cleared dying and dead cells. Various soluble molecules and biophysical properties of the surface of apoptotic cells play significant roles in the appropriate recognition and further processing of dying and dead cells. We exemplarily discuss how Milk fat globule epidermal growth factor 8 (MFG-E8), biophysical membrane alterations, High mobility group box 1 (HMGB1), C-reactive protein (CRP), and anti-nuclear autoantibodies may contribute to the etiopathogenesis of the disease. Up to date knowledge about these key elements may provide new insights that lead to the development of new treatment strategies of the disease.
Clinical Immunology, 2014
Antibodies to DNA (anti-DNA) are the serological hallmark of systemic lupus erythematosus and markers of underlying immune system disturbances. These antibodies bind to both single-stranded and double-stranded DNA, mediating pathogenesis by forming immune complexes. As shown recently, DNA in blood exists in both free and particulate forms, with DNA representing an important component of microparticles. Microparticles are membrane-bound vesicles containing nuclear molecules, released by membrane blebbing during cell death and activation. A panel of monoclonal NZB/NZW F1 anti-DNA antibodies was tested for binding to microparticles generated from apoptotic THP-1 and Jurkat cells. These studies showed that only certain anti-DNA antibodies in the panel, specific for double-stranded DNA, bound to microparticles. Binding to particles was reduced by soluble DNA or DNase treatment. Together, these results indicate that particle binding is a feature of only certain anti-DNA antibodies, reflecting immunochemical properties of the antibodies and the nature of the exposed DNA antigens.